Jsa Jis K 0101 1998 PDF

ICs: 13.060.
25
JIS JAPANESE
INDUSTRIAL
STANDARD
Testing methods for industrial water
I l
‘Translation without guarantee
In the event of any doubt arising, the original
Standard in Japanese is to be evidence
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I l
Translated
by
Japanese Standards Association
1-24, Akasaka 4, Minato-ku Tokyo 107-8440 Japan
O JSA, 1998
PROTECTED BY COPYRIGHT
JAPANESE INDUSTRIAL STANDARD
JIS K 0101: 1998
March, 2000
ERRATA
Page 261 line 1 1
Error: 20 ml of sulfuric acid(1+1)

Correct: 20 ml of sulfuric acid(1+ 10)
Remarks: This erratum is for correcting the first edition of this Standard.
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PROTECTED B Y COPYRIGHT
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JAPANESE INDUSTRIAL STANDARD

JIS K0101: 1998
October, 1999
ERRATA(112)
Page 11, line 17
In Notes( 7), replace “--.andsuspended matters’’ by “*--or

suspended matters”.
Page 17
In 8.2 ( 4 )( c ), V is changed as follows:
Error V :sample used for dilution (mi)

Correct V :sample used for the test (mi)
Page 32, line 4

The numerical value is replaced as follows:
Error x= 0,310 o
Correct x = 0.310 1
Page 59, line 9

Sentences in 16.1( 2 )( a ) are changed as follows:
Error -.*andwash sufficiently with water by suction. Thereafter, place this fiìter
medium(’) on a watch... .
Correct **.andwash sufficiently with water by suction(’). Thereafter, place this filter
medium on a watch-.. .
Page 68, line 19

In 18 ( 3 )( c), replace “heat”by “boil”.
Page 94, line 6

Error phydrazinobenzene diazonium salt
Correct psulfobenzene diazonium salt
Remarks: This erratum is for correcting the first edit,ion of this Standard.
JAPANESE INDUSTRIAL, STANDARD
JIS K0101:1998
October, 1999
ERRATA(BI2)
Page 254, line 3

42.1( 3 )( a ) is changed as follows:
Error Put 10 ml of sample (containing 2 to 50 pg as SO:-) in a precipitation tube for

a centrifuge, and keep it at 20 to 30 OC('>.Add 4 ml-a.
Correct Add water into appropriate sample (containing 2 pg to 50 pg as SO4'-) in a
precipitation tube for a centrifuge to make total 10 ml, and keep it a t 20 "C to
30 Add 4 ml-..
O C ( ' ) .
Page 255, line 10

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42.2( 3 )( a ) is changed as follows:
Error Put 10 ml of sample (containing 50 t,o 500 pg as SO,2-) in a precipitation tube

for a centrifuge, and keep it at 20 to 30 "C('). Add 4 ml**-,
Correct Add water into appropriate sample (containing 50 pg to 500 pg as SO:-) in a
precipitation tube for a centrifuge to make total 10 ml, and keep it a t 20 "C to
30 OC('). Add 4 ml-..
Page 268, line 1
Error chloride ion

Correct bromide ion
Page 268, line 4
Error chlorine
Correct bromine
Page 453
In clause 3, replace Informative referenceVI
'I " by " Informative reference IV".
Remarks: This erratum is for correcting the first edition of this Standard.
K O101 : 1998
Contents
Page
1 Scope .................................................................................................................... 1
2 Common items ................................................................................................... 1
3 Sample ................................................................................................................. 5
3.1 Sampling. sample container. water sampler and water-sampling
operation .......................................................................................................... 5
3.2 Handling of sample ........................................................................................ 5
3.3 Preservation treatment of sample ............................................................... 5
4 Pretreatment of sample .................................................................................... 8

4.1 Boiling with hydrochloric acid or nitric acid ............................................. 8
4.2 Decomposition with hydrochloric acid o r nitric acid ................................ 8
4.3 Decomposition with nitric acid and perchloric acid ................................. 9
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4.4 Decomposition with nitric acid and sulfuric acid ..................................... 10

4.5 Pretreatment in flame atomic absorption method, electric heating
atomic absorption method, ICP atomic emission spectrometry or
ICP mass spectrometry .................................................................................. 10
5 Marking of results ............................................................................................. 11
6 Temperature ....................................................................................................... 12
6.1 Atmospheric temperature .............................................................................. 12
6.2 Water temperature ......................................................................................... 12
7 Appearance ......................................................................................................... 14
8 Odour and threshold odour number (TON) .................................................. 15

8.1 Odour ................................................................................................................ 15
8.2 Threshold odour number (TON) ................................................................... 16
9 Turbidity ............................................................................................................. 18
9.1 Visual-sensation turbidity ............................................................................. 18
9.2 Transmitted-light turbidity .......................................................................... 20
9.3 Scattered-light turbidity ................................................................................ 21
9.4 Integrating-sphere turbidity ......................................................................... 23
10 Colour .................................................................................................................. 26
10.1 Chromaticity according to platinundcobalt ................................................ 26
10.2 Marking . stimulus value Y and chromaticity coordinates x....y
. by .......... 27
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11 pH ......................................................................................................................... 35
11.1 Glass electrode method .................................................................................. 35
12 Electric conductivity ......................................................................................... 42
13 Acid consumption .............................................................................................. 47

13.1 Acid consumption (pH 4.8) ............................................................................ 47
13.2 Acid consumption (pH 8.3) ............................................................................ 48
14 Alkali consumption ........................................................................................... 50

14.1 Alkali consumption (pH 8.3) ......................................................................... 50
14.2 Alkali consumption (pH 4.8) ......................................................................... 52
14.3 Alkali consumption (free acid) ..................................................................... 53
15 Hardness ............................................................................................................. 55
15.1 Total hardness ................................................................................................. 55
15.1.1 Chelatometric titration method ................................................................. 55
15.1.2 Flame atomic absorption method .............................................................. 55
15.1.3 ICP atomic emission spectrometry ........................................................... 56
15.2 Calcium hardness ........................................................................................... 56
15.3 Magnesium hardness ..................................................................................... 57
16 Suspended matters and evaporation residues ............................................. 58
16.1 Suspended matter ........................................................................................... 58
16.2 Total evaporation residue ............................................................................. 60
16.3 Soluble evaporation residue ......................................................................... 61
16.4 Ignition residue ............................................................................................... 61
16.4.1 Ignition residue of suspended matter ...................................................... 61
16.4.2 Ignition residue of total evaporation residue ......................................... 62
16.4.3 Ignition residue of soluble evaporation residue ..................................... 62
16.5 Ignition loss ..................................................................................................... 62
17 ............
Oxygen demand by potassium permanganate a t 100 "C (CODM~) 63
18 Oxygen demand by potassium dichromate (CODcr) .................................... 67
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19 Biochemical oxygen demand (BOD) ............................................................... 70
20 Organic carbon (TOC) ....................................................................................... 79

20.1 Combustion oxidation-infrared system TOC analysis method .............. 79
20.2 Combustion oxidation-infrared system TOC automatic measuring
method .............................................................................................................. 82
21 Total oxygen demand (TOD) ........................................................................... 85
22 Phenols and p-cresols ....................................................................................... 88

22.1 Phenols ............................................................................................................. 88
22.1.1 Pretreatment ................................................................................................. 88
22.1.2 4-amino antipyrine absorptiometry .......................................................... 89
22.2 p-Cresols ........................................................................................................... 94
22.2.1 p-Hydrazinobenzene sulfonic acid absorptiometry ................................ 94
23 Surface active agents ........................................................................................ 97

23.1 Anionic surface active agents ....................................................................... 97
23.1.1 Methylene Blue absorptiometry ................................................................ 97
23.1.2 Ethyl Violet absorptiometry ....................................................................... 104
23.1.3 Solvent extract-flame atomic absorption method .................................. 106
23.2 Nonionic surface active agent ...................................................................... 108
23.2.1 Tetrathiocyanatocobaltate (II) absorptiometry ....................................... 108
24 Dissolved oxygen ............................................................................................... 112

24.1 Winkler method .............................................................................................. 112
24.2 Winkler-sodium azide modification ............................................................. 116
24.3 ......................................................................................
Miller’s modification 119
24.4 Membrane electrodes method ....................................................................... 121
25 Total carbonate .................................................................................................. 125

25.1 Strontium chloride-hydrochloric acid titration method .......................... 125
25.2 Infrared analytical method ........................................................................... 130
26 Hexane extracts ................................................................................................. 132

26.1 Sampling .......................................................................................................... 132
26.2 Extraction method .......................................................................................... 133
27 Missing number ................................................................................................. 137
28 Residual chlorine ............................................................................................... 138

28.1 o-Tolidine colorimetric method ..................................................................... 138
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28.2 Diethyl-p-phenylenediamine (DPD) colorimetric method ....................... 141

28.3 .........................................................................................................
Iodometry 143
28.4 DPD-ammonium iron (II) sulfate titration ............................................... 144
29 Required amount of chlorine ........................................................................... 152
30 Hydroxide ion (OH-) .......................................................................................... 154
31 Fluorine compounds .......................................................................................... 155

31.1 Lanthanum-alizarin complexon absorptiometry ....................................... 155
31.2 Ion selective electrode method ..................................................................... 159
Chloride ion (Cl-) ...............................................................................................
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32 163
32.1 Mercury (II) thiocyanate absorptiometry ................................................... 163
32.2 Mercury (II) nitrate titrimetric method ..................................................... 164
32.3 Silver nitrate titrimetric method ................................................................. 166
32.5 Ion chromatography ....................................................................................... 170
33 Iodide ion (I-) ..................................................................................................... 174

33.1 Iodine extraction absorptiometry ................................................................. 174
33.2 Iodine titrimetric method .............................................................................. 175
34 Bromide ion (Br-) ............................................................................................... 179

34.1 Iodine titrimetric method .............................................................................. 179
35 Cyanide compounds ........................................................................................... 183

35.1 Pretreatment ................................................................................................... 183
35.1.1 Cyanide .......................................................................................................... 183
35.1.1.1 Aeration method (hydrogen cyanide t o be generated a t pH 5.0) ..... 183
35.1.1.2 Method of distillation by heating (hydrogen cyanide to be
generated under existence of zinc acetate a t pH 5 . 5 ) ....................... 185
35.1.2 Total cyanogen (hydrogen cyanide t o be generated a t p H 2 o r
less) ................................................................................................................ 188
35.2 4-Pyridinecarboxylic acid-pyrazolone absorptiometry ............................. 190
36 Ammonium ion (NH4+) ...................................................................................... 196

36.1 Pretreatment ................................................................................................... 196
36.1.1 Aggregate precipitation method ................................................................ 196
36.1.2 Distillation method ...................................................................................... 197
K O101 : 1998
36.2 Indophenol Blue absorptiometry .................................................................. 199

36.3 Acid-basic titrimetric method ....................................................................... 201
36.4 I o n selective electrode method ..................................................................... 203
37 Nitrite ion (Noz-) and nitrate ion (NOS-) ...................................................... 210

37.1 Nitrite ion (NOZ-)............................................................................................ 210
37.1.1 ..............................................
Naphthylethylenediamine absorptiometry 210
37.1.2 Ion chromatography .................................................................................... 212
37.2 Nitrate ion (Nos-) ........................................................................................... 213
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37.2.1 Reducing distillation-Indophenol Blue absorptiometry ........................ 213

37.2.2 Reducing distillation-acid-base titrimetric method .............................. 216
37.2.3 Copper and cadmium column reduction-naphthylethylenediamine
absorptiometry .............................................................................................. 217
37.2.4 Brucine absorptiometry .............................................................................. 221
37.2.5 Ion chromatography .................................................................................... 223
38 Organic nitrogen ................................................................................................ 225

38.1 Pretreatment (Kjeldahl method) .................................................................. 225
38.2 Indophenol Blue absorptiometry .................................................................. 226
38.3 Acid-base titrimetric method ........................................................................ 227
39 Total nitrogen .................................................................................................... 229

39.1 Sum total method ........................................................................................... 229
39.2 Ultraviolet absorptiometry ............................................................................ 231
39.3 Hydrazinium sulfate reducing method ....................................................... 234
39.4 Copper and cadmium column reducing method ........................................ 237
39.5 Thermal decomposition method ................................................................... 239
40 ..................................................................................................
Sulfide ion (S2-) 242
40.1 Methylene-blue absorptiometry .................................................................... 242
40.2 Iodometry ......................................................................................................... 244
41 Sulfite ion so^^-) .............................................................................................. 249

41.1 Iodometry ......................................................................................................... 249
.............................................................................................
42 Sulfate ion (S042-) 252
42.1 Barium chromate-diphenylcarbazide absorptiometry .............................. 252
42.2 Barium chromate absorptiometry ................................................................ 254
42.3 Gravimetry ....................................................................................................... 256
K O101 : 1998
43 Phosphorus compound and total phosphorus ............................................... 259

43.1 Phosphate .....................................................................................
ion (P043-) 259
43.1.1 Molybdenum blue (ascorbic acid reduction) absorptiometry ................ 259
43.1.2 Molybdenum blue [tin (II) chloride reduction] absorptiometry ........... 262
43.2 Hydrolytic phosphorus ................................................................................... 264
43.3 Total phosphorus ............................................................................................ 266
43.3.1 Potassium peroxodisulfate decomposition ............................................... 266
43.3.2 Nitric acid-perchloric acid decomposition method ................................ 270
43.3.3 Nitric acid-sulfuric acid decomposition method .................................... 273
44 Silica (SiOs) ........................................................................................................ 275

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44.1 Ionic silica ........................................................................................................ 275

44.1.1 Molybdenum yellow absorptiometry ......................................................... 275
44.1.2 Molybdenum blue absorptiometry ............................................................ 276
44.1.3 Molybdenum blue extraction absorptiometry ......................................... 277
44.2 Dissolved and colloidal silica ....................................................................... 280
44.3 Total silica ....................................................................................................... 280
44.3.1 Fusion by sodium carbonate ...................................................................... 280
44.3.2 Gravimetry .................................................................................................... 281
45 Boron (B) ............................................................................................................. 284

45.1 Methylene blue absorptiometry ................................................................... 284
45.2 Azomethine H absorptiometry ...................................................................... 286
45.3 ICP atomic emission spectrometry .............................................................. 287
46 Arsenic (As) ........................................................................................................ 289

46.1 Silver diethyldithiocarbamate absorptiometry .......................................... 289
46.2 Hydride-generation atomic absorption method ......................................... 293
46.3 Hydride-generation ICP atomic emission spectrometry .......................... 297
47 Sodium (Na) ........................................................................................................ 300

47.1 Flame emission photometry .......................................................................... 300
47.2 Flame atomic absorption method ................................................................ 301
47.3 Ion-selective electrode method ..................................................................... 302
48 Potassium (K) ..................................................................................................... 306

48.1 Flame emission photometry .......................................................................... 306
K O101 : 1998
49 Calcium (Ca) ....................................................................................................... 309

49.1 Chelatometric titration .................................................................................. 309
50 Magnesium (Mg) ................................................................................................ 314

50.1 Chelatometric titration .................................................................................. 314
51 Copper (Cu) ........................................................................................................ 319

51.1 Diethyldithiocarbamic acid absorptiometry ............................................... 319
51.3 Electric heating atomic absorption method ............................................... 323
51.5 ICP mass spectrometry .................................................................................. 327
52 Zinc (Zn) .............................................................................................................. 331

53 Cadmium (Cd) .................................................................................................... 338

54 Nickel (Ni) .......................................................................................................... 346

54.1 Dimethylglyoxime absorptiometry ............................................................... 346
55 Tin (Sn) ............................................................................................................... 352

55.1 Phenylfluorone absorptiometry .................................................................... 352
55.2 Quercetin absorptiometry ............................................................................. 354
56 Lead (Pb) ............................................................................................................. 358

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57 Mercury (Hg) ...................................................................................................... 365

57.1 Atomic absorption spectrometry by reduction-vaporization ................... 365
57.2 Atomic absorption spectrometry by heating-vaporization ....................... 370
58 Manganese (Mn) ................................................................................................ 373

58.1 Periodic acid absorptiometry ........................................................................ 373
59 Aluminum (Al) ................................................................................................... 382

59.1 Quinolinol absorptiometry ............................................................................ 382
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60 Iron (Fe) .............................................................................................................. 391

60.1 Phenanthroline absorptiometry ................................................................... 391
61 Chromium (Cr) ................................................................................................... 399

61.1 Total chromium ............................................................................................... 399
61.1.1 Diphenylcarbazide absorptiometry ........................................................... 399
61.1.3 Electric heating atomic absorption method ............................................ 403
61.1.5 ICP mass spectrometry ............................................................................... 406
61.2 Chromium (VI) [Cr (VI)] ............................................................................... 408
61.2.1 Diphenylcarbazide absorptiometry ........................................................... 408
61.2.3 Electric heating atomic absorption method ............................................ 410
61.2.5 ICP mass spectrometry ............................................................................... 411
K O101 : 1998
62 Vanadium (V) ..................................................................................................... 413

62.1 N-benzoyl-N-phenylhydroxylamineabsorptiometry .................................. 413
63 Bacterial test ...................................................................................................... 419

63.1 Sampling and collection of bacteria ............................................................ 419
63.2 General bacteria ............................................................................................. 420
63.3 Heterotrophic bacteria ................................................................................... 423
63.4 Escherichia coli group .................................................................................... 426
63.5 Fecal Escherichia coli group ......................................................................... 428
64 Biological test ..................................................................................................... 431

64.1 Biological test .................................................................................................. 431
64.2 Bacteria ............................................................................................................ 433
64.3 Algae ................................................................................................................. 438
64.4 Animal .............................................................................................................. 439
Annex (informative) Supplement ........................................................................... 440

I Transparentness ................................................................................................ 440
II Oxygen demand by alkaline potassium permanganate (CODOH).............. 443
III Cation surface-active agent ............................................................................. 445
IV Ion-selective electrode method for iodide ion ............................................... 448
V Ion-selective electrode method for bromide ion ........................................... 451
VI Ion-selective electrode method for nitrate ion ............................................. 454
.............................................
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VII Ion-selective electrode method for sulfide ion 457

VI11 Barium-sulfate turbidimetry for sulfate ion ................................................. 460
Attached Table 1 Normative references ............................................................... 461
JAPANESE INDUSTRIAL STANDARD JIS K O101 : 1998
1 Scope This Japanese Industrial Standard specifies the testing methods for in-
dustrial water.
Remarks : Normative references to this Standard are shown in Attached Table 1.
2 Common items The common items shall be as follows:
General rule The general items common to the chemical analysis shall be in
accordance with JIS K 0050.
Definitions For the purposes of this Standard, the definitions in JIS K 0102,
JIS K 0211 or JIS K 0215 apply.
In addition, the inductively coupled plasma mass spectrometry is hereafter
referred to as ?ICP mass spectrometry?.
Gas chromatography The general items common t o the gas chromatography

shall be in accordance with JIS K 0114.
Absorptiometry The general items common to the absorptiometry shall be

in accordance with JIS K 0115.
Inductively coupled plasma atomic emission spectrometry The general

items common to inductively coupled plasma atomic emission spectrometry (here-
after referred to as ?ICP atomic emission Spectrometry?) shall be in accordance
with JIS K 0116.
Infrared spectrophotometry The general items common to the infrared spec-

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trophotometry shall be in accordance with JIS K 0117.
Atomic absorption method There are a flame absorption method, an elec-

tric heating system atomic absorption method (hereafter referred t o as ?electric
heating atomic absorption method?) and other atomic absorption methods. The
general items common thereto shall be in accordance with JIS K 0121.
Ion-selective electrode method The general items common to the ion-se-

lective electrode method shall be in accordance with JIS K 0122.
Ion chromatography The general items common to the i o n chromatography

shall be in accordance with JIS K 0127.
(10) Determination range The determination ranges indicated in respective test

methods are expressed by the mass (mg, pg o r ng) in the final solution. How-
ever, in the atomic absorption method, flame emission photometry, ICP atomic
emission spectrometry, ICP mass spectrometry, ion chromatography, ion-selective
electrode method, and testing methods of total organic carbon (TOC), total oxygen
demand (TOD), dissolved oxygen and residual chlorine, the determination range
is expressed by the concentration (mg/Z o r pg/Z) of the final solution.
2
K O101 : 1998
(11) Repeatability The repeatability for the standard solution shall be indicated
by coefficient of variation (%) (1) obtained by repetitive tests within the deter-
mination range of respective testing methods.
0
Note (1) Coefficient of variation (%) = -x 100
2
where, o : standard deviation
-
x : average
(12) Water The water t o be used in this Standard shall be water A l t o A4 speci-
fied in JIS K 0557, however, where it is specified in relevant items, the water
shall be in accordance therewith.
Dissolved-oxygen-free water Transfer water A2 or A3 specified in JIS
K 0557 into a flask, boil for about 5 min to remove the dissolved oxygen,
then connect the gas washing bottle containing alkaline pyrogallol solu-
tion(2) as in Fig. 2.1, and allow t o cool while shielding from oxygen in the
atmosphere. Otherwise, allow to remove dissolved oxygen by blowing high
purity nitrogen grade 2 specified in JIS K 1107 for approx. 15 min instead
of boiling.
A: 1O00 ml Flat bottom flask

B: 250 ml Gas-washing bottle
C: Rubber stopper
D: Rubber tube
E: Alkaline pyrogallol solution
Fig. 2.1 An example for cooling and preservation

of dissolved-oxygen-free water
Carbonic acid-free water Transfer water A2 or A3 specified in JIS K
0557 into a flask. After removing dissolved gas and carbon dioxide by boiling
for approx. 5 min, use the same apparatus as that in Fig. 2.1, put potas-
sium hydroxide solution (250 gll) into a gas washing bottle, shield from carbon
dioxide in the atmosphere, and allow t o cool.
Note (2) Dissolve 6 g of pyrogallol (l,S73-benzenetriol)specified in JIS K
8780 in 50 ml of water, and reserve in a coloured bottle. Sepa-
rately, dissolve 3 0 g of potassium hydroxide specified in JIS K
8574 in 50 ml of water. Mix both liquids in use. 1ml of this solution
absorbs approx. 12 ml of oxygen (approx. 17 mg).
(13) Reagents
(a) I n the case where a n item is designated, JIS marked reagent of the high-
est quality shall be used. Where JIS marked reagent does not exist, re-
agents without impairing the test shall be used(3).
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K 0101 : 1998
For standardization of titrants, reference materials for volumetric analysis

specified in JIS K 8005 shall be used.
The concentration of solution of reagents shall be indicated g/1 or mg/Z for
mass concentration and mol/E or mmolll for mol concentration unless oth-
erwise specified.
For compounds, masses as anhydride are used.
The concentration of standard solution shall be indicated mass in l m l
(mg/ml or pg/ml) except ion-selective electrode method and flame emission
photometry.
The concentration indicated in parentheses after the name of the solution
means the approximate concentration except standard solutions. For ex-
ample, the sodium hydroxide solution (O. 1mol/Z) means the concentration
of sodium hydroxide solution of approx. 0.1 mol/Z.
The concentration indicated before the name of solution means the cor-
rect concentration. In general, it shall be indicated with numerical value
without fraction, and the factor be separately obtained.
The water t o be used for preparation of reagents shall be the water speci-
fied in (12),however, where it is specified in respective items, the water
shall be in accordance with specifications of the item.
When preparing the standard solution of low concentration by diluting the
standard solution, use a 10ml or over transfer pipette unless otherwise
specified.
The names of reagents shall be, as a rule, conformed to the compound no-
menclature determined by The Chemical Society of Japan in accordance
with the inorganic chemical nomenclature and organic chemical nomencla-
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ture of International Union of Pure and Applied Chemistry (IUPAC) and
the names of JIS reagents.
For handling of reagents, waste solution, etc. sufficient cares shall be taken
following laws and regulations relating thereto.
Note (3) For the test using the reagent of very small quantity in such de-
termination method as electric heating atomic absorption method,
ICP mass spectrometry, etc., specially high pure reagents shall
be used.
(14) Appliances Glassware, porcelain crucible, porcelain evaporating dish and filter
paper used in this Standard are as follows.
(a) Glassware specified in JIS R 3503 and JIS R 3505 shall be used. If spe-
cial appliances are required, they are exemplified o r explained in respec-
tive items.
For heating procedures, borosilicate glass-1 specified in JIS R 3503 shall
be used.
Desiccant used in a desiccator shall be silica gel(4) unless otherwise speci-
fied.
4
K O101 : 1998
(b) Porcelain crucible and porcelain evaporating dish specified in JIS R 1301
and JIS R 1302 shall be used.
(c) Filter paper for quantitative analysis specified in JIS P 3801 shall be used.
The type of filter papers shall be specified in respective items.
Note (4) Packaging silica gel desiccant type A grade 1 specified in JIS Z
0701 shall be used.
Remarks : In the case where silica, boron, sodium, potassium, arsenic, zinc,
etc. are tested, cares shall be taken sufficiently on the elution
of these components from borosilicate.
(15) Measurement of absorbance (Absorptiometry) If not specified on absorption

cell, the absorption cell of 10 mm in optical path length shall be used.
( 16) Working curve [absorptiometry,atomic absorption method, flame emis-

sion photometry, ICP atomic emission spectrometry, ICP mass spec-
trometry, ion chromatography, ion-selective electrode method, total
organic carbon (TOC), and total oxygen demand (TOD)] When a work-
ing curve is prepared, divide the determination range given by a test method
into 4 to 6 stages, and take standard solutions so as t o conform thereto. Pre-
pare the working curve within the determination range.
In the tests for atomic absorption method, flame emission photometry, ICP
atomic emission spectrometry, ICP mass spectrometry, ion chromatography,
ion-selective electrode method, total organic carbon (TOC), and total oxygen
demand (TOD),use the working curve newly prepared in testing. When the
same item is continuously tested on many samples, use appropriately standard
solution halfway in testing, and confirm an indication value.
In absorptiometry, allow the preliminarily prepared working curve to be used.
(17) Note, remarks, figure, table, and formula For the note, remarks, figure,
table, and formula, serial number is annexed t o each clause.
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K 0101 : 1998
3 Sample
3.1 Sampling, sample container, water sampler and water-samplingoperation

The sample means the water sampled for carrying out the test. Sampling, sample
container, water sampler and water-sampling operation shall be in accordance with
JIS K 0094.
3.2 Handling of sample Test total quantity contained in a sample unless other-
wise specified. Therefore, where suspended matters are contained in the sample,
sufficiently mix by shaking to make the sample homogeneous. Thereafter, take the
sample to be used for the test. However, for the test of an anion, use the filtrated
sample unless otherwise specified. Where the total quantity is obtained, specify the
handling in each item.
Besides, where only those in dissolved state are tested, immediately after taking
the sample, filter with filter paper of grade 5C(1), discard approx. 50 ml of the ini-
tial filtrate, and take the filtrate thereafter as the sample.
Note (1) Filter paper grade 6 or the filter material with 1 pm o r under in pore
diameter may be used.
3.3 Preservation treatment of sample Perform a test immediately after sam-

pling unless otherwise specified. I n the case where the test can not be performed
immediately and the sample is preserved, carry out the operation as follows in ac-
cordance with 7 (preservation treatment of sample) of JIS K 0094, and test as soon
as possible. In the case where the sample is preserved in a cold place, do not freeze.
(1) Reagents The following reagents shall be used.
Hydrochloric acid As specified in JIS K 8180.

Hydrochloric acid (for analysis of arsenic) As specified in JIS K 8180.
Nitric acid As specified in JIS K 8541.
Sulfuric acid As specified in JIS K 8951.
Phosphoric acid As specified in JIS K 9005.
L(+)-ascorbicacid As specified in JIS K 9502.
Sodium hydroxide solution (200 gll) Dissolve 20 g of sodium hydrox-
ide specified in JIS K 8576 in water to make 100 ml.
Basic zinc carbonate suspension Dissolve 20 g of zinc sulfate hepta-
hydrate specified in JIS K 8953 in water to be mixed with sodium carbon-
ate solution (100 gll) of the same volume as that. Prepare a t service.
Copper (II) sulfate pentahydrate As Specified in JIS K 8983.
Chloroform As specified in JIS K 8322.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
6
K O101 : 1998
(2) Preservation treatment Perform the preservation treatment as follows.
Preserve samples to be used for the tests of oxygen demand by potassium

permanganate ( C O D d a t 100 O C , oxygen demand by potassium dichromate
(CODcr), biochemical oxygen demand (BOD), total organic carbon (TOC),
total oxygen demand (TOD), and surface active agent in a dark place at O
to 10°C.
Add hydrochloric acid o r sulfuric acid to the samples used for the tests of
ammonium ion, organic nitrogen, and total nitrogen, regulate pH a t 2 to 3,
and preserve in a dark place at O t o 10 OC. For a short number of days,
allow the sample to be preserved in a dark place a t O to 10 "C as it is with-
out preservation treatment.
Add about 5 m l of chloroform t o the sample t o be used for the test of ni-
trite ion and nitrate ion per l E of the sample, and preserve in a dark place
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
at O to 10 "C. For a short number of days, allow the sample as it is t o be

preserved in a dark place at O to 10 "C without preservation treatment.
Add sodium hydroxide solution (200glZ) to the sample to be used for the
tests of iodide ion, and bromide ion to make pH approx. 10 and preserve
(allow 2 t o 4 grains of sodium hydroxide per 1E of the sample to be added).
Add sodium hydroxide solution (2OOglZ) t o the sample to be used for the
test of cyanide compound and sulfide ion t o make pH approx. 12, and pre-
serve (allow 4 to 6 grains of sodium hydroxide per 1E of the sample to be
added). In the case where oxidizing matters such as residual chlorine or
the like coexist with the sample t o be used for the cyanide compound test,
after reducing by adding L(+)-ascorbicacid, make pH approx. 12.
Further, for sulfide ion, take a sample into a dissolved oxygen measur-
ing bottle, add basic zinc carbonate suspension at a rate of approx. 2 ml
per 100 ml of the sample to fix as zinc sulfide, and allow t o be preserved.
Add phosphoric acid t o the sample to be used for the phenols test t o make
pH approx. 4, add 1g of copper (II) sulfate pentahydrate per 1 Z of the sample,
mix by shaking, and preserve in a dark place at O to 10 "C.
Perform preservation treatment for the sample to be used for the tests of
phosphide compound and total phosphorus as follows. Add chloroform at a
rate of approx. 5 ml per 1 I of a sample under a state as it is, and preserve
in a dark place at O t o 10°C. In that case, for a short number of days,
allow t o be preserved in a dark place at O t o 10 "C under a state as it is
without preservation treatment. For the sample t o be used for the test of
dissolved phosphide compound, after filtration in accordance with 3.2, add
chloroform at a rate of approx. 5 ml per 1I of the sample, and preserve in
a dark place a t O t o 10 "C. In the case, for a short number of days, allow
t o be preserved in a dark place at O to 10 OC under a state as it is without
preservation treatment.
The sample t o be used for total phosphorus may be preserved making
the pH value about 2 by adding sulfuric acid or nitric acid,
7
K O101 : 1998
(h) Add nitric acid to the samples t o be used for the tests of metallic elements
such as copper, zinc, lead, cadmium, manganese, iron, aluminium, nickel,
cobalt, arsenic, tin, total chromium, mercury, vanadium, etc. t o make pH
approx. 1, and preserve.
In the case where treatments with sulfuric acid and nitric acid, or nitric
acid and potassium permanganate are not performed in testing for the sample
to be used for testing arsenic and organic matters, a great quantity of ni-
trates, nitrites, etc. do not contained, add hydrochloric acid (for analysis of
arsenic) to make pH approx. 1, and preserve.
Preserve the sample to be used for the test of chromium (VI) in a dark
place at O t o 10 O C under a state as it is.
For the sample to be used for the test of dissolved state metal elements,
after filtering a sample in accordance with 3.2, add nitric acid to make pH
approx. 1, and preserve.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
8
K O 1 0 1 :1998
4 Pretreatment of sample Though the pretreatment of a sample is specified in

each test item, since the pretreatment operations in the test of metal element are
almost common independently of the kind of metal elements, the operations are specified
collectively hereafter. However, the pretreatments for the tests of sodium, potas-
sium, calcium, magnesium, arsenic, chromium (VI), mercury, etc. among metal ele-
ments are specified in respective test items.
The pretreatments in the test of metal elements have the purpose mainly to de-
compose the coexisting organic substance, suspended matters and metal complex.
Though the method of heating after adding various acids t o the sample is used, a
suitable method is selected according t o the state of the sample and the kind of test.
4.1 Boiling with hydrochloric acid or nitric acid This method applies t o the
sample containing an extremely small amount of organic substances and suspended
matters.
(a) Hydrochloric acid As specified in JIS K 8180.
(b} Nitric acid As specified in JIS K 8541.
(2) Operation The operation shall be carried out as follows.
(a) Add hydrochloric acid o r nitric acid a t a rate of 5 m l per 1OOml of the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
sample (1).
(b) Boil for approx. 10min by heating.
(c) After standing t o cool, add water t o make the specified amount, as required.
Note (1) In the case of testing dissolved metal elements, the sample fil-
tered in accordance with 3.2 shall be used.
4.2 Decomposition with hydrochloric acid or nitric acid This method ap-
plies t o the sample containing little organic substance and hydroxide, oxide, sulfide,
phosphate, etc., as suspension.

(b) Nitric acid As specified in JIS K 8541.
(a) After mixing the sample(2) sufficiently by shaking, immediately take the
suitable amount of sample in a beaker and add hydrochloric acid or nitric
acid at a rate of 5 ml per 100 ml of the sample.
(b) Heat to concentrate the sample to approx. 15 ml.
(c) When the insoluble matters remain, filter the sample with filter paper of
grade 5B, and then wash with water sufficiently.
(d) After standing to cool, transfer the filtrate and washings into a suitable
capacity of volumetric flask, and dilute t o the marked line with water.
9
K O 1 0 1 : 1998
Note (2) For testing the dissolved metal elements, the sample filtered in
accordance with 3.2 shall be used, and the method of 4.1 be used.
Remarks : For the sample to which decomposition with mixed acid of hy-
drochloric acid and nitric acid is favorable, after performing the
operation to (b), stand t o cool t o room temperature. When
hydrochloric acid is used in (a), add 5 ml of nitric acid, when
nitric acid is used, add 5 ml of hydrochloric acid, cover with a
watch glass, and heat again. When violent reaction ends, re-
move the watch glass, expel nitrogen oxide by heating further,
and concentrate to approx. 5 ml. In the case where acid is in
short supply in this operation, add an appropriate quantity of
hydrochloric acid and nitric acid, heat by the same operation,
and dissolve. When insoluble matters remain, add 15 ml of warm
water, and carry out the operations of ( c ) and (d).
4.3 Decomposition with nitric acid and perchloric acid This method applies
t o the sample containing organic matters that are difficult t o be oxidized.
(a) Perchloric acid As specified in JIS K 8223.
(b) Nitric acid As specified in JIS K 8541.
Mix the sample(2) by shaking thoroughly, immediately take a proper amount
into a beaker o r porcelain evaporating dish.
Add 5 t o 10 ml of nitric acid, heat it on a heating plate gently t o concen-
trate up to about 10 ml(3), and stand to cool.
Add 5 ml of nitric acid and then add 10 ml of perchloric acid(4) little by
little. Continue heating. When the white fume of perchloric acid begins t o
generate, cover the container with a watch glass, and decompose organic
substances while keeping the conditions in which the perchloric acid flows
down along the wall of the container.
When organic substances remain not composed, further add 5 ml of nitric
acid and repeat the operation specified in ( c ) until the organic substances
decompose.
After standing to cool, dilute the solution with water to approx. 50 ml. When
insoluble matters remain, filter the solution with filter paper of grade 5B,
then wash with water. Transfer the filtrate and washings into a proper
capacity of volumetric flask, and add water up to the marked line.
Notes (3) The sample may be transferred i n t o a Kjeldahl flask t o be de-
composed.
(4) The decomposing procedure by heating using perchloric acid may
be dangerous t o explode in some kinds of samples, therefore pay
attention t o the following.
i) For oxidizable organic substances, add nitric acid of (b)be-
fore adding perchloric acid, and decompose sufficiently by
heating procedure.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
10
K 0101 : 1998
ii) When perchloric acid is added, the concentrated liquid shall

be cooled beforehand without fail.
iii) The decomposition by heating shall be carried out under the
coexist state of perchloric acid with nitric acid without fail.
iv) The concentrated solution shall be not made dryness.
4.4 Decomposition with nitric acid and sulfuric acid This method is appli-
cable (5) for various samples.
(a) Nitric acid As specified in JIS K 8541.
(b) Sulfuric acid (l+l) Take one volume of water into a beaker, cool it and add
gradually one volume of sulfuric acid specified in JIS K 8951 while stirring.
Mix the sample(2) thoroughly by shaking, immediately take its proper amount
into a beaker or porcelain evaporating dish and add 5 to 10 ml of nitric acid.
Heat to approx. 10 ml(3) of solution, then add again 5 ml of nitric acid and
10 ml of sulfuric acid (l+l), and heat until the white fume of sulfuric acid
generates t o decompose organic substances.
When the decomposition of organic substances is difficult, add further 10 ml
of nitric acid, and repeat the operation specified in (b) t o decompose the
organic substances.
After standing to cool, dilute the solution with water to approx. 50 ml. If
insoluble substances ( 6 ) remain, filter with filter paper of grade 5B, wash
with water, then transfer the filtrate and washings into a proper capacity
of volumetric flask, and add water up t o the marked line.
Notes (5) In the case of applying the flame atomic absorption method in
which the water solution is sprayed as it is, this method is not
desirable.
(6) Where lead is contained and precipitates are generated, carry
out the operation specified in 4.3 or the following operation:
Carry out the operation specified in (b) to evaporate the so-
lution to almost dryness, add approx. 30 ml of water and 15 ml
of hydrochloric acid, and heat t o dissolve. Where insoluble mat-
ters exist, filter with filter paper of grade 5B, then wash with
warm hydrochloric acid (1+10).After standing t o cool, transfer
the filtrate and washings into a proper capacity of volumetric
flask, and add water up to the marked line.
4.5 Pretreatment in flame atomic absorption method, electric heating atomic

absorption method, ICP atomic emission spectrometry or ICP mass spectro-
metry The pretreatment shall be carried out by selecting the optimum method of
the methods given in 4.1 to 4.4 taking into consideration the quantity of organic sub-
stances and suspensions contained in the sample, their existing state, the method of
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
11
K O101 : 1998
atomic absorption method, ICP atomic emission spectrometry or ICP mass spectrom-
etry to be applied, sufficiently (7) (8).
In the case where the flame atomic absorption method or ICP atomic emission
spectrometry in which the direct spraying of prepared sample is performed, the sample
shall be of hydrochloric acid or nitric acid(9). In the case of electric heating atomic
absorption method or ICP mass spectrometry, it shall be of nitric acid, and the acid
concentration shall be a suitable concentration (10).
Notes (7) The pretreatment in the case where the solvent extraction method
is applied prior to the flame atomic absorption method or ICP atomic
emission spectrometry, unless otherwise designated, shall be as shown
in the text, and the interfering organic substances and other mate-
rials which are likely to interfere shall be thoroughly decomposed.
In the case where the flame atomic absorption method or ICP atomic
emission spectrometry is carried out by spraying the sample as it
is, the following pretreatment may be carried out.
In the case of a sample containing an extremely small amount of
organic substances e b 5 u s p e n d e d matters, carry out the operation 'of wq
specified in 4.1, As general method of pretreatment for the sample
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
containing organic substances and suspended matters, apply the speci-

fications of 4.3 or 4.4. In this case, expel the most part of sulfuric
acid and perchloric acid by generating white fume sufficiently.
For ICP mass spectrometry, the blank test value cannot be ignored
in some kinds and concentrations of acid, so that the effect of the kind
and concentration of acid of elements shall be verified in advance.
It should be judged based on the results of the test which is car-
ried out for recovery test by adding a specified amount of objective
component to the sample, which pretreatment method is to be se-
lected.
(8) Refer to Note (3) of clause 2. For highly purified reagents, there are
nitric acid, hydrochloric acid, perchloric acid and sulfuric acid specified
in JIS K 9901, JIS K 9902, JIS K 9904 and JIS K 9905, respec-
tively, and others.
(9) In the case of an ICP atomic emission spectrometry, since when the
sample is of sulfuric acid, the introduction amount of the sample is
little and the sensitivity becomes occasionally worse, application of
4.4 is limited to an inevitable case.
(10) In the cases of a flame atomic absorption method and an electric heat-
ing atomic absorption method, the concentration shall be 0.1 t o
1 mol/l. In the case of an ICP atomic emission spectrometry, and
tin (Sn) is not for object, the concentration shall be 0.1 to 0.5 moVZ.
And tin (Sn) is for object, it shall be 1 t o 1.5mol/Z. In the case of
ICP mass spectrometry, it shall be 0.1 to 0.5 molíl. However, in these
cases, it shall be almost the same concentration as that in the case
where a working curve is prepared.
5 Marking of results In the case where there are two or more test methods, it
shall be described clearly which method is used.
12
K O101 : 1998
6 Temperature Temperature is divided into atmospheric temperature and wa-

ter temperature. Then, both the atmospheric temperature and the water tempera-
ture shall be measured a t the time of sampling.
6.1 Atmospheric temperature The atmospheric temperature shall be measured

according t o the following:
(1) Implement The implement shall be as follows.
(a) Thermometer 50 "C thermometer of solid-stem general purpose liquid-
in-glass thermometer specified in JIS B 7411.
(a) By use of the etched-stem liquid-in-glass thermometer, read out the scale
when the temperature-sensitive liquid has stopped. The measurement shall
be carried out a t sampling site of good ventilation, avoiding direct rays of
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the sun and strong heat radiation from the surroundings, keeping at a position
1.2 t o 1.5 m high above the ground.
Remarks 1 Use the maximum and minimum thermometer (see Fig. 6.1)
for measuring the maximum and minimum temperature dur-
ing the measuring period.
Fig. 6.1 An example of max. and

min. thermometer
6.2 Water temperature The water temperature shall be measured according to

the following:
(a) Thermometer 50 o r 100 "C thermometer of solid-stem general purpose
liquid-in-glass thermometer specified in JIS B 7411.
13
K O101 : 1998

(a) Immersing the etched-stem liquid-in-glass thermometer directly into the
water on site or into the sample(1) immediately after sampling, and while
keeping the temperature-sensitive liquid under the surface of water, read
out the scale when the temperature-sensitive liquid has stopped.
Note (1) In order to avoid influences of the container and the atmospheric
temperature, a great quantity of the sample shall be taken.
Remarks 2 In the case where Pettenkofer water thermometer (see Fig. 6.2)
is used, after changing the sample in a metal cylinder three
times, fill the cylinder with the sample, and read out the scale
when the temperature-sensitive liquid has stopped.
3 For a thermistor thermometer and a metal resistance ther-
mometer, keep the temperature detecting part in water t o be
measured, and read out the scale when the pointer of an indi-
cation part is stabilized at a constant value.
Metal outside
cylinder
Fig. 6.2 Pettenkofer water thermometer
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14
K O101 : 1998
7 Appearance Observe the appearance of the sample immediately after sampling.

(a) Beaker 300 to 500 ml (colourless)
(2) Operation Carry out the operation as follows.
(a) Take the sample immediately after sampling into a beaker, and observe
the following items with the naked eye:
i) Kind of colour and its degree of the whole sample
ii) Kind of colour and its degree of supernatant water
iii) Kind of colour and degree of quantity of floating matters, suspended
matters, etc.
iv) State and degree of oils, tars, etc.
v) Other particular state such as bubbles, odours, etc. of the sample
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
15
K O101 : 1998
8 Odour and threshold odour number (TON) The test of odour shall be clas-
sified into the detection of odour and the threshold odour number (TON)(i).
The odours of water are caused by the effects of increase and extinction of bacteria,
algae, micro-organisms, etc., of mixing-in of municipal sewage, cattle shed drainage,
and industrial waste-water, of elution of inner surface treating substances of a water
storage tank and piping system, and of residual chlorine due t o chlorine treatment.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Because the test of odour is performed by the personal sense of smell, individual
difference is large. The temperature and humidity, taking food and smoking of the
inspector also affect the results.
Note (1) TON is the abbreviation of threshold odour number, which is the dilu-
tion multiple of odour threshold, i.e. the multiple value of dilution when
odour is apparently sensed.
8.1 Odour The odour shall be tested for its kind and degree by warming the sample
t o approx. 40 OC.
(a) Erlenmeyer flask with ground stopper 300 ml
(a) Transfer 200 ml of the sample into a 300 ml Erlenmeyer flask with ground
stopper, stopper lightly, and warm t o approx. 40 OC.
(b) Take off the stopper while shaking the flask, and immediately test for the
existence of odour and its kind and degree.
(c) The odour shall be indicated as shown in Table 8.1, so that the kind and
degree of odour of the sample can be approximately understood.
Table 8.1 Example of classification and kind of odour
Classification Kind of odour

(1) Fragrant odour Odours of melon, violet, garlic, cucumber, aromatics, spices, etc.
(2) Botanical odour Odours of algae, green grass, timber, seaweed, etc.
(3) Earthy odour, Odours of soil, swamp, mould, etc.
mouldy odour
(4) Fishery odour Odours of fish, liver oil, clam, etc.
( 5 ) Chemical odour Odours of phenol, tar, oil, fats and oils, paraffin, chlorine, hydrogen
sulfide, chlorophenol, pharmacy odour, chemicals odour, etc.
(6) Metallic odour Odours of iron, metal, etc.
(7) Putrescent odour Odours of garbage, sewage, pigpen, putrescence, etc.
(8) Unpleasant odour Unpleasant odours such as strong odours of fish, pigpen,
putrescence, etc.
2 The odour should be tested at the time of sampling without

warming, and the results are recorded. (This is called "odour
at the time of cold".)
16
K O101 : 1998
8.2 Threshold odour number (TON) The threshold odour number means the
intensity of odour, and shall be expressed by multiple of dilution "dilution multiple
of odour threshold" when the sample is added into water maintained a t about 40 "C
and the definitely perceptible odour is sensed. To minimize the individual differ-
ence of the sense of smell, the same sample shall be tested by a t least 5 people,
preferably by about 10 people.
(i) Reagent Use the following reagents.
(a) Odour-free water Pass water A3 specified in JIS K 0557 at a rate of
5 ZN-active carbon h) into the apparatus as shown in Fig. 8.1.
Wa
A: Glass bottle ( 5 1 )
B: Rubber stopper
C: Glass fibre
D: Granular active carbon
E: 3 to 5 mm gravel
I _ -. II"^ " - -
Fig. 8.1 Method for preparation of

odour-free water (an example)
(2) Implements The implements shall be as follows.

(b) Long leg burette 50ml
(cl Water bath The water bath with temperature regulator
(3) Preliminary test Carry out the preliminary test as follows.
(a) Transfer 200, 40, 10 and 4 ml of the sample respectively into Erlenmeyer
flasks with ground stopper, and dilute with odour-free water to 200 ml to
make the sample for the preliminary test.
(b) Separately, transfer 200 ml of odour-free water into an Erlenmeyer flask
with ground stopper as reference water.
(c) Warm the sample far the preliminary test and the reference water t o 40 to
50 "C on the water bath, then shake t o mix the reference water, and smell
the odour generated a t the same time of taking-off the stopper.
(d) Thereafter, operate the same as before-mentioned in the order from the
smaller quantity of the sample, then compare the odour of the sample for
the preliminary test with that of the reference water, and obtain the mini-
mum quantity (ml) of the sample perceptible the odour.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
17
K 0101 : 1998
Obtain the volume of the sample t o be used for the test as the .number of
ml shown in longitudinal series according to Table 8.2 from the quantity of
the sample obtained in (d)of (3).
Transfer each volume of the sample obtained in (a)into separate Erlenmeyer

flasks with ground stopper respectively, and add the odour-free water t o
make 200 ml t o take it as the sample for the main test.
Next, operate the same as in (b) t o (d)of (3) t o obtain the number of ml of
the minimum sample perceptible the odour, and calculate the threshold odour
number ( T O N ) from the following formula:
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
200
TON = -
V
where, TON : threshold odour number (TON)
V : sample used for dilution (ml)
Table 8.2 Volume of sample t o b e used for t e s t

Unit: ml
Volume of sample for 200 40 10 4
the preliminary test
Volume of sample to 200 40 10 4.0
be used f o r the test 100 28.5 8.0 2.9
67 20 6.7 2.0
50 13.3 5.0 1.3
40 10 4.0 1.0
Remarks 3 Where the odour of sample is t o o intensive, carry o u t the

preliminary test and the main test after diluting the sample
with water t o ten times.
4 Wash the Erlenmeyer flask with ground stopper to be used
for the test with water beforehand.
5 The test depends on the environmental conditions in most
cases, and therefore carry out the test in a n odour-free,
quiet room.
6 Avoid smoking, tea drinking, meal, etc. just before the test,
and further keep the hands and fingers free from the odour
of soap, lotion, perfume, etc.
Because the sense of smell is weakened by 4 to 5 times
consecutive tests, repose for (approx.) 15 to 30 min.
Obtain the degree of odour ( P O )from the following formula,
if required.
PO=- '
log 2
x logTON = 3.32 x logTON
18
K O101 : 1998
9 Turbidity Turbidity expresses the degree of turbidity of water, and is indicated

by dividing into visual-sensation turbidity, transmitted-light turbidity, scattered-light
turbidity, and integrating-sphere turbidity. Where the turbidity is measured by
comparing with kaolin standard solution, it shall be expressed by using “degree (ka-
olin)” as the unit, and where the turbidity is measured by comparing with formagen
standard solution, it shall be expressed by using “degree (formagen)” as the unit.
The turbidity varies easily, and therefore it is desirable to test immediately after
sampling.
9.1 Visual-sensation turbidity The visual-sensation turbidity shall be obtained

by comparing the turbidity of the sample with the kaolin standard solution with the
naked eye.
Measuring range: 1 to 10 degrees (kaolin)
(a) Water Filter water A3 specified in JIS K 0557 by using filter of
approx. 0.1 ym pore diameter, and take the filtrate after discarding the first
approx. 200 ml.
(b) Refined kaolin Take approx. 10 g of kaolin into a 500 ml beaker, add t o
it 300ml of water and 0.2g of sodium diphosphate +hydrate (sodium
pyrophosphate + hydrate) specified in JIS K 8785 and mix by stirring vig-
orously with a magnetic stirrer for approx. 3 min. Transfer this into 1O00 ml
measuring cylinder with ground stopper, add water up t o the marked line
of 1O00 ml, stopper it, and mix by shaking violently for approx. 1min. After
standing still at room temperature for approx. 1h, discard the solution of
250 ml from the above part by using syphon, and separately take the solu-
tion up to the next 500 ml.
Centrifugalize the separately taken solution at approx. 3 O00 min-1 (the
number of rotations depends on the radius of rotating part of centrifugal
separator), for approx. 20 min or filter to separate kaolin by using filter of
not more than 1pm pore diameter. Heat the filtered kaolin at 105 to 110 “C
for approx. 3 h, and after standing to cool in a desiccator, preserve in a wide-
mouthed bottle.
(c) Kaolin standard solution [i o00 degrees (kaolin)] After dispersing 1.00 g
of refined kaolin into water of proper amount, transfer into a 1O00 ml
volumetric flask, add approx. 800 ml of water and approx. 10 ml of form-
aldehyde solution specified in JIS K 8872, and furthermore add water up
to the marked line.
(d) Kaolin standard solution [lo0 degrees (kaolin)] After mixing by stir-
ring thoroughly the kaolin standard solution [iO00 degrees (kaolin)], im-
mediately take its 100 ml into a 1O00 ml volumetric flask, and add water
up t o the marked line.

(a) Dark box It is convenient when comparing the turbidity with the naked eye
to use a dark box as shown in Fig. 9.1. The measurement by approaching an
electric lamp t o the under window of the dark box becomes easily visible.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
19
K O101 : 1998
(b) Colorimetric tube The flat bottomed colorimetric tube with a ground stop-
per with a marked line of 100 ml at a height of (270+1.5)mm from'the bottom
as shown in Fig. 9.2 having colourless glass. The tubes aligned in height
of marked lines (k1.5 mm) shall be used.
Unit: mm Unit: mm
,
r Approx.
0 21.7
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
-____ ~ __ _I_ -
-______I
_I -
Fig. 9.1 An example of dark box Fig. 9.2 Colorimetric tube

After mixing with shaking the sample thoroughly, take a proper amount (1)
from it into a colorimetric tube, and add water up to the marked line of
100 ml.
Separately, take 1 t o 10 ml of kaolin standard solution [lo0 deg. (kaolin)]
step by step into a colorimetric tube, add water up t o the marked line of
100 ml, and prepare the kaolin standard solution for nephelometry [i to
10 deg. (kaolin)].
After mixing by shaking thoroughly the colorimetric tube containing the
sample and that containing kaolin standard solution for nephelometry, put
them immediately into a dark box, see through from above t o compare
turbidity, and select the kaolin standard solution for nephelometry corre-
sponding t o the sample.
From the corresponding turbidity [deg. (kaolin)] of the kaolin standard
solution for nephelometry, calculate the visual-sensation turbidity of the
sample [deg. (kaolin)] from the following formula:
100
T=T,x-
V
where, T : visual-sensation turbidity [deg. (kaolin)]
20
K O 1 0 1 : 1998
Ts: corresponding turbidity of kaolin standard solu-

tion for nephelometry [deg. (kaolin)]
V : sample (ml)
Note (1) Where the turbidity of the sample is not more than 10 deg. (ka-
olin), take 100 ml as it is.
9.2 Transmitted-light turbidity Measure the intensity of transmitted-light near

660 nm in wavelength passed through the sample, and obtain the transmitted-light
turbidity from the working curve prepared by using kaolin standard solution or
formagen standard solution.
Measuring range: Where the absorption cell is 50 mm, 5 to 50 deg. (kaolin) or
4 t o 80deg. (formagen), and where the absorption cell is
10 mm, 25 to 250 deg. (kaolin) or 20 to 400 deg. (formagen).

--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Water As described in 9.1 (1) (a).

(b) Kaolin standard solution [lo0 deg. (kaolin)] As described in 9.1 (1)
(d).
(c) Formagen standard solution [400 deg. (formagen)] Take 1.00 g of
hydrazinium sulfate (hydrazine sulfate) specified in JIS K 8992, dissolve
in a proper amount of water, transfer into a 100 ml volumetric flask and
add water up to the marked line.
Separately take 10.0 g of hexamethylenetetramine specified in JIS K 8847
t o dissolve in water of a proper amount, transfer it into a 100 ml volumet-
ric flask, and add water up to the marked line.
Take respectively 10 ml of both these solutions in a 200 ml volumetric
flask, and mix by shaking thoroughly. After standing as it is at (25I3) "C
in solution temperature for approx. 24 h, add water up t o the marked line.
(2) Apparatus The apparatus shall be as follows.

(a) Photometer Spectrophotometer or photoelectric photometer
(3.1) In the case of using kaolin standard solution
(a) After mixing by stirring the sample thoroughly, take it into a 50 mm(2) ab-
sorption cell, and measure the intensity of transmitted light near 660 nm(3)
in wavelength by absorbance.
(b) Obtain the transmitted-light turbidity [deg. (kaolin)] of the sample from
the working curve prepared by using kaolin standard solution.
Working curve Take 5 to 50 ml of kaolin standard solution [lo0 deg. (ka-
olin)] step by step into a 100 ml volumetric flask, and add water up to the
marked line t o prepare the kaolin standard solution [5 t o 50 deg. (kaolin)]
for working curve(4). Hereafter, carry out the operation of (a),and pre-
pare the relation curve between the transmitted-light turbidity [deg. (ka-
olin)] and the absorbance of kaolin standard solution for working curve.
21
K O101 : 1998
Notes (2) Where the transmitted-light turbidity of the sample is 25 to 250

[deg. (kaolin)], use a 10mm absorption cell.
(3) Where the sample has a colour (specially where there is absorp-
tion near 660 nm in wavelength), measure the intensity of trans-
mitted-light amount by absorbance by using the filtrate of the
sample filtered by using film filter of 1pm o r under of pore di-
ameter or by using the supernatant precipitated centrifugally [at
approx. 3 O00 min-1 (the number of rotations depends on the ra-
dius of rotating part of centrifugal separator) for approx. 20 min]
as the reference solution.
(4) Where the 10mm absorption cell is used, take 2.5 to 25ml of
kaolin standard solution [i O00 deg. (kaolin)] of 9.1 (1) (c) stepwise,
and prepare the kaolin standard solution [25 t o 250 deg. (kaolin)]
for working curve.
(3.2) In the case of using formagen standard solution
(a) After mixing by shaking the sample thoroughly, carry out the operation(5)
of (3.1)(a).
(b) Obtain the transmitted-light turbidity [deg. (formagen)] of the sample from
the working curve prepared by using formagen standard solution.
Working curve Take 1 t o 20 ml of formagen standard solution [400deg.
(formagen)] step by step into a 100 ml volumetric flask, and add water up
t o the marked line to prepare(6) the formagen standard solution [4 t o 80 deg.
(formagen)] for working curve. Hereafter, carry out the operation of (a),and
prepare the relation curve between the transmitted-light turbidity [deg.
(formagen)]and the absorbance of the formagen standard solution for working
curve.
Notes (5) Where the transmitted-light turbidity of the sample is 20 t o
400 deg. (formagen), use a 10 mm absorption cell.
(6) Where a 10 mm absorption cell is used, take 5 to 100 ml of for-
magen standard solution [400deg. (formagen)] of 9.2 (1)( c ) step
by step t o prepare the formagen standard solution [20 t o 400 deg.
(formagen)] for working curve.
9.3 Scattered-lightturbidity Measure the intensity of scattered light due to grains

in the sample a t near 660nm wavelength, and obtain the scattered-light turbidity
from the working curve prepared by using kaolin standard solution or formagen
standard solution.
Measuring range: 1 t o 5 deg. (kaolin) o r 0.4 to 5 deg. (formagen) (different
depending on the apparatus)
(b) Kaolin standard solution [lo0 deg. (kaolin)] As described in 9.1 (1)(d).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
22
K O101 : 1998
(cl Formagen standard solution [40 deg. (formagen)] Take 10 ml of

formagen standard solution [400deg. (formagen)] of 9.2 (1)( c ) into a 100 ml
volumetric flask, and add water up to the marked line.
(a) Scattered-light turbidity meter An example of the scattered-light tur-
bidity meter is given in Fig. 9.3.
I Filter A Light receiving part

Collimator lens / ,-///
Light
source
+ Incident light
c3 Transmitted light
-+ Scattered light E l
I -
Fig. 9.3 Structural example of scattered-light

turbidity meter
Take water into an absorption cell, regulate the indication value(7) of a
scattered light turbidity meter a t zero, then use kaolin standard solution
for working curve [5 deg. (kaolin)], and regulate the indication value(7) of
the scattered light turbidity meter at 100 %.
After mixing the sample by shaking sufficiently, take into a n absorption
cell, and measure the indication value (intensity of scattered light) near
660 nm in wavelength.
Obtain the scattered light turbidity [deg. (kaolin)] of the sample from the
working curve prepared by using kaolin standard solution.
Working curve Deal out step by step 1 t o 5 ml kaolin standard solution
[i00deg. (kaolin)] into a 100 ml volumetric flask, add water to the marked
line, and prepare kaolin standard solution [i t o 5 deg. (kaolin)] for work-
ing curve. Hereafter carry out the operation of (a) and (b),and prepare
the relation curve between the scattered light turbidity [i to 5deg. (ka-
olin)] of kaolin standard solution for working curve and the indication value
(intensity of scattered light).
Note (7) For the intensity of scattered light, measurement a t positions of
90" or 270" to incident light are generally carried out.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Take water into an absorption cell, regulate the indication value(7) of a
scattered light turbidity meter at zero, then use formagen standard solu-
tion [5 deg. (formagen)]for working curve, and regulate the indication value (7)
of the scattered light turbidity meter a t 100 %.
23
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`--- K O101 : 1998
After mixing the sample by shaking sufficiently, take into an absorption

cell, and measure the indication value (intensity of scattered light) near
Obtain the scattered light turbidity [deg. (formagen)] of the sample from
the working curve prepared by using formagen standard solution.
Working curve Deal out step by step 1t o 12.5 ml of formagen standard
solution [40 deg. (formagen)] into a 100 ml volumetric flask, add water to
the marked line, and prepare formagen standard solution [i t o 5deg.
(formagen)] for working curve. Hereafter, carry out the operation of (a)
and (b),and prepare the relation curve between the scattered light turbid-
ity [i to 5 deg. (formagen)] of formagen standard solution for working curve
and the indication value (intensity of scattered light).
9.4 Integrating-sphere turbidity Obtain the ratio of the intensity of scattered

light due t o grains in water t o the intensity of transmitted light, and obtain the in-
tegrating-sphere turbidity from the working curve prepared by using kaolin stan-
dard solution or formagen standard solution.
Measuring range: In the case of 50 mm absorption cell, 0.2 t o 5 deg. (kaolin)
or 0.2 to 5deg. (formagen), and in the case of 10mm ab-
sorption cell, 5 to 100 deg. (kaolin) or 5 to 100 deg. (formagen).
(b) Kaolin standard solution [lOOdeg. (kaolin)] As described in 9.1 (1)
(a).
(c) Formagen standard solution [40 deg. (formagen)] As described in 9.3
(1) (cl.
(a) Integratingsphere turbidity meter An example of the integrating-sphere
turbidity meter is shown in Fig. 9.4.
Reflecting
mirror
Light
source
,-Filter - r Standard
Inlet Outlet ,
Lcci \
'Condenser \
Trap
Sample J
cell Light receiving
__ ~
- - . __.
Fig. 9.4 Structural example of integrating-sphere

turbidity meter
24
K O101 : 1998

(a> Put a 50 mm absorption cell containing water and a trap in the optical path,
and adjust the indicting value to O . Then, insert a standard white plate
into the trap, and adjust so that the indicating value becomes 100.
(b) Next, substitute for 50 mm absorption cell containing water, put a 50 mm
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
absorption cell containing the sample mixed by shaking thoroughly and a
trap (containing no standard white plate) in the optical path, and measure
the intensity of scattered light Td.
(c) Successively, insert a standard white plate into the trap, and measure the
intensity Tt of total transmitted light of the sample.
(d) Calculate the value of TdITt x 100, and obtain the integrating-sphere tur-
bidity [deg. (kaolin)] of the sample from the working curve prepared by using
kaolin standard solution.
Working curve Take 0.2 to 5 m l of kaolin standard solution [lOOdeg.
(kaolin)] step by step into a 100ml volumetric flask, and add water up to
the marked line t o prepare(8) the kaolin standard solution [0.2 t o 5 deg.
(kaolin)] for working curve. Hereafter, carry out the operation of (a) t o
(d),and prepare the relation curve between the integrating-sphere turbid-
ity [deg. (kaolin)] of the kaolin standard solution for working curve and
the value of TdITtx 100,
Note (8) In the case of measuring by using a 10 mm absorption cell, pre-
pare the kaolin standard solution [5 to 100 deg. (kaolin)] for working
curve.
Remarks 1 Method without calculation of TaITt x 100 After carrying
out the operation of (a),put a cell containing the sample sub-
stituting for the absorption cell containing water and adjust
so that the indicating value a t that time becomes 100. Suc-
cessively, read out the indicating value with the standard white
plate detached. For working curve, operate in the same man-
ner on the kaolin standard solution (or formagen standard
solution), take the integrating-sphere turbidity [deg. (kaolin)]
or [deg. (formagen)] on the abscissa and the indicating value,
(corresponding to TdTt x 100) on the ordinate to prepare and
obtain the integrating-sphere turbidity of the sample from this
working curve.

(a) Carry out the operation specified in (3.1)(a)t o ( c ) .
(b) Calculate the value of TdITt, and obtain the integrating-sphere turbidity
[deg. (formagen)] of the sample from the working curve prepared by using
formagen standard solution.
Working curve Take 0.5 to 12.5 ml of formagen standard solution [40 deg.
(formagen)] step by step into a 100 ml volumetric flask, and add water up
to the marked line t o prepare the formagen standard solution [0.2to 5 deg.
25
K O101 : 1998
(formagen)] for working curve(9). Hereafter, carry out the operation of (a)
t o prepare the relation curve between the integrating-sphere turbidity [deg.
(formagen)] of the standard solution for working curve and the value of Ta/
Tt x 100.
Note (9) When measuring by using 10mm absorption cell, prepare the
formagen standard solution [5to 100 deg. (formagen)] for work-
ing curve.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
26
K O101 : 1998
10 Colour For colour marking, the method according t o the chromaticity accord-
ing t o platinudcobalt or the method according to the stimulus value Y and chroma-
ticity coordinates x, y shall be used. The chromaticity according t o platinudcobalt
applies only where the sample is of colour system from faint yellow t o yellow brown.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
10.1 Chromaticity according to platinudcobalt The chromaticity according

to platinudcobalt indicates the degree from faint yellow to yellow brown due t o dis-
solved-in-water or colloidal substances and the colour appearing when 1ml of plati-
numícobalt chromaticity standard solution (1 mg of platinum and 0.5 mg of cobalt)
is added in 1I of water shall be taken as platinum/cobalt chromaticity one degree.
(a) Water Filter water A3 specified in JIS K 0557 by using filter of
approx. 0.1 pm pore diameter, and take the filtrate after discarding the first
approx. 200 ml.
(b) Platinumícobalt chromaticity standard solution (1 O00 deg.) Take
2.49 g of potassium hexachloroplatinate specified in JIS K 8163(1),2.00 g
of cobalt II chloride hexahydrate specified in JIS K 8129 and 200ml of
hydrochloric acid specified in JIS K 8180, add water t o dissolve, transfer
into a 1O00 ml volumetric flask, and add water up to the marked line. Pre-
serve it in a brown bottle.
Note (1) Where platinum is used instead of potassium hexachloroplatinate
specified in JIS K 8163, dissolve 1.00 g of platinum in a proper
amount of aquaregia (hydrochloric acid 3 + nitric acid i), add hydro-
chloric acid in excess and evaporate t o dryness on a water bath.
Repeat this operation two or three times t o remove nitric acid,
thereafter dissolve it together with 2.00 g of cobalt II chloride
hexahydrate specified in JIS K 8129 and 200 ml of hydrochloric
acid specified in JIS K 8180, put into a 1O00 ml volumetric flask,
and add water up to the marked line.
(a) Dark box As described in 9.1 (2) (a).
(b) Colorimetric tube 100 ml As described in 9.1 (2) (b).
Filter the sample with filter paper of class 5C or filter of pore diameter of
1pm o r less, or separate the sample centrifugally at about 3 O00 min-1 (the
number of rotations depends on the radius of rotating part of centrifugal
separator) for 20 min t o remove turbidity.
Take a proper amount of this sample into a 100 ml colorimetric tube, and
add water up t o the marked line of 100 ml.
Take step by step 0.1 t o 2.0ml of platinudcobalt chromaticity standard
solution (1O00 deg.) into a 100 ml colorimetric tube, add water up to the
marked line of 100m1, stopper it and mix by shaking thoroughly to pre-
pare platinumícobalt chromaticity standard solution series of 1 t o 20 deg.
27
K O101 : 1998
(d) Place the sample and chromaticity standard solution series on a white pa-
per o r put into a dark box t o see through from above, and compare the colour
of the sample with the platinudcobalt chromaticity standard solution series
to obtain the corresponding platinudcobalt chromaticity standard solution.
(e) Calculate the platinumícobalt chromaticity of the sample from the follow-
ing formula:
100
c=c*x--
V
where, C : platinumícobalt chromaticity
C, : corresponding platinumícobalt chromaticity stan-
dard solution (deg.)
V : sample (ml)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
10.2 Marking by stimulus value Y and chromaticity coordinates x, y The

percentage transmission of the sample a t each wavelength from 400 t o 700nm is
measured using water as reference solution to obtain tristimulus value(2) X,Y,2,
and the chromaticity coordinates x, y is calculated, then the colour is marked with
stimulus value Y and chromaticity coordinates x, y.
Note (2) The value obtained by totalizing the percentage transmission in se-
ries of x,Y,and 2 according to the kinds of wavelength giving re-
spective stimuli, considering that there are three kinds of sensitivities
of the optic nerve t o colour, and the sensitivity t o colour changes by
mixing rate of the three kinds of stimuli t o the optic nerve.
(1) Reagent The following reagent shall be used.
(a) Water As described in 10.1 (1)(a).
(a) Centrifugal separator
(b) Photometer The spectrophotometer capable of using 100 mm absorption
cell and of measuring the total visible field with the wavelength interval
of 10 nm or less, o r equivalent chromaticity meter in performance.
Remove turbidities by filtering the sample with filter paper of class 5C or
filter of 1pm pore diameter or less, or by centrifugal separation at about
3 O00 min-1 (the number of rotations depends on the radius of rotating part
of centrifugal separator) for 20 min.
Transfer a portion of this sample into a 100 mm absorption ce11(3), and
measure the percentage transmission a t each wavelength specified in Table
10.1(4) using water as reference solution.
Notes (3) When the measurement using 100 mm absorption cell is impos-
sible because of the dark colour of the sample, use an absorp-
tion cell of proper length of optical path t o measure. In this case,
measure the absorbance at each wavelength, convert the value
t o the absorbance of 100mm absorption cell, and calculate by
obtaining the percentage transmission form this absorbance.
28
K O101 : 1998
(4) Table 10.1 follows Attached Table 5-1-1 (20nm in wavelength

interval) of the evaluation method of metamerism indexídegree
of illuminant metamerism specified in JIS Z 8719.
Table 10.1 Polyvalues function fx, fy, fi for calculating tristimulus

values X, Y,2 by 20nm in wavelength interval
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Wavelength Specified chromatic light C
(nm) fX fY fz
400 0. 019 -0.003 0. 062
420 2.993 0. 087 14.387
440 7.634 o. 510 38.438
460 6.642 1.382 38.130
480 2.360 3.206 19.545
SOO O . 068 6.907 5.74*6

520 1.196 12.876 1.444
540 5.590 18.261 0 . 356
560 11.751 19.592 0. 073
580 16.795 15.991 O. 026
600 17,897 10.6Y4 0.013

620 14,. 02 1 6.261 0. 002
648 7.453 2.901 o. O00
660 2,731 1.003 o. O00
680 0.756 o. 273 o. 000
700 O. 166 0 . OS9 0. 000
Total 98.072 100.000 118.225

Chromaticity
coordinates x=0.310 1 y =O. 316 2 z=o.373 7
..-I _ _ __
(c) Method for obtaining tristimulus value and chromaticity coordinates
x, y Totalize the percentage transmission at each specific wavelength of X, Y,
and 2 in Table 10.1 respectively, and calculate the tristimulus value X, Y,and
2, and chromaticity coordinates x , y from the following formula,
Y
y = X+Y+Z
where, X : stimulus value X
29
K O101 : 1998
Y : stimulus value Y
2 : stimulus value 2
K : 100.000
fx(3i.I: fx at wavelength h
f~ (A) : fY at wavelength 3L
f i (A): fz at wavelength 3L
z (A) : percentage transmission at wavelength A
(d) Method for obtaining stimulus value Y Stimulus value Y,as it is, is
used as the stimulus value.
Remarks 1 Method for obtaining dominant wavelength and comple-
mentary wavelength The point in the chromaticity (Fig. 10.1)
is non-coloured chromaticity coordinates ( x = 0.310 1, and
y = 0.316 2).
For the colour of which the chromaticity coordinates are rep-
resented by the point Si in the area enclosed by the straight
line RC, the straight line VC, and the spectrum locus, obtain
the wavelength corresponding t o the intersecting point SI’ of
the extension of the straight line CSI and the spectrum locus
from Fig. 10.3. Call this wavelength the dominant wavelength
of the said colour and express by the symbol h.
Further, for the colour of which the chromaticity coordinates
are represented by the point Sz inside the triangle CRV, ob-
tain the wavelength corresponding to the intersecting point
S2” of the extension of the straight line CS2 and the spectrum
locus from Fig. 10.3. Call this wavelength the complementary
wavelength of the said colour, and express by the symbol &.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
0 o.2 0.4 O. 6
X
- .~
Fig. 10.1 Chromaticity diagram
30
K O101 : 1998
Remarks 2 Method for obtaining excitation purity For the colour of

which the chromaticity coordinates are represented by the point
Si in Fig. 10.1, obtain the excitation purity Pe from the fol-
lowing formula(*), and annex % to the said value.
x - x,
P, =- x 100 ........................................................ (i)
Xa -Xe
or
P, =- Y - Ye x 100 ........................................................ (2)

Ya -Yc
where, x, y : coordinates of point SI

xc,y c : coordinates of point C
XA, ya : coordinates of point SI’
Further, for the colour of which the chromaticity coordinates
are represented by the point SZ,obtain the excitation purity
P,from the following formula(*), and annex % to the said value.
x - x,
P, =- x 100 ........................................................ (3)
Xp -Xe
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
or
P, =- Y - Yc x 100 ........................................................ (4)

YP -Ye
where, x, y : coordinates of point S p
xc,yc : coordinates of point C

xp,y p : coordinates of point S2’ (intersecting point of
the extension of straight line CS2 and purple
boundary)
Note (*> In order to calculate Pe, obtain by the formula of which
the absolute value of a denominator or a numerator
is large.
An example of method of obtaining excitation purity and
dominant wavelength If the tristimulus value obtained by
measuring the sample, X is 40.14, Y , 76.50, and 2,34.25, the
chromaticity coordinates x is 0.266, y, 0.507 and the stimulus
value Y , 76.50. Obtain the chromaticity coordinates as a point
(SI) on Fig. 10.2 and then the colourless point as C (x= 0.310 1,
y = 0.316 2). Connect C and SI with a straight line, and obtain
the intersecting point of the extension of straight line and the
spectrum locus as SI’. SI’ is located at 534 nm on the spectrum
locus, i.e. the dominant wavelength & is 534 nm.
Further, since the coordinates of Si’ xn.is 0.200 and Ya is 0.785,
the excitation purity P , becomes x 100 = 40.7 %,
0.785 - 0.316 2
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
31
K 0101 : 1998
Therefore, the colour of the sample is expressed by the domi-

nant wavelength 3,= 534 nm and the excitation purity Y =
76.50 %.
Further, it is convenient that the excitation purity is obtained
by using Fig. 10.4 when the colour of the sample is thin and by
using Fig. 10.5 when the colour of the sample is dense.
X
__
_._ - -- ____
"I_____ - -- - --
Fig. 10.2 Relation diagram between chromaticity diagram
and dominant wavelength according to 2 degree
visual field X Y Z sysem
32
K 0101 : 1998
0.81
0.76
0.60
o. 50
Y
0.40
0.30
0.20
0.10
0
1 .
The curve indicated with wavelength scale mark is the spec-

trum locus, and the curve connecting both the ends of the
spectrum locus is the purple boundary. The point C indicates
the chromaticity coordinates of standard light C (x = 0.310 O,
y = 0.316 2).
Fig. 10.3 Chromaticity diagram according t o 2 degree visual field

X Y Z system (dominant wavelength a n d hue name)
Remarks 3 Dominant wavelength and hue The relationship between
the dominant wavelength and the hue is shown in Table 10.2
and Fig. 10.3.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
33
K 0101 : 1998
Table 10.2 Dominant wavelength and hue name

~
Dominant Hue name Abbreviated Dominant Hue name Abbreviated

wavelength nm symbol wavelength nm symbol
~
498c to 700 to 618 Red R 498 to 482 Blue green BG

618 to 586 Orange O 482 to 435 Blue B
586 to 571 Yellow Y 435 to 400 to 578, Violet V
571 to 531 Yellow green YG 578, to 528c Purple P
531 to 498 Green G 528c to 498c Reddish purple RP
X
~
Fig. 10.4 Excitation purity (10%marking) (in the

case where the colour of sample is thin)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
34
K 0101 : 1998
Fig. 10.5 Excitation purity (100%marking) (in the

case where the colour of sample is dense)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
35
K O101 : 1998
11 pH For measurement of pH, the glass electrode method specified in JIS Z 8802
shall be applied.
Immediately after sampling, pH shall be measured.
11.1 Glass electrode method pH is measured with a pH meter using a glass

electrode.
Water Water A2 or A3 specified in JIS K 0557. In the case where bo-

rate pH standard solution and carbonate pH standard solution are prepared,
carbonic acid-free water of 2 (12)(b) is used.
Potassium trihydrogen dioxalate dihydrate Potassium trihydrogen
dioxalate dihydrate specified in JIS K 8474.
Potassium hydrogen phthalate Potassium hydrogen phthalate speci-

fied in JIS K 8809 for pH standard solution.
Potassium dihydrogenphosphate Potassium dihydrogenphosphate speci-
fied in JIS K 9007 for pH standard solution.
Disodium hydrogenphosphate Disodium hydrogenphosphate specified
in JIS K 9020 for pH standard solution.
Sodium tetraborate Sodium tetraborate 10 hydrate specified in JIS K
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
8866 for pH standard solution.
Sodium hydrogen carbonate Sodium hydrogen carbonate specified in
JIS K 8622 for pH standard solution.
Sodium carbonate Sodium carbonate specified in JIS K 8625 for pH
standard solution.
(2) pH standard solution(1) The following pH standard solutions shall be used.
(a) Oxalate pH standard solution For the prepared pH standard solution,

grind potassium trihydrogen dioxalate dihydrate with an agate mortar,
preserve in a desiccator for 18 h or longer, dissolve its 12.71 g in a small
quantity of water, transfer to a 1O00 ml volumetric flask, and add water
to the marked line. Put it into a polyethylene bottle with ground stopper
o r a borosilicate glass bottle with ground stopper to be preserved. For
specification pH standard solution, use grade 2 of oxalate pH standard solution
specified in JIS K 0018.
(b) Phthalate pH standard solution For the prepared pH standard solu-

tion, preliminarily heat potassium hydrogen phthalate a t 120 "C for 1h.
After standing to cool in a desiccator, take its 10.21 g, dissolve in a small
quantity of water, transfer to a 1O00 ml volumetric flask, and add water
to the marked line. Put it into a polyethylene bottle with ground stopper
or a borosilicate glass bottle with ground stopper t o be preserved. For the
specification pH standard solution, use grade 2 of phthalate pH standard
solution specified in JIS K 0019.
36
K O101 : 1998
Neutral phosphate pH standard solution For the prepared pH stan-

dard solution, preliminarily heat sodium dihydrogenphosphate and diso-
dium hydrogenphosphate a t 110 "C for 2 h. After standing t o cool in a
desiccator, take 3.40g of potassium dihydrogenphosphate and 3.55 g of
disodium hydrogenphosphate , dissolve in a small quantity of water, trans-
fer to a 1O00 ml volumetric flask, and add water t o the marked line. Put
it into a polyethylene bottle with ground stopper or a borosilicate glass bottle
with ground stopper to be preserved. For the specification pH standard
solution, use grade 2 of neutral phosphate pH standard solution specified
in JIS K 0020.
Borate pH standard solution For the prepared pH standard solution,

grind sodium tetraborate 10 hydrate with an agate mortar. After making a
constant quantity by leaving as it is in a desiccator containing the solution
obtained by further adding sodium bromide specified in JIS K 8514 t o so-
dium bromide solution (saturated), take its 3.81 g , dissolve in a small quantity
of carbonic acid free water [according t o 2 (12)(b)], transfer to a 1 O00 ml
volumetric flask, and add the said carbonic acid free water to the marked
line. Fill a polyethylene bottle with ground stopper or a borosilicate glass
bottle with ground stopper to a full state therewith t o be preserved. For
the specification pH standard solution, use grade 2 of borate pH standard
solution specified in JIS K 0021.
Carbonate pH standard solution For the prepared pH standard solu-

tion, make sodium hydrogen carbonate stand in a desiccator for approx. 3 h,
and take its 2.10 g. Separately, preliminarily put sodium carbonate into a
platinum crucible, heat at 600 "C to a constant weight, and take its 2.65 g .
Dissolve both in a small quantity of carbonic acid free water [according to
2 (12)(b)],transfer t o a 1O00 ml volumetric flask, and add the said car-
bonic acid free water t o the marked line. Put it into a polyethylene bottle
with ground stopper or a borosilicate glass bottle with ground stopper, and
preserve in a desiccator containing soda lime specified in JIS K 8603 or
potassium hydroxide specified in JIS K 8574. For the specification pH stan-
dard solution, use grade 2 of carbonate pH standard solution specified in
JIS K 0022.
Note (1) For the pH standard solution, use the prepared pH standard so-
lution o r grade 2 of the specification pH standard solution.
The prepared pH standard solution is prepared in accordance
with the preparation method of each pH standard solution by a test
operator himself. To the pH value at each temperature of each
prepared pH standard solution, the pH value of Table 11.1shall be
applied.
The specification pH standard solution is systematized as the
pH standard solution traceable t o the primary standard substance
(having traceability t o an upper standard) kept by the national
research institute. The pH standard solutions prepared by pH
standard solution manufacturers are submitted t o the public in-
stitute under national superintendence and direction (the insti-
tute having the secondary pH standard solution decided by the
primary pH standard solution owned by the national research
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
37
K O101 : 1998
institute), receives inspection to be accepted. The said accepted

pH standard solution on the market shall be the specification pH
standard solution.
Though these are grade 1 and grade 2 for the specification pH
standard solution, since the grade 1 specification pH standard
solution guarantees the pH value t o three decimal places. For
grade 2 specification pH standard solution to be used for this test,
three decimal places of the pH value of grade 1 is rounded off t o
the second decimal place. The pH value at each temperature of
grade 2 of each specification standard solution is given in Table 11.2.
Table 11.1 pH value of each temperature of

the prepared pH standard solution
Temperature pH value
"C ~~
Neutral
Oxalate Phthalate phosphate Borate Carbonate (9
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
0 1.67 4.01 6.98 9.46 10.32
5 1.67 4,.01 0.95 9.39 (10.25)
10 1.67 4.00 6.92 9.33 10.18
15 1.61 4.00 6.90 9.27 (10.12)
20 1.68 4.00 6.88 9.22 (10.07)
25 1.68 4.01 6.86 9.18 10.02
30 1.69 4.01 6.85 9.14 (9.97)
35 1.69 4.02 6.84 9.10 (9.93)
38 ~ - ~ - 9.91
40 1.70 4.03 6.84 9.07 -
45 1.10 4.04 6.83 9.04 ~
1.71 4.06 6.83 9.01 -

50
1.72 4.08 6.84 8.99 -
55
4.10 6.84 8.96 -
60 1.73
8.93 -.
70 1.74 4.12 6.85
4.16 6.86 8.89 -
80 1.77
90 1.80 4,.20 6.88 8.85 ~
1.81 8.83 -
95 4.23 6.89
__ - _ I__ I -- _ - _I__.
__
Note (*) The values in parentheses indicate secondary inter-
polated values.
38
K 0101 : 1998
Table 11.2 pH value at each temperature of

specification pH standard solution
Temperature pH value
"C
Oxalate Phthalate Neutral phosphate
Grade 2 Grade 2 Grade 2 Grade 2 Grade 2
O 1.67 4.00 6.98 9.46 10.32

5 1.67 4.00 6.95 9.40 10.24
10 1.67 4.00 6.92 9.33 10.18
15 1.67 4.00 6.90 9.28 10.12
20 1.68 4.00 6.88 9.22 10.06
25 (*) 1.68 (*) 4.01 (*) 6.86 ($1
30 1.68 4.02 6.85
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
35 1.69 4.02 6.84
38 1.69 4.. 03 6.84 9.08 -
40 1.69 4.04 6.84 9.07 9.89
45 1.70 4.05 6.83 9.04 9.86
50 1.71 4.06 6.83 9.01 9.83
, I
55 1.72 4,. 08 6.83 -
60 1.72 4.09 , 6.84 I
70 1.74 4.13 6.84 8.92
80 1.77 4.16 6.86 8.88
90 1.79 4.20 6.88 8.85
95 1.81 4.23 6.89 8.83
~~
" I- -----
~ ~
I - ___-
~~
"I
~ ~~ ~
r^II____-___ -
~
-- --- -
Note (*) pH value with the mark (*) at 25 O C is specified as Japanese Indus-
trial Standard for each pH standard solution.
Remarks 1 A pH value at a temperature not stated on Table 11.1and Table 11.2
is obtained by interpolation.
2 When each pH standard solution is preserved for a long period,
its pH value occasionally varies. Therefore, that preserved for a
long time shall not be used. Since especially borate pH standard
solution and carbonate pH standard solution absorb easily carbon
dioxide in the air and the pH value decreases, therefore cares shall
be taken.
3 Each pH standard solution which has been once used or which was
left open in the air shall not be used.

(a) pH meter The pH meter of type II specified in JIS Z 8802 is used(2).
(b) Thermometer 50 or 100 "C thermometer of solid-stem general purpose
liquid-in-glass thermometer specified in JIS B 741l ( 3 ) .
Notes (2) The type of the pH meter is selected according t o the purpose of
a test.
I n JIS Z 8802,the type of the pH meter is classified into 4
stages of O to III. The repeatability of each type pH meter, when
the pH value is measured by using pH standard solution of one
39
K O 1 0 1 : 1998
kind, is specified as k0.005 for type O, f0.02for type I, 10.05 for

type II, and I O . l for type III.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(3) It is the solid-stem general purpose liquid-in-glass thermometer

specified in JIS B 7411 of I1 "C in temperature measuring pre-
cision.
(4) Calibration of pH meter The pH meter shall be calibrated as follows.

Make a power source of a pH meter, and install a detection part [a glass
electrode (4) (5) and a reference electrode (9, thermometer, etc.].
Wash the detection part repetitively 3 or more times with water, and wipe
it off with clean soft paper.
Take neutral phosphate pH standard solution into a beaker and moisten
the detection part. For the pH meter equipped with a temperature com-
pensating dial or digital switch, conform the scale value t o the tempera-
ture of neutral phosphate pH standard solution(') ( 8 ) .
Regulate to conform the regulation dial (asymmetric potential regulation
dial) t o a pH value (Table 11.1 o r 11.2) corresponding t o the temperature
of neutral phosphate pH standard solution.
Wash the detection part repetitively three o r more times with water, and
wipe off with clean soft paper or the like.
In the case where the pH value of a sample is 7 or under, take phthalate
pH standard solution or oxalate pH standard solution into a beaker, and
moisten the detection part. Use a span regulation dial, and conform it t o
a pH value (Table 11.1 or 11.2) corresponding to the temperature of the
used pH standard solution by regulating a span regulation dial(8). In the
case where the pH value of the sample exceeds 7(9), use borate pH stan-
dard solution o r carbonate pH standard solution, and conform it to a pH
value corresponding t o the temperature of the pH standard solution by the
same operation(8).
Carry out again the operation of (b) to (0,and repeat this operation until
the indication value of the pH value conforms to the pH value correspond-
ing t o the temperature of the pH standard solution by f0.05(10).
Notes (4) For a glass electrode which is in a dry state for a long period,
preliminarily dip into water, and use after reaching equilibrium.
(5) In the case where the glass electrode is stained, wash with de-
tergent, hydrochloric acid (1+20), etc. for a short time as required,
and further wash thoroughly with running water. Handle the
electrode in accordance with an instruction manual.
(6) Remove stains of the reference electrode by the same operation
as for the glass electrode. Refer t o the instruction manual for
exchange or the like of inner liquid (potassium chloride solution).
(7) Conform the temperature of pH standard solution t o the tem-
perature of a sample as far as possible.
40
K O101 : 1998
Make fluctuation in the temperature of each pH standard solu-

tion $12O C in calibration.
In the case where the pH value of a sample is 11 o r higher, al-
kali error is generated for an ordinary glass electrode and the
measured value decreases. Especially when the concentration
of alkali metal ion is high, the error increases. Therefore, mea-
surement is performed in accordance with Remarks 4.
Repeat the operation until the indication value of the pH value
conforms to the pH value corresponding t o the temperature of
the pH standard solution by k0.02 for the pH meter of type I
and by IO.1 for the pH meter of type III.

(a) Wash the detection part of a calibrated pH meter repetitively three or more
times with water, and wipe off with clean soft paper or the like.
(b) Take a sample into a beaker, and moisten the detection part. After con-
forming the scale value of the pH meter equipped with a compensating dial
o r digital switch to the temperature(") of the sample, measure a pH value.
(c) Take out the detection part, wash repetitively three or more times with
water, and wipe off with clean soft paper or the like.
(d) Take the sample again into the beaker, and immerse the detection part,
and measure the pH value(11).
(e) Carry out the operation of (e) and (d) again, average the measured value
wherein three times measured values conform to one another by f0.1(12),
and calculate the pH value of the sample.
Notes (11) Since the pH value differs according t o the temperature of a
sample, take fluctuation in the temperature of the sample as
I 2 "C.
(12) Since the pH value varies easily for the sample with low inter-
ference property, the pH value can not occasionally obtained
repeatability of kO.1. In that case, the value a t which the pH
value conforms t o each other by kO.2 is averaged t o calculate
the pH value.
Further, in the case where the pH value fluctuates easily due
to carbon dioxide in the air, the electrode of a flow liquid type
should be used.
Remarks 4 In the case where the pH value of a sample is 11 or higher
and the concentration of alkali metal element is especially high,
alkali error is apt to be generated. Therefore, use the elec-
trode of which the alkali error is little (for instance, lithium
electrode or the like), carry out the calibration of the pH meter
by using 0.1 mol/Z sodium hydroxide solution containing no
carbonate or potassium hydroxide solution (saturated) at 25 O C .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
41
K O 1 0 1 : 1998
Since O. 1mol/Z sodium hydroxide solution or calcium hydrox-

ide solution (saturated) at 25 "C absorbs carbon dioxide in the
air and the pH decreases easily, prepare a t each service.
The pH value at each temperature of 0.1 mol/Z sodium hy-
droxide solution and potassium hydroxide solution (saturated)
is given in Table 11.3.
Table 11.3 pH value at each temperature of 0.1 mol/Z sodium hydroxide

solution and calcium hydroxide solution (saturated)
Temperature 0.1 mol/Z sodium Calcium hydrox- Temperature 0.1 mol/l sodium Calcium hydrox-
hydroxide solu- ide solution hydroxide solu- ide solution
OC tion (saturated) (*) "C tion (saturated) (*)
O 13.8 13.43 35 12.6 12.14
5 13.6 13.21 40 12.4 11.99
10 13.4 13.00 45 12.3 11.84
15 13.2 12.81 50 12.2 11.70
20 13.1 12.63 55 12.0 11.58
25 12.9 12.45 60 11.9 11.45
30 12.7 12.30
Note (*) Potassium hydroxide solution (saturated) at 25 "C.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
42
K O101 : 1998
12 Electric conductivity Electric conductivity corresponds to a reciprocal num-

ber of the electric resistivity ( n e m ) of solution and is expressed by a unit of S/m.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Further, conductance corresponds to a reciprocal number of electric resistance (Q)

of the solution and is expressed by a unit of S.
I n the test of water, the electric conductivity and the conductance a t 25 "C are
used and are respectively expressed by mS/rn(l) and mS(2) by using units of one thou-
sandth of S/m and S. For measurement when the electric conductivity of a sample
is 1 mS/m (25 OC) or under, JIS K 0552 shall be applied.
Notes (1) mS/m shall be read as milli Siemens per meter.
(2) 1 pS/cm expressed by traditional unit corresponds t o 0.1 mS/m (1pS/cm
= 1x 10-6 W10-2m = 1x 10-4 S/m = 0.1 mS/m).
Remarks 1 Traditionally in a test of water, pS and pS/cm were respectively used
as units of conductance and conductivity. Further, cm-1 was used
as a unit of cell constant.
As conductance, when a numerical value expressed by a unit of
mS is multiplied by 1000, a numerical value expressed by a unit of
pS is obtained.
As conductivity, when a numerical value expressed by a unit of
mS/m is multiplied by 10, a numerical value expressed by a unit of
pS/cm is obtained.
Further, as cell constant, when a numerical value expressed by a
unit of m-1 is multiplied by 0.01, a numerical value expressed by a
unit of cm-1 is obtained.
In addition, units and values in { 1 are traditional units, and are
appended for informative reference.

(a) Electric conductance meter The electric conductance meter consists of
a detector and an indicator. The detector consists of cell in which plati-
num electrodes coated with platinum black are assembled. The cells with
the cell constants as given in Table 12.1 shall be prepared. The indicator
in which the Wheatstone bridge circuit or the like is assembled shall be
used. The cell shall be preserved in water(3).
(b) Thermometer 50 "C thermometer of solid-stem general purpose liquid-
Note (3) The cell shall be confirmed periodically by the method as given
in Remarks 2.

(a) Put on preliminarily the power source of electric conductance meter. Use
an electrode with a cell constant as given in Table 12.1 corresponding t o
electric conductivity of a sample, and wash the cell with water two or three
times. [In case where the cell is stained particularly, immerse it into hy-
drochloric acid (1+100), then wash in the running water sufficiently, and
finally wash it with water two or three times.]
43
K O101 : 1998
After rinsing the cell with the sample two or three times, fill the cell with
the sample. Keep the temperature of the cell at (25I0.5) OC(4), and mea-
sure the conductance (5) (6). Repeat the measurement with changing the
sample several times until the measured values coincide within _+3%(7),
and obtain the conductance.
Calculate the conductivity (mS/m) (25 OC) of the sample from the conduc-
tance according to the following formula:
L=JxL,
where, L : conductivity of the sample (mS/m) (25 OC)
J : cell constant (m-1)
L, : measured conductance (mS)
When the high precision is not required particularly, the conduc-
tance meter equipped with a temperature compensating circuit or
temperature conversion formula may be used. The conductivity
changes with temperature at a rate of about 2 % increase of 1“C
rise. However, when the conductivity becomes 1mS/m (10 pS/cm}
or under, influences of hydrogen ion and hydroxide ion t o be gen-
erated by dissociation of water, this conversion formula cannot be
applied.
Where the scale of conductance meter is graduated in electric
resistance (QI, the conductivity shall be calculated according to
the following formula:

J : cell constant (m-1). However, where the resis-
tivity (Q * m) is indicated directly, it shall be taken
as 1.
R , : measured electric resistance (Q)
Where the scale of conductance meter is graduated in conduc-
tance (pS), the conductivity (mS/m) shall be calculated from the
following formula.
L = J’xL,’XO.l
J’ : cell constant (cm-1)
L,’ : measured conductance ($3)
Where the conductivity of a sample is 1 mS/m (25 O C ) or less, since
the measured values may not often coincide within 5-3 %, the test
is carried out in accordance with JIS K 0552, or the flow type
cell is used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
44
K O101 : 1998
Table 12.1 Cell constant and measuring range

Cell constant (*I I Measuring range
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
~~ ~~ ~~
m-i (cm-l) mS/m (pS/crn)
1 0.01 2 max. 20 max.

10 o. 1 0.1 to 20 1 to 200
100 1 1 to 200 10 to 2 O00
1O00 10 10 to 2 O00 100 to 20000
5 O00 50 100 to 20 O00 1 O00 to 200 O00
Remarks 2 Measurement of cell constant or confirmation of cell

constant Though the measurement of a cell constant and the
confirmation are not required t o be performed every time a
sample is tested, potassium chloride standard solutions (A t o
D) are periodically used to confirm the numerical value. The
measurement of the cell constant and the confirmation of the
cell constant shall be as follows.
Water Water A2 o r A3 specified in JIS K 0557 with
conductivity 0.2 mS/m I2 gS/cmI (25 O C ) or less. Use
it adjusted to (20f2)"C in the preparation.
Potassium chloride Grind potassium chloride (for
conductivity measurement) specified in JIS K 8121
with a n agate mortar, heat a t 500 "C for approx. 4 h,
and allow t o stand to cool in a desiccator.
Potassium chloride standard solution (A) Weigh
out 74.246g of potassium chloride, dissolve it in a
little amount of water, transfer t o a 1 O00 ml volu-
metric flask, and add water t o the marked line.
Potassium chloride standard solution (B) Weigh
out 7.437 g of potassium chloride, dissolve in a little
amount of water, transfer t o a 1O00 ml volumetric
flask, and add water to the marked line.
Potassium chloride standard solution (C) Weigh
out 0.744 g of potassium chloride, dissolve i n a little
amount of water, transfer t o a 1O00 ml volumetric
flask, and add water t o the marked line.
Potassium chloride standard solution (D) Take
100 ml of potassium chloride standard solution (C)
into a 1O00 ml volumetric flask, and add water t o
the marked line.
Preserve those potassium chloride standard solutions
in a polyethylene bottle o r a borosilicate glass bottle by
tightly stoppering. The conductivities of those potassium
chloride standard solutions are given in Table 12.2.
45
K O101 : 1998
Table 12.2 Conductivity of potassium chloride

standard solutions (A to D)
Potassium chloride "C mS/m íFS/cml

standard solution
A O 6 518 65 180
18 9 784 97 840
25 I 11 134 1 111340
0 714 7 140
18 1117 11 170
25 1286 12 860
O 77.4 774
18 122.0 1220
25 140.9 1409
25 I 14.7
~~
1 147
~
(2) Operation Carry out the operation as follows,

(a) Measurement of cell constant In the case of mea-
surement or confirmation of cell constant, wash the
cell with water two o r three times, and then after
washing with potassium chloride standard solution
(use potassium chloride standard solutions of the
measuring range as given in Table 12.1 according to
the cell constant) two or three times, fill the cell with
the potassium chloride standard solution. Keep the
cell at (25I0.5)"C and measure the conductance. By
changing the same potassium chloride standard so-
lution several times t o carry out the measurement,
and repeat this procedure until the measured values
coincide within +3 % (where they do not coincide
within +3 %, the platinum black electrode shall be
newly coated with platinum black in accordance with
Remarks 3).
Calculate the cell constant from the measured value
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
according to the following formula:
J = L K C I + Lm0
LXO
where, J : cell constant (m-1)

Lxo : measured conductance (mS), how-
ever where the conductance is gradu-
1
ated in YS, use a value of p S x -
1O00
LKCI: conductance used potassium chloride
standard solution at this tempera-
ture (mS/m)
46
K O101 : 1998
L H ~:Oconductivity of water used for prepa-

ration of potassium chloride stan-
dard solution at this temperature
(mS/m)
Further, when the cell constant is required t o be
obtained by a unit of cm-1 in the case where the con-
ductance is graduated in @,calculate from the fol-
lowing formula.
where, J?: cell constant (cm-1)

LXO? : measured conductance (pS)
LKCI? : conductivity a t this temperature
used potassium chloride standard so-
lution {pS/cm)
LH~o? : conductivity a t this temperature of
water used for preparation of potas-
sium chloride standard solution
CpS/cml
When the cell constants measured with the two
kinds of potassium chloride standard solutions of very
close concentrations in Table 12.1 do not coincide
within fl %, the platinum black coated on the plati-
num electrodes shall be dissolved in accordance with
Remarks 3, and be newly coated with platinum black.
Remarks 3 Platinum black coatings on electrode Carry out the plati-
num black coating on the electrode as follows.
The platinum black coatings can be easily removed by the
electrolysis using platinum black electrode as anode in
hydrochloric acid (l+ll).
Immerse this platinum electrode in the electrolytic solution
consisting of hexachloroplatinic (IV) acid solution (30 g/Z>
and lead acetate solution (0.25 g/Z}, and apply electric cur-
rent of 15 t o 30 kC (Coulomb)/m2 {1.5to 3.0 C/cm2} t o the
electrode at approx. 6 V d.c. and current density of 10 t o
40 A/m2 [ i t o 4 d c r n z } while mixing with stirring the
electrolytic solution with an appropriate method.
Then, put the electrode in sulfuric acid (1+360) for
approx. 30 min changing current direction at times to expel
hexachloroplatinic (IV) acid and chlorine deposited or
absorbed on the platinum plate.
4 As for measuring instruments on the market, the conductiv-
ity is directly indicated in many cases, therefore, the indicated
values should be confirmed by potassium chloride standard
solution as required.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
47
K O 1 0 1 : 1998
13 Acid consumption The acid consumption is the amount of hydrogen ion (amount
of acid) required for neutralization of alkalis contained in water such as hydrogen
carbonates, carbonates, hydroxides, etc. to specified pH, and shall be expressed by
mmol per 11 of the sample o r by converting to the amount of calcium carbonate equiva-
lent t o the hydrogen ion (acid), expressed by mg per 11 of the sample.
The acid consumption is divided into acid consumption (pH 4.8) and that (pH 8.3).
13.1 Acid consumption (pH 4.8) The acid consumption (pH 4.8) shall be obtained
by titration with 10 mmol/Z sulfuric acid by adding to the sample the mixed solution
of Methyl Red-Bromocresol Green as indicator.
(i) Reagents The following reagents shall be used.

(a) Methyl Red-Bromocresol Green mixed solution Dissolve 0.02 g of
Methyl Red specified in JIS K 8896 and 0.1 g of Bromocresol Green speci-
fied in JIS K 8840 in 100 ml of ethanol (95) specified in JIS K 8102.
(b) 50 mmol/Z sulfuric acid Add 3 ml of sulfuric acid specified in JIS K 8951
to a beaker preliminarily put with 100 ml of water, mix by stirring thor-
oughly, and after standing to cool, add water t o make 11.
Standardization Heat sodium carbonate of standard reagent for volu-
metric analysis specified in JIS K 8005 a t 600 "Cfor approx. 1 h, then allow
t o stand to cool in a desiccator, and weigh out its 1.06 g t o the nearest 1mg.
Dissolve it in water, transfer into a 200 ml volumetric flask, and add wa-
ter up t o the marked line.
Take its 20 ml into a beaker, add 3 t o 5 drops of mixed solution of Methyl
Red-Bromocresol Green as indicator, and then titrate with this 50 mmol/Z
sulfuric acid. When the colour of the solution has turned grey violet, expel
the carbon dioxide by boiling, and after standing to cool, continue the titra-
tion until the colour of the solution turns grey violet. Calculate the factor
cf) of 50 mmolíl sulfuric acid from the number of ml of 50 mmol/Z sulfuric acid
required for titration according to the following formula:
b1 20
f=axGX %
xx0.00530
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
where, a : amount of sodium carbonate (g)

b : content of sodium carbonate (%)
x : amount of 50 mmol/Z sulfuric acid required for
titration (mi)
0.005 30 : equivalent (g) of sodium carbonate to 1ml of
50 mmoM sulfuric acid
(c) 10 mmol/Z sulfuric acid Take 200 ml of 50 mmol/Z sulfuric acid into a
1 O00 ml volumetric flask, and add water up to the marked line.
48
K O101 : 1998

Take 100 ml of a sample (where there exists turbidity, filter with filter paper
of grade 5B o r apply centrifugal separation and use its supernatant) into a
beaker and add 3 t o 5 drops(1) of mixed solution of Methyl Red-Bromocresol
Green as indicator.
While mixing this solution by stirring gently, titrate with 10 mmol/Z sulfu-
ric acid until the colour of the solution turns from blue to grey violet (pH
4.8).
Calculate the acid consumption (pH 4.8) according to the following formula:
i) In the case of expressing mmol/Z:
A = u x ~ x Oo0
- x 0.02
V
where, A : acid consumption (pH 4.8) (mmol/Z)
a : 10 mmol/Z sulfuric acid required for titration (mi)
f : factor of 10 mmol/Z sulfuric acid(2)
V : sample (mi>
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
0.02 : hydrogen ion equivalent t o 1ml of 10 mmol/l sul-
furic acid (mmol)
ii) In the case of expressing by mgCaCOdZ:
B = U Xf X- loo0
x 1.001
V
where, B : acid consumption (pH 4.8)(mgCaCOdZ)
f : factor of 10 mmol/Z sulfuric acid(2)
V : sample (mi)
1.001 : calcium carbonate equivalent to 1ml of 10 mmol/Z
sulfuric acid (mg)
Notes (1) Where oxidizing substance such as residual chlorine and the like
coexists, add after reduction by sodium thiosulfate solution
(0.1 mol/Z).
(2) Use the factor of 50mmol/Z sulfuric acid.
Remarks 1 In the case where the change of colour due to indicator is not
clear such as in the case of coloured sample, use a pH meter
instead of a indicator, and prepare the titration curve of pH-
10 mmol/Z sulfuric acid to obtain the number of ml of 10 mmol/Z
sulfuric acid at pH 4.8.
13.2 Acid consumption ( p H8.3) The acid consumption (pH 8.3) shall be obtained
by the titration with 10 mmol/Z sulfuric acid by adding phenolphthalein solution as
indicator t o the sample.
49
K O101 : 1998
(a) Phenolphthalein solution (5 glZ) Take 0.5 g of phenolphthalein speci-

fied in JIS K 8799, dissolve in 50ml of ethanol (95) specified in JIS K
8102,add water t o make 100 ml, and add 20 mmol/Z sodium hydroxide so-
lution drop by drop until the colour of the solution shows slightly red.
(b) 10 mmol/Z sulfuric acid As described in 13.1 ( i )( c ) .

(a) Take 100 ml of the sample into a beaker, and add 3 t o 5 drops of phenol-
phthalein solution ( 5 glZ) as indicator.
(b) While mixing by stirring this solution gently, titrate with 10 mmol/Z sulfu-
ric acid until the red colour of the solution disappears.
(c) Calculate the acid consumption (pH 8.3) according to the following formula:
i) In the case of expressing by mmollZ:
A = a x f X- loo0x o . 0 2
V
where, A : acid consumption (pH 8.3) (me; equivalentll)
a : 10 mmol/E sulfuric acid required for titration (ml)
f : factor(2) of 10 mmolll sulfuric acid
V : sample (mi)
0.02 : hydrogen ion equivalent t o 1ml of 10 mmol/Z sul-
furic acid (mmol)
ii) In the case of expressing by mgCaCOsl2:
B = a x f x -loo0
x 1.001
V
where, B : acid consumption (pH 8.3) (mgCaCOalZ)
f : factor(2) of 10 mmol/Z sulfuric acid
V : sample (ml)
1.001 : calcium carbonate equivalent t o 1ml of 10 mmol/Z
sulfuric acid (mg)
Remarks 2 As described in Remarks 1. However, obtain the number of
ml of 10mmollZ sulfuric acid a t pH8.3.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
50
K 0101 : 1998
14 Alkali consumption The alkali consumption is the amount of hydroxide ion

(amount of alkali) required for neutralization of precipitated metal elements o r the
like as strong acid, carbonic acid, organic acid and hydroxide dissolved in water t o
specified pH and shall be expressed by mmol per 1Z of the sample or by converting
to the amount of calcium carbonate equivalent to hydroxide ion (amount of alkali),
be expressed by mg per 1Z of the sample.
The alkali consumption is divided into alkali consumption (pH 8.3), that (pH 4.8)
and that (free acid).
14.1 Alkali consumption (pH 8.3) The alkali consumption (pH 8.3) shall be ob-
tained by titration with 20 mmoVZ sodium hydroxide solution by adding phenolphthalein
solution as indicator to the sample.
(i) Reagents Use the following reagents.

Phenolphthalein solution (5 g/Z) As described in 13.2 (i)(a).
0.1 mol/Z Sodium hydroxide solution Take approx. 30 ml of water in a
polyethylene bottle, add and dissolve approx. 35 g of sodium hydroxide speci-
fied in JIS K 8576 little by little while cooling, stopper tightly, and allow
t o stand for 4 to 5 days. Take 5 ml of the supernatant liquid into 1Z poly-
ethylene vessel with airtightness, and add water containing no carbonic
acid specified in 2 (12)(b)to make total 1Z, mix, and preserve it with shielding
carbon dioxide.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Standardization Weigh out approx. 0.2 g to the nearest 1mg of amido-

sulfuric acid of volumetric analysis standard reagent specified in JIS K 8005
preliminarily dried by leaving in a desiccator for about 48 h, transfer into a
200 ml Erlenmeyer flask, and dissolve by adding approx. 25 ml of water. Add
to this solution 3 to 5 drops of bromothymol blue solution (iglZ)(i) as indi-
cator, titrate with 0.1 moVZ sodium hydroxide solution, and take the point
when the colour of solution is changed to green as an end point, Calculate
the factor (f)of 0.1 moVZ sodium hydroxide solution according t o the follow-
ing formula:
b 1
=a 100 x x 0.009 71
where, a : quantity of amidosulfuric acid ( g )
b : content of amidosulfuric acid (%)
x : 0.1 mol/Z sodium hydroxide solution required for
titration (mi)
0.009 71 : amidosulfuric acid equivalent to 1ml of 0.1 mol/Z
sodium hydroxide solution ( g )
20 mmol/Z sodium hydroxide solution Take 200 ml of 0.1 mol/Z sodium
hydroxide solution in a 1O00 ml volumetric flask, and add water contain-
ing no carbonic acid specified in 2 (12)(b)to the marked line. Prepare this
solution at the time of use.
Note (1) Dissolve 0.1 g of bromothymol blue specified in JIS K 8842 with
50 ml of ethanol (95) specified in JIS K 8102,and dilute with water
t o make 100ml.
51
K 0101 : 1998
Remarks 1 Instead of the standardization by amidosulfuric acid, the stan-

dardization may be carried out as follows by using 50 mmoVZ
sulfuric acid standardized by using sodium carbonate of volu-
metric analysis standard reagent specified in JIS K 8005.
Standardization Take 20 ml of 50 mmoW sulfuric acid of
13.1 ( i )(b) in a beaker, add 3 to 5 drops of phenolphthalein
solution (5 gll) as indicator, and then titrate with this 0.1 moVZ
sodium hydroxide solution until the colour of the solution shows
faint red. Calculate the factor (fd of 0.1 moVZ sodium hydrox-
ide solution from the number of ml of 0.1 moVZ sodium hydrox-
ide solution according to the following formula:
20xf
f1=-
X
where, f : factor of 50mmollZ sulfuric acid

x : 0.1 moVl sodium hydroxide solution required
for titration (mi)

(a) Measuring cylinder (with ground stopper) 100 ml measuring cylin-
der or 100 ml glass cylinder with ground stopper.

Take 100 ml of sample(2) into a 100 ml measuring cylinder with a stopper,
taking care so as not t o shake the sample container as far as possible.
Add 4 o r 5 drops of phenolphthalein solution (5glZ) as indicator, place it
on a white paper, titrate with 20 mmol/Z sodium hydroxide solution until
the solution shows slight red colour(3) (41, and take it as the preliminary
test.
Take 100 ml of sample into another 100 ml measuring cylinder with ground
stopper in the same operation as in (a), and add 4 o r 5 drops of phenol-
phthalein solution (5glZ) as indicator. Add a t once the 20mmollZ sodium
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
hydroxide solution of the same amount as the amount consumed in the

preliminary test of (b), stopper, mix by shaking gently, further continue
the titration when the red colour of the solution has disappeared, and ti-
trate until slight red colour appears.
Calculate the alkali consumption (pH 8.3) according to the following for-
mula:
i) I n the case of expressing by mmol/Z:
A = a x fix- loo0
x 0.02
V
where, A : alkali consumption (pH 8.3) (mmolll)
a : 20 mmol/Z sodium hydroxide solution required for
titration (mi)
fl : factor(5) of 20 mmol/Z sodium hydroxide solution
52
K O 1 0 1 : 1998
V : sample (mi)
0.02 : hydroxide ion equivalent to 1 ml of 20 mmolll so-
dium hydroxide solution (mmol)
ii) In the case of expressing by mgCaCOslZ:
B = a x f , x- loo0
x 1.001
V
where, B : alkali consumption (pH 8.3) (mgCaCOslZ)
a : 20 mmolll sodium hydroxide solution required for
titration (mi)
f~ : factor(5) of 20 mmolll sodium hydroxide solution
V : sample (mi)
1.001 : calcium carbonate equivalent to 1ml of 20 mmolll
sodium hydroxide solution (mg)
Notes (2) Where there exist colour and turbidity in the sample, take 100 ml
of sample in a measuring cylinder with ground stopper, and ti-
trate by using this as reference solution.
(3) Make the intensity of colouration at the end point the same as
that of colouration when 100 ml of sodium hydrogencarbonate
(10mmoVZ) solution is taken in a 100 ml measuring cylinder with
a stopper, 4 or 5 drops of phenolphthalein solution (5 gll) are added
and mixed by shaking.
(4) Place the measuring cylinder with ground stopper on a white paper,
and observe it from above obliquely.
(5) Use the factor of 0.1 mol/Z sodium hydroxide solution.
14.2 Alkali consumption (pH 4.8) The alkali consumption (pH 4.8) shall be ob-
tained by titration with 20 mmolll sodium hydroxide solution by adding the mixed
solution of Methyl Red-Bromocresol Green as indicator to the sample. In this mea-
suring value, the amount of alkali required for reaction with salts such as iron, alu-
minium, etc. is contained.

(a) Mixed solution of Methyl Red-Bromocresol Green As described in
13.1 (i)(a).
(b) 20 mmol/Z sodium hydroxide solution As described in 14.1 (1)( c ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Take 100 ml of sample(2) in a beaker, and add 3 to 5 drops of mixed solu-
tion of Methyl Red-Bromocresol Green as indicator.
(b) Titrate with 20 mmolll sodium hydroxide solution mixing by stirring gen-
tly until the colour of the solution turns from red to grey violet (pH 4.8).
(c) Calculate the alkali consumption (pH 4.8) according to the following for-
mula:
53
K O 1 0 1 : 1998
i) In the case of expressing by mmol/Z:
A = a x f l X- loo0xo.02
V
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
where, A : alkali consumption (pH 4.8) (mmol/Z)
titration (mi)
fl : factor(5) of 20 mmol/Z sodium hydroxide solution
V : sample (mi)
0.02 : hydroxide ion equivalent t o 1ml of 20 mmol/Z so-
ii) In the case of expressing by mgCaCOd:
V x 1.001
B = a x f l X - IOoo
where, B : alkali consumption (pH 4.8)(mgCaCOslE)

titration (mi)
fl : factor(5) of 20 mmolíZ sodium hydroxide solution
V : sample (ml)
Remarks 2 As described in Remarks 1 of clause 13. However, obtain the
volume (ml) of 20 mmol/Z potassium hydroxide solution.
14.3 Alkali consumption (free acid) Based on the titration of sulfuric acid, hydro-
chloric acid, nitric acid, strong organic acid, etc. dissolved in water with 20mmol/Z
sodium hydroxide solution in the presence of potassium oxalate, the alkali consump-
tion (free acid) shall be obtained from the titration curve of pH-sodium hydroxide so-
lution (mi).
This measuring value does not contain the alkali consumption due to salts of iron,
aluminium, etc. and only the alkali consumption corresponding to the free acid can
be obtained.
(a) Potassium oxalate monohydrate As specified in JIS K 8522.
(b) 20 mmollZ sodium hydroxide solution As described in 14.1 (i)(e).
(a) Magnetic stirrer
(b) pH Meter The pH meter of type II specified in JIS Z 8802.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
54
K O101 : 1998

Take 100ml of the sample into a beaker, and adjust the liquid tempera-
ture to approx. 10 OC.
Add approx. 20 g of potassium oxalate monohydrate, and mix by stirring to
dissolve.
Mix by stirring with a magnetic stirrer, and titrate by 20mmollZ sodium
hydroxide solution while measuring pH with a pH meter.
Before and after the end point (deflecting point), add each 0.1 ml of 20 mmol/Z
sodium hydroxide solution.
Obtain the end point by preparing the titration curve of pH and 20 mmol/Z
sodium hydroxide solution (ml), and calculate the alkali consumption (free
acid) according to the following formula;
i) In the case of expressing by mmol/Z:
A = a x f l x- Oo0 x 0.02
V
where, A : alkali consumption (free acid) (mmol/Z)
titration (ml)
fi : factor(5) of 20 mmol/Z sodium hydroxide solution
V : sample (ml)
0.02 : hydroxide ion equivalent to 1 ml of 20 mmol/Z so-
ii) In the case of expressing by mgCaCOdZ:
~ X - x 1.001
B = U X ~ loo0
V
where, B : alkali consumption (free acid) (mgCaCO3lZ)
titration (mi)
f~ : factor(5) of 20 mmol/Z sodium hydroxide solution
V : sample (mi)
55
K O101 : 1998
15 Hardness The amounts of calcium ion and magnesium ion in water shall be
converted to the corresponding amount of calcium carbonate and the hardness be
expressed by mg per 12 of the sample.
The hardness shall be divided into total hardness, calcium hardness and magne-
sium hardness. Furthermore, depending on the case, the hardness shall be divided
into total hardness, noncarbonate hardness and carbonate hardness.
15.1 Total hardness
15.1.1 Chelatometric titration method
(i) Obtain the amount of 10mmol/Z EDTA solution required for titration in 50.1
(2) concerning the filtered sample, and calculate the total hardness according
t o the following formula:
H = b x - loo0x 1.001
V
where, H : total hardness (mgCaCOdZ)
b : 10 mmol/Z EDTA solution required for titration in
50.1 (2) (ml)
V : sample (ml)
1.001 : calcium carbonate equivalent t o 1ml of 10 mmol/Z
EDTA solution (mg)
Remarks 1 Noncarbonate hardness and carbonate hardness shall be calculated
as follows from the relation between the acid consumption (pH 4.8)
and the total hardness:
where,
acid consumption (pH 4.8) (mgCaCO3/Z) < total hardness
(mgCaC 03/21,
noncarbonate hardness (mgCaCOdZ)= total hardness
(mgCaCOd2) - acid consumption (pH 4.8) (mgCaCO&,
where,
acid consumption (pH 4.8) (mgCaCOdZ) Itotal hardness
(mgC a C Osí2 1,
carbonate hardness (mgCaCO& = total hardness

(mgCaCOdZ)
noncarbonate hardness (mgCaCO3lZ)= O
15.1.2 Flame atomic absorption method
(1) Concerning the filtered sample, obtain the calcium concentration according to
49.2, and magnesium concentration according to 50.2, and calculate the total
hardness according to the following formula:
H = (2.497~CC,)+(4.118~CM~)
where, H : total hardness (mgCaCO&

Cca : calcium concentration (mgCa/Z)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
56
K O101 : 1998
2.497 : coefficient, where the calcium amount is converted

to calcium carbonate equivalent (100.09/40.08)
C M g : magnesium concentration (mgMgl2)
4.118 : coefficient, where the magnesium amount is con-

verted to calcium carbonate equivalent (100.09/
24.305)
Remarks 2 As described in Remarks 1.
15.1.3 ICP atomic emission spectrometry
( i ) Concerning the filtered sample, obtain the concentration of calcium in accor-

dance with 49.3, further obtain the concentration of magnesium in accordance
with 50.3, and calculate total hardness from the formula of 15.1.2 (i).
15.2 Calcium hardness

( i ) Concerning the filtered sample, obtain the amount of 10 mmol/l EDTA solution
required for titration in 49.1 (21,and calculate the calcium hardness according
to the following formula:
Hc,=ax- loo0x 1.001

V
where, Hc,: calcium hardness (mgCaCOd2)
a : 10 mmol/Z EDTA solution required for titration in
49.1 (2)(ml)
V : sample (mi)
EDTA solution (mg)

(i) Concerning the filtered sample, obtain the calcium concentration according to
49.2, and calculate the calcium hardness according t o the following formula:
€€Cu = 2.497 x C'Ca
where, Hc, : calcium hardness (mgCaCOdZ)

Cca : calcium concentration (mgCa/Z)
2.497 : coefficient, where the calcium amount is converted
to calcium carbonate equivalent (100.09/40.08)

( i ) Concerning the filtered sample, obtain the concentration of calcium in accor-
dance with 49.3,and calculate the calcium hardness from the formula of 15.2.2 (i).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
57
K O101 : 1998
15.3 Magnesium hardness

(i) In 50.1 (2),obtain the amount of 10 mmol/Z EDTA solution required for titration
of calcium and magnesium, and in 49.1 (2),obtain the amount of 10 mmol/Z EDTA
solution required for titration of calcium, and then calculate the magnesium hard-
ness according t o the following formula:
HMg= 1"v -
Vca
x 1 O00 x 1.001
where, H M:~magnesium hardness (mgCaCOdZ)

b : 10 mmol/Z EDTA solution required for titration in
50.1 (2) (ml)
V : sample in 50.1 (2)(mi)
a : 10 mmol/Z EDTA solution required for titration in
49.1 (2) (mi)
VC,: sample in 49.1 (2) (ml)
1.001 : calcium carbonate equivalent t o 1 ml of 10 mmoVZ
EDTA solution (mg)
(1) Concerning the filtered sample, obtain the concentration of magnesium accord-
ing t o 50.2, and calculate magnesium hardness according t o the following for-
mula:
HMg = 4.118 x CMg
where, H M:~magnesium hardness (mgCaCOdZ)
C M: ~concentration of magnesium (mgMg/Z)
4.118 : coefficient, where the amount of magnesium is con-
verted to calcium carbonate equivalent ( 100.091
24.305)
(1) Concerning the filtered sample, obtain the concentration of magnesium in ac-
cordance with 50.3, and calculate the hardness of magnesium from the formula
of 15.3.2 (i).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
58
K 0101 : 1998
16 Suspended matters and evaporation residues Suspended matters in wa-

ter and evaporation residues of water shall be tested by dividing as follows:
Suspended matter The substance remaining on the filter, when the sample
is filtered.
Total evaporation residue The remaining substance when the sample is evapo-
rated to dryness.
Soluble evaporation residue The remaining substance when the filtrate, after
the suspended matter is filtered off, is evaporated t o dryness.
Ignition residues The residues obtained by igniting suspended matters, to-

tal evaporation residues, and soluble evaporation residues a t (600+25) OC for
30 min respectively, and are indicated as ignition residue of respective matters.
Ignition loss The reduced amounts of weight when measuring ignition resi-
due in (4), and are indicted as respective ignition losses.
The test shall be carried out immediately after sampling. When it is impos-
sible, the sample shall be preserved in a dark place at O to 10 OC, and the test
shall be carried out as soon as possible.
16.1 Suspended matter The sample is filtered and the substance remained on
the filter is dried at 105 t o 110 "C to measure the mass.
(i) Apparatus The apparatus shall be as follows.

(a) Filter (separate type) An example of a filter is shown in Fig. 16.1.
Filter in detail
A: Upper filter
funnel
B: Filter medium
C: Filter medium
holder
D: Lower filter
funnel
To pressure E: Rubber stopper
reduction F: Metal clamp
device
G: Suction flask
-- I_ -
--
- __.
Fig. 16.1 An example of filter (separate type)
59
K 0101 : 1998
(b) Filter medium Glass-fibre filter paper, organic filter membrane or metal
filter membrane with pore diameter of 1 pm and diameter of 25 t o 50 mm.
Remarks 1 Though glass fibre filter paper is seldom clogged, glass fibre
may be occasionally separated therefrom. Organic filter mem-
branes are different in chemical resistance and heat resistance
depending on the kind, therefore, cares shall be taken.
(2) Operation Carry out the operation as follows,

(a) In the case where glass fibre filter paper is used, preliminarily attach it t o
a filter, and wash sufficiently with water by suction. Thereafter, place this
filter medium(') on a watch glass(2))heat a t 105 t o 110 "C for approx. 1h(3),
and allow t o stand t o cool in a desiccator. Thereafter, weigh its mass.
(b) Attach the filter medium to filter, pour an adequate amount(5) of the sample(*)
into the filter t o filter by suction. Wash down the substances adhered t o
the walls of the sample container and filter funnel on the filter medium
with water, combine it with the remaining substances on the filter medium,
and wash it with water several times.
(c) Remove the residual substances together with filter medium carefully by
means of a pincette or the like from the filter, transfer on the watch glass
used in (a), dry a t 105 t o 110 " C for 2 h t o allow t o cool in the previous
desiccator, and then weigh the mass.
(d) Calculate the suspended matters (mgll) according to the following formula:
1O00
s = ( a- b )x -V
where, S : suspended matter (mgíl)
a : mass of filter medium containing the suspended
matter and watch glass (mg)
b : mass of filter medium and the watch glass (mg}
V : sample (ml)
Notes (1) Glass fibre filter paper, organic filter membrane, and metal fi-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
bre membrane treated by binders such as acrylic resin or the like

may not be washed.
(2) Use a lighter watch glass as far as possible o r use a light con-
tainer of aluminium foil or the like.
(3) There are some organic filter membranes which deform when
heated a t 105 to 110 O C . In this case heat at 90 OC.
(4) Use the sample passed through 2 min sieve, shake and mix suf-
ficiently to make the suspended matters uniform, and thereaf-
ter take i t quickly.
(5) Take the sample so that the weight of suspended matter after
drying becomes not less than 2 mg.
Remarks 2 Where the sample is difficult to filter, take an adequate amount
of it into a beaker, shake and mix sufficiently every time, add
immediately before the completion of filtering the liquid each
60
K O101 : 1998
time, and stop the addition of the sample when the filtering
speed has become extremely slow. Obtain the quantity of the
sample used from the remainder in the beaker.
3 For the sample containing adhesive suspended matter to the
sample container, take a proper amount of the sample and use
the whole amount t o test.
Rub off the suspended matter adhered to the walls of the
sample container by a policeman (glass rod with rubber tube)
or the like t o collect on the filter medium.
4 For determining ignition residue in the suspended matter, use
organic filter membrane.
5 I n the sample containing oils and fats, grease, wax, etc., for
measurement of suspended matter excluding these contents,
filter the sample and dry together with a filter funnel with-
out removing the filter medium. Attach these to the suction
bottle, and thereafter pour each 10 ml of hexane several times
t o remove oils and fats by washing. Consecutively, carry out
the procedure in ( c ) .
16.2 Total evaporation residue

(a) Evaporating dish Platinum, quartz glass or porcelain evaporating dish.
(a) Heat the evaporating dish made of adequate material(6) a t 105 to 110 "C
for approx. 1h, and then allow t o cool in a desiccator t o measure the mass.
Repeat heating, cooling and weighing until the mass becomes constant.
(b) Take a proper amount (7) of the sample (4) into an evaporating dish(8), and
evaporate carefully to dryness (9).
(c) Heat this evaporating dish at 105 to 110 "C for approx. 2 h, and then allow
to cool in the previous desiccator to measure the mass.
(d) Calculate the amount of the total evaporation residue (mg/Z>according to

the following formula:
1O00
R = ( a - b)X -
V
where, R : total evaporation residue (mgll)
a : mass of evaporating dish containing the residue
(mg)
b : mass of the evaporating dish (mg)
V : sample (mi)
Notes (6) Select the material quality of the evaporating dish t o be used
according t o the property of the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
61
K O101 : 1998
For example, in the case of the sample containing chloride

ion and strong oxidizing substance together, the platinum dish
shall not be used.
(7) Take an aliquot so that the mass of total evaporation residue
after drying becomes not less than 5mg.
(8) Where whole sample cannot be accommodated into an evaporat-
ing dish, put it in by dividing it several times.
(9) Use a hot-plate, boiling water bath, infrared lamp, o r the like
for evaporating to dryness so as not to boil during evaporation.
Furthermore, be careful of contamination from the environ-
ment.
7 For determination of ignition residue in succession, use the
evaporating dish preliminarily made a constant mass by igni-
tion a t (600f25)OC.
16.3 Soluble evaporation residue
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Evaporating dish As described in 16.2 (1)(a).
(b) Filter (separate type) As described in 16.1 (i)(a).
(c) Filter medium As described in 16.1 (i)(b).

(a) Attach a filter medium t o a filter, and filter the sample.
(b) For filtrate, operate in accordance with 16.2 (2) (a) t o (d), as appropriate.
Remarks 8 As described in Remarks 1 and 4.
16.4 Ignition residue
16.4.1 Ignition residue of suspended matter

(1) Reagent The following reagents shall be used.
(a) Ammonium nitrate solution (250 gll) Dissolve 25 g of ammonium ni-
trate specified in JIS K 8545 in water t o make 100 ml.

(a) Crucible Platinum or porcelain crucible, 10 t o 20 ml.
(b) Electric furnace The furnace capable of regulating t o (600I25) O C .
(a) Transfer the suspended matter obtained in 16.1 (2)(e) together with the
filter medium into a crucible made a constant mass by ignition at (600I25) O C .
Moisten it by dripping ammonium nitrate solution (250 g/Z)(lO), put it into
62
K O101 : 1998
the electric furnace gradually raising temperature, heat at (600I25) O C for

approx. 30 min to incinerate, and after allowing to cool in a desiccator, weigh
the mass.
(b) Calculate the ignition residue (mg/Z) according t o the following formula:
1 O00
R = (a- b) X -
V
where, R : ignition residue of the suspended matter (mg/Z)
a : mass of the crucible containing ignition residue
(mg)
b : mass of the crucible (mg)
V : sample (ml)
Note (10) In the case where organic filter membrane is nitro compound, it
should be preferable that after moistening by dripping 2-propanol
specified in JIS K 8839, it is incinerated by ignition.
Remarks 9 In the case where glass fibre filter paper is used as a filter
medium, preliminarily heat another glass fibre filter paper of
the same form in an electric furnace at (600k25) OC, obtain a
blank test value, and correct the mass of the glass fibre filter
paper.
16.4.2 Ignition residue of total evaporation residue

(a) For the total evaporation residue obtained in 16.2 (2)(e), operate in accor-
dance with the operation of 16.4.1 (31,as appropriate.
16.4.3 Ignition residue of soluble evaporation residue

(i) Operation Carry out the operation as follows.
(a) For the soluble evaporation residue obtained in 16.3 ( Z ) , operate in accor-
dance with the operation of 16.4.1 (31,as appropriate.
16.5 Ignition loss

(i) Operation Carry out the operation as follows.
(a) Ignition losses of suspended matter, total evaporation residue and soluble
residue shall be calculated according t o the following formula:
E=S-R
where, E : respective ignition losses (mg/Z)
S : respective evaporation residues (mg/Z)
R : respective ignition residues (mg/Z)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
63
K O 1 0 1 : 1998
17 Oxygen demand by potassium permanganate at 100 OC (CODMn) The sample

is made acidic with sulfuric acid, and potassium permanganate is added as oxidiz-
ing agent. The solution is reacted on a boiling water bath for 30 min, then the amount
of permanganate consumed a t that time is obtained t o express by corresponding amount
of oxygen (mgOlZ).
The test shall be carried out immediately after sampling. If it is impossible, the
sample shall be preserved in accordance with 3.3, and shall be tested as soon as
possible.
Determination range: CODMn 0.5 to 11 mgû/z

Water Water A4 specified in JIS K 0557(l)(2).
Sulfuric acid (1+2) Gradually add 1 volume of sulfuric acid specified in
JIS K 8951 t o 2 volumes of water by stirring. Successively add potassium
permanganate solution (3 g/Z) until the colour of the solution becomes light
pink.
Silver nitrate solution (200 gld) Dissolve 20 g of silver nitrate specified
in JIS K 8550 in water to make 100 ml, and put it into a coloured glass
bottle t o be preserved.
Sodium oxalate solution (12.5 mmol/Z) Dissolve 1.8 g of sodium oxalate
specified in JIS K 8528 in water t o make 11. However, prepare the solu-
tion of molar concentration slightly higher than 2.5 times that of 5 mmolll
potassium permanganate solution in (e).
5 mmolld potassium permanganate solution Take 0.8 g of potassium
permanganate specified in JIS K 8247 into a flat-bottomed flask, and add
1 050 t o 1 100 ml of water to dissolve. Boil the solution for 1 t o 2 h gently,
and then allow to stand overnight.
Filter the supernatant liquid through a sintered glass filter G4 (without
washing with water before and after filtration). Put the filtrate into a coloured
glass bottle washed with steam for approx. 30 min t o preserve.
Standardization Preliminarily heat sodium oxalate of reference mate-
rial for volumetric analysis specified in JIS K 8005 a t 200 OC for approx. 1h,
leave it cool in a desiccator, then weigh out approx. its 0.42 g t o the near-
est 1mg, dissolve in a little amount of water, transfer into a 250 ml volu-
metric flask and add water up to the marked line. Take 25 ml of this solution
into an Erlenmeyer flask of 300 ml, dilute it with water to approx. 100 ml,
and add 10 ml of sulfuric acid (1+2).At a liquid temperature of 25 to 30 OC,
add 22 ml of this 5 mmoV2 potassium permanganate solution at once by means
of a burette and leave as it is until pink colour disappears. Heat to 55 t o
60 OC,and titrate with this 5 mmoVZ potassium permanganate solution. Take
the point when the slight pink colour of the solution has kept approx. 30 s
a s the end point.
Calculate the factor (f)(3) of 5 mmol/l potassium permanganate solution
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
64
K O101 : 1998
b 25 1
= a xÖ
¡ Õx 250 x x 0.001675
where, a : amount of sodium oxalate (g)

b : purity of sodium oxalate (%)
x : 5 mmolll potassium permanganate solution
required for titration (ml)
0.001 675 : sodium oxalate equivalent of 1ml of 5 mmolll
potassium permanganate solution (g)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
The water shall not contain the material which gives CODMn.
Verify the water by the procedure below.
Carry out the procedures (3)(a)t o (d) for 100ml of water.
The amount of 5 mmol/Z potassium permanganate solution con-
sumed for this titration is taken as “a ml”. Separately carry out
the procedures (a>to (d) excluding operation of heating for 100 ml
of water, and the amount of that consumed for titration as “bml”.
Obtain (a - b ) ml. If this value is from 0.15 t o 0.2, i t is proper,
and if beyond the value, organic substances are included in the
water (or reagents), and it is not suitable to use.
When water is distilled, make the water slightly acidic by adding
sulfuric acid (1+2), add potassium permanganate solution (3 gll)
to colourate, and distill. Provided that the colouration of perman-
ganic acid is maintained till the end of distillation.
The factor as near as 1 (0.95 to 1.05) should be used.

(a) Water bath The water bath having such a large heat capacity and heat-
ing ability that when the sample is put in, it is capable of maintaining boiling
condition. The water bath shall be provided with a metal net apart from
the bottom so that a 300 ml Erlenmeyer flask will not touch the bottom of
water bath directly.

Take a proper amount(5) of sample(4) in a 300 ml Erlenmeyer flask, and
add water t o make 100 ml. Then add 10 ml of sulfuric acid (1+2) and add
5 ml of silver nitrate solution (200 glZI(6) (7) while mixing by shaking.
Add 10 ml of 5 mmolll potassium permanganate solution, mix by shaking,
immediately put it into the boiling water bath(8)(9) and heat for 30 min(l0).
Take out from the water bath, add 1 0 m l of sodium oxalate solution
(12.5 mmol/Z), and mix by shaking t o react sufficiently(11).
Keep the liquid temperature a t 55 t o 60 O C , and titrate with 5 mmollZ po-
tassium permanganate solution until it appears slight pink colour.
Separately take 100 ml of water into another Erlenmeyer flask of 300 ml,
and carry out the operations of (a) to ( d ) ( 1 2 ) .
65
K O101 : 1998
(0 Calculate C O D M(mgOlZ)
~ according t o the following formula:
CO&, = ( a - b ) x f x -looo x 0.2

V
where, CO&,-, : oxygen consumption by potassium permanganate
a t 100 "C (mgOlZ)
a : amount of 5 mmol/Z potassium permanganate
solution required for titration (mi)
b : amount of 5 mmolll potassium permanganate
solution required for titration for test using wa-
ter (mi)
f : factor of 5 mmolll potassium permanganate solu-
tion
V : sample (ml)
0.2 : oxygen equivalent t o 1 ml of 5 mmolll potassium
permanganate solution (mg)
Notes (4) Where the sample contains suspended matter, sufficiently mix
by shaking t o make the sample homogeneous, and then take the
sample quickly.
(5) Such amount that after heating for 30 min the residue of 5 mmoU
potassium permanganate solution comes 4.5 to 6.5 ml. However,
where CODM,,of the sample is 11mgOll o r less, it shall be 100 ml.
The proper amount of the sample shall be determined by carrying
out the preliminary test according to (3)Operation.
Where the approximate value of C O D Mis~ known, the proper
amount (Vml) of the sample can be obtained according t o the
following formula:
1 O00 x 0.2
V = 4.5 (or 3.5 to 5.5) x
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
anticipated value of CO&, of sample (mgO/ I )
where, V : sampling amount of sample (ml)

4.5 (or 3.5 to 5.5) : anticipated amount 5 mmolíZ potassium perman-
ganate solution (mi)
0.2 : oxygen equivalent t o 1ml of 5 mmolll potassium
permanganate solution (mg)
Instead of silver nitrate solution (200 gil), silver nitrate powder
equivalent to the solution may be added.
Where chloride ion exists in the sample, add silver nitrate solu-
tion until it reaches the equivalent, and further add 5 ml. When
the amount of silver nitrate solution (200 glZ) is required 10 ml
or over due t o much amount of chloride ion, add excess 2 m l of
the equivalent by using silver nitrate solution (500glZ) or add
1g of silver nitrate powder excess of the equivalent, and further
add 5 ml of water.
The equivalent of silver nitrate (AgN03) t o 1g of chloride ion
is 4.8 g. 100 ml of general seawater [chloride ion (18 gil)] and
66
K O101 : 1998
the equivalent silver nitrate are 8.64g and the adding amount
becomes 9.6 g.
If many samples are put all at once, it is liable t o appear devia-
tions of heating time by the required period for adding opera-
tion of sodium oxalate solution (12.5 mmolíl) when being taken
out, as well as the boiling in the water bath ceases. Therefore,
they should be put at appropriate intervals as required.
A ring weight made of lead or iron shall be attached a t the neck
of Erlenmeyer flask to prevent it from toppling down.
In this procedure, keep the solution level of the sample in the
300 ml flask below the level of boiling water.
Occasionally the reaction requires rather longer period of time
due to mixing of manganese (IV) oxide in the silver chloride.
Also when adding 5 ml or more of silver nitrate solution (200 gL)
t o the sample much content of chloride ion, use 5 ml of silver nitrate
solution (200 gll) in the procedure.
Remarks : Instead of 5 ml of silver nitrate solution (200 glZ), 1g of silver
sulfate powder ground well in an agate mortar may be added.
Also 1g of silver sulfate powder is used in ( e ) . In the sample
for much content of chloride ion, add further 1g to the amount
more than 10 % of the equivalent with chloride ion.
Silver sulfate equivalent t o 1g of chloride ion is 4.4 g, and
becomes as follows:
Adding amount of silver sulfate = [chloride ion (g) x 4.4 x
1.1+ 11 (g) = [chloride ion (g) x 4.84 + 11 (g). General seawater
becomes 9.7 g for 100 ml of [chloride ion (18 gíZ)].
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
67
K O101 : 1998
18 Oxygen demand by potassium dichromate (CODcr) Potassium dichromate

and sulfuric acid are added to the sample, after boiling for 2 h with a reflux con-
denser attached, the amount of consumed dichromate shall be obtained and expressed
by the corresponding amount of oxygen (mgO/E).
The test shall be carried out immediately after sampling. If it is impossible t o
carry out immediately, the sample shall be preserved in accordance with 3.3, and
the test shall be carried out as soon as possible.
Water Water A4 specified in J I S K 0557.
Silver sulfate-sulfuricacid solution Dissolve 11g of silver sulfate speci-
fied in J I S K 8965 in 1I of sulfuric acid specified in J I S K 8951. It will
take 1 to 2 days to dissolve completely (it may be dissolved by heating).
Mercuric sulfate (II) As specified in J I S K 8980.
Potassium dichromate solution -mollZ

(Go
sium dichromate specified in JIS K 8517, and dissolve in water to make
1 Weigh out 1.23 g of potas-
11.
1,lO-phenanthrolineiron (II) solution Dissolve 1.48 g of 1,lO-phenan-
throline (o-phenanthroline) monohydrate specified in JIS K 8789(1) and 0.70 g
of iron (II) sulfate heptahydrate specified in J I S K 8978 in water to make
100 ml.
25 mmol/Z ammonium iron (II) sulfate solution(2) Dissolve 10 g of am-
monium iron (II) sulfate t ammonium ferrous sulfate) hexahydrate speci-
fied in JIS K 8979 in approx. 500 ml of water, add 20 ml of sulfuric acid.
After cooling, dilute with water to 1I, Standardize a t the time of use.
Standardization Preliminarily grind potassium dichromate of volumet-
ric analysis standard reagent specified in JIS K 8005 with an agate mor-
tar, heat a t 150 "C for approx. 1h, and allow to stand to cool in a desiccator.
Weigh out 0.246 g of 100 % KLh-207 to the nearest 1mg, dissolve in a small
quantity of water, transfer to a 200 ml volumetric flask, and add water to
the marked line. Take 20 ml of this solution into a 300 ml Erlenmeyer flask,
add water t o make approx. 100 ml, and add 30 ml of sulfuric acid specified
in J I S K 8951. After cooling, add 2 to 3 drops of 1,lO-phenanthroline iron
(II) solution as an indicator, titrate with this 25 mmol/Z ammonium iron (II)
sulfate solution, and take the point when the colour of solution turns from
blue green to a red brown colour as the end point. Calculate the factor (f)
of 25 mmoVZ ammonium iron (II) sulfate solution from the following formula.
b 20 1
= a x ?ÕÖx %Ö
x x 0.001226
where, a : quantity of potassium dichromate (g)
b : content of potassium dichromate (%)
x : 25 mmol/Z ammonium iron (II) sulfate solution
0.001 226 : potassium dichromate equivalent to 1ml of
25 mmol/Z ammonium iron (II) sulfate solution (g)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
68
K 0101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Notes (1) In the case where chloride 1,lO-phenanthrolinium monohydrate

(1,lO-phenanthroline chloride) specified in JIS K 8202 is used,
take 1.78g.
(2) 1ml of 25 mmol/Z ammonium iron (II) sulfate solution corresponds
(A
t o 1ml of potassium dichromate solution -mol/Z .
)
Apparatus The apparatus shall be as follows.
(a) Reflux condenser 300 mm in length Liebig condenser o r Allihn condenser
with interchangeable ground-glass joint.
(b) Round bottom flask or Erlenmeyer flask 250 t o 300 ml in capacity,

and of interchangeable ground-glass joint with reflux condenser in (a).
(c) Hot plate or mantle heater

Take a proper amount(4) of sample(3) in a 250 ml round bottom flask or
Erlenmeyer flask containing 0.4 g of mercuric sulfate (II)(5), dilute with water
t o 20 ml, and then mix with shaking sufficiently.
Add 10 ml of potassium dichromate solution
[2:0 1
-mol/l , further add
of silver sulfate-sulfuric acid solution while mixing with shaking carefully,
30 ml
and then put several boiling tips into it.

Attach a reflux condenser t o the flask, and heat for 2 h.
After cooling, wash the condenser with approx. 10 ml of water, then trans-
fer the washings into the flask, and further dilute with water t o approx.
140 ml.
Add 2 t o 3 drops of 1,lO-phenanthroline iron (II) solution as indicator, and
titrate the excess of dichromate with 25 mmol/l ammonium iron (II) sul-
fate solution until the colour of solution turns from bluish green t o reddish
brown, and take the said point as an end point.
Separately, take 20 ml of water, and carry out the operation specified in
(4 t o (e>.
Calculate CODcr (mgO/Z) according to the following formula:
CODcr = ( b - a)x f x x o .2
V
where, CODcr : oxygen demand by potassium dichromate (mgO/Z)
a : 25 mmol/l ammonium iron (II) sulfate solution
b : 25 mmol/¿ ammonium iron (II) sulfate solution
required for titration for test using water ( m i )
f : factor of 25 mmoM ammonium iron (II) sulfate
solution
69
K 0101 : 1998
V : sample (ml)
0.2 : oxygen equivalent to 1ml of 25 mmol/Z ammonium
iron (II) sulfate solution (mg)
Notes (3) Where suspended matters are contained, take separately the
sample quickly after being mixed homogeneously by sufficiently
shaking.
(4) The amount such as about half of the potassium dichromate so-
lution added first remains after boiling for 2 h.
(5) Though masking of 40mg of chloride ion is carried out, where
the concentration of chloride ion is high like seawater, this method
shall not apply because obstructions can not be removed.
Remarks : Since mercury compound is used in this method, cares shall be
particularly taken on the treatment of waste water after the
test.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
70
K O 1 0 1 : 1998
19 Biochemical oxygen demand (BOD) The biochemical oxygen demand means

the amount of dissolved oxygen consumed by aerobic microorganisms living in wa-
ter. Dilute the sample with dilution water, and the amount shall be obtained from
the amount of dissolved oxygen t o be consumed when the sample is left as it is a t
20 "C for 5 days.
This test shall be carried out immediately after sampling, and when it is impos-
sible to carry out the test immediately, the sample shall be preserved in accordance
with 3.3 and shall be tested as soon as possible.
Water Water A3 Specified in JIS K 0557(1).
Buffer solution (pH 7.2) (solution A) Dissolve 21.75 g of dipotassium
hydrogenphosphate specified in JIS K 9017,8.5 g of potassium dihydrogen-
phosphate specified in JIS K 9007, 44.6 g of disodium hydrogenphosphate
12-water specified in JIS K 9019, and 1.7 g of ammonium chloride specified
in JIS K 8116 in water to make 1 O00 ml. pH of this buffer solution shall
be 7.2.
Magnesium sulfate solution (solution B) Dissolve 22.5 g of magnesium
sulfate heptahydrate specified in JIS K 8995 in water t o make 11.
Calcium chloride solution (solution C) Dissolve 27.5 g of calcium chloride
(anhydrous) specified in JIS K 8123 in water t o make 1Z.
Iron (III) chloride solution (solution D) Dissolve 0.25g of iron (III)
chloride (ferric chloride) hexahydrate specified in JIS K 8142 in water t o
make 11. Prepare it a t the time of use.
Hydrochloric acid (1+11) Prepare by using hydrochloric acid Specified
in JIS K 8180.
Sodium hydroxide solution (40 g/Z) Dissolve 4 g of sodium hydroxide
specified in JIS K 8676 in water t o make 100 ml.
Sodium sulfite solution (12.5 mmol/Z) Dissolve 1.6 g of sodium sulfite
(anhydrous) specified in JIS K 8061 in water to make 11. Prepare it at
the time of use.
Potassium iodide Potassium iodide specified in JIS K 8913.
Dilution water Adjust the water temperature to near 20 O C , and add 1ml
each of solutions of A, B, C and D to 1 1 of water(2) saturated with dissolved
oxygen by aeration. pH of this solution is 7.2 [When this solution does not
indicate pH 7.2, adjust to pH 7.2 by use of hydrochloric acid (1+11) o r so-
dium hydroxide solution (40gll).]. Confirm ( 3 ) preliminarily that the differ-
ence of amount of dissolved oxygen of the dilution water between the
beginning and 5 days later, when being bottled in an incubation bottle and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
left in a thermostatic bath at 20 "C for 5 days, is not more than 0.2 mgOlZ.
Seed liquid Use the supernatant liquid of sewage (4) (51, river water(6),
soil extracts(7), etc.
Seeded dilution water(8) At the time of test, prepare the seeded dilu-
tion water by adding a proper amount(9) of seed liquid to the dilution wa-
ter.
71
K O101 : 1998
Notes (1) The water shall be refined in distillation apparatus made of quartz
glass o r borosilicate glass-1.
(2> I t is preferable that water is saturated with dissolved oxygen by
passing air from which contaminants are removed by washing.
Air is washed by the following method.
Filter air with an activated charcoal filter [for instance, fill a
gas drying tower (300 mm) with granulated active carbon]. Then,
wash with potassium permanganate solution (5 gll) made acidic
by sulfuric acid, and further wash with potassium hydroxide so-
lution (250 g/Z).
Because the biochemical reaction is different according t o the con-
centration of organic substances and the kind of microorganism
contained, it is difficult t o correct by carrying out the blank test
on dilution water. Therefore, use the dilution water of which
oxygen demand for 5 days is 0.2mgOll or less.
The domestic sewage is often used as seed liquid. After allowing
the fresh raw sewage t o stand at 20 "C (or room temperature) for
24 to 36 h, its supernatant liquid is used. It is not desirable t o
use the sewage containing many nitrifying microorganisms (which
oxidize ammonium ion and nitrite ion) and such fresh sewage that
has not reached a sufficient biochemical equilibrium.
For the sample which does not indicate normal BOD when sew-
age is used as seed liquid, soil extract or the incubated liquid
obtained by the adaptation to the sample in a laboratory shall
be used.
Good results may be obtained when the downstream water at
500 t o 1O00 m from the discharge point of the river always re-
ceiving the discharge of this sample is used. Even if the sub-
stance harmful t o biochemical reaction coexists in the sample,
the waters of rivers, lakes o r swamps which are receiving the
discharge of the sample often contain microbial population re-
sistant t o them.
Add about 200 g of soil (where plants are growing) in 2 I of wa-
ter and stir it t o mix, Then use the supernatant liquid.
In the case where no aerobic microorganism, nor bacteria exists
or their population is small, use the seeded dilution water.
Where seeded dilution water is used for the test of BOD, carry
out the correction (seed correction) to the seed liquid used for
preparation of seeded dilution water according t o the following
operation:
Prepare the diluted seed liquids in several stages by diluting
the seed liquid properly with dilution water, and measure dis-
solved oxygen in parallel to the diluted sample. Taking the amount
of dissolved oxygen before incubation of diluted seed liquid as
Bi and that after leaving 5 days as Bz, select that (Bi-Bz) x 100
B,
of which is in the range of 40 t o 70 %, and use ( B I- B 2 ) x f as
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
72
K O 1 0 1 : 1998
the seed correction value [refer to the calculation formula of BOD

specified in (4)(d)3. Do not carry out the correction by obtain-
ing the amount of consumption of dissolved oxygen of seeded
dilution water for 5 days.
(9) Add the seed liquid so that BOD of seeded dilution water be-
comes 0.6 t o 1mgOlZ to ensure the normal activity of microor-
ganisms. The seed liquid is, ordinarily, for 1 1 of dilution water,
5 to 10 ml supernatant liquid of sewage, 10 to 50 ml in river water,
and 20 t o 30 ml in soil extracts.

Incubation bottle A 100 to 300 ml narrow mouth bottle with ground stop-
per having a correctly known capacity, the ground stopper of which is cut
off obliquely. An example is shown in Fig. 19.1.
-~
Fig. 19.1 An example of incubation bottle

Thermostat (incubator specified in JIS T 1702) The thermostat capable
of regulating the temperature to (20+1)OC. In order to prevent carbon dioxide
assimilation (carbonic acid assimilation) by algae in the diluted sample,
the sunlight shall be excluded. The thermostatic water bath of similar
specification may be used.
(3) Pretreatment of sample The pretreatment of a sample shall be carried o u t

as follows.
Where the sample contains acids and alkalis, oxidizing substances such as
residual chlorine, and supersaturated dissolved oxygen o r dissolved gas, the fol-
lowing pretreatment shall be carried out.
Further, where the liquid amount increases by pretreatment, the results shall
be corrected on the increment.
(a) Sample containing alkali or acid Add hydrochloric acid (1+11)o r so-
dium hydroxide solution (40 gll) t o adjust pH of the sample to about 7.
(b) Sample containing oxidizing substance such as residual chlorine
or the like Preliminarily add 0.1 g of sodium azide specified in JIS K
9501 and 1 g of potassium iodide specified in JIS K 8913 to 100 ml of the
sample. After shaking t o mix, add hydrochloric acid (l+l) t o make
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
73
K O101 : 1998
approx. 1pH, and allow to stand in a dark place for several min. Titrate
the liberated iodine with sodium sulfite solution (12.5 mmol/Z) using the
starch solution as indicator until the blue colour disappears. Separately
take the same amount of sample, and after reducing residual chlorine by
adding the calculated amount of sodium sulfite solution (12.5 mmol/l) ob-
tained from the previous titration value, make its pH approx. 7, if required,
by use of sodium hydroxide solution (40 g/Z) o r hydrochloric acid (l+ll).
(cl Sample supersaturated with dissolved oxygen or dissolved gas If
the temperature of the sample of treated water, river water, etc. sampled
during the winter season is 20°C or under, the dissolved oxygen and the
dissolved gas are liable to be supersaturated when the sample is made 20 O C .
Further in the case of river water and swamp water containing a large
amount of algae, oxygen is generated by carbon assimilation, and there-
fore the dissolved oxygen is liable to be supersaturated. Because in these
samples, the dissolved oxygen becomes gas during BOD measurement, and
is apt t o be lost outside the incubation bottle to cause incorrect results, it
is necessary to reduce preliminarily the amount of dissolved oxygen and
dissolved gas t o near the saturation amount a t 20 "C by means of stirring
or aeration.
Remarks 1 Aerobic bacteria which decompose carbonic organic substances
and nitrifying bacteria which oxidize (nitrify) nitride such as
ammonium occasionally increase in a biochemically treated
sample. In such a sample, sum total amount of oxygen de-
mand by acidic decomposition of organic substances and that
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
by nitrifying nitride such as ammonium is measured. The
oxygen demand by this nitrifying does not correspond t o the
amount of nitride in the sample, but varies by the number of
nitrifying bacteria.
In order to measure the oxygen demand under the state
wherein nitrifying action is restrained, the procedure below
shall be carried out:
At the preparation of diluted sample in (4) (a),add 2 mg of
N-(2-propenyl) thiourea (*) o r 10 mg of 2-chloro-6-(trichloro-
methyl) pyridine powder(Z*)in per 1Z of the diluted sample.
Notes (*) Add 2 ml of N-(2-propenyl) thiourea (N-allylthiourea) solution
(1 mg/ml) [Dissolve O , 1g of N-(Z-propenyl)thiourea in water t o
make 100 ml. Preserve it in a dark place a t O t o 10 OC.].
(2*) 2-chloro-6-(trichloromethyl) pyridine is difficult t o be dissolved
in water so that powdered one is added. After adding, it is not
sufficiently dissolved and some part may be floated, therefore,
attention should be paid when it is transferred into the incuba-
tion bottle. There are ones dissolvable in water mixing with
other reagents, which may be used.
(4) Operation The operation shall be carried out as follows,
(a) Preparation of diluted sample Take dilution water o r seeded dilution

water in 1 O00 ml measuring cylinder with ground stopper (in the case of
200 ml or more incubation bottle, use a 2 O00 ml measuring cylinder with
74
K 0101 : 1998
ground stopper) up to approx. a half by means of syphon, taking care not

to introduce air bubbles. Next, add a proper amount of pretreated sam-
ple(l*)(11) (12), and add dilution water or seeded dilution water up to the
marked line of 1O00 ml (in the case of 2 O00 ml measuring cylinder with
ground stopper, up to the marked line of 2 O00 ml). Stopper and mix gen-
tly. Perform the same operation with the amount of sample changed, or
further dilute this solution stepwise by the similar operation t o prepare 4
t o 5 kinds of different dilution multiples(l3) (14) (15).
For each kind of diluted sample prepared, prepare 2 t o 4 incubation bottles,
transfer the diluted sample by use of a syphon into them, and after filling
sufficiently, stopper tightly.
Use one out of each set of incubation bottles different in dilution mul-
tiples for determination of dissolved oxygen prior t o incubation, and put
the other bottles into a thermostatic vessel o r thermostatic water bath
adjusted a t (20i11)"C to incubate for 5 days(l6) (17).
Measurement of dissolved oxygen amount of diluted sample before
incubation Allow the diluted sample to stand for 15 min after prepara-
tion(18), and determine the dissolved oxygen in accordance with 24.2, 24.3
or 24.4. In the case of 24.3 or 24.4, the amount of dissolved oxygen may
be measured by use of the diluted sample remaining in the measuring cyl-
inder.
Measurement of dissolved oxygen after incubation Measure the
amount of dissolved oxygen of the diluted sample incubated for 5 days in a
thermostatic vessel o r water bath by the same method as in (b).
Calculation of BOD From the amount of dissolved oxygen before and
after incubation, calculate BOD (mgOll) of the sample according to the fol-
lowing formula (19):
i) When seeding is not carried out:
BOD =
(Dl -a)
P
ii) When seeded dilution water is used(19):
- (B1- Bz) x f
BOD = (Dl - 0 2 )
P
where, BOD : biochemical oxygen demand (mgOlZ)
Di : dissolved oxygen of diluted sample 15 min after
preparation (mgOlZ)
Dz dissolved oxygen of diluted sample after incu-
bation (mgOlZ)(20)
P : ratio of the sample in the diluted sample (sample/
diluted sample)
Bi : dissolved oxygen before incubation of the diluted
seed liquid a t the time of measuring BOD of the
seed liquid (mgOlZ)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
75
K O101 : 1998
Bz : dissolved oxygen after incubation of the diluted

seed liquid at the time of measuring BOD of the
seed liquid (mgOlZ)
X
f:-
Y
x : seed liquid i n the diluted sample at the time
of measuring BOD of the sample (%)
y : seed liquid in the diluted seed liquid at the
time of measuring BOD of the seed liquid (%)
The amount of consumption of the dissolved oxygen in the di-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
luted sample for obtaining normal BOD value of the sample, (Dl -
Dz), is in the range of 3.5 to 6.2 mgOlZ. If the amount of remaining
dissolved oxygen due to insufficient dilution is 1mgOlZ or less,
or, conversely, the amount of consumption of dissolved oxygen
in 2 mgOlZ or less due t o over dilution, the normal BOD value is
difficult to obtain.
If BOD of the sample can be anticipated from experience or oth-
ers, the aliquot of the sample t o be taken may be obtained as
fol10w s:
For example, the saturation amount of dissolved oxygen at
20 "C is 8.84 mgOlZ, its 40 % is approx. 3.5 mgOlZ, and its 70 %
is approx. 6.2 mgOIZ, therefore, in the case of making 12 by com-
bining the dilution water and the sample, the amount of the sample
to be taken separately (Vml) can be obtained according t o the
following formula:
(3.5 to 6.2) x 1000
V=
anticipated BOD value of sample (mgOlZ)
where, V : aliquot quantity of sample (mi)
3.5 : 40 % equivalent of saturated quantity
(8.84 mgO/Z) of dissolved oxygen at 20 "C
6.2 : 70 % equivalent of saturated quantity
(8.84 mgOlZ) of dissolved oxygen at 20 "C
Where the BOD value of the sample is 5 mgOlZ or less, the
amount of sample to be taken separately shall be 800 ml o r more,
and where the dissolved oxygen is not sufficiently contained, the
test shall be carried out after aeration.
Where the sample contains suspended matters, a proper amount
shall be taken after mixing to combine the suspended matters
homogeneously.
If the preparation of the diluted sample of higher dilution mul-
tiples in succession on the base of the diluted sample remaining
in the measuring cylinder is continued, the labour and time can
be saved.
In the case of diluting 100 times o r more, do not dilute at one
time. Preliminarily, take 50 to 100 ml of the sample into other
76
K O101 : 1998
1 O00 ml measuring cylinder with ground stopper, dilute with

dilution water or seeded dilution water to 1 O00 ml, and prepare
the diluted sample specified in (a>using this diluted sample.
In the case of the sample of BOD value not more than 100 mgO/Z,
it may be directly diluted in an incubation bottle according to the
following method:
Prepare 4 incubation bottles of correctly known capacity, put
preliminarily about half amount of dilution water or seeded di-
lution water into respective bottles, then add the sample of the
calculated amount corresponding to the capacity of the bottle
according to the dilution multiples, and further fill the space in
the bottle with dilution water o r seeded dilution water. Take
care not to introduce air bubbles during this operation.
In the case of using thermostatic water bath, immerse the whole
incubation bottle into the water.
When the incubation bottle is placed with water sealed in a ther-
mostatic vessel, the sealing water evaporates, and, therefore, it
shall be refilled at times.
Where the reducing substances such as sulfides, sulfites, and iron
(II) coexist, the Immediate Dissolved Oxygen Demand for 15 min
(IDOD) shall be distinguished from BOD. For obtaining IDOD,
the following operation shall be carried out:
After preliminary measurement of the dissolved oxygen of the
sample and the dilution water, dilute the sample with the dilu-
tion water at a definite ratio and allow t o stand for 15 min. Then
measure the dissolved oxygen ( D I ) . Calculate the dissolved oxy-
gen (Dc)in the diluted sample from the respective dissolved oxy-
gen (mgOlZ) in the sample and the dilution water preliminarily
measured, and then calculate IDOD (mgOlZ) of the sample ac-
cording to the following formula:
where, IDOD : oxygen demand for 15 min (mgO/Z)

Dc : dissolved oxygen in diluted sample water before
incubation (mgOIZ) = (S x P ) + (Do
xp )
S : dissolved oxygen in the sample (mgO/Z)
Do : dissolved oxygen in dilution water
ímgO/Z)
p : ratio of the dilution water in the diluted
sample (dilution wateddiluted sample)
P : ratio of sample in the diluted sample (sample/
diluted sample)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Di : dissolved oxygen of diluted sample left for 15 min

after preparation (mgO/Z)
77
K 0101 : 1998
(19) The calculation may be carried out according t o the following

method:
BOD = (Dl
-Da)
x nl -(Bi - B a ) xnax
v x ( n - 1)
1
100
where, D I: dissolved oxygen of diluted sample 15 min after
preparation (mgOlZ)
D2 : dissolved oxygen of diluted sample after incu-
bation (mgO/Z)
ni : dilution multiple of diluted sample
diluted sample
sample 1
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
n2 : dilution multiple a t the time of measuring BOD

of the seed liquid
diluted seed liquid in measuring
BOD of seed liquid (ml)
seed liquid in measuring
BOD of seed liquid (mi) I
dissolved oxygen in the diluted seed liquid be-
fore incubation in measuring BOD of seed liq-
uid (mgOlZ)
dissolved oxygen in the diluted seed liquid af-
ter incubation in measuring BOD of seed liquid
t mgO/Z)
percentage of seed liquid in the seeded dilution
water (vol%)
Normally, take the amount as
0.6 x 100
V>
(Bl - &) x nz
(20) The diluted sample of which dissolved oxygen demand during 5
days, (Dl-Oz),is within the range of 3.5 to 6.2 mgOlZ o r of which
D D
value is within the range: - x 100 = 40 to 70 % shall be taken
D,
for the calculation of BOD. It is most desirable t o be near me-
dian of this condition. However, where BOD of the sample is
not more than 3.5 mgOlZ, the dissolved oxygen demand during 5
days, even when it is not diluted, does not become 40 % o r more
of saturation value of the dissolved oxygen. I n such a case, it
shall be calculated from that value.
Remarks 2 Method for preparation of seed liquid The incubation to
adapt microorganisms to the sample shall, preferably, be car-
ried out according to the following method:
Transfer 52 of the sample into the glass water tank
(approx. 6 I),and adjust pH to approx. 7 by using hydrochlo-
ric acid (1+11)or sodium hydroxide solution (40glZ). Then,
add 100 t o 300 ml of seed liquid such as the sewage, river water,
78
K O101 : 1998
etc. which contain a large amount of microorganisms and 10

t o 50ml of the buffer solution (solution A). After mixing by
stirring thoroughly, take a portion of it, and measure CODMn
or the amount of organic carbon. Then, aerate it for 24 t o
48 h continuously, thereafter take its portion again, and mea-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
sure CODMn o r the amount of organic carbon. If a remark-
able change is observed in the values measured before and after,
judge that the biochemical reaction is under progress in the
sample, and further continue the aeration to let the organ-
isms adapted t o the sample increase. If there is no such re-
markable change, take the sample separately, dilute with
dilution water suitably, then carry out the seeding the same
as the above, and aerate it for 24 to 48 h continuously. Then,
test the changes of CODM~, o r the amount of organic carbon
and the amount of suspended matters. If remarkable change
such as the decrease of CODMn, the decrease of amount of organic
carbon, or the increase of amount of suspended matters are
found as the results, it means that the biochemical reaction
is active. According to the composition of organic matters in
the sample, these operations shall be continued for one week
or longer.
Further, in the case where the procedures as mentioned above
with the sample diluted by 10 times or more with dilution water,
if a remarkable change in CODM, or the amount of organic
carbon is observed, it is also necessary t o increase the ratio
of the sample gradually. In this way, incubate microorgan-
isms adapted to the sample, and use it as seed liquid.
3 Method of confirming test operation The following method
is preferable to confirm the adequateness of using seed liq-
uid, seeded dilution water, etc. or test operation:
Transfer 5 to 10 ml of the standard mixture of glucose and
glutamic acid [take 150mg of D(+)-glucose specified in JIS
K 8824 and 150 mg of L-glutamic acid specified in JIS K 9047,
dissolve in water, then transfer into a 1O00 ml volumetric flask,
and add water up t o the marked line1 into a 300ml incuba-
tion bottle of correctly known capacity (where the capacity of
1
the incubation bottle is 100 ml, use - amount of the above-
3
mentioned), fill with seeded dilution water, then stopper tightly,
and measure BOD. BOD of this standard solution shall be
( Z Z O I l O ) mgOlZ. If the deviation from this value is remark-
able, the quality of the dilution water or the activity of the
seeding substances are doubtful.
4 If heavy metal elements such as copper, chromium, mercury,
silver, arsenic, etc. are dissolved in the sample, the correct
value can not be obtained occasionally. In such a case, the
seeding substance well-adapted to these heavy metal elements
shall be incubated in accordance with Remarks 2.
79
K O101 : 1998
20 Organic carbon (TOC) The organic carbon means the carbon in the organic
substance existing in water. To this determination, combustion oxidation-infrared
system TOC analysis method and combustion oxidation-infrared system TOC auto-
matic measuring method apply.
This test shall be carried out immediately after sampling. If the test can not be
carried out immediately, preserve the sample in accordance with 3.3, and the test
shall be carried out as soon as possible.
20.1 Combustion oxidation-infrared system TOC analysis method A small

quantity of a sample together with air with carbon dioxide removed or oxygen is
injected into a high temperature tube for measuring total carbon. After oxidizing
carbon in organic substances and carbon in inorganic substances [inorganic carbon
(principally, carbonates)] t o carbon dioxide, its concentration is measured by a non-
dispersive infrared gas analyzer t o obtain the total amount of carbon.
Separately, the sample is injected into a tube for measuring carbon in inorganic
substances kept a t temperature for the organic substances not to be decomposed,
and the generated carbon dioxide is measured to obtain the amount of inorganic carbon.
The amount of organic carbon is calculated from the amount of total carbon by
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
subtracting that of inorganic carbon.
Determination: C 1 to 150 mgll,
Repeatability: 3 t o 10 % in coefficient of variation (different according t o the
apparatus and measuring conditions).
Water Water A3 or A4 specified in JIS K 0557(l)(2) ( 3 1 , and containing
no carbonic acid(4) shall be used. This water is used for the preparation
and operation of the reagent t o be used in this test. A blank test is carried
out in accordance with ( 5 ) and the adaptability of use shall be confirmed.
TOC standard solution (1 mgClml) Heat potassium hydrogen phthalate,
standard reagent for volumetric analysis specified in JIS K 8005 of 120 "C
for approx. 1h, and allow to stand t o cool in a desiccator. Take its 2.125 g,
dissolve in a small quantity of water, transfer to a 1O00 ml volumetric flask,
and add water to the marked line.
TOC standard solution (0.1 mgC/ml) Take 10 ml of TOC standard so-
lution (1mgC/ml) into a 100 ml volumetric flask, and add water t o the marked
line.
Inorganic carbon standard solution (1 mgC/ml) Allow sodium hydrogen-
carbonate specified in JIS K 8622 t o stand in a desiccator for approx. 3 h,
and take its 3.497 g. Separately, preliminarily heat sodium carbonate (an-
hydrous), standard reagent for volumetric analysis specified in JIS K 8005
at 600 "C for approx. 1h, allow to stand to cool in a desiccator, and take
its 4.412g. Dissolve both in a small quantity of water, transfer t o a 1O00 ml
volumetric flask, and add water t o the marked line.
Inorganic carbon standard solution (0.1 mgClml) Take 10 ml of in-
organic carbon standard solution (1mgC/ml) into a 100 ml volumetric flask,
80
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(0 Total carbon measuring tube Tube filled by catalyst for determination

of total carbon.
(g) Inorganic carbon measuring tube Tube filled by catalyst for determi-
nation of inorganic carbon.
(h) Carrier gas Air removed off carbon dioxide or oxygen specified in JIS K
1101.
Notes (1) Water of which the concentration of TOC is made as low as pos-
sible is used. Since when the refined water is preserved by putting
in a container, it is gradually contaminated and the concentra-
tion of TOC occasionally increases, it should be preferable t o use
immediately after refining.
(2) In order t o decrease the concentration of TOC as low as possible,
take demineralized water or distilled water into a distillation flask,
drip potassium permanganate solution (3 g/Z) until the solution
is coloured, add 2 t o 3 ml of sulfuric acid (l+l)per 1O00 ml of
water, and distill. (Make colouration of potassium permangan-
ate remain until the distillation is completed.) Discard initial
distillation content (corresponds to approx. one fifth of the quantity
of water in the distillation flask), and sample fraction correspond-
ing to three fifths of the middle.
(3) Water refined by suitably combining an ion exchange method, a
distillation method, a reverse osmosis method, an ultraviolet ir-
radiation method, an activated charcoal adsorption filtration
method, an ultrafiltration method, a precision filtration method,
etc. may also be used.
(4) I t is refined in 2 (12) (b).
(a) Microsyringe 20 t o 1501.11
(b) TOC analyzer
(3) Preparatory operation Carry out the preparatory operation as follows.
(a) Warm up the TOC analyzer.
(b) Inject the specified amount(5) (for example 20 pl) of TOC standard solution
(1mgC/ml) or TOC standard solution (0.1 mgC/ml) into a total carbon
measuring tube of the TOC analyzer with a microsyringe t o obtain the
indicating value (height of peak).
(c) Repeat the operation specified in (b) several times and confirm that the
indicating values are constant.
(d) After shaking the sample thoroughly t o mix homogeneously, inject the same
amount as in (b) into the total carbon measuring tube with a microsyringe
to obtain indicating value, and obtain the approximate total carbon con-
centration (mgCII) of the sample by comparing with (b).
Note ( 5 ) Where the concentration of carbon of the sample is low, the in-
jecting amount shall be 100 t o 150 p1. Further, where it is high,
the injecting amount shall be reduced o r diluted by a specified
multiple.
81
K 0101 : 1998
(4) Preparation of working curve Prepare the working curve as follows.

Take TOC standard solution (1mgC/ml) or TOC standard solution (0.1 mgC/
mi) into a 100 ml volumetric flask stepwise so that the approximate con-
centration of carbon of the sample obtained by (3)(d)becomes approximately
in the middle, and add water up t o the marked line.
Inject a definite amount [for example, the same amount as in (3) (b)]of TOC
standard solution prepared according t o (a) at the maximum concentration
into the total carbon measuring tube with a microsyringe and adjust the
sensitivity of TOC analyzer and the injection amount of the standard solu-
tion so that the indicating value becomes approx. 80 % of the maximum scale.
Inject the specified amount [the amount as Specified in (b)]of TOC standard
solution of each concentration prepared by (a)into the total carbon measur-
ing tube in succession with a microsyringe to obtain the indicating value.
Inject the same amount of water as in (e) for the blank test into the total
carbon measuring tube with a microsyringe, obtain the indicated values
and correct the results of (c). Prepare the relation curve between the amount
of organic carbon and the indicated value t o take it as the working curve
of the total carbon.
Using the inorganic carbon standard solution (1mgC/ml) or inorganic car-
bon standard solution (O. 1mgC/ml), prepare the inorganic carbon standard
solutions stepwise so as to contain the same amount of carbon as TOC stan-
dard solution prepared stepwise as in (a).
Inject the carbon standard solution of each concentration specified amount
[the amount as specified in (b)] of inorganic prepared as in (e) in succes-
sion into the inorganic carbon measuring tube with a microsyringe t o ob-
tain the indicted values.
Inject the same amount of water as in (f) for the blank test into the inor-
ganic carbon measuring tube with a microsyringe, obtain the indicated values
and correct the results of (f). Prepare the relation curve between the amount
of inorganic carbon and the indicated value to take it as the working curve
of the inorganic carbon.
(a) Where the suspended matters are contained in the sample, thoroughly mix
by stirring with a homogenizer or mixer to disperse them homogeneously.
(b) Inject the specified amount of the sample(5) [for example, the same amount
as in (4) (b)]into the total carbon measuring tube with a microsyringe t o
obtain the indicated value.
(c) Inject the Specified amount of the sample [for example, the same amount
as in (4) (f)]into the inorganic carbon measuring tube with a microsyringe
to obtain the indicated value.
(d) Where the sample is diluted, for the blank test in (b) and (e), take the same
amount of water with a microsyringe respectively, and carry out the re-
spective operations specified in (b) and (e) to correct the results obtained
on the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
82
K O101 : 1998
(e) Obtain the amount of total carbon and inorganic carbon in the injected sample
from the working curves of the total carbon and inorganic carbon prepared
preliminarily t o calculate respective concentrations (mgCD).
(f) Calculate TOC (mgC/E) of the sample according t o the following formula:
TOC = (Ct- CJ x d
where, TOC : organic carbon (mgC/Z)
Ct : total carbon in the injected sample (mgC/Z)
C; : inorganic carbon (mgC/Z)
d : dilution multiple of the injected sample
Remarks 1 In addition to the method to obtain organic carbon by subtract-
ing inorganic carbon from the total carbon, the following method
may be used: Preliminarily add the hydrochloric acid to the
sample t o make its pH 2 or under, and remove inorganic car-
bon by passing high purity nitrogen grade 2 specified in JIS
K 1107. Then, inject a small portion of the solution into the
high temperature total carbon measuring tube, and then de-
termine the carbon to take it as the amount of organic car-
bon. This method is suitable for the sample containing a
relatively large amount of inorganic carbon. However, where
volatile organic matters are contained, the error becomes larger.
2 For the method t o oxidize the organic carbon t o carbon diox-
ide by TOC analyzer, there is the wet oxidizing method by use
of ampoule other than the combustion method. In this wet
oxidizing method, take 3 t o 10ml of sample in a glass am-
poule, and add potassium peroxodisulfate and phosphoric acid
or potassium dichromate and phosphoric acid t o make the
solution acidic. Thereafter, pass oxygen sufficiently to remove
carbon dioxide. After meltsealing the ampoule, heat for a speci-
fied period of time in an autoclave to oxidize the organic matters.
Break the ampoule in the apparatus, pass carbon dioxide with
generated nitrogen to introduce into the carbon dioxide mea-
suring part.
3 For the determination of carbon dioxide generated, other than
infrared analysis method, thermal conductivity measuring
method is used.
20.2 Combustion oxidation-infrared system TOC automatic measuring

method Adjusting pH to 2 or less by adding acid to a sample supplied continuously
to a measuring instrument. After removing inorganic carbon by aeration, feed its spe-
cific quantity together with a carrier gas into a high temperature total carbon mea-
suring tube, oxidize carbon in organic substances to carbon dioxide, and measure its
concentration with a non-dispersive infrared gas analyzer to obtain the concentration
of organic carbon (TOC).
Determination range: C 0.05 t o 150 mg/Z
Repeatability: 3 t o 10 % in coefficient of variation (different according t o the
type of apparatus and measuring conditions)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
83
K 0101 : 1998

Water As described in 20.1 (1) (a).
TOC standard solution (1 mgC/ml) As described in 20.1 (1) (b).
TOC standard solution (0.1 mgC/ml) As described in 20.1 (1) ( c ) .
Zero calibration liquid The water of (a) is used.
Span calibration liquid Take an appropriate quantity of TOC standard
solution (0.1 mgC/ml) [or TOC standard solution (1mgC/ml)l into a volu-
metric flask, and add water to the marked line. Prepare so as to obtain
the concentration of TOC corresponding t o approx. 80 % of the measuring
range of a measuring instrument by the same operation. Prepare at the
time of use.
Acid solution Prepare a specific concentration by using as low concen-
tration of TOC as possible of phosphoric acid specified in JIS K 9005, hy-
drochloric acid specified in JIS K 8180, or sulfuric acid specified in JIS K
8951.
Carrier gas As described in 20.1 (1) (h).
(a) TOC automatic measuring instrument The combustion oxidation-in-
frared system TOC automatic measuring instrument of which the measur-
ing range is 1O00 pgC/Z or under or 1mgC/Z o r over, as specified in JIS K
0805.

(a) Supply acid solution and carrier gas t o a measuring instrument.
(b) Perform warming up of the measuring instrument to stabilize functions of
respective parts and an indication recording part.
(c) Calibrate the measuring instrument by using zero calibration liquid and
span calibration liquid.

(a) Supply the sample to the measuring instrument, and confirm that the in-
dication value is stabilized.
(b) Obtain the concentration (mgCII) of organic carbon in the sample (TOC)
from the indication value.
Remarks 4 For the method wherein organic carbon is oxidized to carbon
dioxide with a TOC automatic measuring instrument, there
is a method wherein wet oxidation decomposition under high
pressure a t high temperature (for instance, approx. 2 MPa,
200 O C ) is performed by adding oxidizer (peroxodisulfate) in
addition t o a combustion oxidizing method. There are two
methods for this method. The one is the method in which the
sample is made acidic of 2 o r less pH, inorganic carbon is
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
84
K O101 : 1998
removed by aeration, and measurement is performed. The other

one is the method in which the sample is made acidic, is joined
by oxidizer, total carbon is determined, separately the sample
is made acidic, inorganic carbon is determined at a tempera-
ture a t which organic substances are not decomposed
(approx. 130 O C ) , and the quantity of organic carbon is obtained
by subtracting the quantity of inorganic carbon from the quan-
tity of total carbon.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
85
K 0101 : 1998
21 Total oxygen demand (TOD) The total oxygen demand means the amount
of oxygen t o be consumed by the component elements of organic substances in the
sample such as carbon, hydrogen, nitrogen, sulfur, phosphorus, when the sample is
burned. To this test the combustion method applies.
Inject a small amount of a sample together with the inert gas containing a defi-
nite amount of oxygen into a combustion tube a t a high temperature to burn the
organic substances and the like, and then determine the concentration of oxygen in
inert gas to obtain the amount of total oxygen demand from its reduced amount.
Carry out this test as soon as possible after sampling. When it is impossible t o
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
carry out the test immediately, preserve the sample in accordance with 3.3,and carry
out the test as soon as possible.
Determination range: O 10 t o 500 mg/Z
Repeatability: 3 to 10 % in coefficient of variation (different according to the
type of the apparatus and measuring conditions)
Water Water A3 o r A4 specified in JIS K 0557(1) (2) (31, and containing
no dissolved oxygen(*) shall be used. Use this water for the preparation
and operation of the reagent to be used in this test. Carry out a blank test
according t o ( 5 ) , and confirm the adaptability of use.
TOD standard solution (1 mgO/ml) Heat potassium hydrogen phtha-
late, standard reagent for volumetric analysis specified in JIS K 8005 at
120 "C for approx. 1h, and allow to stand to cool in a desiccator. Take its
0.851 g, dissolve in water, transfer to a 1O00 ml volumetric flask, and add
water t o the marked line.
TOD standard solution (0.1 mgO/ml) Take 10 ml of TOD standard so-
lution (1mgO/ml) into a 100 ml volumetric flask, and add water to the marked
line. Prepare a t the time of use.
Carrier gas High purity nitrogen grade 2 specified in JIS K 1107 and
oxygen specified in JIS K 1101,
Notes (1) Water of which the concentration of TOD is made as low as pos-
sible is used. Since when the refined water is preserved by putting
in a container, it is gradually contaminated and the concentra-
tion of TOD occasionally increases, it should be preferable t o use
immediately after refining.
(2) To lower the concentration, carry out the distillating operation
in Note (2) of clause 20.
(3) Refer Note (3) of clause 20.
(4) Refine according t o 2(12)(a).
(a) Microsyringe 10 t o 20 1-11
(b) TOD analyzer
86
K O101 : 1998

Warm up the TOD analyzer.
Inject the specified amount (for example 20 pl) of TOD standard solution
(i mgO/ml) o r TOD standard solution (0.1 mgO/ml) into the TOD analyzer
with a microsyringe, and adjust the sensitivity of the TOD analyzer so as
the indicating value (peak height) t o become approx. 80 % of the maximum
scale.
Repeat the operation specified in (b) several times and confirm that the
indicating values are constant.
Mix by shaking the sample(5) sufficiently t o make it homogeneous, then
inject the specified amount [the sample amount as in (b)]with a microsyringe
t o obtain the indicating value, and obtain the approximate total oxygen
demand of the sample in comparison with ( c ) .
Note (5) For the sample of which TOD is 500mgOIZ or more, test after
diluting adequately with water.

Take TOD standard solution (i mgO/ml) or TOD standard solution (0.1 mgO/
mi) into a 100 ml volumetric flask stepwise so as the approximate value of
TOD of the sample obtained by (3) (d) t o be in the middle, and add water
up to the marked line.
Inject the specified quantity (for instance, 20 pl) of TOD standard solution
of the maximum concentration prepared according t o (a)with a microsyringe,
and adjust the sensitivity so that the indicating value becomes approx. 80 %
of the maximum scale.
Inject the specified amount [the amount as Specified in (b)] of TOD stan-
dard solution of each concentration prepared by (a)with a microsyringe in
succession to obtain the indicating value.
Take the same amount of water as in ( e ) for the blank test with a
microsyringe, and obtain the indicated value by operating the same as in
( c ) t o correct the indicated value in ( c ) . Prepare the relation curve between
each oxygen equivalent of TOD standard solution and the indicated value.
( 5 ) Operation Carry out the operation as follows.

(a) Where suspended matters are contained in the sample, thoroughly mix by
stirring with a homogenizer o r mixer t o disperse them homogeneously.
(b) Inject the specified amount of the sample(6) [for example, the same amount
as in 4(b)] t o the TOD analyzer with a microsyringe t o obtain the indi-
cated value.
(c) Where the sample is diluted, for the blank test, take the same amount of
water as in (b) with a microsyringe, then carry out the operation specified
in (b)t o obtain the indicated value, and correct the results obtained on the
sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
37
K O101 : 1998
(d) Obtain the amount of oxygen demand by the injected sample from the work-
ing curve preliminarily prepared t o calculate TOD of the injected sample
(mgOlZ).
(e) Calculate the concentration of TOD (mgOlZ) of the sample according to the
following formu1a :
TOD = a x d
where, TOD : total oxygen demand (mgO/Z)
a : oxygen demand of the injected sample (mgO/Z)
d : dilution multiple of the injected sample
Note (6) Where the total oxygen demand is high, the sample shall be di-
luted by a specified multiple.
Remarks 1 Coexistence of dissolved oxygen interferes with the measure-
ment; particularly its effect is serious where the total oxygen
demand is small. In this case, correct it by measuring the
amount of dissolved oxygen in the sample separately.
2 Where the sample is acidic and contains sulfate ion, when the
sample is heated at a high temperature, it will be decomposed
as follows to generate oxygen, and a negative error will be
induced.
2HzS04 + 2H20 + 2S02 + O2
However, in the sample in which, when the sample is evapo-
rated, the sulfuric acid becomes alkali metal salts, this reac-
tion does not occur. Therefore, where sulfate ion coexists, add
sodium hydroxide solution (200 glZ) to adjust pH to approx. 11
and then carry out the test.
3 Where nitrate ion coexists, when the sample is heated at a
high temperature, it will be decomposed as follows to gener-
ate oxygen, and a negative error will be induced.
4NaN03 + 2Na20 + 4 N 0 + 302
or
2NaN03 + Na20 + NzO + 202
4 If the sample containing heavy metal ion is measured for a
long period of time, the catalyst in the combustion tube dete-
riorates and the oxidizing ability decreases. In such a case,
exchange or regeneration of the catalyst is necessary.
5 In the case of sea water or the like which contains a large
amount of salts, the base line of the indicated value may some-
times not return to the original place, and therefore repeat
the measurement until the stable indicating value is obtained,
or carry out the test after diluting the sample adequately.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
_*I^._-,-_ . ... ..- . . .... . .. -
88
K O101 : 1998
22 Phenols and p-cresols These are divided into phenols and p-cresols.
22.1 Phenols For the test of phenols, 4-aminoantipyrin absorptiometry shall be

applied t o the sample pretreated (distilled).
Phenols are easily decomposed by phenol decomposing bacteria, and are easily
attacked by oxidizing substances, reducing substances, alkaki, etc. The test shall
be carried out just after sampling. In the case when the test can not be carried out
immediately, the sample is preserved in accordance with 3.3 and shall be tested as
soon as possible.
22.1.1 Pretreatment
(a) Water The water A3 specified in JIS K 0557(l). Preserve it in a boro-
silicate glass bottle.
(b) Phosphoric acid (1+9) Prepare by using phosphoric acid specified in JIS
K 9005.
(c) Copper (II) sulfate solution Dissolve l o g of copper (II) sulfate 5-hy-
drate specified in JIS K 8983 in water t o make 100 ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(d) Methyl orange solution (1 gll) Dissolve 0.1 g of Methyl Orange speci-
fied in JIS K 8893 in 100 ml of hot water.
Note (1) The water shall be refined by using the distillation apparatus made
of quartz glass o r borosilicate glass.
(a) Distillation apparatus The apparatus with ground-glass joint
(3) Operation of distillation Carry out the distillation operation as follows.
(a) Take 250 ml of the sample(2)( 3 ) into a 500 ml distilling flask, add several
drops of Methyl Orange solution (1g/Z), then add phosphoric acid (1+9) until
the colour of Methyl Orange changes to make pH about acid 4, and there-
after add 2.5 ml of copper (II) sulfate solution.
(b) Attach the distilling flask to the distilling apparatus and distill using a
250 ml measuring cylinder (with stopper) as a receiver.
(c) When distillate in the measuring cylinder becomes 225 ml, stop heating once.
(d) After the boiling of the sample in the flask has ceased, add 25 ml of water
to the distilling flask. Again continue the distillation to distil 25 ml fur-
ther and make the total amount of the distillate 250 ml(4).
Notes (2) When the concentration of phenols in the sample is not less than
50 mgil, take a proper amount of the sample, and dilute with water
t o 250ml.
When the approximate concentration of phenols in the sample
is known, make the amount of the sample 100 ml and the added
amount of copper (II) sulfate solution 1ml. Carry out the same
operation to make the total distillate 100ml.
89
K 0101 : 1998
(3) When the concentration of phenols is not more than 25 pgll, take
500 ml of the sample into a 1I distilling flask, and add 5 ml of
copper (II) sulfate solution. When 450 ml is distilled, stop heat-
ing once. After cooling, add 50 ml of water, and continue the dis-
tillation again to distil further 50 ml to make the distillate 500 ml.
(4) When the distillate is turbid, add phosphoric acid (1+9) again to
the distillate t o make acidic pH about 4. Then add 2.5 ml of copper
(II) sulphate solution, and repeat the operation of distillation.
If turbidity does not disappear after redistillation, treat in ac-
cordance with the removing method of oily substances and tars
as specified in 22.1.2 Remarks l ( 3 ) .
22.1.2 4-amino antipyrine absorptiometry The sample preliminarily treated

--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(distillation) is adjusted t o pH of approx. 10, and 4-amino antipyrine (4-amino-1,2-

dihydro-1,5-dimethyl-2-phenyl-3H-pyrazole-3-on) solution and potassium hexacyano
ferrate (III) solution are added, and the absorbance of red antipyrine-dye to be gen-
erated is measured t o determine phenols.
I n this method, other than phenol the phenol derivatives having substituents at
o- and m-positions and the hydroxyl-substituted polycyclic compounds react with 4-
amino antipyrine to generate antipyrine-dye and are determined. Phenol derivative
having substituent a t p-position is difficult to react with 4-amino antipyrine, and
therefore hardly develops colour. The intensity of colouring of antipyrine-dye is different
according t o the kind, position, number, etc. of the substituent.
In this test, phenols shall be expressed as phenol in comparison with the inten-
sity of colour by phenol standard solution.
Determination range: Extraction method C6&0H 2.5 to 50 pg, direct method
CsHsOH 50 t o 500 pg,
Repeatability: 3 t o 10 % in coefficient of variation
(b) Ammonium chloride-ammonia buffer solution (pH 10) Dissolve 67.5 g
of ammonium chloride specified in JIS K 8116 in 570 ml of ammonia wa-
ter specified in JIS K 8085 to make 1I with water. Preserve it in a cold
place with stoppering closely.
(c)
G 1
Potassium bromate solution - mol / I Dissolve 2.78 g of potassium bro-
mate specified in JIS K 8530 and 10 g of potassium bromide specified in
JIS K 8506 in water to make 1Z.
(d) 0.1 mol/Z sodium thiosulfate solution Dissolve 26 g of sodium thiosulfate
pentahydrate specified in JIS K 8637 and 0.2g of sodium carbonate (anhy-
drous) specified in JIS K 8625 in water to make 1I. After standing for at
least two days in an airtight vessel, standardize. This solution is used by
standardization at the time of use.
Standardization Heat potassium iodate (standard reagent for volumetrical
analysis specified in JIS K 8005) a t 130 "C for about 2 h, and allow to cool
in a desiccator. Then, take 0.713 g of potassium iodate, dissolve in a small
quantity of water, transfer into a 200 ml volumetric flask, and add water
90
K O101 : 1998
up to the marked line. Take 20 ml of this solution into a 300 ml Erlenm-

eyer flask with ground stopper, then add 2 g of potassium iodide specified
in JIS K 8913 and 5 m l of sulfuric acid (1+5), and stopper immediately.
Mix by shaking gently, and allow t o stand in a dark place for about 5 min.
Add 100ml of water, and titrate the liberated iodine with this sodium
thiosulfate solution. After the yellow colour of the solution has become pale,
add 1ml of starch solution (10 gll) as indicator, and titrate until the blue
colour of iodine-starch disappears.
Separately, carry out the blank test for water under the same conditions,
Calculate the factor C f ) of 0.1 moll sodium thiosulfate solution from the num-
ber of ml corrected by the blank test according t o the following formula:
b 20 1
=ax 100' %Öx x x 0.003 567
where, a : amount of potassium iodate (g)
b : content of potassium iodate (%o)
x : 0.1 molll sodium thiosulfate solution required for
the titration (mi)
0.003 567 : potassium iodate equivalent to 1ml of 0.1 mol/!! so-
dium thiosulfate solution (g)
Potassium hexacyanoferrate (III) solution Take 9 g of large crystals
of potassium hexacyanoferrate (III) specified in JIS K 8801,wash their
surface with a small amount of water, and then dissolve it in water to make
100 ml. Filter the solution, if required. Prepare the solution every week,
and if the colour of the solution changes to dark red within a week, do not
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
use the solution.

Sodium sulfate (anhydrous) As specified in JIS K 8987.
Potassium iodide As specified in JIS K 8913.
4-amino antipyrine solution (20 glZ) Dissolve 2.0 g of 4-amino antipyrine
specified in JIS
(4-amino-1,2-dihydro-l,5-dimethyl-2-phenyl-3~-pyrazole-3-on)
K 8048 in water to make 100 ml. Prepare this solution a t the time of use.
Starch solution (10 g/Z) Mix 1g of starch (soluble) specified in JIS K
8659 in approx. 5 ml of water, then add it in 100 ml of hot water while mixing
by stirring, and after boiling for 1 min, allow to cool. Prepare the solution
a t the time of use.
Phenol standard solution (i mgCsH~OWm1) Dissolve 1g of phenol(5)
specified in JIS K 8798 in water to make I I . Preserve it in a dark and
cool place.
Standardization Take 50 ml of this solution in a 500 ml Erlenmeyer flask
with ground stopper, and add approx. 100 ml of water. Add to this solution
Go 1
50 ml of potassium bromate solution - mol / l (the amount of reaction is
approx. 40 ml), and further add 5 ml of hydrochloric acid (at this time, white
precipitate of tribromophenol is generated).
91
K O101 : 1998
Stopper tightly, shake the solution gently to mix, and after the liberation
of brown bromine, allow to stand for 10 min. Then, add 1g of potassium io-
dide specified in JIS K 8913, and titrate the liberated iodine with 0.1 mol/Z
sodium thiosulfate solution, and after the yellow colour of the solution has
become pale, add 1ml of starch solution (10 g/Z) as indicator. Continue the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
titration until the blue colour of iodine-starch disappears. Take the number
of ml of 0.1 moVZ sodium thiosulfate solution required for this titration as ( b ) .
(w
Separately, add 20 ml of potassium bromate solution - mol Z
I
l to 100 ml
of water, then operate in the same manner as above and obtain the num-
ber of ml (a) of 0.1 mol/Z sodium thiosulfate solution required for titration.
Calculate the concentration of phenol standard solution (mg/ml) according
1
P ~ ( 2 . -b)
5 ~x f x - X 1.569
50
where, P : phenol standard solution (mgC~H50H/ml)
f : factor of 0.1 molíl sodium thiosulfate solution
1.569 : phenol equivalent to 1ml of 0.1 moVZ sodium thio-
sulfate solution (mg)
(i) Phenol standard solution (10 ygCsH50Wml) Take number of ml of phe-

nol standard solution corresponding to 10 mg of phenol (1rngC~H~ûH/rnl)
in a 1O00 ml volumetric flask, and add water up to the marked line. Pre-
pare this solution at the time of use.
(m) Phenol standard solution ( i pgCsH50H/ml) Take 50 ml of phenol stan-
dard solution (10 pgC6H50H/ml)in a 500 ml volumetric flask, and add water
up t o the marked line. Prepare this solution at the time of use.
Note ( 5 ) When tested by gas chromatography, phenol standard solution of
which retention time corresponding to that of cresols has no peak
shall be used.
The following operation shall be carried out t o be confirmed.
Take 10 ml of phenol standard solution (1mgC6H~OH/ml)into
a separatory funnel, regulate pH at 1to 2 with sulfuric acid (1+3),
add 5 m l of 1,2-dichloroethane specified in JIS K 8465, mix by
shaking, and allow t o stand. Separate the organic solvent layer,
add sodium sulfate (anhydrous), and mix by shaking to be dehy-
drated. Inject 5 p1 of this solution into a gas chromatograph ap-
paratus, and obtain a gas chromatogram.
Conditions for gas chromatograph
Column tube Glass made, 3 mm in inside diameter and 3 m
in length
Column filler Acid-washed refractory brick(*) (180 to 250 pm)
which is impregnated by approx. 2.5 % with ester fixed liquid
phase.
Detector Flame ionization detector
92
K O101 : 1998
Carrier gas High purity nitrogen grade 2 specified in JIS K

1107 is used, of which flow rate is 40 t o 50 ml/min.
Column tank temperature 180 “C
Detector tank temperature 195 “C
A peak appears in the order of o-cresol, phenol, p-cresol, and
rn-cresol under those conditions.
Note (*) Its main component is diatomaceous earth and the resistant tem-
perature is 1100 “C.
Informative reference : For column fillers, “Chromosorb W as a carrier,
and “KG-02”,“FAP-S”o r the like having equiva-
lent performance which is covered with ester fixed
liquid phase are on the market.

(a) Separating funnel 200 ml
(b) Photometer Spectrophotometer o r photoelectric photometer

(a> Take 100 ml of the sample pretreated as specified in 22.1.1(6) (2.5 t o 50 pg
as phenol) into a 200 ml separating funnel, add 3 ml of ammonium chlo-
ride-ammonia buffer solution (pH lo), shake t o mix, and adjust the solu-
tion t o pH 1050.2,
(b) Add 2 ml of 4-amino antipyrine solution (20 g/Z), shake t o mix, and then
add 2 ml of potassium hexacyanoferrate (III) solution. After mixing by shaking
thoroughly, allow t o stand for 3 min(7).
(c) Add 10 ml of chloroform, and after mixing by shaking violently for 1min
or more, stand it still. Filter the chloroform layer with dry filter paper o r
after transferring into a beaker, add approx. 1g of sodium sulfate (anhy-
drous) to dehydrate.
(d) Transfer this solution into an absorption cell, and measure the absorbance
at a wavelength near 460 nm using chloroform of blank test operated in ac-
cordance with (a)to ( c ) separately on 100 ml of water as reference solution.
(e) Obtain the amount of phenol from the working curve, and calculate the
concentration of phenols in the sample (rngC&ûH/Z).
Working curve Take stepwise 2.5 t o 50 ml of phenol standard solution
(1pgC6&ûH/ml) into a 250 ml separating funnel and add water t o make
100 ml. Further, add 3 ml of ammonium chloride-ammonia buffer solution
(pH lo), mix by shaking, adjust pH to 1010.2, and carry out the operations
specified in (b) to (d) t o prepare the relation curve between the amount of
phenol (CsH50H) and the absorbance.
Notes (6) Where the amount of phenol in 100 ml of the sample is 2.5 pg or
less, take 500 ml of the sample to distillate, and take the whole
amount into a 1O00 ml separating funnel. Then add 10 ml of am-
monium chloride-ammonia buffer solution (pH lo), 3 ml of 4-amino
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
93
K O 1 0 1 : 1998
antipyrine solution (20 g/Z), and 3 ml of potassium hexacyanoferrate

(III) solution, after mixing by shaking sufficiently, allow t o stand
for 3 min, and add 10 ml of chloroform t o extract.
Further, prepare the working curve under the same conditions.
(7) Where the colouration at this time is sufficiently strong, trans-
fer this solution into a n absorption cell, and measure the absor-
bance at the wavelength near 510 nm using the blank test solution
operated in accordance with (a) and (b)separately on 100 ml of
water as the reference solution. Obtain the amount of phenol
from the working curve, and calculate the concentration of phenols
(mgCsHsOHIZ).
Working curve Take step by step 5 to 50ml of phenol stan-
dard solution (10 pgCsHsOH/ml) into a 100 ml measuring cylin-
der with ground stopper, and add water up to the marked line
of 100ml. Hereafter, carry out the operations specified in (a)
and (b), and prepare the relation curve between the amount of
phenol and the absorbance.
Remarks 1 In this test, oxidizing substances, reducing substances, metal ion,
aromatic amines, oils and tars, etc. interfere.
Generally, most of such interfering substances can be removed by
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
distillation, if the sample contains oxidizing substances, sulfur com-
pounds, oils and tars, they shall be treated as follows:
Oxidizing substances If the sample contains the oxidizing sub-
stances such as residual chlorine, it liberates iodine when po-
tassium iodide is added under acidic condition. In such a case,
it is necessary to add a slight excess of iron (II) sulfate hepta-
hydrate specified in JIS K 8978 or sodium metaarsenite speci-
fied in JIS K 8046 immediately after sampling.
Sulfur compounds If the sample contains hydrogen sulfide
and sulfite ion, add phosphoric acid to make pH about 4 imme-
diately after sampling. Then, send air carefully in the sample
or mix by stirring the sample, and expel hydrogen sulfide and
sulfur dioxide. Thereafter add copper (II) sulfate pentahydrate
specified in JIS K 8983 to the solution.
Oily matters and tars If the sample contains oily matters and
tars add sodium hydroxide (granular) specified in JIS K 8576
without adding copper (II) sulfate pentahydrate immediately after
sampling t o adjust pH of the solution t o 12 t o 12.5. Transfer it
into a separating funnel, and add chloroform specified in JIS K
8322 to extract oily matters and tars. Then discard the chloro-
form layer. Heat the aqueous layer on the water bath t o expel
the residual chloroform, then add phosphoric acid specified in
JIS K 9005 to adjust pH to 4 o r under, and add 2.5 ml of cop-
per (II) sulfate solution.
2 If the sample is free from colour and turbidity and does not contain
interfering substances, the distillation operation may be omitted and
the test may be carried out directly.
94
K 0101 : 1998
22.2 p-Cresols For the test ofp-cresols, p-hydrazinobenzene sulfonic acid absorp-
..
tiometry shall be applied t o the pretreated (steam distilled) sample.
22.2.1 p-Hydrazinobenzene sulfonic acid absorptiometry The phenols are al-

lowed t o react with Gibbs' reagent a t pH 8.0 t o change into indophenol. p-cresols
which do not react with Gibbs' reagent are distilled by steam under acidic condition
of ascorbic acid. Diazo compound, p-hydrazinobenzene diazonium salt, formed by p-
hydrazinobenzene sulfonic acid and nitrous acid, is allowed to couple with the dis-
tilled p-cresols, and the absorbance of red colour of azo colouring matter produced is
measured. Thus, p-cresols are determined by this method,
Range of determination: p-CH&H*OH 10 t o 150 pg
(1) Reagents Use the following reagents.

Sulfuric acid (1+17) Prepare by using sulfuric acid specified in JIS K
8951.
Sodium hydroxide solution (100g l l ) Dissolve 1 0 g of sodium hydrox-
ide specified in JIS K 8576 t o make 100ml.
Sodium c a r b o n a t e As specified in JIS K 8625.
Sodium chloride As specified in JIS K 8150.
Methyl o r a n g e solution (1g l l ) As described i n 22.1.1 (1)(d).
Chloroform or diethyl ether Chloroform specified in JIS K 8322 o r di-
ethyl ether specified in JIS K 8103.
Gibbs' reagent Dissolve 0.5 g of 2,6-dibromo-N-chloro-p-benzoquinone-
monoimine (2,6-dibromoquinone-chloroimide) specified in JIS K 8491 in 50 ml
of ethanol (95) specified in JIS K 8102. Prepare this reagent a t the time
of use.
L(+)-ascorbic acid As specified in JIS K 9502.
Ammonia water (1+7) Prepare by using ammonia water specified in JIS
K 8085.
p-Hydrazinobenzene sulfonic a c i d solution
Solution A Add 1 g ofp-hydrazinobenzene sulfonic acid 0.5 hydrate specified
in JIS K 9525 and 0.3 g of sodium carbonate (anhydrous) specified in 31s
K 8625 in 80 ml of water, warm in a water bath t o dissolve, then add 9 ml
of hydrochloric acid specified in JIS K 8180 and dilute i t with water to
100 ml. Because this solution deposits crystals a t room temperature, pre-
serve it in the thermostatic bath a t approx. 37 " C . Do not use the solution
for which one week or longer has elapsed.
Solution B Take 4 ml of solution A in a 100 ml volumetric flask, cool to
approx. 10 OC, then add 5 ml of sodium nitrite solution (10 g l l ) (dissolve so-
dium nitrite specified in JIS K 8019 in water t o make 100 ml) and allow to
stand at approx. 10 "C for 3 t o 5 min. Add water preliminarily cooled t o
approx. 10 "C t o the marked line. Prepare this solution a t the time of use.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
95
K O101 : 1998
(k) p-Cresol standard solution (1 mgCH&6H&H/d) Take 1.00 g ofp-cresol

specified in JIS K 8306, and dissolve it in a small quantity of water. Transfer
the solution into a 1O00 ml volumetric flask, and add water up to the marked
line.
(1) p-Cresol standard solution (0.1 mgCHsCsHsOH/ml) Take 20 ml of p -
cresol standard solution (1mgCHsC6H&H/ml) into a 200 ml volumetric flask,
and add water up t o the marked line.
(a) Steam distillation apparatus Small type having ground-glass joint.
(a) Take 500 ml of the sample (containing 0.1 t o 1.5 mg asp-cresol) in a 1O00 ml
separating funnel, and add several drops of Methyl Orange solution (1g/Z)
as indicator. Add, drop by drop, sulfuric acid (1+17)until the colour of the
solution changes to red to acidify the solution [when the sample is acidic,
neutralize with sodium hydroxide solution (100 g/Z), until the colour of so-
lution changes to yellow, then add, drop by drop, sulfuric acid (1+17) t o
acidify the sample again]. Then, add 150 g of sodium chloride and 40 ml of
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
chloroform(*) t o the solution, and mix by shaking vigorously t o extract.
Transfer the chloroform layer t o another 200 ml separating funnel.
(b) Repeat the extraction operation 4 times by using each 25 ml of chloroform
in the same manner as in (a). Combine the chloroform layers with that in
the previous separating funnel.
(c) Back extract it with 4 ml of sodium hydroxide solution (100 g/Z) in this chlo-
roform layer, then repeat the back-extraction twice with each 3m1, and
combine the back-extract solution.
(d) Boil this back-extract solution on the water bath, and after volatilizing the
dissolved chloroform, allow t o cool.
(e) Transfer it into a 100 ml volumetric flask with 20 ml of water. Add 2 g of
sodium carbonate (anhydrous) and further add drop by drop sulfuric acid
(1+17)to adjust pH to 8. Then, add 20 ml of water and 5 ml of Gibbs’ reagent
and allow t o stand for 24 h (if phenols coexist, the colour of the solution
turns blue).
(0 Add 1g of L(+)-ascorbicacid and then add water up t o the marked line.
(g) Take 10 ml of this solution in a distilling flask t o carry out steam distilla-
tion, and distil 30 ml of the distillate in a 50 ml measuring cylinder with
ground stopper.
(h) After diluting the distillate with water t o approx. 40 ml, add 5 ml of solu-
tion B of p-hydrazinobenzene sulfonic acid and mix by shaking. Then, add
1ml of ammonia water (1+7), further add water up t o the marked line of
50 ml, again mix by shaking, and then allow to stand for about 5 min.
96
K O101 : 1998
(i) Transfer a portion of this solution into an absorption cell, and measure the
absorbance at a wavelength near 495 nm.
fi) For the blank test, take 40 ml of water, carry out the operations Specified
in (h)and (i) t o measure the absorbance, and correct the absorbance ob-
tained on the sample.
(k) Obtain the amount ofp-cresols from the working curve, and calculate the
concentration of p-cresols in the sample (mgCH3C6H4OH/l).
Working curve Take stepwise 1 to 15 ml of p-cresol standard solution
(0.1 mgCH3C6HdOH/ml) in the 100 ml volumetric flask. Then, carry out the
operation specified in ( e )on and after the addition of 2 g of sodium carbon-
ate (anhydrous), and the operation specified in (f). Take 10 ml of this so-
lution in a distilling flask, and after obtaining 30 ml of distillate by carrying
out steam distillation, dilute with water t o about 40ml. Thereafter, add
5 ml of solution B of p-hydrazinobenzene sulfonic acid, and carry out the
operations specified in (h) t o U) to prepare the relation curve between the
amount of p-cresol (CH3C6H40H) and the absorbance.
Note (8) Diethyl ether specified in JIS K 8103 may be used instead of
chloroform. In this case, it is not necessary t o add sodium chlo-
ride specified in JIS K 8150.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
97
K O101 : 1998
23 S u r f a c e active agents Surface active agents shall be classified into anionic

surface active agents and nonionic surface active agents.
Since the surface active agents are easily decomposed by microorganisms, the test
shall be carried out immediately after sampling. In the case where the test can not
be carried out immediately, they shall be preserved in accordance with 3.3, and shall
be tested as soon as possible.
23.1 Anionic s u r f a c e active agents To determination of anionic surface active

agents, Methylene Blue-absorptiometry, Ethyl Violet-absorptiometry or solvent ex-
tract-flame atomic absorption method shall be applied.
The anionic surface active agents include sulfate esters of higher alcohol, sulfate
esters of fatty oil, sulfonated anionic surface active agents [alkylaryl sulfonates (straight-
chain alkylbenzensulfonates, LAS), alkyl sulfonates, alkene sulfonates, etc.] etc.
23.1.1 Methylene Blue absorptiometry The ion pair t o be generated by reac-

tion of anionic surface active agent with Methylene Blue [3,7-bis (dimethylamino)
phenothiazine-5-ium chloride1 is extracted with chloroform, and the absorbance is
measured to express it as sodium dodecylsulfate.
Determination range: anionic surface active agent [ N ~ O ~ S O ( C H ~ ) I I C H ~ ]
2 t o 501.18
Repeatability: 5 to 10 % in coefficient of variation
W a t e r Water A3 specified in JIS K 0557.
Sulfuric acid (1+35) Prepare by using sulfuric acid specified in JIS K 8951.
Sodium hydroxide solution (40 g/Z) As described in 19 ( i )(g).
Alkaline sodium tetraborate solution After dissolving 9.54 g of sodium
tetraborate 10-hydrate specified in JIS K 8866 in water, dilute with water
t o 500 ml. Add 50 ml of sodium hydroxide solution (40 g/Z), and dilute the
total quantity with water to 11.
Methylene Blue solution (25g/Z) Dissolve 0.3 g of Methylene Blue (usually
trihydrate) specified in JIS K 8897 in water t o make 1Z.
Absorbent cotton
Anionic surface active agent standard solution [1 mgNa03SO(CHz)~
CHdmll Take 1.00 g of sodium dodecylsulfate (sodium laurylsulfate)(1) as
100 %(2), dissolve in water, transfer to a 1O00 ml volumetric flask, and add
water to the marked line.
Anionic surface active agent standard solution [lo p gNa03SO(CH2)11
CHdml] Take 1 0 m l of anionic surface active agent standard solution
[imgNa03SO(CHdilCH3/m1]into a 1O00 ml volumetric flask, and add water
to the marked line. Prepare at the time of use.
Notes (1) Sodium dodecylsulfate on the market of which the purity and av-
erage molecular weight are known, shall be used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
98
K O101 : 1998
(2) In the case where the purity and average molecular weight are
confirmed, Remarks 4 shall apply.
(b) Photometer Spectrophotometer or photoelectric photometer
Put 50 ml of water, 10 ml of alkaline sodium tetraborate solution, and 5 ml
of Methylene Blue solution (0.25 gll) into a separating funnel (A).
Put 100 ml of water, 10 ml of alkaline sodium tetraborate solution, and
5 ml of Methylene Blue solution (0.25 g/Z) into a separating funnel (B).
Add 10 ml of chloroform to each of them. After mixing by violently shak-
ing for 30 s, allow to stand, and discard the chloroform layer. Repeat this
operation once again.
Add 2 to 3 ml of chloroform to the water layer. After mixing by gently shaking,
allow to stand, and discard the chloroform layer. Repeat this operation
until the chloroform layer becomes colourless.
Add 3 ml of sulfuric acid (1+35)t o the water layer in the separating funnel
(B) which has been washed with chloroform.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Further, when the legs of the separating funnels (A) and (B) are wet,
wipe off with filter paper or the like.
Add a suitable quantity [containing 2 t o 50 pg as Na03SO ( C H ~ ) I I C Hof~ a]
sample(3) t o the water layer in the separating funnel (A) for which the
preparatory operation of (3)has been carried out. However, allow the to-
tal quantity not to exceed 100 ml.
Add 10 ml of chloroform, mix by gently shaking for approx. 1min, allow t o
stand, and transfer the chloroform layer into the separating funnel (B) for
which the preparatory operation of (3)has been carried out.
After mixing the separating funnel (B) by shaking gently for approx. 1min,
allow t o stand. Fill the leg part of the separating funnel with absorbent
cotton, and transfer the chloroform layer to a 25 ml volumetric flask.
Add again 10ml of chloroform t o the separating funnel (A), repeat the
operations of (b) and (c),extract, combine the chloroform layer t o the pre-
ceding 25 ml volumetric flask in the same way as in (cl,and add chloro-
form t o the marked line.
Transfer it to an absorption ce11(4), and measure the absorbance near 650 nm
in wavelength by using chloroform as reference solution.
Use 50ml of water as a blank test, put it into the separating funnel for
which the preparatory operation of (3)is preliminarily carried out, obtain
the absorbance by performing the operations of (a) t o (e), and correct the
absorbance obtained for the sample.
99
K O101 : 1998
(g) Obtain the quantity of anionic surface active agent from the working curve,
and calculate the concentration of anionic surface active agent in the sample
[mgNa03SO(CHd1iCH3/Zl.
Working curve Deal out stepwise 0.2 to 5 m l of anionic surface active
agent standard solution [lo ~~gNa03SO(CH2)1iCH3/ml], put into a separat-
ing funnel for which the preparatory operation of (3)has been preliminar-
ily performed, carry out the operations of (a)to (f),and prepare the relation
curve between the quantity of anionic surface active agent standard solu-
tion [N~O~SO(CHZ)IICH~] and the absorbance.
Notes (3) Adjust pH at approx. 7 with sodium hydroxide solution (40gll)
for acidity or with sulfuric acid (1+35) for alkalinity by using a
pH meter.
(4) When a 50mm absorption cell is used, the anionic surface ac-
tive agent of 0.4 t o 10 pg can be determined.
Remarks 1 If a great amount of ions of nitrate, cyanide, thiocyanate, etc.
exist, the determination is disturbed.
Since a cationic surface active agent is strongly bonded to
an anionic surface active agent, a negative error occurs ac-
cording to the coexisting quantity. However, for ordinary water
its quantity is very little compared with the anionic surface
active agent.
2 In the water near the bottom mud where water earthworms,
brandling earthworms, etc. exist, positive errors are liable to
be caused.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
3 In order to determine sulfonic anionic surface active agent (LAS
or the like), hydrolyze anionic surface active agent of alcohol
series or the like by the following operation, determine the
residual sulfonated anionic surface active agent by the opera-
tion of (4), and express it as sodium dodecylsulfate.
Take a suitable quantity of a sample [containing 4 t o 100 pg
as N ~ O ~ ~ O ( C H Z )into
~~C anHErlenmeyer
~] flask with ground-
glass joint, add 25 ml hydrochloric acid specified in JIS K 8180
and several pieces of boiling tips, and dilute with water to 50 ml
in the quantity of solution. Thereafter, mount a reflux con-
denser, and boil quietly for approx. 2 h.
After standing t o cool, add several drops of phenol phtha-
lein solution (5 gll) as indicator [according to 13.2 ( i )(a)],neu-
tralize by adding sodium hydroxide solution (400 g/Z) initially
and sodium hydroxide solution (40 g/Z) near the point of neu-
tralization until the colour of the solution turns pale pink, and
dilute it with water t o make 100ml.
Hereafter, perform the operations of (3)and (41, obtain the
quantity of sulfonated anionic surface active agent, and cal-
culate the concentration of sulfonated anionic surface active
agent in the sample [ ~ ~ N ~ O ~ ~ O ( C H Z ) I I C H ~ / Z I .
100
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
4 To the measurement for the content and average molecular

weight of sodium dodecylsulfate, the following method shall
be applied.
(a> Sulfuric acid (0.5 moUZ) Gradually add 30 ml of sul-
furic acid specified in JIS K 8951 into 1,? of water.
(b) Ethanol (95) As specified in JIS K 8102.
(c) Phenolphthalein solution (5 gll) As described in
13.2 (i)(a).
(d) Hexane As specified in JIS K 8848.
(e) Sodium sulfate (anhydrous) As specified in JIS
K 8987.
(0 1 mol/Z sodium hydroxide solution Take approx.
60 ml of water into an alkali-resistant container such
as a polyethylene bottle or the like, dissolve by add-
ing little by little 80 g of sodium hydroxide specified
in JIS K 8576 while cooling (wash the surface with a
small quantity of water), tightly stopper, and allow to
stand for 4 to 5 days. Take 50 ml of the supernatant
into a 1 ,? polyethylene airtight vessel, and add 1 ,? of
water containing n o carbonic acid of 2 (12)(b). After
mixed, preserve it with shielding from carbon dioxide.
Standardization Allow amidosulfate, standard re-
agent for volumetric analysis, specified in JIS K 8005
to stand in a desiccator under not more than 2 kPa for
approx. 48 h, and dry. Weigh out approx. 2 g thereof
to the nearest 0.1 mg, put into a 200 ml Erlenmeyer
flask, dissolve by adding approx. 25 ml of water, add
3 to 5 drops of Bromothymol Blue solution ( i gl,?)as an
indicator (according to Note (1) of 14),titrate with this
1molll sodium hydroxide solution, and take the point
when the colour of the solution changes to green as an
end point. Calculate the factor (f)of 1 moll,?sodium
hydroxide solution from the following formula.
b 1
f = a xxx0.097
E X1
where, a : quantity of amidosulfate (g)
b : content of amidosulfate (%)
x : 1molll sodium hydroxide solution re-
quired for titration (mi)
0.0971: amidosulfate equivalent to 1 ml of
1 mol4 sodium hydroxide solution (g)
( g ) Higher alcohol mixture standard solution Weigh
out 0.50 g of 1-decanol, 0.50 g of 1-dodecanol, and 0.50 g
of 1-tetradecanol, and dissolve them in 15 ml of hex-
ane specified in JIS K 8848.
101
K 0101 : 1998

(a) Erlenmeyer flask with reflux condenser Reflux
condenser with cooing part of 200 to 300 mm in length
(Liebig condenser and Allihn condenser), and a 300 ml
Erlenmeyer flask with ground-glass joint.
(b) Separating funnel 300 ml
(c) Gas chromatograph An example of using condi-
tion is shown.
Column tube Made of stainless steel, 3 to 4 mm in-
side diameter, and 1 t o 2 m length
Column filler A refractory brick with 150 t o
250 pm(*) in particle size as a carrier is impregnated
by about 10 % with silicon fixed liquid phase.
Detector flame ionization detector
Carrier gas high purity nitrogen grade 2 specified
in JIS K 1107 o r helium (99.8 vol%)
Sample gasification chamber temperature 290
t o 300°C
Column tank temperature 170 to 180°C
The flow rate of carrier gas is so regulated that higher
alcohols flow out for approx. 30 min.
Note (*) Its main component is diatomaceous earth
and the resistant temperature is 1100 O C .
Informative reference : Column fillers on the market
are “Chromosorb W”as a re-
fractory brick, and a carrier
having equivalent perfor-
mance which is impregnated
with silicon SE-30 or the like
as a fixed liquid phase.
(3) Measuring operation of purity Carry out the opera-
tion as follows.
(a) Weigh out approx. 4 g of sodium dodecylsulfate t o the
nearest 0.1 mg, and put it into a 300 ml Erlenmeyer
flask.
(b) After adding 20 ml of sulfuric acid (0.5 mol/Z), heat
on the water bath with the reflux condenser attached.
Taking care t o the foaming, shake lightly the Erlen-
meyer flask at times, and heat it until the solution
becomes transparent.
(c) Consecutively heat and reflux it on the hot plate for
about 2 h.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
102
K 0101 : 1998
After standing t o cool, wash the inside wall by pour-

ing approx. 30 ml of ethanol (95) from the upper part
of the condenser, and after washing with a proper
amount of water, detach the condenser.
Dilute with water t o approx. 100 ml, then add sev-
eral drops of phenolphthalein solution (5gll) as in-
dictor, titrate with 1mol/Z sodium hydroxide solution,
and take the point when the colour of the solution
shows pale red as an end point.
Separately, carry out the blank test under the same
condition, and calculate the content (%) of sodium
dodecylsulfate according to the following formula:
P = ( a- h ) x f x M
sx 1000
where, P : content of sodium dodecylsulfate (%)
a : 1mol/l sodium hydroxide solution
b : 1mol/Z sodium hydroxide solution
required for titration of blank test (ml)
f : factor of the 1mol/l sodium hydrox-
ide solution
M : average molecular weight of sodium
dodecylsulfate
S : amount of sodium dodecylsulfate (g>
(4) Measuring o p e r a t i o n f o r a v e r a g e molecular weight
of s o d i u m dodecylsulfate Carry out the operation as
follows.
Take 50 ml of the solution obtained by the operation
specified in (3)(e)in a 300 ml separating funnel.
Add 100 ml of the mixture of ethanol and water (2+1>
and 50 ml of hexane, shake to mix, and extract higher
alcohols.
Let stand t o separate the hexane layer, and transfer
the aqueous layer into another 300 ml separating fun-
nel.
Add 50ml of hexane to this aqueous layer, mix by
shaking t o extract, and by allowing t o stand still t o
separate the hexane layer. Discard the aqueous layer,
and combine the hexane layer with the previous hex-
ane layer.
Add 50 ml of water t o this layer, mix by shaking, then
allow t o stand still t o separate hexane layer, and
discard the aqueous layer. Again, carry out the wash-
ing operation, securely separate the aqueous layer and
discard it.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
103
K O101 : 1998
(0 Add approx. 2 g of sodium sulfate (anhydrous) t o de-

hydrate the hexane layer, and then transfer into a
100 ml evaporating dish t o expel hexane on a water
bath o r hot plate.
(g) Add 15ml of hexane t o residual higher alcohol t o
dissolve (it becomes the hexane solution containing
approx. 100 gll of higher alcohol).
(h) Set the gas chromatograph under the most suitable
conditions in accordance with JIS K 0114, then take
11-11of higher alcohol mixture standard solution with
a microsyringe, and inject it into a gas chromatograph
(column). Record the chromatogram of each higher
alcohol, and confirm the position of flowing out.
(i) Then, take 1 1-11 of hexane solution of higher alcohol
obtained as specified in (g)with a microsyringe, then
inject into the gas chromatograph (column),and record
the chromatogram of each higher alcohol.
6 ) Repeat the operation specified in (i) three times,
measure the peak area of each higher alcohol (by the
half-width method o r the like), and obtain the aver-
age value of peak areas.
(k) Obtain the mole percentage of each higher alcohol of
10 to 14 in carbon number from the following formu-
lae, and calculate the average molecular weight of
sodium dodecylsulfate:
cio
mlo= x 100
-+-+-
El0 Ri2 RI4
ClO cl
2 c
14
R,,
ci
2
mi2= x 100
R,,+-+- RI2 R14
ClO c12 c
14
14
m14= x 100
Rio
-I-I- Riz R14
where, m10 to mi4 : mole percentage of each higher

alcohol of 10 t o 14 in carbon
number (%)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
104
K O101 : 1998
CIOto cl4 : molecular weight of each higher

alcohol of 10 to 14 in carbon
number
(Cio = 158.29, Ci2 = 186.34,
ci4 = 214.39)
Rio to RI4 : peak area of each higher alcohol

of 10 t o 14 in carbon number
M : average molecular weight of
sodium dodecylsulfate
102 : correction factor for converting
t o sodium dodecylsulfate (the
value of formula weight of S O D a
subtracted by formula weight of
OW
23.1.2 Ethyl Violet absorptiometry Ion pair t o be generated by reaction of an-

ionic surface active agent on Ethyl Violet [N-[4-[bis [4-(diethylamino) phenyl] meth-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ylene]-2,5-cyclohexadiene-l-ylidene]-N-ethylethaneammonium chloride] is extracted
in toluene. Its absorbance is measured, and is expressed as dodecylsulfate.
Determination range: 0.5 to 12.5 pg anionic surface active agent [NaOsSO
(CHdiiCH31

Water Water A3 specified in JIS K 0557.
Sodium sulfate solution ( i mol/Z) Dissolve 142 g of sodium sulfate speci-
fied in JIS K 8987 in water to make 11.
Acetic acid-EDTA buffer solution (pH 5) Dissolve 7.5 g of ethylene-
diamine-tetraacetate dihydrogen disodium 2-hydrate specified in JIS K 8107
in water to make approx. 700 ml. Add 12.5 ml of acetic acid specified in JIS
K 8355 thereto, use a pH meter, and add sodium hydroxide solution (2 moVZ)
until pH becomes 5. Then, add water to make 11.
Ethyl violet solution (i mmol/Z) Dissolve 0.280 g of Ethyl Violet(5) in
water to make 500ml.
Toluene As specified in JIS K 8680.
Anionic surface active agent standard solution
CH3/ml] As described in 23.1.1 (i)(h).
Anionic surface active agent standard solution
CH3/ml] As described in 23.1.1 (i)(i).
Anionic surface active agent standard solution [0.5 p ~ N ~ O ~ S O ( C H Z ) I ~
CH3/mll Take 1 0 m l of anionic surface active agent standard solution
Cl0 ~gNaO~SO(CH~)llCH~/mll into a 200 ml volumetric flask, and add wa-
ter to the marked line. Prepare at the time of use.
105
K 0101 : 1998
1
Note (5) Double salt of Ethyl Violet combined with y mol zinc chloride is
used. In the case where others than this double salt are used,
a quantity of the sample of which the concentration becomes
1mmolíZ, is taken to be prepared.
Further, in the case where a value of the absorbance when the
blank test of (3)(g) is operated is large (approx. 0.04 or more), it
is prepared again by using Ethyl Violet in another lot.


Take a suitable quantity of a sample [containing 0.5 t o 12.5 pg as Na03SO
(CH2)iiCH3]into a separating funnel, and add water to make 100 ml.
Add 5 ml of sodium sulfate solution (1moll), 5 ml of acetic acid-EDTA buffer
solution (pH 5), and 2 ml of Ethyl Violet solution (1mmol/Z) thereto.
Add 5 ml of toluene, and mix by shaking for 10 min(").
Allow t o stand, and discard approx. 100 ml of the aqueous layer.
Further, allow to stand still, and when the toluene layer is completely sepa-
rated, discard the aqueous layer.
Transfer the toluene layer t o an absorption cell, and measure the absor-
bance near 611 nm in wavelength by using toluene as reference solution.
Use 100 ml of water as blank test, obtain the absorbance by carrying out
the operations of (b)to (0,
and correct the absorbance obtained on the sample,
Obtain the quantity of anionic surface active agent from the working curve,
[rngNaO~SO(CH~)1iCH3/Zl.
Working curve Deal out stepwise 1 t o 25ml of anionic surface active
agent [0.5 ygNaO~SO(CH2)1iCH3/rn1]into separating funnels. After adding
water to make 100m1, perform the operations of (b) t o ( g ) , and prepare
the relation curve between the quantity of anionic surface active agent
[Na03SO(CH2)ilCHd and the absorbance.
Note (6) Since time for mixing by shaking has an influence on the absor-
bance, the time for mixing by shaking shall be strictly kept.
Remarks 5 When a great quantity of chloride ion coexists such as seawa-
ter or a sample mixed with seawater, a part thereof gener-
ates ion pair with Ethyl Violet, and the absorbance increases
due t o extraction by toluene. I n such samples, carry out op-
erations as follows: Even after operations of (3)(d)and (e),
adhesive matter remains on the inside wall, transfer the toluene
layer into a small type separating funnel, add ethyl violet
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
106
K O101 : 1998
(15 pmol/Z)-sodium sulfate (10 g/Z) solution [take 7.5 ml of ethyl

violet (immoLIZ), add 5 g of sodium sulfate and dilute with water
to make 500 mi] 20 ml, and mix by shaking for 1min. After
standing, discard most of water layer, stand again to sepa-
rate toluene layer completely and discard the water layer.
Thereafter, operate (f) to (h).
6 Though nitrate ion does not interfere t o 1rngNO3-lZ degree,
when coexisting by more than the said quantity, positive er-
rors are generated. Other ions contained in ordinary river water
or the like do not interfere.
Though cationic surface active agent is strongly combined
with anionic surface active agent, negative errors are gener-
ated according to their coexisting quantity. However, its quan-
tity is very little in ordinary water compared with anionic
surface active agent.
23.1.3 Solvent extract-flame atomic absorption method Ion pair is made by

anionic surface active agent and dibenzo-18-crown-6 having taken in potassium, which
is extracted with 4-methyl-2-pentanone. Potassium in the extracted solution is de-
termined by a flame atomic absorption method, which is expressed by sodium
dodecylsulfate.
Determination range: 2.5 to 50 pg anionic surface active agent [NaOsSO
(CHdiiCH31
Repeatability: 2 to 10 % coefficient in variation

Potassium sulfate (20 mmol/Z)-ammonium acetate (50mmol/Z) mixed
solution Dissolve 3.5 g of potassium sulfate specified in JIS K 8962 and
3.9g of ammonium acetate specified in JIS K 8359 in water to make
approx. 700 ml, use a pH meter, regulate pH at 5 by adding sulfuric acid
(1+35),and make 1Z with water.
Potassium sulfate (4 mmol/Z)-ammonium acetate (10 mmol/Z) mixed
solution Dilute 200 ml of potassium sulfate (20 mmol/Z)-ammonium ac-
etate (50mmol/Z) mixed solution with water to 12.
Dibenzo-18-crown-6 4-methyl-2-pentanone solution (0.5 mmol/Z)
Dissolve 90 mg of refined dibenzo-18-crown-6 (7) in 500 ml of 4-methyl-2-
pentanone specified in JIS K 8903.
Anionic surface active agent standard solution [imgNa03SO(CHz)ii
CHdml] As described in 23.1.1 (i)(h).
Anionic surface active agent standard solution [5 pgNaO&O(CH&i
CHJml] Take 5 m l of anionic surface active agent standard solution
[imgNa0~SO(CHs)11CHdmll into a 1O00 ml volumetric flask, and add water
Note (7) Dibenzo-18-crown-6 shall be refined as follows.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
107
K 0101 : 1998
Add approx. 2.5 g of dibenzo-18-crown-6 into approx. 200 ml of

benzene specified in JIS K 8858, and dissolve by heating on a
water bath. Filter it by suction with a sintered glass filter (1G3).
Though crystals are generated immediately when the filtrate is
cooled, dissolve by heating on a water bath again, and filter by
suction. After performing this operation until the crystal becomes
white (2 t o 3 times), cool the filtrate, and filter by suction.

(b) Frame atomic absorption analyzer
(c) Potassium hollow cathode lamp
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Take a suitable quantity of a sample [containing 2.5 to 50pg as Na03SO
(CH2)iiCH31 into a 100 ml separatory funnel, add 10 ml of potassium sul-
fate (20 mmol/Z)-ammonium acetate (50 mmol/Z) mixed solution, and dilute
the quantity of solution with water t o 50ml.
Add 10 ml of dibenzo-18-crown-6 4-methyl-2-pentanone solution (0.5 mmoVZ),
and mix by shaking for 1 min.
After standing still, discard the aqueous layer, add 25 ml of potassium sulfate
(4 mmol/Z)-ammonium acetate (10 mmol/Z) mixed solution, mix by shaking,
allow to stand still, and discard the aqueous layer.
Spray 4-methyl-2-pentanone layer into the acetylene-air flame in accordance
with the operation described in 6 (measuring operation) of JIS K 0121,
and read the indication value(8) of 766.5 nm in wavelength.
Obtain the quantity of anionic surface active agent from the working curve,
[mgNaO3SO(CH~)iiCHdZl.
Working curve Deal out step by step 0.5 to 10 ml of anionic surface active
agent [ 5 ygNa03SO(CHz)liCH3/ml]into separating funnels, perform the op-
erations of (a) t o (d), and prepare the relation curve between the quantity
of anionic surface active agent [Na03SO(CH&iCH3] and the indication value.
Prepare the working curve when the sample is measured.
Note (8) Absorbance or its proportional value.
Remarks 7 Though calcium and magnesium coexist by 500mg, they do
not give influences. When even by coexistence of several tens
mg of sodium, its fraction is taken in dibenzo-18-crown-6,then,
is extracted by making a pair of anionic detergent and ion,
and if it is sprayed a s i t is, a negative error is provided.
However, by mixing by shaking of the solvent layer after sepa-
ration by extraction and potassium sulfate (4 mmol/Z)-ammo-
nium acetate (10 mmol/Z) mixed solution of (3) (c), sodium is
substituted by potassium, and interference is removed.
108
K O101 : 1998
Since a cationic surface active agent is bonded strongly to

an anionic surface active agent, a negative error is provided
according to its coexisting quantity. However, for ordinary water
its quantity is very little compared with an anionic surface ac-
tive agent. Even though a nonionic surface active agent coex-
ists by approx. 400 pg, there is no interference.
23.2 Nonionic surface active agent There are polyoxyethylene alkyl ethers,
polyoxyethylene alkylphenol ethers, polyoxyethylene alkyl esters, polyoxyethylene
sorbitan alkyl esters, etc. for nonionic surface active agents.
For determination of nonionic surface active agents, a tetrathiocyanatocobaltate
(II) absorptiometry shall be applied to the sample which is processed by pretreat-
ment (ion-exchange separation).
23.2.1 Tetrathiocyanatocobaltate (II)absorptiometry Extract complex of non-

ionic surface active agent and ammonium tetrathiocyanatocobaltate (II) with ben-
zene, measure the absorbance of an ultraviolet part, and express it as heptaoxyethylene
dodecylether.
Determination range: O . 1 to 2 mg of nonionic surface active agent [CHs (CH&O
(CHzCHzOhHI
(a) Water Water A3 specified in JIS K 0557.
(b) Hydrochloric acid (1+11) Prepare by using hydrochloric acid specified
in JIS K 8180.
(c) Sodium hydroxide solution (40 g/Z) As described in 19 (1)(g).
(d) Ammonium tetrathiocyanatocobaltate (II) solution Dissolve 310 g of
ammonium thiocyanate specified in JIS K 9000 and 140 g of cobalt nitrate
(II) hexahydrate specified in JIS K 8552 in water to make 500 ml. Trans-
fer it t o a 1 O00 ml separating funnel, add 50 ml of benzene specified in
JIS K 8858, mix by violently shaking, and allow to stand. Discard the
benzene layer, add 50 ml of benzene again, mix by shaking, and allow t o
stand. Discard the benzene layer, filter the aqueous layer with dry filter
paper, and remove fine foams of benzene.
(e) Sodium chloride As specified in JIS K 8150.
(f) Sodium sulfate As specified in JIS K 8987.
(g) Ethanol (95) As specified in JIS K 8102.
(h) Ethanol (l+l) Add 1 volume of ethanol (95) specified in JIS K 8102 to 1
volume of water t o prepare.
(i) Benzene As specified in JIS K 8858.
(j) Strong acidic cation exchange resin It is of low linking degree (con-
tent of divinylbenzene is 4 t o 6 %o> and is 300 to 1 180 pm in particle size,
R-Na' form. It is used by refining as follows.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
109
K O 1 0 1 : 1998
Pour 250 ml of strong acidic cation exchange resin together with water
into a column of 40 t o 50 mm in inner diameter and approx. 1O00 mm in
height (made of glass o r acrylic resin) and fill that not t o be mixed with air
bubbles. After making 2 I of hydrochloric acid (1+11)flow at approx. 5 U(Z-
resin. h), and wash by making 1Z of water flow in the same way. Then
make 1I of sodium hydroxide solution (40 g/Z) flow at approx. 5 Z/(Z-resin h),
and wash by making 1I of water flow in the same way. Further wash by
making 1I of hydrochloric acid (1+11)and 1I of sodium hydroxide solution
(40glZ) flow in the same way. Then, wash with water [make it flow a t a
rate of approx. 20 ZN-resin h)] until the red colour of phenolphthalein so-
lution ( 5 glZ) [refer t o 13.2 (i)(a)] is hardly recognized,
(k) Strong basic anion exchange resin (I form) I t is of low linking de-
gree (content of divinyl benzene is 4 to 6 %) and is 300 to l 180 Frn in par-
ticle size. R-C1-form. It is used by refining as follows.
Pour 500 ml of strong basic anion exchange resin (I form) together with
water into a column (made of glass or acrylic resin) of 40 to 50 mm in in-
ner diameter and approx. 1O00 mm in height, and fill that not to be mixed
with bubbles. After making 2 I of sodium hydroxide solution (40 g/Z) flow
a t approx. 5 ZN-resin h), wash by making approx. 2 I of water flow in the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
same way. Then, after making 2 I of hydrochloric acid (1+11)flow at

approx. 5 ZN-resin h), wash by making approx. 2 I of water flow in the same
way. Further wash by making 2 I of sodium hydroxide solution (40 g/Z) and
2 I of hydrochloric acid (1+11)flow in the same way. Then, wash with water
[make it flow at a rate of approx. 20 Il(,!-resin = h)] until the solution turns
blue with Methyl Red-Bromocresol Green mixed solution [refer to 13.1 (i)(a)].
(i) Nonionic surface active agent standard solution [0.1 mgCH3(CH2)l10
(CHzCH20)7H/ml] Weigh out 0.100 g of heptaoxyethylene dodecyl ether(9)
to its 100 %, dissolve in water, transfer into a 1O00 ml volumetric flask,
and add water t o the marked line. Prepare at the time of use.
Note (9) In the case where quality is confirmed, the test method specified
by The Chemical Society of Japan (CSJ) shall apply.
(2) Apparatus The following apparatus shall be used.
(b) Ion exchange resin column An example is given in Fig. 23.1.
Preparation of ion exchange resin column Take strong acid cation
exchange resin and strong basic anion exchange resin (I form) by 1 to 2 in
volume ratio. Fill the glass tube as given in Fig. 23.1 by adding water
sufficiently mixing not t o be mixed with air bubbles, and adjust the height
of an ion exchange resin column to approx. 200 mm. Pass 100 ml of etha-
nol (l+l).Allow this ion exchange resin column to be used repetitively
several times.
110
K O101 : 1998
Unit: mm
30 in inner diameter
10 in inner diameter
Glass filter plate G2
-- 4 to 5 in inner diameter
-
Fig. 23.1 An example of ion exchange resin column
(c) Photometer Spectrophotometer
(d) Absorption cell That made of quartz glass o r that equivalent in quality.
(3) Pretreatment Carry out the pretreatment as follows.
(a) Take 100 ml of a sample(10), add 100 ml of ethanol (95), and mix by shaking.
(b) Pass this solution a t a rate of 10 to 15 Z/(Z-resin h) through an ion exchange
resin column, and receive the effluent into a 500 ml beaker.
(c) When the surface of solution approaches the upper part of the ion exchange
resin pillar of the ion exchange resin column, add little by little 100 ml of
ethanol (l+l), and make the sample in the ion exchange resin column flow
out. Joint the effluent t o the 500 ml beaker of (b).
(d) Evaporate the effluent on a water bath to approx. 30 ml.
(e) After standing to cool, transfer this solution into a 100 ml volumetric flask,
Note (10) Regulate pH to approx. 7 by using a pH meter with sodium hy-
droxide solution (40 g/Z) for acidity and with hydrochloric acid
(1+11)for alkalinity.
(a) Take an appropriate quantity of the solution of (3)(e)[containing 0.1 t o
2 mg as CH3(CHa)llO(CH2CH20)7H]into a 200 ml separating funnel, and
dilute it with water to 100 ml.
(b) Add 15 ml ammonium tetrathiocyanatocobaltate (II) solution and 35 g of
sodium chloride(11). After mixing by shaking for approx. 1min, allow to
stand for approx. 15 min.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
111
K O101 : 1998
(c) Add 25 ml of benzene(121, mix by violently shaking for 1min, and allow to
stand.
(d) Discard the aqueous layer, transfer the benzene layer t o a beaker, add
approx. 5 g of sodium sulfate (anhydrous), and mix by shaking t o dehydrate.
(e) Transfer it to an absorption cell, use the benzene for which the operation
of (b)to (d)is performed on 100 ml of water as reference solution, and measure
the absorbance near 322nm in wavelength.
(0 As a blank test, take the same quantity as (a) of the solution of (3)( e )into
a 200 ml separating funnel, make 100 ml with water, use 15 ml of water
instead of 15 ml of ammonium tetrathiocyanatocobaltate (II) solution. Af-
ter performing the operation of (b) t o (d), use benzene as reference solu-
tion, obtain the absorbance near 322nm in wavelength, and correct the
absorbance obtained on the sample.
(g) Obtain the quantity of nonionic surface active agent from the working curve,
and calculate the concentration of nonionic surface active agent in the sample
[~~CH~(CH~)~I~(CH~CH~~)~H/ZI.
Wbrking curve Deal out step by step 1 t o 20 ml of nonionic surface ac-
tive agent standard solution [O. 1mgCH~(CH2)~~O(CH~CH~O)~Wmll into 200 ml
separating funnels, add water t o make 100m1, perform the operation (b)
to (e), and prepare the relation curve between the quantity of nonionic surface
active agent [CH3(CH2)liO(CH2CH20)7H]and the absorbance.
Notes (11) Potassium chloride may be used.
(12) 1,2-Dichloroethane may be used.
Remarks 8 When polyethylene glycol coexists, it is included in a determi-
nation value as nonionic surface active agent to make an error.
Therefore, after preliminarily extracting with 1-butanol speci-
fied in JIS K 8810 o r 2-butanone (ethyl methyl ketone) speci-
fied in JIS K 8900 to be removed, the pretreatment of (3)is
performed,
9 In the case where anionic surface active agent and cationic
surface active agent do not coexist, the pretreatment of (3)may
be omitted.
10 In the case where the concentration of nonionic surface active
agent is 1mg of C H ~ ( C H ~ ) I ~ O ( C H Z C H ~
or~ under,
) ~ H / Zconcen-
trate as follows, and operation is performed.
Add at a rate of 50 g of sodium chloride and 2.5 g of sodium
carbonate (anhydrous) Specified in JIS K 8625 per 500 ml of a
sample, dissolve, transfer to a 1O00 ml separating funnel, add
25 ml of ethyl acetate specified in JIS K 8361, mix by violently
shaking for approx. 2 min, and allow t o stand. Transfer the sepa-
rated ethyl acetate layer t o a beaker, add 25 ml of ethyl acetate
to the aqueous layer, and repeat extraction again. Join the sepa-
rated ethyl acetate layer into the preceding beaker. Heat the
ethyl acetate layer on a water bath, remove ethyl acetate by va-
porization, dissolve by adding a small quantity of methanol, and
add water to make a specified volume. Thereafter, perform the
pretreatment of (3),and determine.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
112
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
24 Dissolved oxygen The Winkler method, Winkler-sodium azide modification,

Miller's modification, or membrane electrode method applies t o the determination of
dissolved oxygen. This test shall be carried out immediately after sampling.
24.1 Winkler method Manganese (II) sulfate and alkaline potassium iodide are
added, the generated manganese (II) hydroxide is oxidized by the dissolved oxygen
t o form manganese (III) hydroxide. Then dissolve the precipitate by adding sulfuric
acid, and titrate the free iodine with sodium thiosulfate solution t o determine the
dissolved oxygen.
Determination range: O 0.1 mgll or more

Alkaline potassium iodide solution Dissolve 700 g of potassium hydrox-
ide specified in JIS K 8574 and 150 g of potassium iodide specified in JIS
K 8913 respectively in water, and mix to dilute with water t o 11. Pre-
serve this solution in a coloured bottle.
Iodine solution (50 mmol/Z) Dissolve 40 g of potassium iodide specified
in JIS K 8913 in a small amount of water, add to it 12.7 g of iodine speci-
fied in JIS K 8920 to dissolve, and dilute with water t o 11.
Iodine-alkaline potassium iodide solution Take 125 ml of alkaline po-
tassium iodide solution in a 250 ml volumetric flask, add 10 ml(1) of iodine
solution (50 mmol/l), and then add alkaline potassium iodide solution up
to the mark.
Manganese (II) sulfate solution Dissolve 480 g of manganese (II) sul-
fate 5-hydrate specified in JIS K 8997 in water t o make 11.
Sulfuric acid (3+1) Take 250 ml of water into a beaker. Add gradually
750 ml of sulfuric acid specified in JIS K 8951 shaking it while the beaker
is being cooled. Cool to room temperature, and then, add water to make
11.
Starch solution (10 g/Z) As described in 22.1.2 (1)(i).
50 mmol/Z Sodium thiosulfate solution Dissolve 12.5 g of sodium thio-
sulfate 5 hydrate Specified in JIS K 8637 and 0 . 2 g of sodium carbonate
specified in JIS K 8625 in water, add water to make 11. Stand for two
days. Use this solution after standardization a t the time of use.
Standardization Heat the potassium iodate (standard reagent for volu-
metric analysis) specified in JIS K 8005 a t 130 "C for approx. 2 h, and al-
low t o cool in a desiccator. Take its 0.357 g, dissolve i t in a small quantity
of water, transfer into a 200 ml volumetric flask, and add water up to the
mark. Take its 20 ml in a 300 ml Erlenmeyer flask with ground stopper,
add 2 g of potassium iodide specified in JIS K 8913 and 5 ml of sulfuric
acid (1+5), immediately stopper tightly to mix by shaking gently, and al-
low to stand in a dark place for 5 min.
Add t o this solution 100 ml of water, and titrate the liberated iodine with
this sodium thiosulfate solution. When the yellow colour of the solution
has become pale, add 1ml of starch solution (10 gll) as indicator, and ti-
trate until the blue colour of the generated iodine starch disappears.
113
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
K O101 : 1998
Separately carry out the blank test under the same condition for water
to correct, and calculate the factor (f> of 50 mmol/Z sodium thiosulfate so-
lution from the correct numeric of ml according t o the following formula:
b 20 1
= a x?ÖÖx%Öx x x 0,001783
where, x : 50 mmol/Z sodium thiosulfate solution required for

titration (corrected value) (mi)
a : amount of potassium iodate (9)
b : content of potassium iodate (%)
0.001 783 : potassium iodate equivalent to 1ml of 50 mmol/Z
sodium thiosulfate (g)
(h) 5 mmol/Z Sodium thiosulfate solution Take 25 ml of 50 mmol/Z sodium
thiosulfate solution in a 250 ml volumetric flask, and add water up to the
mark. Prepare this solution at the time of use, and do not use the solution
for which not less than 12 h has elapsed after preparation.
Note (1) For ordinary test, prepare it by adding 10 ml of iodine solution
(50 mmol/Z) to alkaline potassium iodide solution. If required, obtain
the amount of reducing substances such as sulfite ion, sulfide ion,
etc., and determine the adding amount by calculating the amount
of iodine corresponding thereto.
For example, add 16 ml of iodine solution (50 mmol/E) t o alka-
line potassium iodide solution for 1 mg of sulfite ion (SO32-) and
0.2 mg of hydrazinium ion (NzHs+).
(a) Sampler Two samplers with capacity of (500+5) ml as shown in Fig. 24.1.
114
K 0101 : 1998
Unit : mm
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
- "_ I _ -
Sampler (500 mi) Details of part A (part B)

Fig. 24.1 An example of sampler
(b) Magnetic stirrer
(3) Sampling from piping and apparatuses Carry out the sampling as follows.
Direct outlets of two samplers upward, assemble so that the inlets of Sam-
plers can be held a t the higher position than the sampling mouth of the
piping, and connect the lower end of the sampler t o sampling mouth with
soft polyvinyl chloride tube (or thick rubber tube) and Y-tube (to the outlet
of sampler, connect nothing).
Where the temperature of sample is higher than room temperature, pro-

vide a proper cooling spiral tube(2) in the sample piping s o as the tempera-
ture of sample to be lower than room temperature by 1 to 2 O C .
Adjust the flow rate of sample so that the both two samplers are filled in
40 t o 60 s at the same time, and then s o sufficiently make the sample flow
continuously so that the original sample in the sample piping is substi-
tuted completely(3).
115
K O101 : 1998
(d) Close the cocks of parts A of two samplers, immediately close cocks of parts
B of both two samplers, detach the connecting tubes, and confirm that there
is no air bubbles completely with the samplers reversing. If there are any
air bubbles, newly take sample in both samplers.
(e} Use one sampler for main test, and the other for blank test.
Notes (2) Where the cooling spiral tube is used, set a cooling water ad-
justing valve at the inlet of cooling spiral tube to make the cool-
ing water overflow, and set a sample flow-rate adjusting valve
a t the outlet of cooling spiral tube.
(3) Where the sample piping is used even intermittently, make the
sample flow for a time required for substituting completely the
original sample in the sample piping and cooling spiral tube.

Drip off the water in the tube of part A of sampler for test and fill iodine-
alkaline potassium iodide solution up to the base line at the uppermost
part of part A tube. Where there are any air bubbles in the tube, remove
them with a clean copper wire.
Open the cock of part A, and put the iodine-alkaline potassium iodide so-
lution until the solution surface aligns with the base line of lower part while
adjusting with a cock of part B. Then, close both cocks, and wash the in-
side of tubes of part A and part B of the sampler by using washing bottle.
Reverse the sampling mouth, fill manganese (II) sulfate solution up t o the
base line of the uppermost part of part B tube in accordance with the op-
eration of (a) as appropriate, and put manganese (II) sulfate solution in
accordance with the operation of (b)as appropriate.
After washing the inside of tubes of part A and part B of the sampler by
using washing bottle, overturn the sampler repeatedly for about 1min t o
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
mix sufficiently and allow t o stand for a while.

Again, overturn the sampler t o suspend homogeneously the precipitates gen-
erated in the sampler, quickly fill sulfuric acid (3+1)up t o the base line of
uppermost part of part B tube, add sulfuric acid (3+1)in accordance with
(a) as appropriate and overturn t o combine with mixing,
Carry out accurately the adding operation of reagent, and carry out the
operation specified in (a) to (e) within 15 min after sampling.
Discharge 4 to 10 ml into a 10 ml measuring cylinder from the part A tube
of sampler, record the amount and discard.
Transfer the residual solution into a porcelain evaporating dish, titrate with
5 mmol/Z sodium thiosulfate solution while stirring with a glass rod o r
magnetic stirrer to mix, after the yellow colour of the solution has become
pale add 3 ml of starch solution (10 gll) as indicator to mix, and titrate until
the blue colour of iodine starch disappears.
116
K O 1 0 1 : 1998
(i) For the sample of sampler for blank test, put-in iodine-alkaline potassium
iodide solution from the part A tube with the same operation as in (a) and
(b),and then add sulfuric acid (3+1) from the part B tube according to the
operation of (e). After mixing t o combine, put-in the manganese (II) sulfate
solution with the same operation as in (cl,and mix to combine sufficiently.
ci) Using the same porcelain evaporating dish as in the case of sample, titrate
with the operations specified in ( g ) and (h).
(k) Calculate the concentration of dissolved oxygen in the sample (mgOIZ) ac-
o= ---
a;[ ;b]xloooxfno.oa-o.olo4
where, O : dissolved oxygen (mgO/Z)

a : 5 mmol/Z sodium thiosulfate solution required for
titration (mi)
b : 5 mmolll sodium thiosulfate solution required for
titration of blank test (mi)
Va : titrated sample (ml) (the amount subtracted by the
discarded amount before titration from the capacity
of sampler)
v
b : titrated sample for blank test (ml) (the amount sub-
tracted by the discarded amount before titration
from the capacity of sampler)
f : factor of 5 mmol/Z sodium thiosulfate solution(4)
0.04 : oxygen equivalent to 1ml of 5 mmol/Z sodium thio-
0.010 4 : corrected value of dissolved oxygen in the added
reagent
Note (4) Use the factor of 50 mmoVZ sodium thiosulfate solution.
Remarks 1 Where there exists not less than 1mgOlZ of dissolved oxygen,
25 mmolll sodium thiosulfate solution may be used.
2 For converting (mgOlZ) of dissolved oxygen to {mlO/Z), multi-
ply 0.7004.
24.2 Winkler-sodium azide modification The nitrite ion interfering the Winkler
method is decomposed by adding sodium azide, and then the dissolved oxygen is
determined.
Determination range: O 0.5 mgll o r more
(a) Alkaline potassium iodide-sodium azide solution Dissolve respectively
350 g of potassium hydroxide specified in JIS K 8574 (or 250 g of sodium
hydroxide specified in JIS K 8576) and 75 g of potassium iodide specified
in JIS K 8913 in water, mix, and add water to make 500 ml. Separately
dissolve 5 g of sodium azide specified in JIS K 9501 in 20 ml of water, and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
117
K O101 : 1998
mix them also. Put into a light shielded polyethylene bottle t o be preserved
in a dark place.
Manganese (II) sulfate solution As described in 24.1 (i)(d).
Starch solution (10 glZ) As described in 22.1.2 (1) (i).
25 mmol/Z Sodium thiosulfate solution Take 100 ml of 50 mmol/Z so-
dium thiosulfate solution of 24.1 (i)(g)into a 200 ml volumetric flask, dis-
solve 0.1 g of sodium carbonate specified in JIS K 8625 in water, add it
thereto, and further add water to the marked line. Prepare this solution
immediately before the use, and never use it 12 h or longer elapsed.
(a) Dissolved oxygen measuring bottle As described in 19 (2)(a).
(3) Sampling Carry out the sampling as follows.
In the case of using sampler In the case of using Vandorn sampler,
insulation sampler, etc., connect the soft vinyl chloride tube to the flow-
ing-out opening of the sampler, and let this soft vinyl chloride tube enter
t o the bottom of the dissolved oxygen measuring bottle. Let flow the sample
1
into the dissolved oxygen measuring bottle quickly up t o about - with taking
3
care so as the air bubbles not to be generated, and then wash the measur-
ing bottle. In the same operation, let the sample enter newly the measur-
ing bottle, and let overflow the sample of 25 to 50 % of the capacity of bottle,
then take out the soft vinyl chloride tube gently, and stopper tightly so
that no air bubble remains.
In the case of sampling from piping and appliances Attach a soft
vinyl chloride tube to the sampling valve fixed to the piping and appliances,
and continuously pass water at a rate of approx. 1 Umin. Enter the soft
vinyl chloride tube to the bottom of dissolved oxygen measuring bottle,
overflow the sample of the amount about 5 times the capacity of the mea-
suring bottle, then take out the soft vinyl chloride tube, and stopper tightly
so that no air bubble remains.
In the case of direct sampling In the case of direct sampling of the
surface water of rivers, drain, effluent reservoir, etc. in the dissolved oxy-
gen measuring bottle, at first wash the measuring bottle sufficiently with
the sample, and put the measuring bottle under the water surface. Let the
sample flow gently into the bottle to fill completely, and stopper tightly so
that no air bubble remains. In the case of sampling with a bucket, let flow
into the bucket with the same operation and stopper tightly.
(a) Take out the stopper of the dissolved oxygen measuring bottle, add quickly
to it 1 ml of manganese (II) sulfate solution and 1ml of alkaline potassium
iodide-sodium azide solution per 100 ml of sample by immersing the tip of
pipette into the sample respectively, and stopper tightly so that no air remains
in the bottle.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
118
K 0101 : 1998
Repeat the inversion of bottle for about 1min, and mix thoroughly so that
the formed precipitates disperse in the whole bottle.
Allow to stand still for a while t o settle the precipitates, and after carrying
out the operation specified in (b), allow t o stand still.
When the precipitates have settled and the supernatant liquid has become
1
about - of the whole bottle, open the stopper gently, and add 1 ml of sulfuric
2
acid per 100 ml of sample with a pipette along the neck of the bottle. Again
stopper tightly, and invert the bottle several times t o dissolve the precipi-
tates.
Take separately a proper amount (it may be the whole amount) of this solution
and transfer into an Erlenmeyer flask.
Titrate with 25 mmol/l sodium thiosulfate solution, after the yellow colour
of the solution has become pale, add 1ml of starch solution (10 g/Z) as in-
dicator, and titrate until the blue colour of the iodine starch disappears.
Calculate the concentration of dissolved oxygen in the sample (mgO/Z) ac-
where, O : dissolved oxygen (mgOlZ)

titration (mi)
vi : capacity of the dissolved oxygen measuring bottle
when the ground stopper is applied (ml)
vz : the sample taken from the dissolved oxygen mea-
suring bottle to titrate (mi)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
2): sum of alkaline potassium iodide-sodium azide
solution and manganese (II) sulfate solution (ml)
f: factor of 25 mmol/Z sodium thiosulfate solution(5)
0.2 : oxygen equivalent to 1ml of 25 mmol/Z sodium thio-
Note (5) Use the factor of 50 mmol/Z sodium thiosulfate solution specified
in 24.1 (1)(g).
Remarks 3 In the case of sample containing oxidizing substances
Where the water contains residual chlorine o r the like, take
the sample for blank test by using another dissolved oxygen
measuring bottle in accordance with the operation specified
in (3)Sampling, then add t o this sample alkaline potassium
iodide-sodium azide solution and sulfuric acid, stopper tightly,
and repeat mixing with inverting the bottle. Then, add man-
ganese (II) sulfate solution, stopper tightly, and repeat mix-
ing with inversion. Titrate this solution by the operation
specified in (4) (e) t o (g),and correct the amount of dissolved
oxygen.
119
K O 1 0 1 : 1998
4 In the case of sample containing reducing substances It

is preferable to use iodine-alkaline potassium iodide solution
specified in 24.1 (i>(c) instead of alkaline potassium iodide-
sodium azide solution. In this case also carry out blank test
by using another dissolved oxygen measuring bottle the same
as in Remarks 3, and correct the amount of dissolved oxygen.
5 In the case of sample of sea water Because the sea water
contains microorganisms in many cases, accelerate the reac-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
tion t o test quickly. For accelerating the reaction, add respec-
tively twice quantity of alkaline potassium iodide-sodium azide
solution and manganese (II) sulfate solution, and then add twice
quantity of sulfuric acid after inversion.
6 In the case where iron (III) coexists in sample If 1ml of
potassium fluoride solution (300 gll) per 100 ml of sample is
added before adding sulfuric acid, presence of 100 to 200mglZ
of iron (III) does not interfere.
24.3 Miller’s modification By shielding the sample from air with liquid paraf-
fin, sodium potassium tartrate-sodium hydroxide solution and 3,7-bis (dimethylamino)
phenothiazine-5-iumchloride(Methylene Blue) solution are added, and then the so-
lution is titrated with ammonium iron (II) sulfate solution to determine the dissolved
oxygen.
Determination range: O 1mglZ or more
Sodium potassium tartrate-sodium hydroxide solution Dissolve 350 g
of sodium potassium (+)-tartrate 4 hydrate specified in JIS K 8536 and
100 g of sodium hydroxide specified in JIS K 8576 in water t o make 11.
Methylene Blue solution Dissolve 0.1 g of Methylene Blue (usually
trihydrate) in 100 ml of water.
Liquid paraffin As specified in JIS K 9003.
Ammonium iron (II) sulfate solution Add 5 ml of sulfuric acid specified
in JIS K 8951 to 100 ml of water, add to this solution 5.4 g of ammonium
iron (II) sulfate 6 hydrate specified in JIS K 8979 to dissolve, and add water
containing no dissolved oxygen specified in 2 (12) (a>to make 11.
Standardization For obtaining the corresponding amount of dissolved
oxygen of this solution, titrate with this solution in accordance with (3)
Operation, using the water of which concentration of dissolved oxygen is
obtained by 24.2 as standard, and calculate it according to the following
formula:
where, f : equivalent amount of dissolved oxygen to 1ml of

ammonium iron (II) sulfate solution (mgOj
a : dissolved oxygen in the w a t e r used (mgOlE)
120
K O101 : 1998
b : ammonium iron (II) sulfate solution required for

titration (mi)
Carry out this standardization at the time of use.
Sampler A 5 0 m l syringe which is connected with a glass tube of
approx. 1.5 mm in inner diameter and 250 to 300 mm in length by ground
glass joint or by rubber tube at the tip.
Dissolved oxygen measuring test tube A test tube of approx. 30 mm
in outer diameter and of approx. 200 mm in height.
Stirring rod A glass rod of approx. 3 mm in diameter and of approx. 250 mm
in total length the lower part of which is bent by approx. 10 mm o r is made
spiral.
Long stem burette The burette which has a capacity of 5 to 10 ml, and
the stem’s tip of which reaches the bottom of the test tube.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Add 2 drops of Methylene Blue solution, 5 ml of sodium potassium tartrate-
sodium hydroxide solution and approx. 5 ml of liquid paraffin to the dis-
solved oxygen measuring test tube.
(b) Suck the sample into the sampler, rinse i t twice, and then suck so gradu-
ally that no air bubble enters t o take 50 ml of the sample. At this time,
keep the conditions filled with the sample in the glass tube.
(cl Immerse the tip of glass tube of the sampler gently under the liquid paraf-
fin layer, and inject 50 ml of sample into it with taking care not to disturb
the liquid paraffin layer.
(d) Gently insert a stirring rod in the solution, and insert the tip of the long
stem burette under the liquid paraffin layer.
(e) Titrate with the ammonium iron (II) sulfate solution until the blue colour
of the Methylene Blue disappears.
(0 Calculate the concentration of dissolved oxygen in the sample (mgOlZ) ac-
1 O00
O=axfx-
50
where, O : dissolved oxygen (mgO/Z)
a : ammonium iron (II) sulfate solution required for
titration (ml)
f : equivalent amount of dissolved oxygen to 1 ml of
ammonium iron (II) sulfate solution (mg)
121
K 0101 : 1998
24.4 Membrane electrodes method Dissolved oxygen in the sample is measured

by use of membrane electrodes.
Determination range: O 0.5 mg/Z or more
(a> Sodium sulfite solution Dissolve approx. 25 g of sodium sulfite speci-
fied in JIS K 8061 in water, and dilute with water t o 500 ml. Prepare this
solution at the time of use. Use this solution for zero adjustment(".
(b) Dissolved oxygen saturated water(8) Pass through water the air washed
by potassium hydroxide solution (250 g/Z) a t a flow rate of approx. 1Zlmin
by use of spherical o r plate-like sintered glass filter(7) to saturate the dis-
solved oxygen(". Prepare this solution immediately before carrying out
the span adjustment operation.
Notes (6) If a trace of cobalt (II) chloride 6 hydrate specified in JIS K 8129
is added the dissolved oxygen is easily reduced by sodium sulfite.
(7) Usually, in the case of 200ml of water, air is passed for 5 t o
10 min, and in the case of 500 ml, for 10 t o 20 min.
(8) Prepare the dissolved oxygen saturated water coincident with the
temperature of the sample within 20.5 O C . Obtain the concen-
tration of dissolved oxygen in this solution from Table 24.1. Since
the concentration of dissolved oxygen differs also by fluctuation
in atmospheric pressure, allow the atmospheric pressure t o be
preferably corrected.
Further, in the case where measuring the concentration of dis-
solved oxygen in the sample of high salts concentration, prepare
the dissolved oxygen saturated water to which sodium chloride
specified in JIS K 8150 is added conforming to the molar con-
centration of salts in the sample.
(a) Dissolved oxygen measuring container A glass container of 100 to
300ml attached with rubber stopper, to which sample injection tube for
taking the sample and a syphon for discharging the sample(9) are attached.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
An example of it is shown in Fig. 24.2.
122
K O101 : 1998
Sample injection
opening
I
A: Rubber tube
B: Glass container
C : Rotor
D: Magnetic stirrer
E: Thermometer
F: Discharge tube
G: Rubber tube
H: Pinch cock
I: Membrane elec-
trode
Fig. 24.2 An example of measuring container

Thermometer 50 "C thermometer of solid-stem general purpose liquid-
Dissolved oxygen meter Generally, the dissolved oxygen meter into which
a temperature compensating circuit is incorporated is used.
Magnetic stirrer
Note (9) The dissolved oxygen measuring bottle, incubation bottle, etc. may
be used.
(3) Preparatory operation Carry out the preparatory operation as follows,

Connect the electrode of the dissolved oxygen meter, and apply current for
approx. 30 min.
Inject sodium sulfite solution at the same temperature as that of the sample
into a measuring container, immerse electrodes while stirring gently t o mix
with a magnetic stirrer, and after the indicating value has been stabilized(l9,
adjust the indicating value to zero by rotating the zero adjuster dial.
Take out the electrodes and the thermometer, wash thoroughly with wa-
ter(11), and insert them into another measuring container.
Inject the dissolved oxygen saturated water(l2) gently from an end of injec-
tion tube to the bottom of the measuring container, and after flowing out
the water by 25 t o 50 % of the capacity of measuring container, close the tip
of syphon.
While mixing with a magnetic stirrer (19, wait the stabilization of indicat-
ing value of dissolved oxygen meter. Then, read out the temperature, and
obtain the corresponding dissolved oxygen saturation amount from Table
24.1. Then rotate the span adjusting dial t o adjust the indicating value.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
123
K O101 : 1998
(0 Repeat the operation specified in (b) to ( e ) two o r three times, and confirm
that the indicating values coincide with zero and the saturation amount of
dissolved oxygen respectively.
Notes (10) Usually, it requires 2 to 5 min.
(11) At the time of changing the operation from (b)to ( c ) , wash the
electrodes particularly thoroughly.
(12) Dissolved oxygen saturated water may be prepared in the con-
tainer by aeration.
(13) Since the indicating values are different according t o the speed
of stirring to mix, keep the same condition as that a t the time
of span adjusting operation as far as possible.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Inject gently the sample to the bottom of measuring container so that no
bubble enters in accordance with (3) (d)(14).
(b) While mixing with magnetic stirrer(l3), confirm the scale of the thermom-
eter(8) and after waiting the stabilization of indicating value of dissolved
oxygen meter, read out the indicating value (mgOll).
Note (14) When a dissolved oxygen measuring bottle, an incubation bottle
or the like is used as measuring container, take the sample gen-
tly into a container by using a syphon, and immediately immerse
the electrodes and a thermometer t o measure.
Remarks 7 The indicating value increases by approx. 5 % each tempera-
ture rise of 1O C .
124
K O101 : 1998
Table 24.1 Saturated amount of dissolved oxygen in water
Tempera- Amount of dissolved

ture oxygen to be sub-
0 5 000 10 000 15 000 20 o00 tracted for every
"C
100 mgCl-ll of
Amount of dissolved o gen mgûil chloride ion rnczOli
0 14.16 13.40 12.63 11.87 11.10 0.015 3
1 13.77 13.03 12.29 11.53 10.80 0.0148
2 13.40 12.68 11.97 11.25 10.52 0.0144,
3 13.04, 12.35 11.65 10.95 10.25 0.0140
4 12.70 12.03 11.35 10.67 9.99 0.013 5
5 12.37 11.72 11.O6 10.40 9.74 0.013 1
6 12.06 11 .42 10.79 10.15 9.51 0.012 8
7 11.75 11.15 10.52 9.90 9.28 0.0124
8 11.47 10.87 10.27 9.67 9.06 0.0120
9 11.19 10.61 10.03 9.44 8.85 0.011 7
10 10.92 10.36 9.79 9.23 8.66 0,011 3
11 10.67 10.12 9.57 9.02 8.4,7 0.011 o
12 10.43 9.90 9.36 8.82 8.29 0.010 7
13 10.20 9.68 9.16 8.64 8.11 0.0104
14' 9.97 9.47 8.97 8.46 7.95 0.010 1
15 9.76 9.27 8.78 8.29 7.79 0.0099
16 9.56 9.06 8.60 a. 12 7.63 0.0096
17 9.37 8.90 8.44 7.97 7.49 0. 009 e 4
18 9.18 8.73 8.27 7.82 7.36 0.009 1
19 9.01 8.57 8.12 7.67 7.22 0.008 9
20 8.84 8.41 7.97 7.54 7.10 0.008 7
21 8.68 8.26 7.83 7.4,0 6.97 0.008 6
22 8.53 a . ii 7.70 7.26 6.85 0.008 4
23 8.39 7.98 7.57 7.16 6.74 0.008 2
24, a.25 7.85 7.44 7.04 6.65 0.008 1
25 a. i1 7.72 7.32 6.95 6.52 0.007 Y
26 7.99 7.60 7.21 6.82 6.42 0.007 8
27 7.87 7.48 7.10 6.71 6.32 0.007 7
28 7.75 7.37 6.99 6.61 6.22 0.007 6
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
29 7.6% 7.26 6.88 6.51 6.12 0.007 6

30 7.53 7.16 , 6.78 6.41 6.03 0.007 5
31 7.43 7.06 j 6.66 6.31 5.93 0.007 5
32 7.32 6.96 6.59 6.21 5.84 0.0074
33 7.23 6.86 ~ 6.49 6.12 5.75 0.00ï 4
34 7.13 6.77 ~ 6.40 6.03 5.65 0.0074
I
35 7.04 6.67 I 6.30 5.93 5.56 0.0074
125
K O101 : 1998
26 Total carbonate The total carbonate is a combined amount of carbonic acid,

hydrogen carbonate ion and carbonate ion, and shall be expressed by the amount of
carbon dioxide (COS). For the determination of total carbonate, the strontium chlo-
ride-hydrochloric acid titration method or the infrared analytical method shall be used.
25.1 Strontium chloride-hydrochloric acid titration method Add the sample

to sodium hydroxide solution to transform the total carbonate to carbonate ion. Then,
add strontium chloride to generate precipitate of strontium carbonate. Add hydro-
chloric acid to neutralize the excessive sodium hydroxide, and further add a definite
amount of hydrochloric acid to dissolve the precipitate. After removing the liber-
ated carbon dioxide by passing air, titrate the excessive hydrochloric acid with so-
dium hydroxide solution and obtain the amount of consumed hydrochloric acid to
determine the total carbonate.
Determination range: CO2 1 to 40mg
Repeatability: 2 to 10 96 in coefficient of variation
(a) Water As described in 20.1 (i)(a).
(b) Strontium chloride solution Dissolve 17 g of strontium chloride 6 hy-
drate specified in JIS K 8132 in water to make 100 ml.
(c) Phenol phthalein solution (5 g/Z) As described in 13.2 ( i )(a).
(d) 0.1 mol/¡! hydrochloric acid Take 10 ml of hydrochloric acid specified
in JIS K 8180 into a 1 O00 ml beaker contained 100 ml of water, and add
water to make 1 1 .
Standardization Heat sodium carbonate (standard reagent for volumet-
ric analysis) specified in JIS K 8005 at 600 "C for approx. 1 h, and allow to
cool in a desiccator. Weigh out approx. 1 g of this to the nearest 1mg, dis-
solve in a small quantity of water, put it into a 200ml volumetric flask,
and add water up to the mark. Take 20 ml from this solution, transfer into
a 200 ml Erlenmeyer flask, add 2 or 3 drops of Methyl Red-Bromocresol Green
mixed solution(1) as indicator, and titrate with this 0.1 mol/Z hydrochloric
acid until the colour of the solution indicates grayish purple (pH 4.8).
Boil at near end point to expel carbon dioxide, and after cooling, again
titrate until it indicates grayish purple. Calculate the factor Cfl) of 0.1 mol/
I hydrochloric acid according to the following formula from the number of
ml of 0.1 moVZ hydrochloric acid required for titration:
f1=CJX-
b X-
20 1
100 200 x x 0.005 30
where, a : amount of sodium carbonate ( g )

b : content of sodium carbonate (96)
x : 0.1 mol/l hydrochloric acid required for titration
(ml1
0.005 30 : equivalent amount of sodium carbonate t o 1ml of
0.1 mol/Z hydrochloric acid (9)
Note (1) As described in 13.1 ( l ) ( a ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
126
K O101 : 1998
(e) 40 mmol/Z hydrochloric acid Take 100 ml of 0.1 mol/Z hydrochloric acid
in a 250 ml volumetric flask, add water up to the mark. For factor of this
solution, use the factor of 0.1 mol/Z hydrochloric acid.
(0 40 mmol/Z sodium hydroxide solution Take approx. 30 ml of water i n
a polyethylene bottle, dissolve approx. 35 g of sodium hydroxide by adding
little by little, while cooling, and tightly stopper to allow to stand for 4 to
5 days. Take 2 ml of this supernatant liquid in a 1I airtight polyethylene
vessel, and add water containing no carbonic acid of 2 (12)(b) t o make 11.
Put this solution into a reagent bottle (made of polyethylene) attached
with automatic burette (50ml in capacity), and attach a tube filled with
soda lime specified in JIS K 8603 o r potassium hydroxide grains specified
in JIS K 8574 at the inlet and outlet openings of air to store.
Standardization Take 20 ml of 40 mmol/Z hydrochloric acid specified in
( e ) in a 200 ml Erlenmeyer flask, add 2 to 3 drops of phenolphthalein solu-
tion (5 g/Z) as indicator, and titrate with this 40 mmol/Z sodium hydroxide
solution until the colour of the solution indicates faint red.
Calculate the factor (fz) of the 40 mmol/Z sodium hydroxide solution ac-
cording t o the following formula from the number of ml of 40mmol/Z so-
dium hydroxide solution required for titration:
where, fl : factor of 40 mmol/Z hydrochloric acid

x : 40 mmol/Z sodium hydroxide solution required for
titration (ml)
(a) Total carbonate measuring apparatus An example is shown in Fig. 25.1.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
127
K O101 : 1998
‘i
\
Unit: mm
A: 500 ml Erlenmeyer flask C: Gas washing bottle

B: 300mm Gas drying tower c : Put with sulfuric acid
a: Put with glass wool D: One-way cock
b: Put with soda lime or E: Rubber tube
potassium hydroxide grains
Fig. 25.1 An example of total carbonate m e a s u r i n g apparatus

(3) O p e r a t i o n Carry out the operation as follows.
Add 50 ml of 40 mmol/Z sodium hydroxide solution and 0.2 ml of phenol-
phthalein solution (5g/Z) as indicator to a 500 ml Erlenmeyer flask.
Add 200 ml of the sample(2) (when the total carbonate in the sample is of
large amount, take a suitable amount so as t o become 40 mg o r less, and
add water to make 200 ml), and connect a 500 ml Erlenmeyer flask as shown
in Fig. 25.1 to allow to stand for several min.
Add 10 ml of strontium chloride solution, again connect to shake sufficiently
t o mix, and allow to stand for approx. 10 min.
Add 40 mmoVZ hydrochloric acid dropwise, and neutralize until colourlessness
is kept for 1min(3).
Add 50 ml of 40 mmol/Z hydrochloric acid and again connect.
Pass air a t a rate of 1 Umin for 5 min.
Remove the 500ml Erlenmeyer flask, and wash the air introducing tube
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
with water. Next, titrate gradually with 40 mmol/Z sodium hydroxide so-
lution, and take the point when the colour of the solution indicates faint
pink as the end point.
For blank test, take 200 ml of water into a 500 ml Erlenmeyer flask, and
carry out the operation of (a) to (g).
128
K O101 : 1998
(i) Calculate the concentration of total carbonate in the sample (mgCO2lZ)

c = ( b - a)x f 2 x -
*Oo x 0.880 2
V
where, C : total carbonate (mgCO2lZ)
titration (mi)
b : 40 mmol/Z sodium hydroxide solution required for
f z : factor of 40 mmol/Z sodium hydroxide solution
V : sample (mi)
0.880 2 : equivalent amount of carbon dioxide to 1 ml of
40 mmol/Z sodium hydroxide solution (mg)
Notes (2) The solution in the Erlenmeyer flask shall be kept for pH so as
not to be 12 or less. pH shall be confirmed by using alkali blue
pH test paper. Where the sample is acidic, such amount of
40mmol/Z sodium hydroxide solution that it is able to neutral-
ize to approx. 7 in pH shall be added.
(3) Where a large amount of magnesium ion coexists, t h e dis-
colouration at the time of neutralization may be difficult to find,
and therefore gradually titrate when approached t o the end point.
Remarks 1 This method is applicable where the limits shown in the fol-
lowing for respective components are not exceeded.
200 mgMgíZ
25 mgFelZ
5 mgP043-lZ
Fe2+
Pod3-
1 coexist
5 mgFe/Z
10 mgP02-/Z
Fe3+ 1
2.5 mgFelZ
coexist
Pod3- 5 mgP043-lZ
Furthermore, the coexistence of large amounts of aluminium,
ammonium ion, silica, etc. becomes interferent.
2 Calculation of concentrations of carbonic acid, hydrogen car-
bonate ion and carbonate ion.
The respective concentrations of carbonic acid, hydrogen car-
bonate ion and carbonate ion can be calculated from the con-
centration of total carbonate and pH of the sample according
to the following formula and Table 25.1 o r Fig. 25.2.
H2co3 = C X a X 1.409
HC0,- = C x b x 1.387
c03'- = C X c X 1.364
where, H2C03 : carbonic acid (mgH&O3/Z)

HC03- : hydrogen carbonate ion (mgHCO3-íZ)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
129
K O101 : 1998
cos2-: carbonate ion (mgC032-/Z)

C: total carbonate (mgCo&
a: mole ratio of carbonic acid t o total carbon-
ate
b: mole ratio of hydrogen carbonate ion t o to-
tal carbonate
c: mole ratio of carbonate ion t o total carbon-
ate
1.409 : coefficient in the case where the amount of
total carbonate (COZ)is converted t o carbonic
acid equivalent (62.03/44.01)
total carbonate (COZ) is converted t o
hydrogencarbonate ion equivalent (61.02/
44.01)
total carbonate (COZ) is converted to carbon-
ate ion equivalent (60.0u44.0i)
Table 25.1 Concentration distribution of total carbonate relative to pH
Concentration distribu Concentration distribution (25 "C)

b (HC0,-) a (H,CO,) 1 b (HCO,-)
2.0 I.0000 - 0.022 6 0.972 8 0.0046

2.5 0.9999 0.000 1 0.00'7 2 0.9783 0.0145
3.0 0.9996 0.000 4 0.002 2 0.9530 0.0448
3.5 0.998 6 0.001 4 0 . O00 6 0.870 1 0.1293
4c.0 0.993 7 0.0043 0 . 000 2 0.680 1 0.3197
4.5 0.986 6 0.0134 - 10.5 0 , O00 0 0.4022 0.507 8
5.0 0.958 7 0,041 3 __ 11.0 ~ 0.1754 0.824 6
5.5 0.880 0 0,1200 -
11.5 - 0.063 0 0.937 O
6.0 0.6988 0.301 2 0.000 0 12.0 0.020 8 0.979 2
0.423 2 0.576 7 0.000 1 12.5 - 0.006 7 0,993 3
6.5
- 7.0 0 . 188 3 0.811 3 0.0004 1 13.0 0.002 1 0.9979

7.5 0.0683 0.930 3 0.001 4. I
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
130
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Curve a: H2C03, b: HC03-, c: co32-

These indicate respective mole ratios of carbonic acid,
hydrogencarbonate ion and carbonate ion relative to total car-
bonate in Table 25.1, and Fig. 25.2 and in pH (25 OC) of the
sample.
Fig. 25.2 Concentration distribution of total carbonate
relative to pH (25 O C )
25.2 Infrared analytical method This is a method t o determine the total car-
bonate by operating as same as in the case of inorganic carbon t o be measured in
determining the organic carbon (TOC) according to the infrared analytical method.
Determination range: COZ 3 t o 450mgCOnlZ
(a) Water Use the water of 20.1 (1)(a).
(b) Carbonate standard solution (0.5 mgCO2lmZ) Allow sodium hydrogen-
carbonate specified in JIS K 8622 to stand in a desiccator for approx. 3 h,
and take its 0.478 g. Separately preliminarily heat sodium carbonate, stan-
dard reagent for volumetric analysis specified in JIS K 8005 at 600 OC for
approx. 1 h, allow t o stand t o cool in a desiccator, and take its 0.602 g . Dis-
solve both in a small amount of water, transfer into a 1O00 ml volumetric
flask, and add water to the mark.
(c) Carbonate standard solution (0.1 mgCOalmZ) Take 20 ml of carbonate
standard solution (0.5 mgCOalm1) in a 100 ml volumetric flask, and add water
up to the mark.
(d) Inorganic carbon measuring tube As described in 20.1 (1)(g).
(e) Carrier gas As described in 20.1 (1)(h).
(a) Microsyringe 10 pl and 150 pl
(b) Total carbonate determination apparatus As described in 20.1 (2)(b).
131
K 0101 : 1998

(a) Inject 20 p.1 carbonate standard solution (0.1 mgCOdml) [or carbonate stan-
dard solution (0.5 mgCO2/ml)l into a total carbonate determination appa-
ratus with a microsyringe in accordance with 20.1 (3)(a)to (e)as appropriate,
and read a peak height to be indicated with an indicator [an infrared gas
analyzer (recorder)].
(b) Hereafter, carry out the operation of 20.1 (3)(d).
(a) Prepare the working curve by using carbonic acid standard solution
(0.1 mgCO2lml) [or carbonic acid standard solution (0.5 mgCOz/rnl)l in ac-
cordance with 20.1 (4) (e) t o (g) as appropriate.
(a) Operate in accordance with 20.1 (51, as appropriate, inject the same amount
of sample as that of carbonate standard solution used for preparation of
working curve into the total carbonate determination apparatus with the
microsyringe two o r three times and read out the corresponding peak height.
(b) Obtain the concentration of total carbonate (mgCOdZ) in the sample from
the preliminarily prepared working curve.
Remarks 3 Where 20.1 (1) (e) is used as the standard solution for prepa-
ration of working curve, calculate the concentration of total
carbonate (mgCO2lE) according t o the following formula:
Total carbonate (COZ)(mgCOdZ)
= inorganic carbon (mgC/Z)x 3.664
where, 3,664 : coefficient in the case where the amount of
carbon is converted to the total carbonate
equivalent (COS)
4 In the case where the concentrations of carbonic acid, hydro-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
gen carbonate ion and carbonate ion in the sample are calcu-
lated, carry out the operation in accordance with Remarks 2.
132
K O101 : 1998
26 Hexane extracts Hexane (n-hexane) extracts mean the substances which re-
main when hexane is volatilized at about 8 0 ° C after extracting with hexane from
slightly acidic sample solution.
This test applies mainly t o the determination of mineral oil, and animal and veg-
etable fats and oils which are difficult t o volatilize. However, these which are ex-
tracted by hexane and difficult to volatilize are contained in the determination value.
To this test, the extraction method shall be applied.
26.1 Sampling The sampling shall be carried out as follows.

Reagents The following reagents shall be used.
(a) Hydrochloric acid (1+1) Prepare by using hydrochloric acid specified in
JIS K 8180.
(b) Methyl Orange solution (1 gll) As described in 22.1.1 (1) (d).
Sample container In the case of surface layer water and falling water,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
use a 1 t o 2 1 wide-mouthed glass bottle with ground stopper. In the case
of lower layer water, use a 1 to 2 1 glass bottle with ground stopper ca-
pable of being attached to a water sampler. Either of those bottles shall
be washed with hexane prior t o use.
Water sampler A Heyroth type water sampler o r a proper water sam-
pler similar thereto.
Sampling method The sampling method shall be as follows.
Sampling of falling water In the case of sampling water falling from
waterway, weir, channel, pipe, etc., receive the sample directly in the sample
container, and stop sampling so that the proper space remains(1).
Sampling from piping or the like under condition of passing water In
the case of piping, apparatus, etc. under passing condition of water, open
the sampling valve t o let flow out the amount approx. five times the vol-
ume of water remaining in the sampler piping a t a rate of about l Zlmin,
then receive it in a sample container, and stop the sampling so that the
proper space remains in the container(1) (2) (3) (4).
Sampling from deep waterway, water tank, etc. In the case of sam-
pling in the deep waterway and water tank, use the water sampler capable
of sampling the all layer samples and take the samples of all layers. In
the case of Heyroth water sampler, attach the sample container to the frame
of the sampler, then lower the container to near the bottom, draw up the
water sampler while sampling the water at a constant speed, and take the
sample when it has reached the water surface so that the proper space
remains in the sample container(5).
Sampling from reservoir, lake, river, etc. By use of a water sampler
attached with the sample container, take the sample at a n arbitrary depth
in accordance with (cl,as appropriate, corresponding t o the purpose of test.
Notes (1) In this case, do not wash the sample container with the sample.
133
K O101 : 1998
(2) I n the case of sampling from the piping apparatus, etc. under
conditions at a high temperature and high pressure or at nega-
tive pressure, carry out as follows:
In the case of high temperature water, attach the cooler t o
the sampling piping, and cool to room temperature or below. In
the case of high pressure water [the pressure of 1.96 MPa or more],
take the sample after reducing the pressure with a pressure
reducer provided, and if it is at a high temperature, cool down
to room temperature or below by passing through the cooler. In
the case of negative pressure water, take the sample after rais-
ing to the atmospheric pressure by pressure riser (in the case of
negative pressure water at a high temperature, make atmospheric
pressure after cooling to room temperature with the cooler pro-
vided before the pressure riser) [Refer t o 4.3 of JIS K 00941.
(3) If the apparatus or the like is under stopped condition, oily sub-
stances, in most cases, are separated from water in the piping
and apparatus, and therefore, the concentration of oily substance
varies according t o the water passing speed and passing period.
If there is a fear that the oily substances are attached in the
sampling valve and piping, open the sampling valve fully to pass
water approx. 10 min, and further pass water for 10 min at a rate
of approx. 1Zlmin. Wash by repeating this operation.
(4) The flow rate, just before the sampling, shall not be changed.
(5) The sampling shall be in accordance with the sampling from the
all layer sample of JIS K 2251, as appropriate.
(4) Handling of sample The sample shall be handled as follows.
(a) The sample taken according to (3)shall not be transferred into another
container, and shall not be taken as aliquot. The total amount of it shall
be used for the test.
(b) Obtain the amount of sample from the mass of the container containing
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the sample by subtracting the mass of the sample container, o r by mark-

ing the level of water in the sample container when taking the sample, then
adding water up to the mark at the time of completion of test, and by tak-
ing the volume of the added water as the amount of the sample.
(c) Where it is necessary to preserve o r to transport the sample, add several
drops of Methyl Orange solution (1glZ) as indicator, then add hydrochloric
acid (l+l) until the colour of the solution turns red, and stopper tightly(6).
Note (6) When oily substances are floating, these ooze out easily due t o
vibration during transportation, and therefore, take care for the
handling.
26.2 Extraction method After adjusting the sample to p H 4 or under with hy-
drochloric acid, the extraction is carried out with hexane. Then hexane is expelled
a t 80 O C , and the mass of the remaining substance shall be measured t o determine
the amount of hexane extracts.
Determination range: 5 t o 500mg
134
K O101 : 1998

(a) Water Use water A3 specified in JIS K 0557.
(b) Hydrochloric acid (l+l)Prepare by using hydrochloric acid specified in
JIS K 8180.
(c) Sodium sulfate As specified in JIS K 8987.
(d) Methyl Orange solution (1 gll) As described in 22.1.1 (1)(d).
(e) Hexane As specified in JIS K 8848.
(f) Nitrogen High purity grade 2 nitrogen specified in JIS K 1107,
Separating funnels The funnels having 200 ml and 1O00 t o 3 O00 ml in
capacity and short legs. Wash with hexane prior to use. Do not apply
lubricant onto the cock.
Dryer Capable of adjusting t o (80+5)O C .
Hot plate or mantle heater Capable of adjusting to (80+5) "C. The water
bath capable of adjusting temperature may be used.
Distilling apparatus The one with an interchangeable ground joint ca-
pable of connecting with a distilling flask (50 to 100 ml capacity), " b " type
connecting tube and Liebig condenser (300 mm length). Wash them with
hexane prior to use.
Evaporating vessel Aluminium foil dish, platinum dish, beaker having
capacity of 50 t o 100ml with mass as small as possible. Wash with hex-
ane thoroughly respectively prior to the use, heat at (80k5) O C for approx.
30 min, allow to stand to cool in a desiccator, and measure the mass pre-
liminarily t o the nearest 0.1 mg.
Transfer the sample(7) taken in accordance with 26.1 into a 1 O00 t o 3 O00 ml
separating funnel(9, add 2 or 3 drops of Methyl Orange solution (1g/Z) as
indicator, and then add by dripping hydrochloric acid ( l + l )until the colour
of the solution changes to red.
Wash the sample container twice with approx. 20 ml each of hexane, and
add the washings into the 1O00 to 3 O00 ml separating funnel. Shake vig-
orously to mix for approx. 2 min, and then allow to stand(9).
Return the aqueous layer in the sample container, and swirl the 1O00 t o
3 O00 ml separating funnel gently t o separate the remaining aqueous layer
as much as possible(l0). Then, return it into the sample container, and
transfer the hexane layer into the 200 ml separating funnel.
Transfer the aqueous layer in the sample container into the 1O00 t o 3 O00 ml
separating funnel used in (a),then separate the hexane layer from the aqueous
layer by carrying out the operation in (b) and ( c ) again, and combine the
hexane layer in the 200 ml separating funnel.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
135
K O 1 0 1 : 1998
Wash the 1 O00 t o 3 O00 ml separating funnel with a small amount of hex-
ane, and combine the washings in the 200 ml separating funnel.
Swirl the 200 ml separating funnel gently, and after standing still, remove
the mixed-in water thoroughly with taking care not t o lose hexane(10).
Add 20 ml of water to hexane layer, shake to mix for approx. 1 min, then
allow t o stand, and discard the aqueous layer. Repeat this operation of
washing several times until the washings turn t o yellow with Methyl Or-
ange. Remove the aqueous layer as much as possible.
Add 3 t o 5 g of sodium sulfate t o hexane layer, and mix with shaking to
remove the water(l1).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Wipe the leg of 200 ml separating funnel with dry filter p a p e r ( 9 , and fil-
ter the hexane layer through absorbent cotton or filter paper(l2). Transfer
the hexane layer into a distilling flask of a distilling apparatus(l3).
Wash the 200 ml separating funnel with a small amount of hexane, filter
the washings by the operation as in (i),and transfer into the distilling flask
of the distilling apparatus. Wash the used absorbent cotton o r the filter
paper twice with approx. 5 ml each of hexane, and combine the washings
also t o the distilling flask.
Put the distilling flask in a mantle heater, and after connecting the flask
with a " b type connecting tube and a Liebig condenser, adjust the tem-
perature of the heater to approx. 80 O C , distill hexane a t a rate of one drop
per second and receive the distilled hexane in a receiver(l4). Continue the
distillation until the amount of the liquid in the flask attains t o about 2 ml.
Feed high purity grade 2 nitrogen specified in JIS K 1107 t o the " I- " type
connecting tube from its upper mouth t o attain room temperature.
Transfer the residue in the distillation flask into a mass known evaporat-
ing vessel. Wash the distillation flask three times with a small amount of
hexane, and add the washings in the evaporating vessel. Put the evapo-
rating vessel in a hot plate or mantle heater kept at approx. 80 "C to volatize
hexane (15).
Wipe the outside of the evaporating vessel with a clean wet cloth o r the
like, then with a clean dry cloth or the like. Transfer it into a dryer ad-
justed t o (80+5)"C and heat for approx. 30 min, Then, transfer the evapo-
rating vessel in a desiccator, and allow to cool for approx. 30 min. Accurately
weigh its mass to the nearest 0.1 mg, and subtract the mass of the evapo-
rating vessel t o obtain the mass (mg) of hexane extracts.
For the blank test, take the same amount of hexane as that of total hexane
used in this test into the distillation flask(l3), and carry out the operation
specified in (k)to (m) t o obtain the mass (mg) of the residue.
Calculate the concentration (mgll) of hexane extracts in the sample according
1 O00
P = (a- b) x -
V
where, P : hexane extracts (mg/Z)
136
K O101 : 1998
a : mass of hexane extracts in the test operation (mg)

b : mass of the residue in the blank test (mg)
V : sample (ml)
Notes (7) Usually, approx. 1 I of the sample is enough. However, take the
sample so that the mass of hexane extracts becomes 5 to 500 mg.
Use the total amount.
Select the separating funnel of proper size corresponding to the
amount of sample.
According t o the nature of the sample, the emulsion is sometimes
formed in hexane layer, or hexane layer sometimes becomes turbid.
In such a sample, return the aqueous layer in the separating funnel
into the original sample container as much as possible, and add
approx. 10 g of sodium chloride specified in JIS K 8150 or am-
monium sulfate specified in JIS K 8960 (not containing substances
soluble in hexane) then attach the interchangeable ground Liebig
condenser or Dimroth condenser of approx. 300 mm to the mouth
of separating funnel, immerse the separating funnel into the
constant temperature water bath kept a t approx. 80 OC, and the
emulsion is sometimes broken down by refluxing hexane for
approx. 10 min. In addition to the heating reflux, add approx. 10 g
of sodium chloride specified in JIS K 8150 o r ammonium sul-
fate specified in JIS K 8960 to the hexane layer and the emul-
sion layer in the separating funnel. After mixing by shaking,
transfer t o a centrifugal separating tube with a small amount of
water. When centrifugally separated at 8 O00 min-1 or over for
approx. 5 min, the emulsion layer becomes little, and the hex-
ane layer can be made easy t o be separated. Return it t o the
separating funnel, and follow the operation of ( c ) and after.
Continue the operation until the amount of aqueous layer t o be
separated attains t o 1ml or less. When the sample contains a
large amount of greases or solid fats, add the hexane addition-
ally prior to the separation of the aqueous layer.
Hexane layer sometimes becomes turbid. In such a case, after
separating the aqueous layer as much as possible, when it is
dehydrated by adding sodium sulfate, it may become transpar-
ent. In some cases, sodium chloride specified in JIS K 8150 o r
ammonium sulfate specified in JIS K 8960 may be more effec-
tive. However, the reagents containing substances soluble in
hexane shall not be used.
That removed of extracts by washing thoroughly with hexane.
When filtering, filter shall be wetted preliminarily with a small
amount of hexane.
When the hexane layer can not be contained at once in the dis-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
tillation flask, distill hexane by dividing in 2 o r 3 times.

The hexane distilled by distillation can be reused when distilled
again.
137
K O 1 0 1 : 1998
(15) Sufficient caution shall be taken not to catch fire. The solvent
shall be recovered as much as possible without discarding by
evaporation. After the evaporation of hexane, if water is observed
in the evaporating vessel, add acetone and repeat the evapora-
tion to remove water. Since residuals of salts in water cause
errors, therefore caution shall be taken. If salts remain, carry
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
out the operation of (m). After obtaining the mass (mg) of hex-
ane extracts, dissolve the hexane extracts by adding a small
amount of hexane, and separate. Repeat this procedure. After
removing the hexane extracts, obtain the mass (mg) of remain-
ing substances by performing the operation of (m), and correct.
Remarks 1 In the case where the mass of hexane extracts is 5 mg or less
and its determination is difficult, it is recommended t o carry
out the test according t o 24.3 or 24.4 in JIS K 0102.
2 For the sample which is marked in turbidity or is liable to
generate emulsion, a Soxhlet extracting method should be
preferably applied. Add 2 to 3 drops of Methyl Orange solu-
tion (1 gil) to the sample as an indicator, add hydrochloric acid
(l+l) until the colour of the solution turns t o red, and adjust
pH t o 4 or under. Then, lay two sheets of class 5A filter pa-
per by piling onto a Buchner funnel, add 100 ml of diatoma-
ceous earth suspension thereto, and filter by suction. Wash
diatomaceous earth with approx. 1E of water under pressure
reduction condition. Add the sample made acidic to this fil-
trate and filter by suction. After sucking thoroughly, wind
the filter medium together with the filter paper, transfer t o
cylindrical filter paper, wipe the wall of funnel, container,
stirring rod, etc. off with the filter paper washed by hexane,
put them into the same cylindrical filter paper, and dry in a
dryer a t (80k5) "C for approx. 30 min.
Transfer the cylindrical filter paper t o the Soxhlet extrac-
tor. After drying the sample container thoroughly, wash twice
with approx. 20 ml of hexane, and pour washings onto the cy-
lindrical filter paper. Then, move t o extracting operation, and
repeat the extraction approx. 20 times. Hereafter, carry out
the operation of 26.2 (3) (k)t o ( o ) ,and calculate the concen-
tration of hexane extracts in the sample (mgll).
27 Missing number
138
K O101 : 1998
28 Residual chlorine The residual chlorine means hypochlorite to be generated

by dissolving a chlorine agent in water and chloroamine to be generated by combin-
ing it with ammonia. The former is called free residual chlorine, the latter is called
combined residual chlorine, and both are collectively called residual chlorine.
For the determination of residual chlorine, in the case of low concentration, o-
tolidine colorimetric method or diethyl-p-phenylenediamine (DPD) colorimetric method
applies. In the case of comparatively high concentration, iodometry applies.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Further, in the case where monochloroamine, dichloroamine, etc. in combined re-
sidual chlorine a r e determined, DPD-ammonium iron (II) sulfate titration or
amperometric titration applies. This test shall be carried out immediately after
sampling.
28.1 o-Tolidine colorimetric method 3,3-dimethylbenzidine (o-tolidine) solution

is added to the sample, and the residual chlorine is determined by comparing the
colour yellow developed by reaction with the residual chlorine with that of the re-
sidual chlorine standard colour solution. By treating with sodium arsenite solu-
tion, it is able to classify into the residual chlorine, free residual chlorine, and combined
residual chlorine.
Determination range: C1 0.01 to 2.0mglZ
o-Tolidine solution Dissolve O . 14 g of 3,3'-dimethylbenzidinium dichlo-
ride (o-tolidine dihydrochloride) specified in JIS K 8669 in 50 ml of water,
and add this solution while mixing by stirring into 50ml of hydrochloric
acid (3+7). Preserve it in a coloured bottle. Do not use the solution for
which six months or more have elapsed.
Phosphate buffer solution (pH 6.5) Dissolve 22.86 g of disodium
hydrogenphosphate specified in JIS K 9020 which is dried at 110 "C for
approx. 2 h and allowed to cool in a desiccator and 46.14g of potassium
dihydrogenphosphate Specified in JIS K 9007 in water to make 11. When
precipitation occurs, filter, take 200 ml of this solution, and then dilute to
1I with water.
Potassium chromate-potassium dichromate solution Dissolve 3.63 g
of potassium chromate specified in JIS K 8312 and 1.21g of potassium
dichromate Specified in JIS K 8517 in phosphate buffer solution (pH 6.51,
transfer it into a 1O00 ml volumetric flask, and add phosphate buffer solu-
tion (pH6.5) up to the marked line.
Residual chlorine standard colour solution Take potassium chromate-
potassium dichromate solution and phosphate buffer solution (pH 6.5) in
the ratio as shown in Table 28.1 according to the concentration of corre-
sponding residual chlorine (mgCl/Z) into a 100 ml colour comparison tube,
and mix them. Preserve the residual chlorine standard colour solution in
a dark place, and do not use if precipitates are formed.
Sodium arsenite solution (5 gll) Dissolve 0.5 g of sodium metaarsenite
specified in JIS K 8046 in water to make 100 ml.
139
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Colour comparison tube 100 ml, Flat bottom tube with a marked line
of 100 ml at a height of (200tr1.5)mm from the bottom.
(b) Colour comparison tube stand For 100m1, that the bottom and side
surface of which a milky white plate is attached.
Residual Potassium chromate- Phosphate buffer Residual Potassium chromate- Phosphate buffer
Chlorine potassium dichromate solution (pH 6.5) chlorine potassium dichromate solution (pH 6.5)
m&u[ solution ml ml mgcul solution ml ml
0.01 0. 18 99.82 0.70 7.48 92.52
0.02 0.28 99.72 0.80 8.54 91.46
0.05 0.61 99.39 0.90 9.60 90.40
0.07 0.82 99. ia 1.00 10.66 89.34
0.10 1.13 98.87 1.10 12.22 87.78
0.15 1.66 98.3% 1.20 13.35 86.65
0.20 2.19 97.81 1.30 14.48 85.52
0.25 2.72 97.28 1.40 15.60 84.40
0.30 96.75 1.50 16.75 83.25
0.35 3.78 96.22 1.50 17.84 82.16
3.25
'
~
0.40 4.31 95.69 1.70 18.97 81.03

0.45 4.84 95.16 1.80 20. 09 79.91
0.50 5.37 94.63 1.90 21.22 78.78
1
I,
0.60 6.42 93.58 2.00 22.34 77.66
(3) O p e r a t i o n Carry out the operation as follows.

(a) Take 5 ml of o-tolidine solution into a colour comparison tube, add a proper
amount of sample (1) (containing not more than 0.2 mg of residual chlorine),
and further add water up to the marked line of 100 ml. Then stopper quickly,
and mix by shaking.
(b) Allow t o stand in a dark place for 5 min(2).
(c) See through from the above t o compare with the residual chlorine stan-
dard colour solution, obtain the corresponding residual chlorine standard
colour solution, and record the concentration of corresponding residual chlorine
a (mgCl/Z).
(d) Take 5 ml of o-tolidine solution in another colour comparison tube, add t o
this solution the sample of the same amount as that in the operation of
(a),and stopper quickly t o mix by shaking.
(e) Within 5 s, add 5 ml of sodium arsenite solution ( 5 g/Z>and mix by shaking.
Further, add water up to the marked line of 100 ml, and mix by shaking.
(f) Compare the solution with residual chlorine standard colour solution, and
obtain the corresponding residual chlorine standard colour solution thereto.
Then, record the concentration of corresponding residual chlorine b (mgCl/Z).
140
K O101 : 1998
For the blank test, take 5 ml of sodium arsenite solution ( 5 g/Z) in a 100 ml
colour comparison tube, then add to this solution the sample of the same
amount as that in the operation in (a),and mix by shaking.
Add 5 ml of o-tolidine solution, mix by shaking, further add water up to
the marked line of 100 ml, and mix by shaking.
Within 5 s, compare the solution with the residual chlorine colour solution
to obtain the corresponding residual chlorine standard colour solution, and
record the concentration of corresponding residual chlorine cl (mgCl/Z).
After further allowing t o stand for 5 min in a dark place, compare with the
residual chlorine standard colour solution t o obtain the corresponding re-
sidual chlorine standard colour solution, and record the concentration of
corresponding residual chlorine cz (mgCl/Z).
Obtain the concentration of residual chlorine, free residual chlorine and
combined residual chlorine according t o the following formulae:
100
A = (U - cZ) X-
V
100
B = ( b- Ci)x -
V
C=A-B
where, A : residual chlorine (mgCl/Z)
u : residual chlorine obtained in ( c ) (mgCl/Z)
cz : residual chlorine obtained in í j ) (mgCl/Z)
V : sample (mi)
B : free residual chlorine (mgCVE)
b : residual chlorine obtained in ( f ) (mgCl/Z)
c1 : residual chlorine obtained in (i) (mgCl/Z)
C : combined residual chlorine (mgCl/Z)
Notes (1) When the sample is alkaline, use pH meter, and add hydrochlo-
ric acid (1+5) t o make pH approx. 7.
Further, the pH at the time of colouration shall always be
1.3 o r under.
(2) Combined residual chlorine of residual chlorine requires to reach
the maximum colouring 6 min at O O C , 3 min at 20 OC, and 2 min
and 30 s a t 25 "C.
Remarks 1 In case where no blank test is carried out, if iron of 0.3 mg/Z
min., manganese of 0.01 mg/Z min., and nitrite ion of 0.3 mg/Z
min., are contained, it will be interfered. In order to prevent
interfering of iron and manganese, add 1,Z-cyclohexanediamine
tetraacetate solution (10 gil) at a rate of 3 ml per 100 ml of the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
sample.
2 In the case of using residual chlorine measuring apparatus
on the market, preliminarily confirm that there is no error by
comparing with residual chlorine standard colour solution.
141
K O101 : 1998
3 For the water t o be used in this test, confirm neither exist-

ence of residual chlorine nor consumption of chlorine in ac-
cordance with 29 (3).
28.2 Diethyl-p-phenylenediamine (DPD) colorimetric method Add N, N-di-

ethyl-p-phenylenediammonium (DPD) sulfate t o a sample, and determine by com-
paring the colour from rose pink t o rosy red to be generated by reaction with residual
chlorine with the colour of residual chlorine standard colour solution.
Determination range: C1 0.05 to 2 mgll
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Potassium iodide As specified in J I S K 8913.

(c) DPD dilution powder Grind 1.0 g of N , N-diethyl-p-phenylenediammonium
sulfate (N, N-diethyl-p-phenylenediaminesulfate)in an agate mortar. Add
24 g of sodium sulfate specified in JIS K 8987 thereto, knead thoroughly,
put crystalline grains into a coloured glass bottle, and preserve in a dark
place a t O t o 10 "C avoiding humidity. Do not use the coloured one.
(d) Phosphate buffer solution (pH 6.5) Take 100 ml of potassium dihydrogen
phosphate solution (0.2 molí0 (dissolve 27.2 g of potassium dihydrogen
phosphate specified in JIS K 9007 in water to make 1I ) , add sodium hy-
droxide solution (0.2 mold) (dissolve 8 g sodium hydroxide specified in J I S
K 8576 in water t o make 1I) until pH becomes 6.5 by using a pH meter,
add O. 13 g of 1,2-~yclohexanediaminetetraacetate monohydrate, and dis-
solve.
(e) C.I. Acid Red 265 solution Heat C.I. Acid Red 265 [I-(4-methylbenzene-
sulfonamide)-7-(2-methylphenyl-azo)-8-hydroxy-3,6-naphthalene disulfonate
disodium] a t 105 t o 110°C for 3 t o 4h, and allow t o stand t o cool in a
desiccator. Dissolve its 0.329 g weighed to the nearest 1mg in a small amount
of water, transfer into a 1O00 ml volumetric flask, and add water t o the
marked line. Take 50ml of this solution into a 50ûml volumetric flask,
and add water to the marked line. Preserve it in a dark place at O to 10 OC,
and do not use that for which 6 months or a longer time has elapsed.
(0 DPD residual chlorine standard colour solution Take C.I. Acid Red
265 solution into a 50 ml volumetric flask in accordance with Table 28.2,
and add water to the marked line. Respectively transfer it to a colour com-
parison tube. Tightly stopper, and preserve in a dark place a t O to 10 "C.
Do not use that for which 6 months or a longer time has elapsed.
142
K O101 : 1998
Table 28.2 DPD Residual chlorine standard

colour solution (in 50 mi)
Residual chlorine CI. Acid Red 265 solution
mgll ml
0.05 0.5
0.1 1.0
0.2 2.0
0.3 3.0
0.4 4.0
0.5 5.0
0.6 6.0
0.7 7.0
0.8 8.0
0.9 9.0
1 .o 10.0
1.2 12,o
1.4 14.0
1.6 16.0
1.8 18.O
2.0 20. o
_ -

(a) Colour comparison tube 50 ml, that having a flat bottom attached with
the 50 ml marked line at a height of (150I1) mm from the bottom.
(b) Colour comparison tube stand For 50 ml. That equipped with a milky
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
white plate on its bottom and side face.
Take 2.5 ml of phosphate buffer solution (pH 6.5) into a 50 ml colour com-
parison tube, and add 0.5 g of DPD dilution powder thereto. Then, add a
suitable amount of the sample(3) (containing 0.1 mg or under of residual
chlorine) and further add water t o the marked line.
Stopper tightly, mix by shaking thoroughly, see through the colouration from
the side face within 1min(4) t o compare with the DPD residual chlorine
standard colour solution, obtain the concentration of residual chlorine cor-
responding t o this (mgCl/Z) from the corresponding DPD residual chlorine
standard colour solution to take it as the residual free chlorine, and calcu-
late the concentration of free residual chlorine in the sample (mgCl/Z).
After completion of operation of (b), add approx. 0.5 g of potassium iodide,
and stopper to dissolve by shaking to mix. After allowing to stand for approx.
2 min, compare the colouration with the residual chlorine standard colour
solution the same as in (b), obtain the concentration of residual chlorine
corresponding thereto (mgCl/Z) from the corresponding residual chlorine
standard colour solution, and calculate the concentration of residual chlo-
rine in the sample.
143
K O 1 0 1 : 1998
(d) Calculate the concentration of combined residual chlorine (mgCll1) accord-

ing to the following formula:
Concentration of combined residual chlorine (mgCl/Z)
= residual chlorine (mgCl/Z) - residual free chlorine (mgCl/Z)
Notes (3) Where the acidity or alkalinity of the sample is strong, adjust
pH t o approx. 6.5 by using sodium carbonate solution (50 gll) o r
hydrochloric acid (l+ll).
(4) Time for mixing by shaking is contained. Sodium sulfate in DPD
dilution powder may not be thoroughly dissolved.
28.3 Iodometry The iodine liberated by the reaction of residual chlorine and po-
tassium iodide is titrated with sodium thiosulfate solution t o determine the residual
chlorine. If oxidizing substances t o liberate iodine coexist, it will be determined as
residual chlorine.
Determination range: C1 0.1 mg or more

(b) Potassium iodide As specified in JIS K 8913.
(c) Acetic acid ( l + l j Prepare by using acetic acid specified in JIS K 8355.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(d) Starch solution (10 g/2) As described in 22.1.2 (1) (i).
(e) 10 mmol/Z sodium thiosulfate solution Take 25 ml of 0.1 mol/Z sodium
thiosulfate solution of 22.1.2 (1)(d)into a 250 ml volumetric flask, and add
water up to the marked line. Prepare this solution at the time of use and
do not use the solution for which 12 h or more have elapsed after prepara-
tion. Use the factor of 0.1 mol/Z sodium thiosulfate solution as this factor.

Take a proper amount of sample (1) (containing 0.1 to 7 mg of residual chlorine)
into a 500ml Erlenmeyer flask with ground stopper, add water to make
approx. 300 ml, and add 1 g of potassium iodide and 5 ml of acetic acid (l+l).
Stopper, and mix by shaking, and then allow t o stand in a dark place for
approx. 5 min.
Titrate the liberated iodine with 10 mmol/Z sodium thiosulfate solution. After
the yellow colour of the solution has became pale, add 1ml of starch solu-
tion (10 glZ) as indicator, and continue the titration until the blue colour of
generated iodine starch disappears.
For the blank test, take 100 ml of water, and carry out the operation speci-
fied in (a) t o ( c ) .
Calculate the concentration (mgC1lZ) of residual chlorine in the sample
144
K O101 : 1998
A = ( a - b ) x f x -'Ooo x 0.354 5
V
where, A : residual chlorine (mgCl/Z)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
titration (mi)
b : 10 mmol/l sodium thiosulfate solution required for
blank test (mi)
f : factor of 10 mmol/Z sodium thiosulfate solution
V : sample (ml)
0.354 5 : amount of residual chlorine equivalent to 1ml of
10 mmol/Z sodium thiosulfate solution (mg)
Remarks 4 In the case where the colouration and turbidity of a sample
are remarkable and the test is difficult t o be carried out, sepa-
rate residual chlorine by the method of Remarks 4, of 33 (re-
sidual chlorine) in JIS K 0102,and determine.
28.4 DPD-ammonium iron (11) sulfate titration Titrate liberated residual chlo-
rine with ammonium iron (11) sulfate solution by using N,N-diethyl-p-phenylene-
diammonium (DPD) sulfate as indicator, determine, further add potassium iodide to
separate monochloroamine and dichloroamine of combined residual chlorine, and
determine.
Determination range: C1 20 to 500yg

Phosphate buffer solution (pH 6.2) Dissolve 24 g of disodium hydrogen-
phosphate specified in JIS K 9020 and 46 g of potassium dihydrogenphosphate
specified in JIS K 9007 in approx. 800 ml of water, add 0.8 g of dihydrogen
disodium ethylenediamine tetraacetate dihydrate specified in JIS K 8107
after dissolving in approx. 100 ml of water and further add water to make
1I in total amount.
DPD solution Dissolve 1.1g of N , N-diethyl-p-phenylenediammonium sul-
fate ( N ,N-diethyl-p-phenylenediamine sulfate) in water t o which 8 ml of
sulfuric acid (1+3) and 0.2 g of dihydrogen disodium ethylenediamine-
tetraacetate dihydrate specified in JIS K 8107 are added, and dilute with
water to make 1I in total amount.
Put this solution into a coloured glass bottle with ground-in stopper, and
preserve in a cool dark place(5).
Potassium iodide solution Dissolve 0.5 g of potassium iodide specified
in JIS K 8913 in 100 ml of water. Prepare a t the time of use.
Barium diphenylaminesulfonate solution (1 g l l ) Dissolve O. 1g barium
diphenylaminesulfonate in 100 ml of water.
145
K O101 : 1998
(g) mol/Z potassium dichromate solution (for standardization)

300
Preliminarily grind potassium dichromate reference material for volumet-
ric analysis, specified in JIS K 8005 in an agate mortar, heat a t 150 "C for
approx. 1h, and allow t o stand t o cool in a desiccator. Take 0.981 g t o the
said K2Cr207 as 100 %, dissolve in a small amount of water, transfer into a
1O00 ml volumetric flask, and add water t o the marked line.
(h) 2.82 mmol/2 ammonium iron (II) sulfate solution Dissolve 1.11g of
ammonium iron (II) sulfate hexahydrate specified in JIS K 8979 in water
to which 8 ml of sulfuric acid (1+3) is added, and add water t o make 1Z in
total amount, Standardize a t the time of use.
Standardization Take 100 ml of this 2.82 mmoM ammonium iron (II) sul-
fate solution into an Erlenmeyer flask, add 10 ml of sulfuric acid (1+5)and
5 ml of phosphoric acid specified in JIS K 9005, and add 2 ml of barium di-
phenylamine sulfonate solution (1g/Z) as an indicator. Titrate this solution
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
1
with 300 moW potassium dichromate solution (for standardization), and take
the point when the colour of solution turns to blue purple to be kept as it is
for approx. 30 s as an end point. Calculate the factor cf) of 2.82 mmol/Z am-
1
monium iron (II) sulfate solution from the number of ml of 300 mol4 potas-
sium dichromate solution (for standardization) required for titration according
to the following formula.
~ x 1 o o o x x
f= 300
J -
2.82 x 100
1
where, x : the number of ml of 300 mol/Z potassium dichro-
mate (for standardization) required for titration
Note (5) Approx. 0.5 g of DPD dilution powder of 28.2 (1)( c ) may be used
instead of DPD solution.
(2) Operation Carry out the operation as follows. However, carry out the opera-
tion of (a)to ( c ) for determination of residual chlorine, the operation of (d) for
determination of liberated residual chlorine, the operation of (e) for determina-
tion of monochloroamine, and the operation of (f) for determination of
dichloroamine.
(a) Take a suitable quantity of a sample(3) (containing 20 t o 500 yg as residual
chlorine C1) into a 300 ml Erlenmeyer flask, add water to make approx. 100 ml,
add 5 ml of phosphate buffer solution (pH 6.2) and 5 ml of DPD solution(6),
and mix by shaking.
(b) Add approx. 1g of potassium iodide, dissolve, and allow t o stand still for
approx. 2 min t o make it colour red.
(cl Titrate with 2.82 mmol/Z ammonium iron (II) sulfate solution, and take the
point when the colour of red disappears as an end point. Take a (mi) as
the titre.
(d) Separately take the same amount of the sample as that sampled in (a)into
a 300 ml Erlenmeyer flask, add water to make approx. 100 ml, add 5 ml of
phosphate buffer solution (pH 6.2) and 5 ml of DPD solution, and mix by
146
K O101 : 1998
shaking. Quickly titrate with 2.82 mmol/Z ammonium iron (II) sulfate so-
lution, and take the point when the colour of red disappears as an end point.
Take b (ml) as the titre.
Then, add 0.1 ml of potassium iodide solution(7) (equivalent to two drops),
mix by shaking, titrate with 2.82 mmol/E ammonium iron (II) sulfate solu-
tion, and take the point when the colour of red disappears as an end point.
Take c (ml) as the titre.
Further add approx. 1g(8) of potassium iodide (crystalline), dissolve, and
allow to stand still for approx. 2 min t o colour red.
Titrate with 2.82 mmol/Z ammonium iron (II) sulfate solution, and take the
point when the colour of red disappears as an end point. Take d (mi) as
the titre.
Calculate the concentration (mgCl/Z) of residual chlorine, liberated residual
chlorine, monochloroamine, and dichloroamine in the sample according t o
the following formulas.
A = a x f x - IOoo x o . l
V
xo.l
B = b x f x - loo0
V
xo.1
C = c x f x - loo0
V
D = d x f x- loo0x o . l
V
where, A : residual chlorine (mgC1íZ)
B : liberated residual chlorine (mgCl/Z)
C : monochloroamine (mgCl/Z)
D : dichloroamine (mgCl/Z)
f : factor of 2.82 mmolll ammonium iron (II) sulfate
solution
V : sample (ml)
0.1 : residual chlorine equivalent to 1 ml of 2.82 mmoVI
ammonium iron (II) sulfate solution (mg)
Notes (6) In the case of using DPD dilution powder of 28.2(1)(c), add
approx. 0.5 g.
('1 0.5 mg of fine crystal of potassium iodide specified in JI$ K 8913
may be joined.
(9 In the case where the concentration of dichloroamine is 1mglZ
o r over, though allowed t o stand still for 2 min or longer, colora-
tion is liable to be imperfect. In that case, the additional amount
of potassium iodide specified in JIS K 8913 is made approx. 0.5 g .
Remarks 5 In the case of a sample containing nitrogen trichloride, nitro-
gen trichloride is also contained in the determined value of
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
147
K O101 : 1998
dichloroamine. The fractional determination is carried out as

fol10ws.
Take a suitable amount of a sample (containing 20 t o 500 pg
as residual chlorine C1) into a 300ml Erlenmeyer flask, add
water t o make 100 ml, add 0.1 ml of potassium iodide solu-
tion or approx. 0.5 mg of the pulverized crystal of potassium
iodide specified in JIS K 8913, and mix by shaking. Put 5 ml
of phosphate buffer solution (pH6.2) and 5 m l of DPD solu-
tion (or approx. 0.5 g of DPD dilution powder) into another bea-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ker, mix, and add t o the said Erlenmeyer flask.
Quickly titrate with 2.82 mmolíl ammonium iron (II) sulfate
solution, and take the point when the colour of red disappears
as an end point. Take e (mi) as the titre. Allow the amount of
liberated residual chlorine to be contained in this titre. Cal-
culate the concentration of nitrogen trichloride and the concen-
tration of dichloroamine according t o the following formulas.
E -
= 2 (e - a)x f x loo0
V
xo.l
where, E : nitrogen trichloride (mgCl/Z)

others : as described in (2) (h)
6 In the case where iodide ion or bromide ion coexists, iodide
ion and bromide ion are also contained in the determined value
of liberated residual chlorine. In order t o correct the deter-
mined value of liberated residual chlorine, it is corrected by
the following operation.
Obtain liberated residual chlorine by the operation of (2) (d).
Separately take 100 ml of a sample into a 300 ml Erlenmeyer
flask, and add 1ml of aminoacetate solution [dissolve 20 g of
aminoacetate (glycine) specified in JIS K 8291 in 100ml of
water].
Then add 5 ml of phosphate buffer solution (pH 6.2) and 5 ml
of DPD solution, titrate with 2.82 mmol/Z ammonium iron (II)
sulfate solution, and take the point when the colour of red dis-
appears as an end point. Subtract this from the titre of (2) (d),
and calculate the concentration of liberated residual chlorine.
7 Chlorine (IV) dioxide is contained in the value of residual
chlorine and liberated chlorine.
8 pH in titration is adjusted within a range of 6.2 to 6.5. When
pH is low, a part of monochloroamine comes to be contained in
the determined value of liberated residual chlorine. Further,
when pH is increased, the solution is coloured by dissolved
oxygen.
148
K O101 : 1998
9 When temperature of the solution in measurement is increased,

combined residual chlorine comes to be contained in the de-
termined value of liberated residual chlorine, and colouration
of DPD is inclined to become pale.
10 I n this test, since EDTA is contained in the phosphate buffer
solution (pH 6.2), copper does not give influences to an extent
of 10 mgll. In the case where 2 mgll or over of chromate ion
coexists, since the end point of titration becomes obscure, barium
chloride is preliminarily joined t o the sample, and barium
chromate is precipitated to remove interference.
11 Amperometric titration This is a method to determine the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
residual chlorine according t o amperometric titration by
phenylarsenoxide solution. This method is able t o separate
and to determine the liberated residual chlorine, combined re-
sidual chlorine, monochloroamine and dichloroamine.
Further, carry out the operation of (3) (a) to (0 of Remarks
11 for determination of residual chlorine, the operation of (3) (g)
t o (i) of Remarks 11 for determination of liberated residual
chlorine, the operation of (3) ( g ) to (k)of Remarks 11 for de-
termination of monochloroamine in combined residual chlorine?
and the operation of (3) (g) t o (m) of Remarks 11for determi-
nation of dichloroamine. Further, iron, manganese, nitrite ion,
etc. do not interfere.
Determination range: C1 0.04 mg or more
Phosphate buffer solution (pH 7) Take 25.4 g of
potassium dihydrogenphosphate specified in JIS K
9007 and 34.1 g of disodium hydrogenphosphate speci-
fied in JIS K 9020 into a beaker, dissolve in 800 ml
of water and add water to make 12.
Acetic acid-acetate buffer solution (pH 4) Take
480 g of acetic acid specified in JIS K 8355 and 243 g
of sodium acetate 3 hydrate specified in JIS K 8371
into a beaker, dissolve in 400ml of water and add
water to make I I .
Potassium iodide solution (50 gll) Dissolve 5 g of
potassium iodide specified in JIS K 8913 in water
to make 100 ml. Prepare at the time of use and put
it into a coloured glass bottle.
60mmol/Z iodine solution Dissolve 8 g of potas-
sium iodide specified in JIS K 8913 in approx. 20 ml
of water, add to it 2.6 g of iodine specified in JIS K
8920 to dissolve, dilute with water t o make 200m1,
and transfer into a coloured glass bottle.
149
K O101 : 1998
Standardization Take 20 ml of this iodine solution

into a 300 ml Erlenmeyer flask with ground stopper,
titrate with 0.1 mol/Z sodium thiosulfate solution [as
described in 22.1.2 (1) (d)],when the yellow colour of
the solution has become pale, add 1ml of starch so-
lution (10 g/Z) as indicator, and titrate further until
the blue colour of iodine starch disappears. Calcu-
late the factor c f ) of 50 mmoVZ iodine solution according
to the following formula from the number of ml (x)
of O. 1 mol/,! sodium thiosulfate solution for titration:
f = x x =f 0
where, f o : factor of 0.1 mol/Z sodium thiosulfate

solution
(f) 2.82 mmolíl iodine solution (for standardization)
Dissolve 10 g of potassium iodide specified in JIS K
8913 in approx. 20 ml of water, transfer into a 200 ml
volumetric flask, add 56.41f ml (f: factor of 50 mmol/Z
iodine solution) of 50 mmol/Z iodine solution, and add
water up t o the marked line. Take 20 ml of this solu-
tion into a 100 ml volumetric flask, and add water up
to the marked line. Prepare this solution a t the time
of use.
(g) 5.64 mmol/Z phenylarsenoxide solution Dissolve
0.8 g of phenylarsenoxide in 150 ml of sodium hydrox-
ide solution (12 g/Z),put its 110 ml into 800 ml of water
to mix thoroughly by stirring, adjust pH to 6 t o 7 by
using hydrochloric acid (l+ll),and after adding 1 ml
of chloroform, add water t o make 11.
Standardization Take 5 ml of 2.82 mmol/Z iodine
solution (0.2 mgCl/ml) (for standardization) into a
300 ml beaker, add water t o make 200 ml, titrate with
this phenylarsenoxide solution with amperometric ti-
tration apparatus, and take the point when the indi-
cating value of ammeter does not lower as the end
point. Obtain the number of ml (x)of phenylarsenoxide
solution required therefor, and calculate the factor VI)
of this phenylarsenoxide solution according t o the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
following formula:
5
f1=-
X
(a) Amperometric titration apparatus
Direct current ammeter Standard rated value of
5 PA, grade 1 ammeter of electrical indicating instru-
ment specified in JIS C 1102-1 and JIS C 1102-2
Platinum electrode
150
K O101 : 1998
Reference electrode
Take a proper amount of sample (containing 0.04 to
2 m g as C1) into a 300ml beaker, and add water to
make about 200ml.
Add 1 ml of potassium iodide solution (50 gíZ), and
further add 1ml of acetic acid-acetate buffer solu-
tion (pH 4) (adjust pH t o approx. 4).
Immerse the platinum electrode and reference elec-
trode into the sample, and connect the platinum elec-
trode t o the positive terminal of the direct current
ammeter and the reference electrode to the negative
terminal.
Mix by stirring with a magnetic stirrer to such an
extent that no air is drawn into.
Add dropwise each definite amount of 5.64mmollZ
phenylarsenoxide solution until the indicating value
of the ammeter does not lower.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Prepare the titration curve by reading the number

of ml of 5.64 mmol/Z phenylarsenoxide solution and
the indicating value of the ammeter, and obtain the
end point of titration t o take it as a (ml).
Separately take a proper amount of the sample (con-
taining 0.04 to 2 mg as Ci) into a 300 ml beaker, and
add water to make about 200ml.
Add 1ml of phosphate buffer solution (pH 7).
Carry out the operations of (cl to (f),and obtain the
end point of titration t o take it as b (ml).
Add 0.2 ml of potassium iodide solution (50 gil) to the
sample after titration.
If the indicating value of the ammeter has changed,
carry out the operations of ( c ) t o (f)t o obtain the end
point of titration, and take it as c (mi).
Then, add 1ml of acetic acid-acetate buffer solution
(pH4) to the sample after titration.
If the indicating value of the ammeter has changed,
carry out the operations of ( e ) and (f) t o obtain the
end point of titration, and take it as d (ml).
Calculate the concentrations (mgCVZ) of residual chlo-
rine, liberated residual chlorine, monochloroamine and
dichloroamine in the sample according t o the follow-
ing formulas:
151
K O101 : 1998
A = a x f l x - Oo0 x 0.2
V
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
B = b X f i X - loo0x 0 . 2
V
x 0.2
C = c x f , x -loo0
V
D = d x f , x -loo0
x0.2
V
where, A : residual chlorine (mgCl/U
B : liberated residual chlorine (mgCl/Z)
C : monochloroamine (mgC1/1)
D : dichloroamine (mgCl/E}
f l: factor of 5.64 mmoVZ phenylarsenoxide
solution
V : sample (ml)
0 . 2 : residual chlorine equivalent to 1ml
of 5.64 mmol/Z phenylarsenoxide solu-
tion (mg)
152
K O101 : 1998
29 Required amount of chlorine The required amount of chlorine means the

additive amount of chlorine required for making the definite concentration of the
residual chlorine after reaction of a definite time by adding chlorine to the sample.
As increasing the additive amount of chlorine, the concentration of residual chlo-
rine after a definite time increases, but in the case of the sample containing a large
amount of ammonium ion, organic nitrogen, etc., the concentration of residual chlo-
rine decreases when it reaches a definite additive amount. If add furthermore the
chlorine, the concentration of residual chlorine begins t o increases. This relation is
drawn, and the concentration of additive chlorine for the residual chlorine t o be-
come a definite concentration is obtained to take it as the required amount of chlo-
rine.
(a) Water Water A3 specified in JIS K 0557. Preserve in a borosilicate glass
bottle.
(b) Chlorine standard solution (0.1 mgCVml) Dilute the sodium hypochlorite
solution (effective chlorine: 7 to 12 %) with water so that the effective chlorine
becomes approx. 0.1 mgCl/ml. At the time of use, measure the concentra-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
tion by carrying out the operation of 28.3 (2).
Take each 200 ml of sample into several 300 ml Erlenmeyer flasks with ground
stopper, add to it stepwise 1 to 20 ml of chlorine standard solution (0.1 mgCV
ml)(l) with taking care so as not to adhere the inside wall of the flask, stopper
tightly, and after mixing by shaking, allow it to stand(2) in a dark place.
After 1 h, measure the concentrations of respective residual chlorine ac-
cording t o the method of 28.1, 28.2, 28.3 or 28.4.
Then, take the concentration of residual chlorine on the ordinate of a sec-
tion paper, and take the chlorine-additive concentration (the concentration
immediately after adding aqueous chlorine) in (a) on the abscissa t o draw
as in Fig. 29.1(3).
O a b c d
Chlorine-additive
concentration
-__.__I_ I (mgCVZì __.
Fig. 29.1 Method of obtaining required amount

of chlorine by drawing
153
K O 1 0 1 : 1998
(d) I n the case such as Type II, take the chlorine additive concentration a
(mgCVZ) for the residual chlorine concentration to indicate the specified value
k (4) (mgCVI) as the required amount of chlorine.
(e) In the case such as Type III, take the chlorine additive concentration d
(mgCl/Z) indicate the specified value K (mgCl/Z) after passing, for the re-
sidual chlorine concentration, the point indicating the minimum value c as
the required amount of chlorine.
Notes (1) The concentration of chlorine standard solution in the range of
0.05 to 0.1 mgCVm1 is sufficient. However, where the required
amount of chlorine is not less than 6 mg/Z, the concentration of
0.2 mgCl/ml is adequate.
(2> Retain the same temperature as the water temperature a t the
time of taking water as far as possible.
The standing period of time shall, as a rule, be the period re-
quired to reach the specified place of the facility from the chlo-
rine-pouring point, and, in general, approx. 1 h is employed.
(3) The sample having no substance to react with chlorine shows,
theoretically, Type I. The actual sample shows Type II or Type
III.
(4) The value of k , in the case of aiming the removal of iron and
manganese, is practical to be in the range of 0.5 t o 1.0 (mgCl/Z).
In the case of service water, usually the water of 0.1 (mgCl/Z)is
used.
Remarks 1 In the case of the service water test, the value of required
amount of chlorine, a, b, etc. subtracted by the value of k (or-
dinarily 0.1) is used.
In addition, the value of b subtracted by k (0.1)is called as
the consumption amount of chlorine. I n the case of Type II,
the required amount of chlorine is equal t o the consumption
amount of chlorine.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
154
K O101 : 1998
30 Hydroxide ion (OH-) Acid-base titration is applied to determination of hy-

droxide ion. After the coexisting carbonate ion, phosphate ion, etc. are precipitated
by addition of strontium chloride as strontium salts, it is titrated with sulfuric acid
using phenolphthalein as indicator and the hydroxide ion is determined.
Determination range: OH- 0.1 mg or more
(a) 10 mmol/Z Sulfuric acid As described in 13.1 (1) ( c ) .
(b) Strontium chloride solution Dissolve 4.5 g of strontium chloride 6 hy-
drate specified in JIS K 8132 with water t o make 1E .
(c) Phenolphthalein solution ( 5 glZ) As described in 13.2 (1)(a).
(d) Nitrogen High purity grade 2 nitrogen specified in JIS K 1107.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

Take 100 ml of sample (where it contains 15 mg o r more as OH-, take its
proper amount, and add water t o make 100 mi) into a 300 ml Erlenmeyer
flask with ground stopper.
Add 1ml of strontium chloride solution per each 1mg of coexisting carbon-
ate ion and phosphate ion, further add 4 ml in excess, and shake gently t o mix.
Heat while passing air or nitrogen(') removed of carbon dioxide, and after
boiling for approx. 30 s, cool with water.
After cooling, add 2 t o 3 drops of phenolphthalein solution (5 g/l) as indica-
tor, and titrate with 10 mmol/Z sulfuric acid until the red colour of solution
disappears.
Calculate the concentration of hydroxide ion in the sample (mgOH-/Z) ac-
H = a x f x - *Oo x 0.3402
V
where, H : hydroxide ion (mgOH-/Z)
f : factor of 10mmol/Z sulfuric acid
V : sample (mi)
0.340 2 : hydroxide ion equivalent t o 1 ml of 10 mmol/Z sul-
furic acid tmg)
Note (1) Pass the air or nitrogen washed with potassium hydroxide solu-
tion (220glZ) and water through the liquid surface.
Remarks 1 Where the concentrations of carbonate ion and phosphate ion
are unknown, it is necessary t o put in the strontium chloride
solution sufficiently in excess.
2 When the strontium chloride is added in excess, white turbidity
of strontium sulfate may be caused, but it does not interfere
the titration.
155
K O 1 0 1 : 1998
31 Fluorine compounds The fluorine compounds are generic term for fluoride
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ion, metal fluoride, etc., and expressed as fluoride ion. To the determination of fluo-
ride ion the lanthanum-alizarin complexon absorptiometry or ion selective electrode
method applies.
31.1 Lanthanum-alizarin complexon absorptiometry By the measurement of

the absorbance of blue complex formed by the reaction of fluoride ion with the com-
plex of lanthanum (III) and alizarin complexon, fluoride ion is determined.
Determination range: F- 4 to 50 pg
Remarks 1 Most anions do not interfere with this method, however cations,
especially aluminium, cadmium, cobalt, iron, nickel, beryllium, lead,
etc. interfere seriously, and therefore fluoride ion shall be sepa-
rated by distillation preliminarily.

Perchloric acid Perchloric acid specified in JIS K 8223 is heated to gen-
erate white fume, and it is allowed to stand to cool.
Phosphoric acid As specified in JIS K 9005.
Sodium hydroxide solution (100gll) As described in 22.2.1 (1)(b).
Silicon dioxide Silicon dioxide specified in JIS K 8885 of 100 to 150 pm
in grain size(1).
Phenolphthalein solution ( 5 gll) As described in 13.2 (1) (a).
Lanthanum-alizarin complexon solution Dissolve 0.192 g of alizarin
complexon ( 1,2-dihydroxyanthraquinon-3-ilmethylamine-N,N-diacet ate
dihydrate) in 4 ml of aqueous ammonia (1+10) and 4 ml of ammonium ac-
etate solution (200 g/Z), add it into sodium acetate solution (dissolve 41 g
of sodium acetate 3 hydrate specified in JIS K 8371 in 400ml of water
and add 24 ml of acetic acid specified in JIS K 8355)while mixing by stir-
ring. Add 400 ml of aceton specified in JIS K 8034 t o this solution gradu-
ally while mixing by stirring, then add lanthanum solution [dissolve 0.163 g
of lanthanum (III) oxide in 10 ml of hydrochloric acid (1+5) by heating] and
mix by stirring. After standing to cool, adjust pH to approx. 4.7 with ace-
tic acid or aqueous ammonia specified in JIS K 8085 using pH meter, and
then add water to make 11.
Fluoride ion standard solution (0.1 mgF-/ml) Take sodium fluoride (stan-
dard reagent for volumetric analysis) specified in JIS K 8005 in a plati-
num dish, and heat at 500 "C for approx. 1h. Then, allow to cool in a
desiccator. Then, weigh out 0.221 g of NaF for its 100 % purity and dis-
solve it in a small quantity of water. Transfer the solution into 1O00 ml
volumetric flask and add water up to the marked line. Put this solution
into a polyethylene bottle and preserve. Otherwise, use fluoride ion stan-
dard solution F- 100 specified in JIS K 0030.
Fluoride ion standard solution (2 pgF-/ml) Take 10 ml of fluoride ion
standard solution (0.1 mgF-/ml), transfer it into a 500 ml volumetric flask
and add water u p to the marked line.
156
K 0101 : 1998
Notes (1) Crystalline silicon dioxide is used. In the case where its quality
is not judged, heat in a platinum crucible at 1 150 "C or higher
for approx. 1h, and allow to stand t o cool in a desiccator. In
this case, take 1 0 m l of fluoride ion standard solution (2ygF-/
ml) and the recovery shall be confirmed by performing (b) to (e)
of (3)and (a) to (e) of (4).
(2) Those on the market may be used.
Informative reference : When Alufusone on the market is used, dissolve
2.5 g in water t o make 50 ml. Prepare when used.
(a) Distillation apparatus An example is shown in Fig. 31.1.
(b) Photometer Spectrophotometer o r photoelectric photometer.
(3) Distillating operation Carry out the distillating operation as follows.
Take a proper amount of sample(3) (containing 30 pg o r more as F-) in a por-
celain evaporating dish or beaker, and add 2 to 3 drops of phenolphthalein
solution (5 g/Z). Then add, dropwise, sodium hydroxide solution (100 gll) t o
make slightly alkaline, and thereafter heat the solution to concentrate t o
approx. 30 ml.
Transfer the solution while washing by approx. 10 ml of water into the
distillation flask shown in Fig. 31.1. Then add approx. 1g of silicon diox-
ide, 1ml of phosphoric acid and 40 ml of perchloric acid [or 30 ml of sulfu-
ric acid(4) specified in JIS K S9511(5). Add 20 ml(6) of water t o a 250 ml
volumetric flask of the receiver and keep the tip of back-flow stopper un-
der water surface.
Heat directly a distillation flask(% After temperature of solution in the
distillation flask reaches approx. 140 O C , pass steam.
Adjust distillation temperature at (145+5) "C and distillating speed to 3 t o
5 mumin, and continue distillation until the solution amount in the receiver
reaches approx. 220 ml.
Remove the condenser and the back-flow stopper, wash the inner tube of
the condenser and the inside and outside of the back-flow stopper with a
small amount of water, add washings t o the receiver, and further add water
to the marked line.
When the dissolved fluoride ion is tested, filter the sample with
filter paper class 5C, discard the initial approx. 50 ml, and use
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the filtrate thereafter as the sample.

Put sulfuric acid specified in JIS K 8951 into a beaker, and gen-
erate white fume vigorously by heating. Thereafter, allow to stand
to cool.
Put approx. 10 boiling tips of 2 to 3 mm diameter into a distilla-
tion flask.
In the case where a great amount of halides other than fluoride
ion are contained in the sample, add preliminarily several drops
157
K 0101 : 1998
of sodium hydroxide solution (40g/Z) and several drops of phe-

nolphthalein solution (5 g/Z). Dropwise add sodium hydroxide
solution (40 gll) as occasion demands so that the solution in the
receiver keeps faint pink until distillation is completed.
Further, in this case, drip sulfuric acid (1+35)t o the distillated
solution until faint pink disappears after distillation is completed,
and carry out the operation of (e) hereafter.
(7) Adjust the flame so as to be able t o heat the distillation flask t o
the liquid surface therein. Allow an oil bath, a glycerin bath,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
etc. t o be used.
Remarks 2 A double tube type distillation flask may be used instead of
the distillation flask. In the case, put 1,1,2,2-tetrachloroethane
(boiling point: 146.3 OC) specified in JIS K 9620 into its outer
casing, heat directly the outer casing and after l,l,Z,Z-
tetrachloroethane starts boiling, pass steam. When 1,1,2,2-
tetrachloroethane is used for a long time, it is decomposed and
is coloured and besides, its boiling point descends. In that
case, distill, and use the fraction at 146 OC.
Further, when 1,1,2,2-tetrachloroethaneafter use is dis-
carded, take cares so as not t o cause environmental contami-
nation.
158
K O101 : 1998
Unit: mm
_
50
, Is04
A: steam generating flask, 1O00 ml I: exchangeable spherical ground joint

B: connection and inducing tube J: presser bar spring
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
C: trap K: 200 "C thermometer
D: distillation flask, 500 ml L: rubber tube
E: Liebig condenser, 300 mm M: pinch cock
F: back-flow stopper (approx. 50 ml) N: stopper for inserting the therm ometer
G: receiver (250 ml volumetric flask) O: trap sphere (Kjeldahl type)
H: exchangeable ground joint
Fig. 31.1 A n example of distillation apparatus
(a) Take a proper amount not more than 30 ml from the distillate obtained in
(3)distillating operation (containing 4 to 50yg as F-) into a 50ml volu-
metric flask.
(b) Add 20 ml of lanthanum-alizarin complexon solution(8), then add water up
to the marked line to mix by shaking, and allow to stand for 1h.
(c) Separately, take 30 ml of water into a 50 ml volumetric flask, and carry
out the operation specified in (b).
159
K 0101.: 1998
(d) Transfer a portion of the solution obtained as in (b) on the sample into an
absorption cell, and measure the absorbance a t a wavelength near 620 nm
using the solution in ( c ) as reference solution.
(e) Obtain the amount of fluoride ion from the working curve, and calculate
the concentration of fluoride ion in the sample (mgF-ll).
Working curve Take, stepwise, 2 t o 25 ml of fluoride ion standard solu-
tion (2 pgF-lml) in 50 ml volumetric flasks, carry out the operation speci-
fied in (a)t o (d)to measure the absorbance, and prepare the relation curve
between the amount of fluoride ion (F-) and the absorbance.
Note (8) In the case of using Alufusone solution prepared as in Informa-
tive reference of 31.1 (l), after adding 5 ml of the solution and
10 ml of acetone specified in JIS K 8034 to the sample solution,
31.2 Ion selective electrode method After distilling fluorine compounds t o sepa-
rate in pretreatment, the buffer solution (total ion intensity regulating solution) is
added to adjust pH to 5.0 t o 5.5, and the fluoride ion is determined by the measure-
ment of potential by using fluoride ion selective electrode as the indication electrode.
Determination range: F- 0.1 t o 100mglZ
Buffer solution (pH 5.2)(9) Dissolve 58 g of sodium chloride specified in
JIS K 8150 and l g of diammonium hydrogen citrate specified in JIS K
8284 in 500 ml of water, and add 50 ml of acetic acid specified in JIS K
8355. Adjust pH of the solution to 5.2 using pH meter by dropwise adding
sodium hydroxide solution (200 glZ), and then add water to make 11.
Fluoride ion standard solution (100 mgF-IZ) As described in 31.1 (1)(g).
Fluoride ion standard solution (10mgF-ll) Take 20 ml of fluoride ion
standard solution (100 mgF-ll) into a 200 ml volumetric flask, and add water
t o the marked line. Prepare at the time of use.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Fluoride ion standard solution (1mgF-IC) Take 20 ml of fluoride ion

standard solution (10 mgF-l1) into a 200 ml volumetric flask, and add wa-
ter to the marked line. Prepare a t the time of use.
Fluoride ion standard solution (0.1 mgF-IC) Take 20 ml of fluoride ion
standard solution (1 mgF-l1) into a 200 ml volumetric flask, and add water
Note (9) As buffer solution, the solution of the following composition may
be used:
(i) Add 57 ml of acetic acid specified in JIS K 8355,58 g of sodium
chloride specified in JIS K 8150, and 4 g of 1,2-cyclohexane
diamine tetraacetate monohydrate in 500 ml of water t o dis-
solve, and add, dropwise, sodium hydroxide solution (200 gll).
After adjusting pH t o 5.0 to 5.5 by use of pH meter, dilute with
water to 11.
160
K O101 : 1998
(ii) Add 57 ml of acetic acid specified in JIS K 8355, 58 g of sodium

chloride specified in JIS K 8150, and 0.3 g of trisodium citrate
dihydrate specified in JIS K 8288 to 500 ml of water t o dissolve,
and then add sodium hydroxide solution (200 g/Z). After adjust-
ing pH to 5.0 t o 5.5 by use of pH meter, dilute with water to
11.
(a) Potentiometer High input resistance potentiometer of 1mV minimum
scale (for example, digital type pH-mV meter, pH-mV meter with magnify-
ing span, potentiometer for ion selective electrode, etc.).
(b) Indication electrode Fluoride ion selective electrode
(c) Reference electrode Double liquid-junction type (or salt bridge) refer-
ence electrode (double junction sleeve type reference electrode or ceramics
type reference electrode with small resistance). Put potassium chloride so-
lution (3mol/l or saturated solution) in as inside cylinder solution. Put
potassium chloride solution (3 moL4 or saturated solution) or potassium nitrate
solution (100 gll) in as the outside cylinder solution.
(d) Magnetic stirrer Not giving the change of liquid temperature by heat
generation due to rotation.

Take 100 ml of fluoride ion standard solution (0.1 mgF-/Z) into a 200 ml beaker,
and add 10 ml of buffer solution (pH 5.2)(1*).
Immerse the indication electrode (11) (12) and reference electrode (13) (14) into
this solution, and stir with a magnetic stirrer(l5)vigorously to mix, but bubbles
are not t o touch the electrode(l6).
Measure the temperature of the solution and measure the potential with a
potentiometer (17).
Take respectively 100 ml of fluoride ion standard solution (1mgF-/Z), 100 ml
of fluoride ion standard solution (10 mgF-/Z), and 100 ml of fluoride ion
standard solution (100 mgF-l1) into a 200 ml beaker, and add 10 ml of buffer
solution (pH 5.2). Adjust temperature of each fluoride ion standard solu-
tion (1 t o 100 mgF-/Z) within fl "C of solution temperature of (c)(17), carry
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
out the operation of (b) and (cl, and measure the potential(l*).
Take the concentration of fluoride ion on the logarithmic axis, and the
potential on the linear axis on a semilogarithmic section paper, and pre-
pare the relation curve between the concentration of fluoride ion (mgF-ll)
and the potential (18).
Notes (10) The buffer solution (pH5.2) is added in order t o adjust pH to
approx. 5.2 at the time of measurement and to keep the ionic
strength constant.
(11) Immerse the indication electrode (the fluoride ion selective elec-
trodes) into the fluoride ion standard solution (0.1 mgF-/E) at
the time of use, and measure the potential after the indication
value has become stable.
161
K O101 : 1998
If the sensing film of the indication electrode flaws, the slope

of the working curve (potential slope) decreases and the response
speed lowers, and therefore cares shall be taken.
Further, if the sensing film of the indication electrode is
stained, the response speed becomes slow. Wipe off the stains
with absorbent cotton or with soft paper moistened with ethanol,
and then wash the electrode with water.
Select the reference electrode of small resistance, and use, in
general, that of sleeve type or ceramic type.
The sleeve type reference electrode has small resistance and
is optimum when sufficient care is taken on the handling. If
the sleeve is tightened in excess, the resistance becomes large,
and if it is too loose, the outside cylinder solution flows out in
large amount, and therefore the proper tightening is necessary.
Because there is a product of large resistance among the ceram-
ics types, use that for ion selective electrode. Take care that, if
the ceramics type is dried o r stained, the resistance becomes large.
Immerse the reference electrode in the same solution as the
outside cylinder solution in either cases. In the case of the sleeve
type, adjust the tightening of sleeve a t the time of use.
In the case of using potassium chloride saturated solution as
inside and outside cylinder solutions, if the temperature of the
solution lowers, potassium chloride crystals may deposit and ad-
here to make the resistance greater, and therefore care shall
be taken.
If the magnetic stirrer is used for a long time, it generates heat
and may change the temperature of the solution, and therefore
care shall be taken for the change of temperature of the solution.
Where the indication of potentiometer becomes unstable by the
stirring speed, the resistance of reference electrode has, in many
cases, become large.
The response time of fluoride ion selective electrode is approx.
1 min at concentration of 0.1 mgF-/Z of fluoride ion, and approx.
30 s at 1mgF-/Z or more in solution temperature 10 t o 30 "C.
The difference of potentials between fluoride ion standard solu-
tion (1mgF-/Z) and fluoride ion standard solution (100 mgF-/Z)
becomes within the range of 110 to 120 mV (25 OC),and the work-
ing curve from concentration of 0.1 t o 100 mgF-/E of fluoride ion
becomes straight line.
(a) Take 100 ml of distillate obtained by distillation operation of 31.1 (3) into
a 200 ml beaker, add 10 ml of the buffer solution (pH 5.21, and adjust the
temperature of the solution t o within fl "C of the solution temperature
specified in (3)( c ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
162
K O101 : 1998
(b) Carry out the operation of (3)(b)and (cl to obtain the concentration of fluoride
ion from the working curve, and calculate the concentration of fluoride ion
in the sample (mgF-4).
Remarks 3 In the case of ion concentration meter, use fluoride ion standard
solution (1mgF-/Z) and that (100 mgF-/O, carry out the opera-
tion of (3) (b) and (e),and so adjust the indicating values of ion
concentration meter as t o become 1mgF-/Z and 100 mgF-4, re-
spectively. Further, confirm the indication of the ion concentra-
tion meter by using fluoride ion standard solution (0.1 mgF-/Z)
and that (10 mgF-/Z).
4 Since only fluoride ion can be measured by the ion selective
electrode method, all fluorine compounds are preliminarily
converted t o fluoride ions by distillating operation, and they
are measured.
The allowable limits of principal coexisting substances are
shown by the maximum ratio as follows.
HC03-, Cl-, Nos-, I-, Br-, HP042-: lo3
SOP: 104
Though OH-, Al3+,and Fe3+interfere measurement, since
they are removed by distillating separation, there is no influ-
ence.
5 Potentiometric titration by fluoride ion selective
electrode Take 100 ml of the distillate obtained by distillating
1 1
operation of 31.1 (3)into a beaker, titrate 300 to 30 mol/Z lan-
thanum (III) nitrate solution while measuring the potential in
accordance with the operation of (3)(b)and (e) as appropri-
ate, draw the titration curve, obtain the end point of titration,
1
and calculate the amount of fluoride ion. 1 ml of 30 moVZ lan-
thanum (III) nitrate solution is equivalent to 1.899 mg of F-.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
163
K O 1 0 1 : 1998
32 Chloride ion (Cl-) To determination of chloride ion the mercury (II) thiocy-
anate absorptiometry, mercury (II) nitrate titrimetric method, silver nitrate titri-
metric method, ion selective electrode method or ion chromatography shall apply.
32.1 Mercury (II) thiocyanate absorptiometry The chloride ion is determined

by measuring the absorbance of orange red complex generated by the reaction of
iron (III) and thiocyanate ion substituted by chloride ion when mercury (II) thio-
cyanate and ammonium iron (III) sulfate are added to the sample.
Determination range: C1- 20 to 500pg
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Ammonium iron (III) sulfate solution Dissolve 60 g of ammonium iron

(III) sulfate 12 hydrate specified in JIS K 8982 in 1I of nitric acid (5 molíl)
(add 600 ml of water to 380 ml of nitric acid specified in J I S K 8541, cool to
room temperature and further add water to make 11). If turbidities exist,
filter them and preserve in a brown bottle.
Ethanol mercury (II) thiocyanate solution Dissolve 1.5 g of mercury
(II) thiocyanate specified in J I S K 9519 in 500 ml of ethanol (95) specified
in JIS K 8102 and preserve in a coloured glass bottle.
Chloride ion standard solution ( 1 mgCl-/ml) Preliminarily heat sodium
chloride (standard reagent for volumetric analysis) specified in J I S K 8005
at 600 "C for approx. 1h, and allow to cool in a desiccator. Then, take its
1.648 g to NaC1 as 100 % to dissolve in a small amount of water, transfer it
into a 1 O00 ml volumetric flask, and add water up to the marked line. Oth-
erwise, use chloride ion standard solution C1- 1O00 specified in JIS K 0029.
Chloride ion standard solution (10 pgCl-/ml) Take 10 ml of chloride
ion standard solution (1mgCl-/ml) into a 1O00 ml volumetric flask, and add
water up to the marked line.
(a) Glassware Wash with water prior to use.
(a) Filter the sample with a filter paper of class 5C, discard approx. 50 ml of
the initial filtrate, and take 50ml of the next filtrate (where it contains
not less than Cl- 0.25 mg, take a proper amount and dilute with water to
50 mi) into a 50 ml measuring cylinder (with stopper).
(b) Add 10 ml of ammonium iron (III) sulfate solution and 5 ml of ethanol mercury
(II) thiocyanate solution, and stopper to mix thoroughly by shaking.
(c) Keep the temperature of the solution a t approx. 20 OC(') and allow to stand
for approx. 10 min.
(d) As a blank test, carry out the operation of (a) to ( c ) on 50ml of water.
164
K O101 : 1998
(e) Transfer the solution of ( c ) to the absorption ce11(2),use the solution of the
blank test of (d) as a reference solution, and measure the absorbance near
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(0 Obtain the amount of chloride ion from the working curve, and calculate
the concentration of the chloride ion in the sample (mgC1-A).
Working curve Take stepwise 2 t o 50 ml of chloride ion standard solution
(10pgCl-/ml) into a 50ml measuring cylinder (with stopper), add water to
make 50m1, and then carry out the operations of (b)to (e) to prepare the
relation curve between the amount of chloride ion (Cl-) and the absorbance.
Notes (l) The colouring speed is different depending upon the temperature,
and therefore the temperature difference at colouring shall be
within f2 O C .
(2) Where absorption cell of 20 mm in optical path length is used it
is suitable for determination of C1- 10 to 250pg, where that of
50mm in optical path length is used for determination of C1- 5
t o lOOpg, and where that of 100mm is used for determination
of C1- 2.5 t o 50 pg.
Further, where absorption cell of 100 mm in optical path length
is used, take 100 ml of the sample, and use the reagents of two
times amount.
Remarks 1 Bromide ion, iodide ion, cyanide ion, etc. are interferent. Fur-
ther, thiosulfate ion, sulfide ion and sulfite ion are also
interferent, and therefore these shall be oxidized preliminarily.
2 Because the chloride ion exists widely, take care for the con-
tamination from the sweat on hand, or the like and for the
pollution from the air in the laboratory or the like.
3 Because the mercury compound is used, care shall be espe-
cially taken for the treatment of waste solution.
32.2 Mercury (II) nitrate titrimetric method Chloride ion shall be determined
by titration with mercury (II) nitrate solution after adjusting pH of the sample to 2.5.
Iodide ion and bromide ion are determined as chloride ion. The reducing sub-
stances such as sulfite ion, hydrazinium ion (hydrazine), hydroxylamine interfere
with the determination, however, they do not interfere when oxidized with hydro-
gen peroxide preliminarily. Chrome (VI) and iron (III) with 10 mg/Z o r less do not
interfere, respectively.
Determination range: C1- 0.1 to 5 mg
(a) Nitric acid (1+65) Prepare by using nitric acid specified in JIS K 8541.
(b) Hydrogen peroxide (l+l> Prepare by using hydrogen peroxide specified
in J I S K 8230.
(c) Mixed indicator Weigh out 0.50 g of diphenylcarbazone specified in JIS K
8489, 0.05 g of Bromophenol Blue specified in J I S K 8844 and 0.12 g of xy-
lene cyano1 FF specified in JIS K 8272, dissolve them in 100 ml of ethanol (95)
specified in JIS K 8102,and put into a coloured glass bottle to preserve.
165
K 0101 : 1998
(d) Chloride ion standard solution (0.5mgC1-/ml) Take 100 ml of sodium

chloride standard solution (1mgCl-/ml) of 32.1 (1) (c) into a 200 ml volu-
metric flask and add water to the marked line.
(e) 7.05 mmol/l Mercury (II) nitrate solution Take 2.5 g of mercury (II)
nitrate n-hydrate specified in JIS K 8558, add 20 ml of water containing
0.5ml of nitric acid specified in JIS K 8541 to dissolve, transfer into a
1O00 ml beaker, and add water to make 11.
Standardization Take 20 ml of chloride ion standard solution (0.5 mgC1-
/ml) into a beaker, add water to make 100 ml, add 5 drops of mixed indica-
tor, add dropwise nitric acid (1+65) until the colour of the solution turns
from blue to bluish green, and further add 1ml to make pH approx. 2.5.
Then, titrate with this 7.05 mmol/Z mercury (II) nitrate solution, and take
the point when the colour of the solution.turnsfrom yellowish green through
grey to purple as the end point. Separately, take 100 ml of water, and carry
out the blank test t o correct the titrated value. Calculate the factor (f) of
7.05 mmol/Z mercury (II) nitrate solution from the corrected ml number (x),
according t o the following formula:
f=-
20
X
(a) If turbidity appears in the sample, filter the sample with a filter paper of
class 5C(3), discard approx. 50 ml of the initial filtrate, and then take 100 ml
of the next filtrate (where it contains C1- 5 mg or more, take a proper amount,
and dilute with water to 100 ml) into a beaker.
(b) Where reducing substances such as sulfite ion, hydrazinium ion, hydroxyl-
and mix by
amine, etc. coexist, add drop by drop hydrogen peroxide (l+l)
stirring t o decompose.
( c ) Add 5 drops of mixed indicator solution, and after adding drop by drop nitric
acid (1+65) until the colour of the solution turns clearly from blue to blu-
ish green or yellowish green, further add its 1 ml(4).
(d) Titrate with 7.05 mmol/Z mercury (II) nitrate solution, and take the point
when the colour of the solution has turned from yellowish green through
grey to purple as the end point.
(e) As a blank test, take 100 ml of water to carry out the operation (cl and (d).
(0 Calculate the concentration of chloride ion in the sample (mgCl-/l) accord-
ing t o the following formula:
C = (a - b) x f x -
'Ooo x0.5
V
where, C : chloride ion (mgCl-/Z)
a : 7.05 mmol/Z mercury (II) nitrate solution required
for titration (ml)
b : 7.05 mmol/Z mercury (II) nitrate solution required
for titration of blank test (ml)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
166
K O101 : 1998
f : factor of 7.05 mmoll2 mercury (II) nitrate solution

V : sample (mi)
0.5 : chloride i o n equivalent t o 1ml of 7.05 mmolll
mercury (II) nitrate solution (mg)
Notes (3) In the case where remarkable turbidity in the sample is not con-
firmed, the filter operation may be omitted.
(4) Where the acidity is strong, adjust pH t o approx.2.5 with so-
dium hydroxide solution (40 gL).
32.3 Silver nitrate titrimetric method Chloride ion shall be determined by the
titration with silver nitrate solution using uranine (sodium fluoresceine) [9-(2-
carboxyphenyl)-6-hydroxy-3H-xanthene-3-onedisodiumsalt (named by IUPAC)] so-
lution as indicator by adjusting pH of the sample t o approx. 7.
Determination range: C1- 1 mg or over
Remarks 5 If bromide ion, iodide ion, cyanide ion, etc. coexist, these are de-
termined as chloride ion. Sulfite ion, thiosulfate ion, and sulfide
ion interfere with the determination, however, they do not inter-
fere when oxidized with hydrogen peroxide preliminarily.
Nitric acid (1+65) As described in 32.2 (1)(a).
Sodium carbonate solution (50 glZ) Dissolve 5 g of sodium carbonate
Sodium fluoresceine solution (2 glE) Dissolve 0.2 g of uranine (sodium
fluoresceine) specified in JIS K 8830 in water t o make 100 ml.
Dextrin solution Dissolve 2 g of dextrin specified in JIS K 8646 in wa-
ter t o make 100 ml. Prepare this solution a t the time of use.
Chloride ion standard solution ( i mgC1-lml) As described in 32.1 (i)(cl.
28.2 mmol/Z silver nitrate solution Dissolve 4.8 g?ofsilver nitrate specified
in JIS K 8550 in water to make 11. Preserve i t in a coloured glass bottle.
Standardization Take 20 ml of chloride ion standard solution (i mgC1-l
mi) into a beaker, and add water t o make the amount of solution approx.
50 ml. Add to this solution 5 ml of dextrin solution and 1 t o 2 drops of so-
dium fluoresceine solution (2 glZ), and while gently stirring t o mix, titrate
with this silver nitrate solution. Take the point when yellowish green fluo-
rescence disappears and slight red colour appears as the end point. Calcu-
late the factor (f)of 28.2 mmoll1 silver nitrate solution from the number of
ml of silver nitrate solution required (XI according t o the following formula:
f=-
20
X
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
167
K O101 : 1998

When turbidities exist in the sample, filter with a filter paper of class 5C(3),
discard approx. 50 ml of the initial filtrate, and take 50 ml of the next fil-
trate (in the case of C1- 20 mg or more, take a proper amount and dilute
with water t o 50ml) into a beaker.
Adjust pH t o approx. 7 with sodium carbonate solution (50 gll) when the
sample is acidic, and with nitric acid (1+65) when the sample is alkaline.
Add 5 ml of dextrin solution and 1 t o 2 drops of sodium fluoresceine solu-
tion (2 g/Z) and stir t o mix.
While stirring gently t o mix, titrate with 28.2 mmol/l silver nitrate solu-
tion. Take the point when the yellowish green fluorescence disappears and
pale red colour appears as the end point.
Calculate the concentration of chloride ion in the sample (mgCl-/Z) accord-
ing t o the following formula:
1 O00
C=axfx-
V
where, C : chloride ion (mgC1-/Z)
a : 28.2 mmol/Z silver nitrate solution required for
titration (mi)
f : factor of 28.2mmollZ silver nitrate solution
V : sample (mi)
1: chloride ion equivalent to 1 ml of 28.2 mmol/Z sil-
ver nitrate solution (mg)
32.4 Ion selective electrode method Chloride ion shall be determined by the
measurement of the potential by using chloride ion selective electrode as indication
electrode by adjusting pH t o approx. 5 by adding acetate buffer solution to the sample.
Remarks 6 Sulfide ion and the like interfere by this method.
Determination range: C1- 5 t o 1O00 mglZ
(a) Acetate buffer solution (pH 5) Dissolve 100 g of potassium nitrate speci-
fied in JIS K 8548 and 50ml of acetic acid specified in JIS K 8355 into
500 ml of water. Add to this solution sodium hydroxide solution (100 glZ),
adjust pH to 5 by use of a pH meter, and add water t o make 11.
(b) Chloride ion standard solution (1 O 0 0 mgC1-/I) As described in
32.1 ( i )(cl.
(c) Chloride ion standard solution (100 mgCl-/Z) Take 20 ml of chloride
ion standard solution (iO00 rngCl-/Z) into a 200 ml volumetric flask, and
add water t o the marked line.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
168
K O101 : 1998
(d) Chloride ion standard solution (10mgCl-lZ) Take 20 ml of chloride ion

standard solution (100 mgC1-/Z) into a 200 ml volumetric flask, and add water
(e) Chloride ion standard solution (5 mgCl-/Z) Take 10 ml of chloride ion
standard solution (100 mgC1-/Z) into a 200 ml volumetric flask, and add water
(a) Potentiometer As described in 31.2 (2)(a).
(b) Indication electrode Chloride ion selective electrodes
( c ) Reference electrode As described in 31.2 (2)( c ) . However, use potas-
sium nitrate solution (100 g/Z) for outside cylinder solution.
(d) Magnetic stirrer As described in 31.2 (2)(d).
(3) Preparation of working curve
Transfer 100 ml of chloride ion standard solution (5 mgCl-/Z) into a 200 ml
beaker, and add 10 ml of acetate buffer solution (pH 5)(5).
Immerse the indication electrode(6) (7) and reference electrode (8) (9) into the
solution, and stir so strongly that bubbles do not touch the electrode by
using a magnetic stirrer(l0)(11).
Measure the temperature of the solution, and measure the potential by a
potentiometer (12).
Take respectively 100 ml of chloride ion standad solution (10 mgCl-/Z), 100 ml
of chloride ion standard solution (100 mgCl-/Z), and 100 ml of chloride ion
standard solution (1000mgCl-íZ) into a 200ml beaker, and add 10ml of
acetate buffer solution (pH 5 ) ( 5 ) ) . Adjust temperature of each chloride ion
standard solution (10 t o 1 O00 mgCl-/Z) within +1"C t o the liquid tempera-
ture of ( c ) , carry out the operation of (b) and ( c ) , and measure the poten-
tial of chloride ion standard solution.
Take the concentration of chloride ion on the logarithmic axis and take the
potential(l3) on the linear axis of semilogarithmic section paper to prepare
the relation curve between the concentration of chloride ion (mgCl-/Z) and
the potential.
Notes (5) Acetic acid buffer solution (pH 5) is used for adjusting pH to
approx. 5 a t the time of measurement and for making the ionic
strength constant.
(6) Immerse the electrode in the chloride ion standard solution
(5mgCl-/Z), and after the indicating value has become stable,
measure the potential.
(7) As described in Note (12) of 31.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
169
K O101 : 1998
(9) Use potassium chloride solution (3 moVZ t o saturate) as inside-

cylinder solution of reference electrode, and potassium nitrate
solution (100 g/Z) as outside-cylinder solution. In the case of using
potassium chloride solution (saturated) as inside-cylinder solu-
tion, if the temperature of the solution lowers, crystals of potas-
sium chloride may deposit and adhere t o make the resistance
greater, and therefore, care shall be taken.
Because the potassium chloride solution of the inside-cylin-
der solution flows into the potassium nitrate solution (100 g/Z)
of the outside-cylinder solution, the outside-cylinder solution also
shall be exchanged periodically.
(12) If the concentration of chloride ion is 5 mgC1-/Z or more at the
solution temperature of 10 t o 3 0 ° C , the response time of chlo-
ride ion selective electrode is within 1min.
(13) The potential difference between the chloride ion standard solution
(10 mgCl-/Z) and that (iO00 mgC1-/Z) is within the range from 110
to 120 mV (25 O C ) , and the working curve at 5 t o 1O00 mgC1-/Z of
chloride ion concentration is a straight line.
(a) Take 100 ml of the sample(l4) (15) into a 200 ml beaker, add 10 ml of acetic
acid buffer solution (pH 5 ) , and adjust the temperature of the solution within
I1 OC t o that of the solution specified in (3) ( c ) .
(b) Carry out the operation specified in (3)(b)and ( c ) t o obtain the concentra-
tion of chloride ion from the working curve, and calculate the concentra-
tion of chloride ion in the sample (mgCl-/Z).
Notes (14) Adjust pH t o approx. 5 by sodium hydroxide solution (40glZ) for
the acidic sample, and by acetic acid (1+10)for the alkaline sample,
in advance.
(15) For the sample containing sulfide ion, preliminarily add zinc
acetate solution (100 gíl) to fix sulfide ion. Filter off the precipi-
tate with a filter paper, and adjust pH of the filtrate t o approx. 5.
Remarks 7 In the case of ion densitometer, use chloride ion standard solu-
tion (10 mgCl-/Z) and that (iO00 mgCl-/Z), and carry out the op-
eration specified in (3) (b)to ( c ) to adjust the indicating values
of ion densitometer t o 10 mgCl-/l and 1O00 mgCl-/Z. Further
confirm the indication of the ion densitometer by use of other
chloride ion standard solution ( 5 mgCl-/Z) and that (100 mgCl-/Z).
8 The allowable limits of principal coexisting components are
shown by the maximum ratio as follows.
Nos-, so42-,PO^-: 104
F-: lo2
Br-:
I-, CN-, s2-: 10-3
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
170
K O101 : 1998
9 Potential difference titrimetric method by ion selective

electrode Take the 100 ml of a sample into a beaker, adjust
pH of the sample at 7, use chloride ion selective electrode o r
silver ion selective electrode, titrate with 10 to 100 mmol/Z sil-
ver nitrate solution while measuring the potential in accordance
with the operation of (3)(b),draw the titration curve, and obtain
the end point of titration. The inflection point of the titration
curve becomes the order of iodide ion, bromide ion, and chlo-
ride ion. Obtain the end point from each inflection point, and
calculate the concentration of each ion.
1ml of 10 mmol/Z silver nitrate solution is equivalent to
1.269 mg of I-, 0.799 mg of Br-, and 0.354 5 mg of Cl-.
32.5 Ion chromatography Chloride ion in a sample is determined by an ion

chlomatography.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Determination range: C1- 0.1 t o 25 mg/Z(16>
Repeatability: 2 t o 10 % in coefficient of variation (is different according t o the
apparatus and measuring conditions.)
Note (16) For the system of being combined with a suppressor, the determina-
tion range is 0.05 t o 25 mg/Z of Cl-.
( i ) Reagents The following reagents shall be used.
(a) Water Water A2 or A3 specified in JI$ K 0557.
(b) Eluent Since the eluent(l7) is different according to the type of appara-
tus and the class of anion exchanger filled in a separating column, the con-
ditions of separation of chloride ion, nitrite ion, bromide ion, nitrate ion
and sulfate ion are preliminarily confirmed by the operation of Note (21).
(c) Reclaiming solution Though the reclaiming solution(l8)is used when the
suppressor is used, the reclaiming solution is different according t o the type
of apparatus and the class of the suppressor. The operation of Note (21) is
performed by preliminarily combining with the separating column, and the
performance of reclaiming solution is confirmed.
(d) Chloride ion standard solution (imgCl-/mi) As described in 32.1 (i) ( c ) .
(e) Chloride ion standard solution (0.1mgCl-/ml) Take 10 ml of chloride
ion standard solution (i mgCl-/ml) into a 100 ml volumetric flask, and add
water to the marked line.
(0 Anion mixed standard solution [(0.1mgCl-, 0.5 mgNOz-, 0.5 mgBr-,
0.5 mgNOs-, 1 mgS0d2-/ml1 Take respectively 10 ml of chloride ion stan-
dard solution ( i mgC1-/ml) of 32.1 (1)( c ) , 10 ml of nitrite ion standard so-
lution (5 mgNOz-/ml) of 37.1.2 (1)(d),10 ml of bromide ion standard solution
(5 mgBr-/ml) of 34.2 (1)(d), 10 ml of nitrate ion standard solution (5 mgNOs-
/ml) of 37.2.5 (1)(d),and 10 ml of sulfate ion standard solution (10 rngS0d2-
/mi) of 42.4 (i) (d)into a 100 ml volumetric flask, and add water to the marked
line. Prepare at the time of use.
171
K O101 : 1998
An example of the preparation method of eluent is given as fol-

lows.
Example for using suppressor
[Sodium hydrogen carbonate solution (1.7 mmol/Z)-sodium
carbonate solution (1.8 mmol/Z)l Dissolve 0.143 g of sodium
hydrogen carbonate specified in JIS K 8622 and 0.191 g of so-
dium carbonate specified in JIS K 8625 in water t o make 11.
[Sodium hydrogen carbonate solution (0.3mmol/Z)-sodium
carbonate solution (2.7 mmol/Z)l Dissolve 0.025 g of sodium
hydrogen carbonate specified in JIS K 8622 and 0.286 g of so-
dium carbonate specified in JIS K 8625 in water to make 11.
Sodium carbonate solution (3 mmol/Z) Dissolve 0.318 g of
sodium carbonate specified in JIS K 8626 in water t o make 11.
Example for not using suppressor
[Potassium gluconate solution (1.3 mmol/Z)-sodium tetrabo-
rate solution (1.3 mmol/Z)-boric acid solution (SO mmol/Z)-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
acetonitrile solution (100g/Z)-glycerol solution (5 g/Z)]

Dissolve 0.31 g of potassium gluconate, 0.5 g of sodium tetraborate
decahydrate specified in JIS K 8866, 1.86 g of boric acid speci-
fied in JIS K 8863, 100 g (128 mi) of acetonitrile specified in JIS
K 8032 and 5 g (4ml) of glycerol specified in JIS K 8295 in wa-
ter to make 11.
[Phthalic acid solution (2.5 mmol/Z)-2-amino-2-hydroxy-
methyl-1,3-propanediol solution (2.4 mmol/Z)] Dissolve
0.415 g of phthalic acid and 0.291 g of 2-amino-2-hydroxymethyl-
lY3-propanedioltris(hydroxymethy1)aminomethane specified in
JIS K 9704 in water to make 11.
An example of the preparation method of reclaiming solution is
shown as follows.
Sulfuric acid (12.5 mmol/Z) Dissolve sulfuric acid (0.5 mol/Z)
(add 30 ml of sulfuric acid specified in JIS K 8951 little by little
in 500 ml of water, cool, and make to 11 with water) 25 ml with
water to make 1E .
(2.1) Ion chromatograph Though there are the ion chromatograph of the combi-
nation system of a separating column and a suppressor(19) and of the separat-
ing column single system, the ion chromatograph shall conform to the following
conditions and be able to separate and determine chloride ion, nitrite ion, bro-
mide ion, nitrate ion, sulfate ion, etc.
(a) Separating column The separating column is made of stainless steel or
synthetic resin(29, which is filled with strongly basic anion exchanger (surface
layer covering type o r all porous silica type, etc.)(21).
(b) Detector Electric conductivity detector
172
K O101 : 1998
(2.2) Recording part As described in 4.2 (6) of JIS K 0127.

Notes (19) It aims for converting cation in eluent to hydrogen ion, which is filled
by a cation exchange membrane (there are membrane type and elec-
trical dialysis type) having a n enough ion exchange capacity for the
concentration of cation in the eluent or a cation exchanger having
the same performance. It is used by being combined with reclaim-
ing solution. For the electrical dialysis type, however, use the emu-
ent from the detector (solution discharged from detector) as the
reclaiming solution.
(20) There are, for example, the separating columns made of tetrafluoro-
ethylene resin, polyether ether ketone, etc.
(21) Make eluent flow at a constant flow rate (for example, 1to 2 ml/min),
inject a specific amount of anion mixed standard solution [(lo pgC1-,
10 pgNOz-, 10 pgBr-, 10 pgNOs-, 10 pgS042-)/ml]into the separating
column of an ion chromatograph, obtain a chromatogram, and use the
separating column which can separate each anion (separation degree
1.3 or more) by above-mentioned procedure.
Further, allow preferably the performance of the separating col-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
umn to be confirmed periodically.

Prepare the anion mixed standard solution [( 10 pgC1-, 10 pgNOz-,
10 pgBr-, 10 pgNOs-, 10 pgS042-)/m11 as follows.
Take respectively each 5 ml of chloride ion standard solution
(1mgC1-/ml), nitrite ion standard solution (1 mgNOz-/ml), bromide ion
standard solution (i mgBr-/ml), nitrate ion standard solution
(1 mgNOs-/ml), and sulfate ion standard solution (i mgS042-/ml) into
a 500 ml volumetric flask, and add water t o the marked line.
(a) Filter the sample with a 0 . 4 5 hole ~ ~ diameter filter membrane or class
5C filter paper (or class 6 filter paper), discard approx. 50 ml of the initial
filtrate, and take the filtrate thereafter.
(b) In the case where the electric conductivity of the sample is 10 mS/m (100 pS/
cm} (25 OC) or over, dilute at a specific ratio with water, so that the elec-
tric conductivity becomes 10 mS/m o r under.
(a) Make the ion chromatograph a state capable of operating, and allow the
eluent to flow to the separating column at a specific flow rate (for example,
1 to 2 ml/min). For the apparatus requiring a suppressor, make prelimi-
narily the reclaiming solution flow thereto at a specific flow rate.
(b) Inject a specific amount of the sample (for example, a constant amount of
50 to 200 pl) for which the preparatory operation of (3)has been performed
into the ion chromatograph and record the chromatogram.
(c) As for the peak corresponding to chloride ion on the chromatogram, read
its indicated value(22).
173
K O 1 0 1 : 1998
(d) When the sample is diluted, the blank test with the same amount of water
as the sample shall be carried out the operations (a)t o ( c ) , and correct the
indicated value (22) obtained from the sample.
(e) Obtain the amount of chloride ion from the working curve, and calculate
the concentration of chloride ion in the sample (mgCl-/Z).
Working curve Deal out step by step 0.1 to 25 ml of chloride ion stan-
dard solution (0.1 mgC1-/ml)(23)into a 100 ml volumetric flask, and add water
t o the marked line. Carry out the operation of (a)to ( c ) for this solution,
obtain the peak corresponding t o each chloride ion and read the indicated
value(22). Separately, carry out the operation of (a)t o ( c ) for water as a
blank test, and correct the indicated value corresponding to each chloride
ion. Thereafter, prepare the relation curve between the amount of chlo-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ride ion (Cl-) and the indicated value. The working curve is prepared a t
the time of measurement of the sample.
Notes (22) Indicated value means peak height or peak area.
(23) In the case where anions other than chloride ion are simulta-
neously tested, anion mixed standard solution [(O. 1 mgCl-,
0.5 mgNOz-, 0.5 mgBr-, 0.5 mgNOs-, 1mgS042-)/m11shall be used.
Remarks 10 If nitrite ion is 200 mgll or under when the concentration of
chloride ion is 1mg/Z, it does not interfere.
11 If a separating column is used continuously, its performance
decreases, the operation of Note (21) shall be carried out peri-
odically to confirm.
When the performance decreased, prepare solution of which
concentration is ten times the eluent, pour it in the separating
column, wash it and confirm by carrying out of the operation of
Note (21). If it does not recover, replace by a new one.
The separating column is contaminated by suspension matter
and organic matter (protein, oils, surface active agent, etc.) in
the sample, and its performance gradually decreases. There-
fore, the sample containing suspension matter shall be tested
after removing them by the preparatory operation of (3).
For the sample containing organic matter, it is filtered with
an ultrafilter membrane, and after the organic matter is re-
moved as much as possible, it shall be tested.
If anions of strong affinity with filler of the separating col-
umn (for example, iodide ion, chromate ion, etc,) exist in the
sample, they are adsorbed in the filler, and the separation
performance gradually decreases. Therefore, the solution of
5 t o 10 times the concentration of eluent is prepared, and the
separating column shall be washed by injecting thereinto in
the same way as for the sample.
Further, if oxidizing matter and reducing matter coexist, the
separation performance of the separating column decreases. In
such a case, when the sample is diluted at a specific rate with
water to be tested, effects can be prevented to a certain extent.
174
K O101 : 1998
33 Iodide ion (I-) To the determination of iodide ion, the iodine extraction
absorptiometry o r the iodine titrimetric method applies.
33.1 Iodine extraction absorptiometry Iodide ion is allowed t o react with ni-
trite ion in the solution acidified with sulfuric acid and the liberated iodine is ex-
tracted with chloroform, and the absorbance of the solution is measured to determine
the iodide ion.
Determination range: I- 0.1 t o 5 mg
(1) Reagents The following reagents shall be used,
Sulfuric acid ( l + l ) As specified in 4.4 (i)(b).
Sodium nitrite As specified in JIS K 8019.
Urea solution (10 g/Z) Take 1g of urea specified in JIS K 8731, dissolve
in water to make 100 ml.
Sodium sulfate As specified in JIS K 8987.
Iodide ion standard solution ( i mgI-/ml) Take 1.31g of potassium io-
dide specified in JIS K 8913, dissolve in a small quantity of water, transfer
it into a 1O00 ml volumetric flask, and add water up t o the marked line.
Iodide ion standard solution (0.1 mgI-/ml) Take 20 ml of iodide ion
standad solution (i mgI-/ml) in a 200 ml volumetric flask, and add water
up to the marked line. Prepare this solution at the time of use.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

Take a proper amount of sample(1)( 2 ) (containing 0.1 t o 5 mg as I-) in a
100 ml separating funnel, and add water to make approx. 50 ml.
Add 1ml of sulfuric acid (l+l) [if the sample is alkaline, add sulfuric acid
(l+l)to neutralize] and 0.5 g of sodium nitrite and mix by shaking.
Add 10 ml of chloroform, and after vigorously shaking to mix for approx. 2' min,
allow to stand still,
Transfer the chloroform layer into another 100 ml separating funnel. Again
add 10 ml of chloroform t o the aqueous layer t o extract, and then combine
this chloroform layer to the previous chloroform layer.
Add 50 ml of urea solution (10 g/Z) t o the separating funnel containing the
chloroform, and shake vigorously to mix for approx. 2 min, and wash the
chloroform layer.
175
K 0101 : 1998
After allowing t o stand for about 5 min, transfer the chloroform layer into
a 30 ml Erlenmeyer flask with ground stopper containing approx. 1g of sodium
sulfate, shake to mix and remove water.
Transfer a portion of chloroform layer into an absorption cell, and measure
the absorbance at a wavelength near 515 nm using chloroform as reference
solution.
For the blank test, take 50 ml of water, and carry out the operations speci-
fied in (b) to ( g ) t o correct the absorbance obtained on the sample.
Obtain the amount of iodide ion from the working curve, and calculate the
concentration of iodide ion in the sample (mgI-/Z).
Working curve Take stepwise 1 t o 50 ml of iodide ion standard solution
(0.1 mgI-/ml) in a 100 ml separating funnel, and carry out the operations
specified in (a) t o (h)t o measure the absorbance and prepare the relation
curve between the amount of iodide ion (I-) and the absorbance.
Notes (1) In the case where the concentration of iodide ion is not more than
2 mgI-/Z, take a proper amount of sample, add sodium hydroxide
solution (200 g/Z) t o make the solution alkaline, and heat gently
to concentrate. When the sample becomes turbid, filter it, and
carry out the operation specified in (a) and thereafter.
(2) In the case where organic matters are present in a large amount,
take 200ml of sample, add 2 t o 3 m l of potassium aluminium
sulfate solution (dissolve 5 g of potassium aluminium sulfate 12-
water specified in JIS K 8255 in water to make 100ml) to it,
and then add sodium hydroxide solution (50 gil) until precipitates
of aluminium hydroxide appear. After allowing t o stand for approx.
5 min, filter it, and taking a proper amount of the filtrate, carry
out the operation specified in (a) and thereafter.
Remarks 1 When the sample containing iodate ion is acidified with sul-
furic acid, iodine is liberated by the reaction with iodide ion,
therefore a part of or total iodate ion is determined as iodide
ion. Bromide ion does not interfere.
33.2 Iodine titrimetric method Iodide ion is oxidized with hypochlorous acid
to iodate ion at pH 1.3 t o 2.0, After the decomposition of the excessive hypochlorous
acid with sodium formate at pH 3 t o 7, potassium iodide is added, and the liberated
iodine is titrated with sodium thiosulfate solution to determine the iodide ion.
Determination range: I- 0.1 mg or more
( i ) Reagents Use the following reagents
(a) Hydrochloric acid ( l + l ) Prepare by using hydrochloric acid specified in
JIS K 8180.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Hydrochloric acid (1+1i) Prepare by using hydrochloric acid specified

in JIS K 8180.
(c) Sodium hypochlorite solution (effective chlorine 35 gll) Determine
the amount of effective chlorine in the sodium hypochlorite solution (effec-
tive chlorine 7 to 12 % ) ( 3 ) , and dilute with water for the effective chlorine
t o become 35glZ. Prepare this solution at the time of use.
176
K O101 : 1998
(d) Sodium formate solution (approx. 400 gll) Dissolve 40 g of sodium for-
mate specified in JIS K 8267 in water t o make 100 ml.
(e) Potassium iodide As specified in JIS K 8913.
(0 Methyl Orange solution (i gll) As described in 22.1.1 (i)(d).
( g ) Starch solution (10 gll) As described in 22.1.2 (1)(i).
(h) 10 mmolld sodium thiosulfate solution As described in 28.3 ( i )(e).

Note (3) Take 10ml of sodium hypochlorite solution (effective chlorine 7
to 12 %) into a 200 ml volumetric flask, and add water t o the marked
line. Take its 10 ml into a 300 ml Erlenmeyer flask with ground
stopper, and add water to make approx. 100ml. Add 1 t o 2 g of
potassium iodide and 6 m l of acetic acid (l+l>, tightly stopper,
sufficiently mix by shaking, and allow t o stand in a dark place
for approx. 5 min. Thereafter, titrate with 50 mmoVZ sodium thio-
sulfate solution [as described in 24.1 (1) ( g ) ] .When the yellow of
the solution becomes pale, add 1 ml of starch solution (10 glZ) as
indicator, and titrate until the generated blue colour of iodine starch
disappears. Separately, take 10 ml of water for a blank test, and
correct the titre by performing the same operation. Calculate the
amount of effective chlorine by the formula below.
N=axfx-x-2oo 'Oo ~0.001773

10 V
where, N : amount of effective chlorine ( g d )
a : 50 mmol/Z sodium thiosulfate solution re-
quired for titration (mi)
f : factor of 50 mmolll sodium thiosulfate solu-
tion
0.001 773 : effective chlorine equivalent to 1 ml of
50 mmol/Z sodium thiosulfate solution (g)
V : sodium hypochlorite solution (effective chlo-
rine 7 to 12 %) (mi)
(a) Take a proper amount of sample(1)(2) (containing 0.1 t o 5 mg of I-) in a
300 ml Erlenmeyer flask with ground stopper, add one drop of Methyl Orange
solution (i gíZ) as indicator, add dropwise hydrochloric acid (1+11) until the
colour of the solution turns pale red, and then add water to make approx.
50 ml.
(b) Add 1ml of sodium hypochlorite solution (effective chlorine 3.5 g/Z), then
add hydrochloric acid (1+11) to adjust pH to 1.3 t o 2.0 and immerse the
solution into boiling water bath for approx. 5 min.
(cl Add 5 ml of sodium formate solution (400 g/Z)(4), and again immerse into
the boiling water bath for approx. 5 min t o decompose the excessive hypochlo-
rous acid.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
177
K O101 : 1998
After standing t o cool, add 1 g of potassium iodide and 6 ml of hydrochloric

acid (l+l), stopper tightly, mix by shaking, and allow to stand in a dark
place for approx. 5 min.
Titrate the liberated iodine with 10 mmol/Z sodium thiosulfate solution, After
the yellow colour of the solution has turned to pale, add 1 ml of starch solution
(10 gll) as indicator, and titrate until the generated blue colour of iodine
starch disappears.
For the blank test, take 50 ml of water in a 300 ml Erlenmeyer flask with
ground stopper, and carry out the operations specified in (a) to (e).
Calculate the concentration of iodide ion in the sample (mg1-/Z) according
t o the following formula:
c = (u - b) x f x -
'Ooo x0.2115
V
where, C : iodide ion (mg1-/Z)
U : 10 mmol/Z sodium thiosulfate solution required for
titration (ml)
b : 10 mmol/Z sodium thiosulfate solution required for
titration in the blank test (mi)
f : factor of 10 mmol/Z sodium thiosulfate solution(5)
V : sample (mi)
0.211 5 : iodide ion equivalent to 1 ml of 10 mmol/Z sodium
thiosulfate solution (mg)
Notes (4) Decomposition of hypochlorous acid by sodium formate solution
(400glZ) shall be carried out a t p H 3 to 7. When pH becomes
2.7 o r less, iodate ion is reduced, and when pH becomes 7 or more,
the decomposition of hypochlorous acid becomes incomplete.
(5) Factor of 0.1 mol/Z sodium thiosulfate solution shall be used.
Remarks 2 In this method, iron (II, III) interferes. 0.2 mg o r over man-
ganese and 1mg or over arsenate ion interfere. Hydrogen
sulfide and a great amount of organic matter also interfere.
Removal of interference shall be as follows.
Iron and manganese Make alkaline the sample by add-
ing sodium hydroxide solution (200 g/U, and after standing
for approx. 1 h, filter. Combine the filtrate and washings,
and concentrate it by boiling on the water bath to 30 to
50 ml. If the turbidity and the precipitates exist, filter and
wash. Combine the filtrate and washings, then carry out
the operation specified in (2) Operation (a>and thereafter.
Arsenate ion Add 1 ml of iron (III) chloride solution [dis-
solve 5 g of iron (III) chloride 6 hydrate specified in JIS K
8142 in 10 ml of hydrochloric acid (l+l), and add water to
make 100 mi] per 500 ml of sample, and operate hereafter
in the same manner as in (i). However, use aqueous am-
monia (l+l) instead of sodium hydroxide solution (200 glZ).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
178
K O101 : 1998
(3) Hydrogen sulfide Add 2 t o 3 ml of zinc sulfate solution

(dissolve 10 g of zinc sulfate 7 hydrate specified in JIS K
8953 in water t o make 100 mi) per 500 ml of sample, and
mix by stirring thoroughly. Thereafter, operate in the same
manner as specified in (i).
3 In this method, also the iodate ion is determined as iodide ion.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
179
K O101 : 1998
34 Bromide ion (Br-) The iodine titrimetric method or ion chromatography ap-
ply t o determination of bromide ion.
34.1 Iodine titrimetric method Bromide ion is oxidized t o bromate ion with hy-
pochlorous acid at pH 6.5 t o 8.0. After decomposing excessive hypochlorous acid with
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
sodium formate at pH 3 to 7, potassium iodide is added, and liberated iodine is titrated
with sodium thiosulfate solution to determine the bromide ion. Because iodide ion reacts
in the same manner, determine it separately and subtract the amount of it.
Determination range: Br- 0.1 mg or more
Hydrochloric acid (l+l) Prepare by using hydrochloric acid specified in
JIS K 8180.
Hydrochloric acid (1+11) Prepare by using hydrochloric acid specified
in JIS K 8180.
Sodium hydroxide solution (40 gld) As described in 19 ( i )( g ) .
Sodium dihydrogen phosphate solution (500 g l l ) Dissolve 65 g of so-
dium dihydrogen phosphate dihydrate specified in JIS K 9009 in water t o
make 100ml.
Sodium hypochlorite solution (effective chlorine 35 gll) As described
in 33.2 (i)( c ) .
Sodium formate solution (400 g l l ) As described in 33.2 ( i )(d).
Methyl Orange solution (igll) As described in 22.1.1 (i)(d).
Starch solution (10 gll) As described in 22.1.2 (i)(i).
10 mmolll sodium thiosulfate solution As described in 28.3 (i)(e).
Take a proper amount of sample(1) (containing 0.1 to 3 mg as Br-) in a 300 ml
Erlenmeyer flask with ground stopper, add one drop of Methyl Orange solution
(ig/Z) as indicator, and add drop by drop hydrochloric acid (1+11)until the
colour of the solution t u r n s slightly red. Then add water to make
approx. 50 ml.
Add 2 ml of sodium dihydrogen phosphate solution (500gll) and 3 ml of sodium
hypochlorite solution (effective chlorine 35 g/Z), and after adjusting pH from
6.5 to 8.0 with sodium hydroxide solution (40 g/Z) o r hydrochloric acid (l+ll),
boil for approx. 10min.
Add 3 ml of sodium formate solution (400 g/Z)(2), and boil for approx. 5 min
t o decompose excessive hypochlorous acid.
After standing to cool, add 1g of potassium iodide and 6 ml of hydrochloric
acid (l+l),stopper tightly t o mix by shaking, and then allow to stand in a
dark place for approx. 5 min.
180
K O101 : 1998
Titrate the liberated iodine with 10 mmol/Z sodium thiosulfate solution, and
after the yellow colour of the solution has become pale, add 1 ml of starch
solution (10 g/Z) as indicator. Thereafter titrate until the generated blue
colour of iodine starch disappears.
For the blank test, take 50 ml of water in a 300 ml Erlenmeyer flask with
ground stopper, and carry out the operations specified in (a)to (e).
Separately, determine the concentration of iodide ion in the sample (mgI-/Z)
in accordance with 33.1 o r 33.2.
Calculate the concentration of bromide ion in the sample (mgBr-/Z) accord-
ing to the following formula:
B = (a- b) x f x -
'Ooo x 0.133 2 -C x 0.629 6
V
where, B : bromide ion (mgBr-/Z)
titration (ml)
b : 10 mmol/Z sodium thiosulfate solution required for
titration of blank test (ml)
f : factor of 10 mmol/Z sodium thiosulfate solution (3)
0.133 2 : bromide ion equivalent to 1 ml of 10 mmoVZ sodium
V : sample (ml)
C : concentration of iodide ion (mgI-/Z)
0.629 6 : coefficient in the case of converting the amount of
iodide ion to the equivalent amount of bromide ion,
Notes (1) In the case where the concentration of bromide ion is 2 mg/Z or
less, operate in the same manner as in Note ( 1 ) of 33.
(2) Decomposition of hypochlorous acid by sodium formate solution
(400 g/Z) shall be carried out at pH 3 to 7. When pH becomes
2.7 or less, bromate ion will be reduced, and when pH becomes
7 or more, decomposition of hypochlorous acid will be incomplete.
(3) Factor of 0.1 mol/Z sodium thiosulfate solution shall be used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 1 Iron (II, III) interferes with this method. 0.2 mg or more man-
ganese and 1mg o r more arsenate ion coexist, they interfere.
Hydrogen sulfide and a great amount of organic matter also
interfere. Removal of the interference shall be in accordance
with Remarks 2 of 33.2.
181
K O101 : 1998
34.2 Ion chromatography Bromide ion in the sample is determined by an ion

chromatography.
Determination range: Br- 0.5 to 40 mg/E (4)
Repeatability: 2 t o 10 % in coefficient of variation (differs according t o the ap-
paratus and measuring conditions.)
Note (4) In the case of the system combined with a suppressor, the determina-
tion range is 0.1 t o 40 mglZ.
(i) Reagents The following regents shall be used.
Water Water A2 or A3 specified in JIS K 0557.
Eluent As specified in 32.5 ( i )(b).
Reclaiming solution As described in 32.5 (i)(c).
Bromide ion standard solution (5 mgBr-/ml) Heat potassium bromide
specified in JIS K 8506 a t 105 "C for approx. 4 h, and allow to stand t o
cool in a desiccator. Take its 0.745 g, dissolve in a small amount of water,
transfer t o a 100 ml volumetric flask, and add water t o the marked line.
Bromide ion standard solution (0.5 mgBr-/ml) Take 10 ml of bromide
ion standard solution (5 mgBr-íml) into a 100 ml volumetric flask, and add
water t o the marked line. Prepare this solution at the time of use.
Anion mixed standard solution L(O.1 mgCl-, 0.5 mgNOz-, 0.5 mgBr-,
0.5 mgNO3-, 1 mgS042-)/ml] As described in 32.5 ( i )(f).
(2) Apparatus The apparatus shall be in accordance with 32.5 (2). However, the
ultraviolet absorption detector may be used.
(3) Preparatory operation Carry out the preparatory operation as specified in

32.6 (3).
Perform the operation of 32.5 (4)(a) and (b).
As for the peak corresponding to the bromide ion on the chromatogram,
read the indicated value (5).
When the sample is diluted, carry out the operations (a) and (bj as blank
test for water the same amount as the sample, and correct the indicated
value (5) obtained from the sample.
Obtain the amount of bromide ion from the working curve, and calculate
the concentration of bromide ion in the sample (mgBr-4).
Working curve Deal out step by step 0.1 to 8 ml of bromide ion standard
solution (0.5 mgBr-/ml)(6) into a 100 ml volumetric flask, add water to the
marked line, perform the operation of (a) and (b) for this solution, and read
the indicated value(5) corresponding t o each bromide ion. Separately, as a
blank test, perform the operation of (a)and (b>for water, and correct the
indicated value corresponding to each bromide ion. Thereafter, prepare the
relation curve between the amount of bromide ion (Br-) and the indicated
value(6). Prepare the working curve when the sample is measured.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
182
K O101 : 1998
Notes (5) The indicated value means peak height or peak area.
(6) In the case where anions other than bromide ion are simulta-
neously tested, anion mixed standard solution [ ( O . 1mgCl-,
0.5 mgNOn-, 0.5 mgBr-, 0.5 mgNOs-, 1mgS042-)/ml]shall be used.
Remarks 2 In the case where the concentration of bromide ion is 1mg/2,
if nitrite ion is 200 mglZ or under, it does not interfere.
3 As described in Remarks 11 of 32.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
183
K O 1 0 1 : 1998
35 Cyanide compounds The cyanide compound is a generic term for the cya-
nide, cyanocomplex, etc. in the water, and shall be classified into cyanide ion and
total cyanogen.
The cyanide compound shall be converted to cyanide ion by pretreatment, and t o
the determination the 4-pyridine carboxylate-pyrazolone absorptiometry or ion se-
lective electrode method shall apply.
Since cyanide compound is liable to vary, the test shall be carried out just after
sampling. If the test can not be performed immediately, it is preserved in accor-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
dance with 3.3, and shall be tested as soon as possible.
35.1 Pretreatment Make the sample slightly acidic, and collect the hydrogen cya-
nide generated by aeration or heating.
35.1.1 Cyanide In this pretreatment the hydrogen cyanide is generated almost

completely from the cyanide ion and cyano-complexes of zinc and cadmium of small
formation constant, and partially from the cyano-complexes of nickel, copper, etc.
However, no hydrogen cyanide is generated from the cyano-complexes of iron (II)
and iron (III).
35.1.1.1 Aeration method (hydrogen cyanide to be generated at pH 5.0) The

pH of the sample is adjusted to 5.0, and with holding at 40°C in a thermostatic
water bath, air is passed at a rate of approx. 1.2 Zlmin. Hydrogen cyanide generated
shall be collected in the sodium hydroxide solution.
(a) Acetic acid (l+l) Prepare by using acetic acid specified in JIS K 8355.
(b) Acetic acid (1+49) Prepare by using acetic acid specified in JIS K 8355.
(c) Sodium hydroxide solution (200 gll) Dissolve 20 g of sodium hydrox-
ide specified in JIS K 8576 in water to make 100 ml.
(d) Sodium hydroxide solution (20 gll) Dilute the sodium hydroxide solu-
tion (200glZ) in ( c ) by 10 times with water.
(2) Apparatus the apparatus shall be as follows.
(a) Aeration apparatus An example is shown in Fig. 35.1.
184
K O101 : 1998
- "
A: Gas washing bottle 250 ml F: Flow meter

Contain 100 ml of sodium hydroxide G: Soft polyvinyl chloride tube
solution (200 g í l ) or silicone rubber tube
B: Gas washing bottle 250ml a: Glass filter plate G2
Pack the glass wool lightly b: Capillary tube
C: Gas washing bottle 250ml (for sample)
D: Gas washing bottle with filter plate
250 ml
(for absorption of hydrogen cyanide)
E: Thermostatic water bath (40f2 "Cl
Fig. 35.1 An example of aeration apparatus
(3) Operation for aeration Carry out the operation for aeration as follows.
{a) Assemble the aeration apparatus as shown in Fig. 35.1, and add 40 ml of
water and 20 ml of sodium hydroxide solution (20 gll) for absorption of
hydrogen cyanide into the gas washing bottle with filter plate (DI.
(b) Take preliminarily 100 ml of sample into a 300 ml beaker, and add drop by
drop acetic acid ( l + l ) and acetic acid (1+49) or sodium hydroxide solution
(20glZ) by means of a pH meter until pH becomes 5.010.2t o obtain the
amount.
(c) Take 100 ml of the sample(1) ( 2 ) in a gas washing bottle (Cl, and add a proper
amount of acetic acid (l+l) and acetic acid (1+49)or sodium hydroxide solution
(20 glZ) obtained in (b), and connect the tubes as shown in Fig. 35.1.
(d) Keeping the temperature of the thermostatic water bath at (40f2) O C , aer-
ate for 1h at a rate of approx. 1.2 Elmin.
(e) After aeration, transfer the sodium hydroxide solution (absorbing solution)
in the gas washing bottle with filter plate (D) into a 100 ml volumetric flask,
wash the gas washing bottle with filter plate (D) with water, then transfer
also the washings, and add water up to the marked line.
Notes (1) The aliquot of sample shall be the optimum amount obtained from
the determination range described in the respective methods as
specified in 35.2 to 35.3.
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K O101 : 1998
(2) In the case where the sample contains oxidizing substance such
as residual chlorine and oils, reducing substance such as sulfide,
preliminarily remove them in accordance with the methods as
shown in Remarks 1 to 3.
Remarks 1 In the case where the sample contains a large amount of fats
and oils, preliminarily add acetic acid o r sodium hydroxide to
adjust pH from 6 to 7, and transfer into a separating funnel.
Add hexane or chloroform of approx. 2 vol % of the sample, mix
by gently shaking, and after separating fats and oils by standing
still, carry out the operation specified in 35.1.1.1.
2 I n the case where the sample contains oxidizing substance
such as residual chlorine, reduce by adding L(+)-ascorbicacid
(100 glZ} [dissolve 10 g of L(+)-ascorbicacid specified in JIS K
9502 in water to make 100 mi], o r sodium arsenite (100 g/Z)
(dissolve 10 g of sodium metaarsenite specified in JIS K 8046
in water to make 100ml).
3 In the case where the sample contains sulfide, add preliminar-
ily 2 ml of zinc acetate solution (100 gll) [as specified in 35.1.1.2
(1)(d)]. One milliliter of zinc acetate solution (100 g/Z) corre-
sponds to approx. 14 mg of sulfide ion.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
35.1.1.2 Method of distillation by heating (hydrogen cyanide to be gener-
ated under existence of zinc acetate at pH 5.5) Add zinc acetate to the sample,
adjust pH a t 5.5, distill by heating, and collect hydrogen cyanide to be generated in
the sodium hydroxide solution.
(a) Acetic acid (1+1) As described in 35.1.1.1 (1)(a).
(b) Acetic acid (1+49) As described in 35.1.1.1 (1)(b).
(c) Sodium hydroxide solution (20 gll) As described in 35.1.1.1 (1)(d).
(d) Zinc acetate solution (100gil) Dissolve 12 g of zinc acetate dihydrate
186
K O101 : 1998
Unit: mm
A: Distillation flask 1O00 ml (or

500 mi)
B: Connecting introduction tube
C: Ground-glass cock
D: Injection funnel
E: Kjeldahl type trap sphere
F: Liebig condenser 300mm
G: Back-flow stopper (approx. 50 mi)
H: Receiver [measuring cylinder (with
stopper) 250ml (or 100ml)l
I: Interchangeable ground joint
J: Interchangeable spherical ground
joint
K: Fixing spring
Fig. 35.2 An example of distillation apparatus
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187
K O101 : 1998
(3) Distilling operation Carry out the distilling operation as follows.

In the case where the sample is strongly alkaline, take 500 ml of sample(3)
into a 1000ml beaker. Add drop by drop acetic acid (l+l)t o make pH
approx. 7 by use of pH meter, and obtain the added amount required for
neutralization.
Add t o this solution 20ml of zinc acetate solution (lOOg/Z), and add drop
by drop again acetic acid (1+49) t o adjust pH to 5.5 by use of pH meter.
Obtain the added amount of this acetic acid (1+49)(4).
Transfer 500 ml of sample into a 1O00 ml distillation flask, and put approx. 10
pieces of boiling tips of 2 t o 3 mm in diameter(&).
Add the amount of acetic acid (l+l) obtained in (a),and connect the distil-
lation flask to the distillation apparatus as shown in Fig. 35.2.
Use a 250 ml measuring cylinder with stopper for the receiver of distilla-
tion apparatus, put 20 ml of sodium hydroxide solution (20 g/Z) into it, and
connect the receiver as shown in Fig. 35.2,
Add 20ml of zinc acetate solution (lOOg/Z) from the injection funnel, and
further add acetic acid (1+49) obtained as specified in (b).
Heat the distillation flask, and adjust the distilling rate(6) from 2 t o 3 ml/
min to distill until the amount of solution becomes approx. 230 ml(7).
Detach the condenser and back-flow stopper, wash the inner tube of the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
condenser and in- and out-side of back-flow stopper with a small amount
of water, then add the washings also in the receiver, and further add wa-
ter up to the marked line of 250 ml.
Notes (3) Obtain the optimum amount of the sample from the determina-
tion range described in respective methods in 35.2 to 35.3.
(4) Obtain the adding amount of each reagent as correctly as pos-
sible.
(5) A capillary tube one end of which is sealed may be used.
(6) Do not make the distilling rate not less than 3 ml/min, because
the recovery of hydrogen cyanide decreases.
(7) Adjust the height of the 250 ml measuring cylinder with stopper
so that the tip of the back-flow stopper be kept always a t
approx. 15 mm under the liquid surface.
Remarks 4 In the case where the sample contains a large amount of fats
and oils, carry out the same operation as in Remarks 1.
5 In the case where the sample contains oxidizing substances
such as residual chlorine or the like, carry out the same op-
eration as in Remarks 2.
6 In the case where the sample contains reducing substances,
carry out the distilling operation as i t is, and after oxidation
treatment shown in the following for the distillate, carry out
the distilling operation again to remove.
188
K O 1 0 1 : 1998
Transfer the distillate and washings in the receiver obtained

by the distilling operation as specified in 35.1.1.2 into the dis-
tillation flask again. Add 2 or 3 drops of phenolphthalein solu-
tion (5glZ) as indicator as specified in 13.2(1)(a), then
neutralize it with acetic acid (l+l), and further add about 30 ml
of nitric acid (50 mmoVZ). Then add drop by drop the potassium
permanganate solution (3 glZ), further add 1ml in excess over
the point where the permanganate shows pale red or over the
point where brown turbidity of manganese dioxide is generated,
and add water to make approx. 300 ml. Connect the distillation
flask to the distillation apparatus as shown in Fig. 35.2, use a
100 ml measuring cylinder with stopper as the receiver, pour
into it 20 ml of sodium hydroxide solution (20 g/Z), and connect
the receiver as shown in Fig. 35.2. Heat the distillation flask
t o adjust the distilling rate t o 2 t o 3 ml/min, and stop the dis-
tillation when the amount of the solution in the receiver has
become approx. 90 ml. Then, detach the condenser and back-
flow stopper, wash the inner tube of the condenser and the in-
and out-side of back-flow stopper with a small amount of water,
and after adding also the washings in the receiver, add water
up t o the marked line of 100 ml.
35.1.2 Total cyanogen (hydrogen cyanide to be generated at pH 2 or less) Add

phosphoric acid to a sample to make pH 2 or less, add disodium dihydrogen ethylene-
diaminetetraacetate, distill by heating, and collect the generated hydrogen cyanide
in sodium hydroxide solution.
Remarks 7 Cyanide ion and almost all of cyanogen in the cyano-complex are
distilled by the pretreatment. If distilled under the coexisting
condition of oxidizing substances, thiocyanic acid, 2-propenenitrile
(acrylonitrile), etc. are decomposed t o generate hydrogen cyanide.
Therefore, the oxidizing substances shall be preliminarily reduced.

(a) Phenolphthalein (5 gll) As described in 13.2 (1) (a).
(b) Sodium hydroxide solution (20 glZ) As described in 35.1.1.1 (1) (d).
(c) Ammonium amidosulfate solution (100 gil) Dissolve 10 g of ammonium
amidosulfate specified in JIS K 8588 in water t o make 100 ml.
(d) EDTA solution Dissolve 10 g of disodium dihydrogen ethylenediamine-
tetraacetate dihydrate specified in JIS K 8107 in water t o make 100 ml.
(e) Phosphoric acid As specified in JIS K 9005.

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189
K O 1 0 1 : 1998
(3) Distilling operation Carry out the distilling operation as follows.

(a) Take a suitable amount ( 3 ) of the sample in a 500 ml distillation flask, add
water t o make approx. 250 ml, and put into it approx. 10 pieces of boiling
tips of 2 t o 3 mm in diameterW. Add one drop of phenolphthalein solution
( 5 g i l ) as indicator.
(b) In the case where the solution is alkaline, add drop by drop phosphoric
acid until the red colour of the solution disappears($).
(c) Then, add 1ml of ammonium amidosulfate solution (100 g/Z)(9).
(d) Connect a distillation flask as shown in Fig. 35.2,and use a 100 ml mea-
suring cylinder with stopper for receiver, put 20ml of sodium hydroxide
solution (20 g/Z) into this receiver and connect it as shown in Fig. 35.2.
(e) Add 10 ml of phosphoric acid to the distillation flask from the injection funnel,
then add 10 ml of EDTA solution, wash the funnel with a small amount of
water, and add the washings to the distillation flask.
(0 After standing for several min, heat the distillation flask to distill at a
distilling rate(6) of 2 t o 3 ml/min until the amount of solution in the re-
ceiver becomes approx. 90 ml (7).
( g ) Detach the condenser and the back-flow stopper, wash the inner tube of
the condenser and in- and out-side of the back-flow stopper with a small
amount of water, and after adding the washings t o the receiver, add water
up to the marked line of 100 ml.
Notes (8) It is sufficient to add phosphoric acid t o make weak acidic solu-
tion.
The addition of ammonium amidosulfate solution (100 g/Z) is for the
(9)
purpose of removing the interference of nitrite ion in the sample.
In the case where this solution is not added, if nitrite ion exists, it
reacts to EDTA at the distillation t o generate hydrogen cyanide.
One milliliter of ammonium amidosulfate solution (100 g/Z) corre-
sponds to approx. 40 mg of nitrite ion (NOZ-). In the case where
40 mg or more of nitrite ion coexists, the adding amount shall be
increased in proportion t o the amount of nitrite ion.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
In the some special samples, substances other than nitrite ion

react to EDTA to generate hydrogen cyanide, and cannot be re-
moved the interference even though adding ammonium amido-
sulfate solution (100 g/Z).
Also there are organics which react similarly to those other than
EDTA.
Remarks 8 Removal of fats and oils shall be operated in the same man-
ner as in Remarks 1.
9 In the case where oxidizing substance such as residual chlo-
rine or the like is contained, operate in the same manner as
in Remarks 2.
10 In the case where reducing substance such as sulfide is con-
tained in the sample, carry out the operation of Remarks 6
on the distillate solution obtained by carrying out the distill-
ing operation of total cyanogen.
190
K O101 : 1998
35.2 4-Pyridinecarboxylic acid-pyrazolone absorptiometry A portion of cya-

nide ion solution obtained by pretreatment is taken t o be neutralized with acetic
acid. Then, chloramine T solution is added to this solution to make cyanogen chlo-
ride, and mixed solution of 4-pyridinecarboxylic acid and pyrazolone solution is added.
The absorbance of blue colour generated shall be measured to determine the cya-
nide ion.
Determination range: CN- 0.5 t o 9 p g
Acetic acid (1+8) Prepare by using acetic acid specified in JIS K 8355.
Phenolphthalein solution (5 gll) As described in 13.2 (1)(a).
Phosphate buffer solution (pH 7.2) Dissolve 17.8 g of disodium hydro-
gen phosphate specified in JIS K 9020 in approx. 300 ml of water, and add
potassium dihydrogen phosphate (200 g/Z) till pH 7.2, then dilute with water
to 500ml.
Chloramine T solution (10 gll) Dissolve 0.62 g of sodium p-toluene
sulfonchloroamide 3-hydrate (chloramine T) specified in JIS K 8318 in water
to make 50 ml. Prepare this solution a t the time of use.
4-Pyridinecarboxylic acid-pyrazolone solution Dissolve 0.3 g of 3-me-
thyl-1-phenyl-5-pyrazolone specified in JIS K 9548 in 20 ml of N,N-dimethyl
formamide specified in JIS I( 8500. Separately dissolve 1.5 g of 4-pyridine-
carboxylic acid in approx. 20 ml of sodium hydroxide solution (40 g/Z), and add
drop by drop hydrochloric acid (1+10)to make pH approx. 7 (10). Combine both
solutions, add water to 100 ml, and preserve the solution in a dark place a t
10 "C or under, and don't use the solution for which 20 days o r more have
elapsed.
0.1 mol/Z silver nitrate solution Dissolve 17 g of silver nitrate speci-
fied in JIS K 8550 in water to make 11. Transfer t o a coloured glass bottle
and preserved.
Standardization Heat sodium chloride, reference material for volumet-
ric analysis, specified in JIS K 8005 a t 600 "C for approx. 1h, and allow
to cool in a desiccator. Then take its 1.169 g for NaC1 100 %, dissolve in a
small amount of water, transfer to a 200 ml volumetric flask, and add water
up t o the marked line. Take its 20 ml, and add water t o make the amount
of solution approx. 50 ml. Add 5 ml of dextrin solution [as descried in
32.3 (1)(d)] and 3 t o 4 drops of sodium fluorescein solution (2 g/Z) [as de-
scribed in 32.3 (1)(c)]as indicator. Titrate this solution with 0.1 mol/2 sil-
ver nitrate solution and take the point when the yellow green fluorescence
disappears and slight red colour appears as the end point. Calculate the
factor (f)according t o the following formula:
b 20 1
xax-x-
100 200 xx0.005844
where, a : mass of sodium chloride (g)
b : content of sodium chloride (%)
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191
K O101 : 1998
x : 0.1 mol/Z silver nitrate solution required for titra-

tion (mi)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
0.005 844 : sodium chloride equivalent to 1ml of 0.1 mol4 silver

nitrate solution (g)
Cyanide ion standard solution (i mgCN-/ml) Dissolve 0.63 g of potas-
sium cyanide specified in JIS K 8443 in a small amount of water, add 2.5 ml
of sodium hydroxide solution (20 g/U, and add water to make 250 ml. Pre-
pare this solution at the time of use. Obtain the concentration according
to the following method:
Take 100ml of this solution, add 0.5ml of acetone solution of p -
dimethylamino-benzylidenerhodanine(0.2 g/Z) [dissolve 20 mg of p-dimethyl-
amino-benzylidenerhodanine specified in JIS K 8495 [5-(4-dimethylamino
benzylidene)-2-thioxo-4-thiazolidinonel in 100 ml of acetone specified in JIS
K 8034.1 as indicator, and titrate with 0.1 moll silver nitrate solution. Take
the point when the colour of the solution turns from yellow to red as the
end point. Calculate the concentration of cyanide ion standard solution
(mgCN-/ml) according t o the following formula:
1
c = a x f x 5.204 x- 100
where, C : concentration of cyanide ion standard solution
(mgCN-/ml)
a : 0.1 molli silver nitrate solution required for titra-
tion (ml)
f : factor of 0.1 mol/Z silver nitrate solution
5.204 : cyanide ion equivalent t o 1ml of 0.1 mol/Z silver
nitrate solution (mg)
Cyanide ion standard solution ( i pgCN-/ml) Put 10 ml of cyanide ion
standard solution (i mgCN-/ml) into a 1 O00 ml volumetric flask; and after
adding 100 ml of sodium hydroxide solution (20 g/Z), add water up to the
marked line. Take 10 ml of this solution in a 100 ml volumetric flask, and
add water up t o the marked line. Prepare this solution at the time of use.
Calculate the concentration of the solution from the concentration of the
cyanide ion standard solution (i mgCN-/ml).
Note (10) The solution obtained by dissolving 1.8g of sodium 4-pyridine
carboxylate in approx. 50 ml water may be used instead of this
solution.
(a) Take a proper amount (containing 0.5 to 9 pg as CN-) from the cyanide ion
solution obtained by the pretreatment specified in 35.1 into a 50 ml volu-
metric flask.
192
K O101 : 1998
Add one drop of phenolphthalein solution (5 gil) as indicator, and add drop
by drop acetic acid (1+8)while mixing by gently shaking, neutralize and
add 10 ml of phosphate buffer solution (pH 7.2)(11).
Add 0.5 ml of chloramine T solution (10 giz), and allow t o stand for about
5 min in a water bath at 25 OC.
Add 10 ml of 4-pyridine carboxylic acid-pyrazolone solution, further add
water up t o the marked line, stopper tightly, mix by shaking gently, and
allow to stand in a water bath at approx. 25 "C for approx. 30 min.
Transfer a part of the solution into an absorption cell, and measure the
absorbance a t a wavelength near 638 nm.
For the blank test, take 10 ml of water into a 50 ml volumetric flask, add
10 ml of phosphate buffer solution (pH 7.2). Thereafter, measure the ab-
sorbance by performing the operation of ( c ) t o (e), and correct the absor-
bance obtained on the sample.
Obtain the amount of cyanide ion from the working curve, and calculate
the concentration of cyanide ion in the sample (mgCN-íl).
Working curve Take step by step 0.5 t o 9 m l of cyanide ion standard
solution (1pgCN-/ml) into a 50 ml volumetric flask, and make the quantity
of solution approx. 10 ml with water. Carry out the operation specified in
(b)to (f) to prepare the relation curve between the amount of cyanide ion
(CN-) and the absorbance.
Note (11) pH a t the time of colour development shall be in the range of 7
t o 8.
35.3 Ion selective electrode method For the cyanide ion solution (pH 12 t o 13)
t o be obtained by the pretreatment, measure the potential by using a cyanide ion
selective electrode as an indication electrode, and determine the cyanide ion.
Determination range: CN- 0.1 to 100 mgll

Sodium hydroxide solution (0.1 mol/Z) Dissolve 4 g of sodium hydrox-
ide specified in JIS K 8576 in water t o make 1E.
Cyanide ion standad solution (100 mgCN-ll) Transfer 20 ml of cyanide
ion standard solution specified in 35.2 (1) (g) (1mgCN-/ml) into a 200 ml
volumetric flask, and add sodium hydroxide solution (0.1 molíl) up t o the
marked line(12). Prepare this solution at the time of use. Calculate the
concentration of the solution from the concentration of the cyanide ion stan-
dard solution (1 mgCN-/mU.
Cyanide ion standard solution (10 mgCN-lZ) Take 20 ml of cyanide ion
standard solution (100 mgCN-ll) into a 200 ml volumetric flask, and add
sodium hydroxide solution (0.1 mol/l) t o the marked line(l2). Prepare at
the time of use. Calculate the concentration of this solution from the con-
centration of the cyanide ion standard solution (100 mgCN-/l).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
193
K O101 : 1998
Cyanide ion standard solution (1 mgCN-ll) Take 20 ml of cyanide ion

standard solution (10 mgCN-ll) into a 200 ml volumetric flask, and add sodium
hydroxide solution (0.1 mol/Z) t o the marked line(12). Prepare a t the time
of use. Calculate the concentration of this solution from the concentration
of the cyanide ion standard solution (10 mgCN-ll).
Cyanide ion standard solution (0.1 mgCN-ll) Take 20 ml of cyanide
ion standard solution (1mgCN-lZ) into a 200 ml volumetric flask, and add
sodium hydroxide solution (0.1 mol/l) t o the marked line(l2). Prepare at
the time of use. Calculate the concentration of this solution from the con-
centration of the cyanide ion standard solution (1mgCN-ll).
Note (12) pH of each cyanide ion standard solution becomes approx. 13.

(a) Potentiometer As described in 31.2 (2) (a).
(b) Indication electrode Cyanide ion selective electrode
(c) Reference electrode As described in 31.2 (2)(c).
(d) Magnetic stirrer As described in 31.2 (2) (d).

Take 100 ml of cyanide ion standard solution (0.1 mgCN-lE) into a 200 ml
beaker, immerse the indication electrode (13) (14) and the reference elec-
trode(15) (IC), and mix by intensively stirring with a magnetic stirrer(l7) t o
an extent that bubbles do not touch the electrodes(l8).
Measure the temperature of the solution, and measure the potential with
a potentiometer(l9).
Take 100 ml of cyanide ion standard solution (1mgCN-lZ), 100 ml of cya-
nide ion standard solution (10 mgCN-lZ), and 100 ml of cyanide ion stan-
dard solution (100 mgCN-lZ) respectively into a 200 ml beaker, and hereafter
carry out the operation of (a).
Adjust the temperature of each cyanide ion standard solution (1to 100 mgCN-
/ I ) within the temperature of the solution of (b)+1OC, and measure the po-
tential of the cyanide ion standard solution (1 t o 100 mgCN-íZ).
Take the concentration of cyanide ion on the logarithmic axis of semiloga-
rithmic section paper and the potential on its linear axis, and prepare the
relation curve between the concentration of cyanide ion (mgCN-íl) and the
potential (20).
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194
K O101 : 1998
Notes (13) The cyanide ion selective electrode shall be immersed in the cya-
nide ion standard solution (0.1 mgCN-ll) at the time of use, and
after their indicating value has been stabilized, the potential shall
be measured. The response time of the cyanide ion selective
electrode is approx. 1 min when the concentration of cyanide ion
is 0.1 mgll and approx. 30 s when it is 1mgll o r more at the so-
lution temperature of 10°C to 30°C.
(19) Because silver iodide is often used for the cyanide ion selective
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
electrode, direct sunlight causes large variation of potential to
give a positive error, but the effect of indoor lighting is small.
(20) The potential difference between the potential of cyanide ion
standard solution (0.1 mgCN-ll) and that (10 mgCN-ll) is within
the range of 110 to 120 mV (25 OC), and the working curve be-
tween the concentrations of cyanide ion from 0.1 to 100 mgll
becomes a straight line.
(a) Transfer 100 ml of cyanide ion solution obtained by the pretreatment specified
in 35.1 into a 200 ml beaker, and adjust the temperature of the solution to
within +1"C of the solution temperature specified in (3)(b) and carry out
the operation of (3)(a).
(b) Carry out the same operation as specified in (3) (b) and obtain the concen-
tration of cyanide ion from the working curve to calculate the concentra-
tion of cyanide ion in the sample (mgCN-A).
Remarks 11 In the case of ion densitometer, use the cyanide ion standard
solution (0.1 mgCN--/Z)and that (10 mgCN-lZ), carry out the
operation specified in (3) (a)and (b),and adjust the indicat-
ing value of the ion densitometer to be 0.1 mgCN-ll and
10 mgCN-ll respectively. Furthermore, confirm the indica-
tions of the ion densitometer by use of other cyanide ion stan-
dard solution (1mgCN-ll) and that (100 mgCN-ll).
12 Interferences due to sulfide ion and mercaptoacetic acid
(thioglycollic acid) shall be removed by pretreatment. When
the sulfite ion is within 103 times the cyanide ion, it does
not interfere, and therefore the oxidation treatment described
in Remarks 6 may be omitted. Formaldehyde gives a nega-
tive interference.
The allowable limits of main coexisting substances are
shown by the maximum ratio as follows:
195
K O101 : 1998
Cl-, F-, Nos-, CrO42-, K', Na' : lo4

Br-, SCN-, HCOS-, COS2-, S0s2-, Sod2-, Pod3-: lo3
S20s2-, Ag': 10
I- : 0.1
13 Potentiometric titration method of ion selective electrode
Transfer 100 ml of cyanide ion solution obtained by the pre-
treatment specified in 35.1 into a beaker. While measuring
the potential by use of the indication electrode (cyanide ion
selective electrode o r silver ion selective electrode) in accor-
dance with the operation in (3)as appropriate, titrate with
1 to 100mmol/Z silver nitrate solution. Draw the titration
curve t o obtain the titration end point, and calculate the con-
centration of cyanide ion. 1ml of 100 mmollE silver nitrate
solution is equivalent t o 5.204 mg of cyanide ion.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
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K 0101 : 1998
36 Ammonium ion (NH4+) To determination of ammonium ion, Indophenol Blue

absorptiometry, acid-base titrimetric method, ion selective electrode method, o r ion
chromatography shall apply.
After separating interfering substances by aggregate precipitation treatment or
distillation treatment of a sample, the sample is determined by the Indophenol Blue
absorptiometry or the ion selective electrode method. If the concentration of ammo-
nium ion is high, the acid-base titrimetric method or the ion selective electrode method
shall apply.
When ion chromatography is applied, the test shall be carried out immediately
after sampling omitting the pretreatment of sample and 3.3 Preservation treatment
of sample.
Since ammonium ion is liable to vary, the test shall be carried out just after Sam-
pling. In the case where the test can not be immediately carried out, the sample
shall be preserved in accordance with 3.3, and shall be tested as soon as possible.
36.1 Pretreatment Aggregate precipitation method or steam distillation method

shall apply t o the pretreatment.
36.1.1 Aggregate precipitation method The turbidity, colour and metal elements
are removed by the aggregate precipitation with zinc sulfate or sodium carbonate
and sodium hydroxide.
(a) Water Water A3 specified in JIS K 0557. Use this water for the prepa-
ration of the reagents and the operation.
(b) Sodium hydroxide-sodium carbonate solution Dissolve 30 g of sodium
hydroxide specified in JIS K 8576 in approx. 60 ml of water. Separately
dissolve 25 g of sodium carbonate specified in JIS K 8625 in approx. 100 ml
of water, combine both solutions and dilute with water to 200 ml.
(c) Sodium carbonate solution (250glE) Dissolve 25 g of sodium carbon-
ate specified in JIS K 8625 in water t o make 100 ml.
(d) Sodium hydroxide solution (250g l l ) Dissolve 25 g of sodium hydrox-
ide specified in JIS K 8576 in water t o make 100 ml.
(e) Zinc sulfate solution Dissolve l o g of zinc sulfate 7 hydrate specified
in JIS K 8953 in water to make 100ml.

(a) Glassware Wash with water thoroughly prior t o use.
(3.1) Aggregate precipitation by zinc sulfate This method applies to the sample
having comparatively small amount of calcium ion and magnesium ion. Where
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the turbidity and colour remain even when treated with this method, treat ac-
cording to 36.1.2.
(a) Add 1ml of zinc sulfate solution to 100 ml of the sample, mix by stirring
thoroughly, then add sodium hydroxide-sodium carbonate solution (usually
197
K O101 : 1998
0.3 t o 0.5 ml) t o adjust pH t o approx. 10.5, and again mix by stirring thor-
oughly to allow t o stand for a while.
(b) Centrifugally separate or filter (1) the supernatant solution (discard
approx. 25 ml of the initial filtrate) t o make the solution clear, transpar-
ent, transfer to the Erlenmeyer flask with ground stopper and stopper tightly.
(3.2) Aggregate precipitation by sodium carbonate and sodium hydroxide
This method applies t o the sample containing a comparatively large amount of
calcium and magnesium. Where the turbidity and colour remain even when
treated with this method, pretreat according t o 36.1.2.
(a) Add 1 ml of sodium carbonate solution (250 gll) and 0.7 ml of sodium hy-
droxide solution (250 g/I)(2) to the sample [where it is acidic, neutralize it
by using sodium hydroxide solution (250 gll) t o pH approx. 71 and mix by
stirring thoroughly.
(b) Transfer this solution into a 200 ml measuring cylinder with stopper, and
add water up t o the marked line of 200 ml.
(c) Stopper to mix by shaking thoroughly, then allow t o stand in a cool and
dark place for 2 h or more, decant or filter(1) the supernatant solution (discard
approx. 25 ml of the initial filtrate) to make the solution clear, transpar-
ent and stopper tightly.
Notes (1) Use the filter paper of class 5A and after washing with water
thoroughly prior to use.
(2) This adding amount corresponds t o 90 mg of calcium, and 40 mg
of magnesium.
36.1.2 Distillation method Add magnesium oxide t o the sample t o make weak
alkalinity, distill, and collect by absorbing the distilled ammonia in sulfuric acid
(25 mmol/Z).
(a) Water Water of 36.1.1 (1)(a).
(b) Sulfuric acid (25mmol/Z) Add approx. 1.4 ml of sulfuric acid specified
in JIS K 8951 in a beaker containing 100 ml of water, mix by stirring thor-
oughly and dilute with water to I I .
(c) Sulfuric acid (1+35) Prepare by using sulfuric acid specified in JIS K
8951.
(d) Sodium hydroxide solution (40 gil) As described in 19 (1) (g).
(e) Magnesium oxide Prior t o use, heat magnesium oxide specified in JIS
K 8432 a t 600 "C for approx. 30 min and allow to cool in a desiccator.
(a) Distillation apparatus An example is shown in Fig. 36.1. Wash thor-
oughly the glassware with water of (1)(a) prior to use.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
198
K O101 : 1998
I t
I
i
A: distillation flask
500 ml
B: branched connecting \-
tube
C: ground-glass cock
D: injection funnel
E: trap sphere
(Kjeldahl type)
F: Liebig condenser 300 mm
G: back-flow stopper (approx. 50 ml)
H: receiver (measuring cylinder with stopper 200 ml)
I: interchangeable ground joint
J: interchangeable spherical surface ground joint
K: fixing spring
Fig. 36.1 An example of distillation apparatus
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Take a proper amount of sarnple(31, and adjust pH to approx. 7 with so-
dium hydroxide solution (40gll) or sulfuric acid (1+35)when the sample is
not neutral.
(b) Transfer this solution into a distillation flask, and add 0.25g of magne-
sium oxide, several pieces of boiling tips (grain diameter 2 to 3 mm) and
water to make the amount of solution approx. 350 ml.
199
K O101 : 1998
(c) Assemble the distillation apparatus as shown in Fig. 36.1, and put 50 ml
of sulfuric acid (25 mmol/Z) into a 200 ml measuring cylinder with stopper
of the receiver(4).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(d) Heat the distillation flask, and carry out the distillation at a distillation
rate of 5 t o 7 ml/min(5).
(e) When the amount of distillate has reached about 140 ml, stop the distilla-
tion.
(0 Then detach the condenser and back-flow stopper, and wash the inside tube
of condenser and in- and out-side of back-flow stopper with a small amount
of water. Put the washings into a 200 ml measuring cylinder with stopper
of the receiver(6), and add water up t o the marked line of 200 ml.
Notes (3) The sample shall be so taken as to contain 40 pg or over as NH4+
for determination by Indophenol Blue absorptiometry, 0.3 to 40 mg
as NH4+ for acid-base titrimetric method, and 40pg or over as
NH4+ for ion selective electrode method.
(4) In the case where distillate solution is used for acid-base titri-
metric method, use a 500ml Erlenmeyer flask as the receiver,
add accurately 50 ml of sulfuric acid (25 mmol/Z) thereto, and add
5 t o 7 drops of the mixed solution of Methyl Red-Bromocresol
Green as indicator [as described in 13.1 (1) (a)].
Keep the tip of the tube of the condenser always approx. 15 mm
under the solution surface.
(9 In the case where the distillate solution is used for acid-base
titrimetric method, join washings from the inside tube of the
condenser and the in- and out-side of the back-flow stopper into
a 500 ml Erlenmeyer flask, and use all quantity for titration.
Remarks 1 Steam distillation method may be used as the distillation
method. In that case, assemble the apparatus so as t o feed
steam t o the distillation flask as given in Fig. 36.1, and heat
the distillation flask. When boiling starts, send steam t o the
distillation flask, distill a t a distillation rate of 3 t o 5 ml/min
and when approx. 140 ml is distilled, stop the distillation.
36.2 Indophenol Blue absorptiometry Ammonium ion shall be determined by

measuring the absorbance of Indophenol Blue generated by the reaction of ammo-
nium ion with phenol in the coexistence of hypochlorite ion.
Determination range: NH4+ 5 to 1OOpg
Repeatability: 2' to 10 % in coefficient of variation
(b) EDTA solution Dissolve 5 g of disodium dihydrogen ethylenediamine
tetraacetate dihydrate specified in JIS K 8107 in water t o make 100 ml.
(c) Sodium hydroxide solution (200 g/Z) As described in 35.1.1.1 (1) (c). Pre-
pare this solution a t the time of use.
200
K O101 : 1998
Sodium phenoxide solution Take 55 ml of sodium hydroxide solution

(200 g l l ) of (cl into a beaker, and dissolve by adding little by little 25 g of
phenol specified in JIS K 8798 while cooling in cold water. After standing
t o cool, add 6 ml of acetone specified in JIS K 8034 and add water to make
200 ml. Preserve in a dark place at 10 "C or under, and do not use that for
which 5 days or a longer time has elapsed.
Sodium hypochlorite solution (effective chlorine 10 gll) Determine the
concentration of effective chlorine in the sodium hypochlorite solution (effec-
tive chlorine 7 to 12 %)(7), and dilute with water to obtain the solution of
approx. 10 glE effective chlorine. Prepare this solution at the time of use,
Ammonium ion standard solution (i mg"4+/ml) Allow ammonium chlo-
ride specified in JIS K 8116 t o stand in a desiccator [containing magne-
sium perchlorate (for drying) specified in JIS K 82281 for 16 h or longer,
take its 2.97 g, dissolve in water, transfer t o a 1O00 ml volumetric flask,
and add water t o the marked line. Otherwise, use ammonium ion "4' 1O00
of reference material-standard solution specified in JIS K 0034.
Ammonium ion standard solution (10 pgNHk+/ml) Take 10 ml of ammo-
nium ion standard solution (imgNHd+/rnl)in a 1O00 ml volumetric flask, and
add water up to the marked line. Prepare this solution at the time of use.
Note (7) As described in Note (3) of 33.
(a) Glassware Wash thoroughly with water of ( i )(a) prior t o use.
Take a proper amount of sample pretreated as specified in 36.1 (contain-
ing 5 t o 100 pg as "4)' in a 50 ml volumetric flask(8), and add water t o
make approx. 25 ml.
Add 1ml of EDTA solution(9) and 10 ml of sodium phenoxide solution and
mix by shaking.
Add 5 ml of sodium hypochlorite solution (effective chlorine 10 g/Z), then
add water up to the marked line, and stopper to mix by shaking.
Keeping the liquid temperature a t 20 t o 25 OC, allow t o stand for approx.
30 min(10).
Transfer a portion of this solution into an absorption cell, and measure the
absorbance at a wavelength near 630 nm.
For the blank test, take 25 ml of water, carry out the operation specified
in (b)t o (e) t o obtain the absorbance, and correct the absorbance obtained
on the sample.
Obtain the amount of ammonium ion from the working curve and calcu-
late the concentration of ammonium ion in the sample (mgNH4+/Z).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
201
K O 1 0 1 : 1998
Working curve Take step by step 0.5 t o 10ml of ammonium ion stan-
dard solution (10ygNH4+/ml)into a 50ml volumetric flask, add water to
make 25m1, carry out the operation specified in (b)t o (f) t o measure the
absorbance, and prepare the relation curve between the amount of ammo-
nium ion (NH4+)and the absorbance.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Notes (8) When the operation described in 36.1.1 (3.2)(b) was carried out,
neutralize by using hydrochloric acid (l+l)(until pH approx. 7).
The strength of colour development becomes the maximum at
pH 11.5 to 12.5.
(9) Because the colour development of Indophenol Blue becomes
slightly weak when EDTA solution is added, add the same amount
of EDTA solution in preparing the working curve.
(10) When the temperature of the solution is 20 to 25 OC,the colouring
reaches the maximum in approx. 30 min, and is stable for
approx. 30 min thereafter.
Remarks 2 In the case where a trace amount of ammonium ion is deter-
mined, 1ml of disodium pentacyanonitrosylferrate (III) solu-
tion [O. 15 g of sodium pentacyanonitrosylferrate (III) dihydrate
specified in JIS K 8722 is dissolved in water to make 100 mi]
may be added following 10ml of sodium phenoxide solution
by the operation of (3)(b).
The determination range in this case becomes 2.5 to 50 pg
as NH4+. The working curve shall be prepared by the same
operation.
3 In the case where ammonium ion is expressed by ammonium
nitrogen, the following converting formula shall be used.
Ammonium nitrogen (mgNH,+-N/Z)
= ammonium ion (mgNH4+/Z)x 0.776 6
4 Iron (II) and copper (II) up t o 0.15 mg/l each do not interfere
with this method. If these contents are not more than 1mg/Z
respectively, the interference can be removed by adding EDTA
solution.
Aliphatic amines do not interfere, but some aromatic amines
develop coloured substance by the oxidation with hypochlorite
to interfere. Further, the substance such as p-amiophenol
generates Indophenol Blue by the reacting with phenol in al-
kaline solution t o interfere. p-Hydroquinone does not inter-
fere. Hydroxylamine interferes, but the interference can be
removed by the quantitative oxidation by adding hydrogen
peroxide specified in JIS K 8230.
36.3 Acid-basic titrimetric method After absorbing ammonia generated by pre-

treatment (distillation) into a definite amount of sulfuric acid (25 mmol/Z), the re-
sidual sulfuric acid is titrated with 50 mmol/Z sodium hydroxide solution to determine
ammonium ion.
Determination range: "4' 0.3 to 40 mg
202
K O101 :1998

(b) Sulfuric acid (25mmol/Z) As described in 36.1.2 (i)(b).
(c) Methyl Red-Bromocresol Green mixture As described in 13.1 (1)(a).
(d) 50 mmol/Z Sodium hydroxide solution Take 100 ml of 0.1 mol/Z sodium
hydroxide solution of 14.1 (1)(b) in a 200 ml volumetric flask, and add car-
bonic acid-free water of 2 (12)(b) t o the marked line. Prepare when used.
(a) Use total amount of the distillate solution obtained by the pretreatment of
36.1.2 (31,and titrate with 50 mmol/Z sodium hydroxide solution until the
colour of the solution turns t o gray purple (pH 4.8).
(b) Separately accurately take 50 ml of sulfuric acid (25 mmol/Z) into a 500 ml
Erlenmeyer flask, add 5 t o 7 drops of the mixed solution of Methyl Red-
Bromocresol Green, titrate with 50 mmol/Z sodium hydroxide solution until
the colour of the solution turns to gray purple (pH 4.8), and obtain the number
of ml of 50 mmol/Z sodium hydroxide solution equivalent of 50 ml of sulfu-
ric acid (25 mmol/Z).
(c) Calculate the concentration of ammonium ion (mgNH4+/Z)in the sample ac-
cording to the following formula.
A = ( b - a)x f x V
E x 0.902
where, A : ammonium ion (mgNHd+/l)

b : 50 mmol/Z sodium hydroxide solution
equivalent to 50 ml of sulfuric acid
(25 mmol/Z) (ml)
a : 50 mmol/Z sodium hydroxide solution re-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
quired for titration (ml)

f : factor of 50 mmol/Z sodium hydroxide so-
lution (use the factor of 0.1 mol/Z sodium
hydroxide solution)
V : sample (ml)
0.902 : ammonium ion equivalent of 1ml of
50 mmol/Z sodium hydroxide solution (mg)
Remarks 5 Boric acid solution (saturated) may be used instead of sulfu-
ric acid (25mmol/Z) of 36.1.2 (3)(e). In that case, the opera-
tion shall be carried out as follows.
Add 50 ml of boric acid solution (saturated) (add 1 Z of wa-
ter to 50 g of boric acid specified in JIS K 8863,mix by shak-
ing, and use the supernatant solution.) to a 500 ml Erlenmeyer
flask, add 5 t o 7 drops of the mixed solution of Methyl Red-
Bromocresol Green as indicator, and carry out the operation
of 36.1.2 (3)(d) and (e).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
203
K O101 : 1998
Then, titrate with 25 mmol/Z sulfuric acid [take 100 ml of

50mmol/2 sulfuric acid of 13.1 (l)(b)into a 200ml volumet-
ric flask, and add water t o the marked line] until the colour
of the solution turns to gray purple (pH 4.8). Separately take
50 ml of boric acid solution (saturated) into a 500 ml Erlenm-
eyer flask as the blank test. Add 150 ml of water, add 5 to 7
drops of the mixed solution of Methyl Red-Bromocresol Green
as indicator, hereafter titrate in the same way as for the sample,
and calculate the concentration of ammonium ion in the sample
(mgNH4+/Z)according to the following formula.
A = (a- b) x f x -Oo0 x 0.902

V
where, A : ammonium ion (mgNH4+/Z)
a : 25 mmol/Z sulfuric acid required for titra-
tion (ml)
b : 25 mmol/Z sulfuric acid required for blank
test (mi)
f : factor of 25 mmol/Z sulfuric acid (use the
factor of 50 mmol/Z sulfuric acid.)
V : sample (mi)
0.902 : ammonium ion equivalent of 1 ml of
25 mmol/Z sulfuric acid (mg)
36.4 Ion selective electrode method Sodium hydroxide solution is added to the
pretreated sample t o make pH 11 to 13 t o change ammonium ion t o ammonia, and
the potential is measured by use of the indication electrode (ammonia electrode) to
determine ammonium ion,
Determination range: NH4+ 0.1 to 100mg/Z
( i ) Reagent The following reagents shall be used.
W a t e r A3 specified in JIS K 0557.
Sodium hydroxide solution (100 g/Z) As described in 22.2.1 (i)(b).
Ammonium ion standard solution (100 mgNH4+/Z) Take 20 ml of am-
monium ion standard solution (imgNH4+/ml)specified in 36.2 (i)(f) in a
200 ml volumetric flask, and add water up t o the marked line.
Ammonium ion standard solution (10mgNH4'lZ) Take 20 ml of am-
monium ion standard solution (100 mgNHd+/Z)into a 200 ml volumetric flask,
Ammonium ion standard solution (imgNH4+/2) Take 20 ml of ammo-
nium ion standard solution (10 mgNH4+/Z)i n t o a 200 ml volumetric flask,
and add water t o the marked line. Prepare a t the time of use.
Ammonium ion standard solution (0.1 mgNH4+/2) Take 20 ml of am-
monium ion standard solution (imgNH4+/Z)into a 200 ml volumetric flask,
and add water to the marked line. Prepare a t the time of use.
204
K O101 : 1998

(a) Potentiometer As described in 31.2 (2)(a).
(b) Indication electrode Ammonia electrode
(c) Magnetic stirrer As described in 31.2 (2)(d).

Transfer 100 ml of ammonium ion standard solution (0.1mgNHd+/Z)into a
200ml Erlenmeyer flask(ll), and. add 1 m l of sodium hydroxide solution
(100 g/Z)(12).
Immerse the indication electrode (I3) (14) into this solution, and stir so strongly
that no bubble contacts the electrode by a magnetic stirrer(l5) (16).
Measure the liquid temperature and measure the potential with the po-
tentiometer (17).
Respectively take 100 ml of ammonium ion standard solution (imgNH4+/Z),
100 ml of ammonium ion standard solution (10 mgNH4+/Z),and 100 ml of
ammonium ion standard solution (100 mgNH,+/Z) into a 200 ml Erlenmeyer
flask(li), and add 1 ml of sodium hydroxide solution (100 g/Z)(12), Adjust the
temperature of the solution to within +1"C of the solution temperature of (c),
carry out the operation of (b) and (c), and measure the potential of each
ammonium ion standard solution (i to 100 mgNH$/Z) (18).
Take the concentration of ammonium ion (mgNH,+/Z)on the logarithmic axis,
and the potential(lg), on the linear axis of semilogarithmic section paper,
and prepare the relation curve between the concentration of ammonium
ion and the potentialPo).
Ammonia is easily volatilized, and therefore the narrow-mouthed
container is used as far as possible. It is preferable t o use such
a container that can be closed tightly, as required.
pH becomes approx. 12. At pH 11 o r over the ammonium ion
changes to ammonia. Because the ammonia is easily volatil-
ized, sodium hydroxide solution (100 gil) shall be added just before
the potential measurement.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
If the indication electrode is membrane electrode, care shall be

taken t o the fact that the membrane of the electrode, if the
membrane is pressed strongly to the glass film surface of inner
glass electrode, is damaged.
Further, if the glass film surface is separated in excess from
the membrane, the response time becomes long.
If the membrane of the electrode is contaminated, the poten-
tial becomes unstable and the response time, longer.
As described in Note (14) of 31.
Too strong stirring causes the covering of membrane with foams
to generate errors, and therefore cares shall be taken thereto.
205
K O101 : 1998
(17) The response time of ammonia electrode is 3 t o 5 min for am-

monium ion standard solution (0.1 mgNH4"lZ) and 2 t o 3 min for
ammonium ion standard solution (1mgNH4+lZor more) in the
case of 10 to 30 "C in the solution temperature.
(18) The measurement of potential shall be carried out in the order
of lower t o higher concentrations. If it transits from higher con-
centration to lower concentration, the response time becomes
slow. Therefore wash the electrode with water to remove am-
monia, immerse in the ammonium ion standard solution
(0.1 mgNH4'll) added with 1ml of sodium hydroxide solution
(100 glZ) specified in (3)(a),and measure after the indicating
value has become stable.
(19) The difference of potential between the ammonium ion standard
solution (0.1 mgNHd+lZ)and that (10 mgNHa+lZ)comes t o be in
the range of 110 to 120 mV (25 OC).
(20) The working curve is linear from the ammonium ion concen-
tration near 0.1 mgll to 100 mgll.
(a) Take a suitable amount (containing 0.02 to 20 mg as NH4+)of the sample
processed by the pretreatment of 36.1, and add water t o make approx. 100 ml.
Thereafter, in the case where the pretreatment is carried out in accordance
with 36.1.2, drip sodium hydroxide solution (100 g/O, adjust pH a t approx. 8,
transfer into a 200 ml volumetric flask, and add water to the marked line.
Take 100 ml of this solution into a 200 ml Erlenmeyer flask, and add 1ml
of sodium hydroxide solution (100 glO(12).
(b) Adjust the solution temperature t o within -tl O C of the solution tempera-
ture of (3)( c ) .
(c) Carry out the operation specified in (3)(b) and ( c ) to obtain the concentra-
tion of ammonium ion (mgNH4+/Z) from the working curve and calculate the
concentration of ammonium ion in the sample (mgNH4+/Z).
Remarks 6 In the case of ion densitometer, use ammonium ion standard
solution (0.1 rngNH4'IZ) and that (10 mgNH$lZ), and carry out
the operation specified in (3)(a)to ( c ) t o adjust the indicat-
ing values of the ion densitometer t o be 0.1 mgNH4+/Zand
10 mgNH4'll. Furthermore, confirm the indicating values of
ion densitometer by using ammonium ion standard solution
(1mgNH4'/I) and that (100 mgNH4+ll).
7 Where the concentration of ammonium ion in the sample is
0.1 mgll, hydrazinium ion (hydrazine) of not more than 1mglZ
does not interfere, but if 10 mgíl and 100 mgll coexist, they cause
errors of +35 % and +lo0 %, respectively.
Where the concentration of ammonium ion in the sample is
not less than imgll, the coexistence of hydrazinium ion of
100 mgll does not interfere. Where the concentration of ammo-
nium ion in the sample is 0.1 mgll, coexisting morpholine
(tetrahydro-1,4-oxazine CdHsONH) up to 10 mglZ does not inter-
fere, but if its 100 mgll coexists, it causes an error of +lo0 %.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
206
K O101 : 1998
Where the concentration of ammonium ion in the sample

is not less than 1mg/Z coexisting morpholine of 100 mg/Z does
not interfere.
36.5 Ion chromatography Ammonium ion in a sample is determined by ion chro-

matography. In the case where this method is applied, the sample shall be tested
just after sampling omitting 3.3 Preservation treatment.
Determination range: "4' 0.1 t o 30 mg/I (21)
Repeatability: 2 to 10 % in coefficient of variation (differs according to the ap-
Note (21) For the system of combining with a suppressor, the determination
range is almost the same.
(i) Regents The following reagents shall be used.
Water Water A2 o r A3 specified in JIS K 0557.
Eluent Since eluent (22) is varied according to the class of apparatus and
the class of cation exchanger filled in a separation column, preliminarily
confirm separation of ammonium ion, sodium and potassium by the opera-
tion of Note (9.
Reclaiming solution Though the reclaiming solution(23) is used when a
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
suppressor is used, the reclaiming solution varies according t o the class of
apparatus and the class of the removing column. Preliminarily combining
with the separation column, perform the operation of Note (9,and con-
firm the performance of the reclaiming solution.
Ammonium ion standard solution ( i mgNH4+/ml) As described in
36.2 ( i )(a.
Ammonium ion standard solution (0.1 mgNH4+/ml) Take 10 ml of am-
monium ion standard solution (i mgNH4+/ml)into a 100 ml volumetric flask,
Mixed standad solution of alkali metal element-ammonium ion
[(0.1mgNH4+,0.1 mgNa, 0.1 mgK)/mll Take respectively 10 ml of ammo-
nium ion standard solution (i mgNHd+/rnl)of 36.2 ( i )(f), 10 ml of sodium
standard solution (i mgNa/ml) of 47.1 ( i )(a),and 10 ml of potassium stan-
dard solution (i mgWm1) of 48.1 ( i )(a) into a 100 ml volumetric flask, and
add water to the marked line. Prepare a t the time of use.
Notes (22) An example of the preparation method of eluent is given as
follows.
For using suppressor (example)
Methanesulfonate solution (20 mmol/Z) Dissolve 192.3 g
(approx. 123 ml) of methane sulfonate in water to make 11.
Dilute 10 ml of this solution in water to make 11,
Nitric acid (5 mmol/Z) Dilute 10 ml of nitric acid (0.5 mol/I)
(dissolve 36 ml of nitric acid specified in JIS K 8541 in water
to make 1I) in water t o make 1E .
207
K O101 : 1998
For not using suppressor (example)

[2,6-pyridinedicarboxylatesolution (1 mmol/Z)-tartaric
acid solution ( 5 mmol/Z)l Dissolve O . 16 g of 2,6-pyridine-
dicarboxylate and 0.750 g of L(+)-tartaricacid specified in JIS
K 8532 in water to make 11.
(23) An example of the preparation method of reclaiming solution is
shown as follows.
Tetramethylammonium hydroxide solution (40 mmol/Z)
Dissolve 7.25 g of tetramethylammonium hydroxide Penta-
hydrate in water to make 1 2 .
Sodium hydroxide solution (50 mmol/Z) As descried in
36.3 (1)(d). However, omit the standardization.

(2.1) Ion chromatograph For ion chromatograph, there are a system of combin-
ing a separation column with a suppressor(24) and an individual system of the
separation column. Both systems shall satisfy the conditions described in the
following (a) and (b) and shall be capable of separating and determining am-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
monium ion, sodium and potassium.

(a) Separation column The separation column is made of stainless steel or
synthetic resin ( 2 5 ) , which is filled by cation exchanger (surface layer coated
type or all porous silica type, etc.)(26).
(b) Detector Electric conductivity detector
Notes (24) The suppressor herein is for the purpose of converting anion in
eluent to hydroxide ion, which is filled by an anion exchange
membrane (there are membrane type and electrical dialysis type)
having an enough ion exchange capacity for the concentration
of the anion in the eluent or the anion exchanger having the
performance equal thereto. I t is used by being combined with
reclaiming solution. For electrical dialysis type, however, use
effluent from the detector.
(25) There are, for example, the separation columns made by
tetrafluoroethylene resin, polyether ether ketone, etc.
(26) It is preferable to confirm periodically the performance of the
separation column as follows.
Inject a specific amount of the mixed standard solution of alkali
metal elements-ammonium ion [(lo ygNH4+, 10 pgNa, 10 pgK)/
mi] into the ion chromatograph, obtain the chromatogram by
making eluent flow at a specific flow rate (for instance, 1to 2 mY
min), and use that capable of separating each cation (separa-
tion degree 1.3 o r more).
Prepare the mixed standard solution of alkali metal elements-
ammonium ion [(lo pgNH4+, 10 pgNa, 10 pgK)lml] as follows.
Take respectively 5 ml of ammonium ion standard solution
(1mgNH4+/ml>of 36.2 (1)(f), 5 ml of sodium standard solution
(1 O00 mgNa/Z) of 47.1 (1)(a), and 5 ml of potassium standard
208
K O101 : 1998
solution (i O00 mgWZ) of 48.1 ( i )(a) into a 500 ml volumetric

flasks, and add water to the marked line.
(2.2) Recording part As described in 4.2 (6)of JIS K 0127.
(a) Filter the sample with a filter membrane of 0.45 pm pore diameter o r class
5C filter paper (or class 6 filter paper), discard approx. 50 ml of the initial
filtrate, take the filtrate thereafter.
(b) In the case where the electric conductivity of the sample is 10 mS/m { 100 pS/
cm) (25 OC) o r over, dilute with water at a specific rate so that the electric
conductivity becomes 10 mSím or under.
Make the ion chromatograph a state capable of being operated, and make
eluent flow through the separation column a t a specific flow rate (for ex-
ample, 1 t o 2mllmin). For the apparatus requiring a suppressor, make
the reclaiming solution flow a t a specific flow rate.
Inject a specific quantity (for example, a constant quantity of 50 to 200 pl)
of the sample for which the preparatory operation of (3)was performed into
the ion chromatograph, and record a chromatogram.
Read the indicated value(27) on the peak corresponding to ammonium ion
on the chromatogram.
When the sample is diluted, carry out the operation (a) to (e), as blank
test, for the water of the same quantity as the sample, and correct the in-
dicated value (27) obtained on the sample.
Obtain the concentration of the ammonium ion from the working curve, and
calculate the concentration of ammonium ion in the sample (mgNH4'lI).
Working curve Deal out step by step 0.1 t o 30 ml of ammonium ion stan-
dard solution (0.1 mgNH4+/ml)(28)into a 100 ml volumetric flask, and add
water to the marked line. Carry out the operation of (a)to (c) on this solu-
tion, and read the indicated value (27) corresponding to each ammonium ion.
Separately carry out the operation of (a)to (e) on water as a blank test,
and correct the indicated value corresponding to each ammonium ion. There-
after, prepare the relation curve between the amount of ammonium ion ("*+)
and the indicated value. Prepare the working curve a t measurement.
Notes (27) The indicated value means peak height o r peak area.
(28) In the case where sodium and potassium are simultaneously
tested, use the mixed standard solution of alkali metal elements-
ammonium ion I(O.1 mgNH4+,0.1 mgNa, 0.1 mgK)/ml].
Remarks 8 When the concentration of ammonium ion is 1mgll, if sodium
and potassium are 100 mgll or under, they do not interfere,
9 If the separation column is used continuously, the performance
decreases, therefore confirm it by carrying out the operation
Note ( 2 6 ) periodically. When its performance decreased pre-
pare the solution of 20 to 200 times the concentration of the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
209
K O101 : 1998
eluent, inject it t o the separation column to wash, and con-

firm the performance. If the performance does not recover,
replace by new one.
The separation column is contaminated with suspended mat-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ter, organic matter, etc. in a sample, and its performance gradu-
ally decreases. Therefore, for the sample containing suspended
matter, after removing by the preparatory operation of (3),i t
is tested.
For the sample containing organic matter (protein, oil and
fat, surface active agent, etc.), it is filtered with an ultrafilter
membrane, and the organic matter is removed as much as
possible. Thereafter, it is tested.
Further, if cations having strong affinity with the filler of
the separation column (for instance, calcium, magnesium, etc.)
exist in the sample, they are absorbed in the filler, and the
separation performance gradually decreases. Therefore, the
solution of 20 to 200 times the concentration of the eluent is
periodically prepared, and the separation column should pref-
erably be washed by injecting the said solution i n t o the sepa-
ration column in the same way as for the sample.
Besides, if oxidizing substance and reducing substance co-
exist, the separation performance of the separation column de-
creases. In such a case, if the sample is diluted a t a specific
rate with water to be tested, an effect can be prevented t o some
extent.
10 If methanesulfonate solution [2,6-pyridinedicarboxylate-L(+>-
tartaric acid], etc., as cation exchange column and eluent of
carboxylic acid type, is used for eluent, elution and determi-
nation of bivalent cation of calcium, magnesium, etc. are pos-
sible.
210
K O 1 0 1 : 1998
37 Nitrite ion (NOZ-)and nitrate ion (Nos-)
37.1 Nitrite ion (NOZ-) To determination of nitrite ion, the naphthylethylenediamine

absorptiometry o r the ion chromatography applies. Since the nitrite ion is easily
varied, the test is carried out just after sampling. When it can not be performed
immediately, the sample is preserved in accordance with 3.3,and is tested as soon
as possible. However, when the ion chromatography applies, the sample is tested
just after sampling without performing the preservation treatment of 3.3.
37.1.1 Naphthylethylenediamine absorptiometry Add sulfanilamide (4-amino-

benzenesulfonamide) t o a sample, diazotize with nitrite ion, measure the absorbance
of the red colour azo compound to be generated by adding N - 1-naphthylethylenediamine
(N-1-naphthylethylenediammoniumdihydrochloride), and determine the nitrite ion.
Determination range: NOZ- 0.6 to 6 p g
( i ) Reagents The following reagents shall be used.
4-Aminobenzenesulfonamidesolution Dissolve 2 g of sulfanilamide (4-
aminobenzenesulfonamide) Specified in JIS K 9066 in the mixed solution
of 60 ml of hydrochloric acid specified in JIS K 8180 and approx. 80 ml of
water and further add water t o make 200ml.
N-1-Naphthylethylenediammoniumdihydrochloride solution Dissolve
0.2 g of N-1-naphthylethylenediamine dihydrochloride (N-l-naphthyl-
ethylenediammonium dihydrochloride) specified in JIS K 8197 in water t o
make 200 ml. Preserve in a coloured glass bottle, and do not use the solu-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
tion for which one week or a longer time has elapsed.
Nitrite ion standard solution ( i mgNOg-/ml) Heat sodium nitrite speci-
fied in JIS K 8019 a t 105 t o 110 "C for approx. 4 h. After standing to cool
in a desiccator, obtain the purity of sodium nitrite(l), then take sodium
nitrite corresponding t o 1.50 g as NaN02 100 %, and dissolve in a small
amount of water. Transfer the solution into a 1O00 ml volumetric flask,
and add water up to the marked line. Prepare this solution a t the time of
use. Otherwise, use nitrite ion standard solution NOZ- 1O00 specified in
JIS K 0032.
Nitrite ion standard solution (20 pgNOZ-/ml) Take 10 ml of nitrite ion
standard solution (imgNOz-/ml) in a 500 ml volumetric flask, and add water
up to the marked line. Prepare this solution a t the time of use.
Nitrite ion standard solution (2 pgNOz-íml) Take 20 ml of nitrite ion
standard solution (20 ygNO~-/ml)in a 200 ml volumetric flask, and add water
up to the marked line. Prepare this solution at the time of use.
Note (1) As described in 6 ( i )of JIS K 8019.
211
K O101 : 1998

Filter the sample using filter paper of class 5C ( o r 61, discard the initial
approx. 50 ml, then take a proper amount of the next filtrate(2) (contain-
ing 0.6 to 6 pg as NOZ-)in a 10 ml measuring cylinder (with stopper), and
add water to make 10ml.
Add 1ml of 4-aminobenzenesulfonamide solution, mix by shaking, and al-
low t o stand for approx. 5 min. Thereafter, add l ml of N-l-naphthyl-
ethylenediammonium dihydrochloride solution, mix by shaking, and allow
to stand a t room temperature for approx. 20 min.
Transfer a portion of this solution into an absorption cell, and measure the
absorbance a t a wavelength near 540 nm.
For the blank test, take 10ml of water into a 10ml measuring cylinder
(with stopper), carry out the operation specified in (b) and ( c ) to measure
the absorbance, and correct the absorbance obtained on the sample,
Obtain the amount of nitrite ion from the working curve, and calculate the
concentration of nitrite ion in the sample (mgNOz-4).
Working curve Deal out step by step 3 t o 30 ml of nitrite ion standard
solution (2 pgNO2-lrnl) into a 100 ml volumetric flask, and add water to the
marked line. Take respectively 10 ml therefrom into a 10 ml measuring
cylinder (with stopper), carry out the operation of (b) to (d),and prepare
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the relation curve between the quantity of nitrite ion (NOZ-)and the absor-
bance.
Note (2) In the case where the colour and turbidity remain after filtration,
remove them in accordance with aggregate precipitation method
by zinc sulfate specified in 36.1.1 (3) (3.1) or that by aluminium
sulfate.
In the case of aggregate precipitation method by aluminium sul-
fate, add 2 ml of aluminium potassium sulfate solution (dissolve
5 g of aluminium potassium sulfate 12-water specified in JIS K
8255 in water to make 100 mi) per 100 ml of sample and sodium
hydroxide solution (40g/Z) to form flocks of aluminium hydroxide,
and allow t o stand for several minutes. Then, filter (discard
approx. 20 ml of the initial filtrate) to make the solution clear,
transparent.
When aggregates, colouring lowers due to partial adsorption of
nitrite ion to aluminium hydroxide, therefore separately take, step
by step, nitrite ion standard solution (2 pgNO2-íml) and prepare
the working curve using the series of standard solutions treated
in the same manner as in the test t o determine.
Remarks 1 In general nitrite ion does not coexist with oxidizing substances
such as residual chlorine, but if residual chlorine, chloroamines
(monochloroamine, dichloroamine, nitrogen trichloride), etc.
exist, it colours t o red without the existence of nitrite ion, and
may be mistaken as nitrite ion.
For confirming oxidizing substance in the sample, operate
as follows:
212
K O 1 0 1 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Take 100 ml of the sample, and add 1ml of potassium fluo-

ride solution (3OOglZ) and 0.5g of sodium azide specified in
JIS K 9501. Add hydrochloric acid (l+l) to make acidic (pH
approx. i),then add 1g of potassium iodide Specified in JIS
K 8913, and mix by stirring. Then allow t o stand in a dark
place for several minutes. Add starch solution (10 g/Z) [as de-
scribed in 22.1.2 (i)(i)]as indicator, and when the blue colour
of iodine starch is recognized, the oxidizing substance exists.
When testing nitrite ion in succession, titrate with sodium sulfite
solution (50 mmol/Z) until the blue colour of iodine starch dis-
appears, and obtain the amount of sodium sulfite solution
(50 mmol/Z) equivalent to the amount of the oxidizing substance
in the sample from the amount of consumption of sodium sulfite
solution (50 mmoVZ). Add this amount to the sample, and then
carry out the operation specified in (a) to (e).
2 In the case where the expression is performed by nitrite ni-
trogen, the following conversion formula shall be used.
Nitrite nitrogen (mgN02- - N/Z)
= nitrite ion (mgNOz-/Z) x 0.304 5
37.1.2 Ion chromatography Nitrite ion in a sample is determined by the ion

chromatography. When this method applies, the sample shall be tested just after
sampling without performing the preservation treatment of 3.3.
Determination range: NOZ- 0.5 to 40 mglZ(3)
Repeatability: 2 to 10 % in coefficient of variation (differs according to the ap-
Note (3) For the system of combination with a suppressor, the determination
range is 0.1 t o 40 mgNOz-/Z.
Eluent As described in 32.5 (i) (b).
Reclaiming solution As described in 32.5 (i)( e ) .
Nitrite ion standard solution ( 5 mgNOz-/ml) Heat sodium nitrite speci-
fied in JIS K 8019 a t 105 t o 110 "C for approx. 4 h. After standing to cool
in a desiccator, obtain the content of sodium nitrite(l), take sodium nitrite
equivalent to 0.750 g as 100 % NaN02, dissolve in a small amount of wa-
ter, transfer to a 100 ml volumetric flask, and add water to the marked
line. Prepare a t the time of use.
Nitrite ion standard solution (0.5 mgNOz-/ml) Take 10 ml of nitrite
ion standard solution (5 mgNOz-íml) into a 100 ml volumetric flask, and add
water t o the marked line. Prepare a t the time of use.
Anion mixed standard solution L(O.1 mgCl-, 0.5 mgNOs-, 0.5 mgBr-,
0.5 mgNO3-, 1 mgS0~2-)/mllAs described in 32.5 (i)(f).
213
K O101 : 1998
Apparatus The apparatus shall be in accordance with 32.5 (2). However, as

the detector, the ultraviolet absorption detector may be used.
Preparatory operation Carry out the preparatory operation as described in
32.5 (3).
Operation Carry out the operation as follows.
(a) Perform the operation of 32.5 (4)(a) and (b).
(b) Measure indicated value(4) on the peak corresponding to nitrite ion on the
chromatogram.
(c) When the sample is diluted, carry out the operation (a)and (b), as blank
test, for the water of the same quantity as the sample, and correct the
indicated value (4) obtained on the sample.
(d) Obtain the concentration of nitrite ion from the working curve, and calcu-
late the concentration of nitrite ion in the sample (mgNOz-ll).
Working curve Deal out step by step 0.1 to 8 ml of nitrite ion standard
solution (0.5 mgNOz-/ml)(5) into a 100 ml volumetric flask, add water to the
marked line. Perform the operation of (a)and (b) on this solution, and
read the indicated value (4) corresponding to each nitrite ion. Separately,
perform the operation of (a) and (b) as a blank test, and correct the indi-
cated value (4) corresponding t o each nitrite ion. Thereafter, prepare the
relation curve between the quantity of nitrite ion (Noz-) and the indicated
value. Prepare the working curve at the time of measurement of the sample.
(5) In the case where anion other than nitrite ion is simultaneously
tested, use the anion mixed standard solution [(O. 1 mgCl-,
0.5 mgNOz-, 0.5 mgBr-, 0.5 mgNOa-, 1 mgS042-)/m1].
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 3 If chloride ion is 50 mglZ or under, bromide ion is 200 mgll or

under and sulfate ion is 500mglZ or under when the concen-
tration of nitrite ion is 1mgll, they do not interfere.
37.2 Nitrate ion (NOS-) To determination of nitrate ion, reducing distillation-

Indophenol Blue absorptiometry, reducing distillation-acid-base titrimetric method,
copper and cadmium column reduction-naphthylethylenediamine absorptiometry, bru-
cine absorptiometry, or ion chromatography shall apply.
This test is carried out just after sampling. When it can not be carried out imme-
diately, the sample is preserved in accordance with 3.3, and is tested as soon as
possible.
37.2.1 Reducing distillation-Indophenol Blue absorptiometry Add sodium

hydroxide to the sample, and distill. After removing ammonia generated by decom-
position of ammonium ion and a partial organic nitrogen compound, add Devarda’s
alloy, reduce nitrite ion and nitrate ion t o ammonia, distill, and absorb the distilled
ammonia in sulfuric acid (25 mmol/Z). Then, determine ammonium ion in the distil-
late solution by Indophenol Blue absorptiometry, obtain the total sum of nitrate ion
and nitrite ion, subtract the separately determined nitrite ion from the said value,
and calculate the quantity of nitrate ion.
214
K O101 : 1998
Determination range: NOS- 17 to 340 pg

Sulfuric acid (25mmol/Z) As described in 36.1.2 (1)(b).
8951.
Sodium hydroxide solution (40 g/Z) As described in 19 ( i )(g).
Sodium hydroxide solution (300gll) Dissolve 30 g of sodium hydrox-
ide specified in JIS K 8576 in water to make 100 ml. Prepare at the time
of use.
Devarda’s alloy As specified in JIS K 8653. Powdery one.
Sodium phenoxide solution As described in 36.2 (i)(d).
Sodium hypochlorite solution (effective chlorine 10 gil) As described
in 36.2 ( i )(e).
Ammonium ion standard solution (i mgNH4+/ml) As described in
36.2 (i)(f).
Ammonium ion standard solution (10pgNH4+/ml) As described in
36.2 (1) (g).
(a) Glassware Wash sufficiently with water prior to use.
(b) Distillation apparatus As described in 36.1.2 (2)(a). Wash sufficiently
with water prior to use.
(c) Photometer Spectrophotometer or photoelectric photometer
Take a suitable amount (containing 0.14 mg o r over as NOS-) of a sample.
In the case where the sample is not neutral, adjust pH to approx. 7 with
sodium hydroxide solution (40 glZ) or sulfuric acid (1+35).
Wash it with water to be transferred to a distillation flask, add 1 0 m l of
sodium hydroxide solution (300 g/Z) and several pieces of boiling tips (grain
diameter 2 to 3 mm), add water to make approx. 350 ml, remove ammonia
by performing the distilling operation of 36.1.2 (3)(c) to (e)(S),detach the
condenser and the back-flow stopper, and wash sufficiently the inner tube
of the condenser and the in- and outside of the back-flow stopper.
Allow the residual solution in the distillation flask to stand t o cool.
Use another 200 ml measuring cylinder (with stopper) as the receiver, and
put 50 ml of sulfuric acid (25 mmol/Z) thereinto.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
215
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Quickly add 3 g of Devarda's alloy t o the residual solution in the distilla-

tion flask, and add water t o make approx. 350 ml. Thereafter, assemble
the apparatus.
Carry out the operation of 36.1.2 (3)(d) t o (f)(7).
Partially take a suitable amount (containing 5 t o 1OOpg as "4)' of the

distillate solution obtained in (0into a 50ml volumetric flask, and add
water t o make approx. 25 ml.
Measure the absorbance by performing the operation of 36.2 (3)(b) t o (e).
Take approx. 100 ml of water into a distillation flask as a blank test, and
add 10 ml of sodium hydroxide solution (300 g4). Thereafter, carry out the
operation of (d) to (f).
Partially take the same amount as ( g ) on the distillate solution obtained
in (i) into a 50 ml volumetric flask, measure the absorbance by performing
the operation of (h),and correct the absorbance obtained on the sample.
Obtain the amount of ammonium ion (mgNH4+)in the aliquot distillate
solution from the working curve of 36.2 (3).
Separately, obtain the concentration of nitrite ion in the sample (mgNO2-lZ)
in accordance with 37.1.1 o r 37.1.2.
Calculate the concentration of nitrate ion in the sample (mgNOs-lZ) accord-
ing t o the following formula.
N=ax3.437x-x--
'Ooo 2oo Cx1.348
VI vz
where, N : nitrate ion (mgNOJ-/Z)
U : amount of ammonium ion in the distillate solution
obtained in (k)(mgNH4')
3.437 : coefficient when ammonium ion is converted t o
vi :
nitrate ion equivalent -
sample (mi)
c::3
v
2 : distillate solution partially taken in ( g ) (ml)
C : nitrite ion in the sample obtained in (1) (mgNOz-íZ)
1.348 : coefficient when nitrite ion is converted to nitrate
ion equivalent -
(46.3
Notes (9 In this operation, the receiver may contain water instead of sul-
furic acid (25 mmol/Z).
(7) When bubbling is vigorous at the start of distillation, weaken
heating and after approx. 10min has elapsed and bubbling be-
comes quiet, distill again.
Remarks 5 The ammonia distilled by the operation of (3)(b) occasionally
contains those generated by decomposition of organic nitro-
gen compound other than ammonium ion having existed in the
sample. Therefore, ammonium ion in the sample shall not be
determined by using this distillate solution.
216
K O101 : 1998
6 In the case where the expression is performed by nitrate ni-

trogen, use the following conversion formula.
Nit rat e nitrogen (mgNOS-NIZ)
= nitrate ion (mgNOs-/E) x 0.225 9
37.2.2 Reducing distillation-acid-base titrimetric method Add sodium hy-

droxide t o a sample, distill, and remove ammonia generated by decomposition of am-
monium ion and a partial organic nitrogen compound. Thereafter, add Devarda’s
alloy, reduce nitrite ion and nitrate ion t o ammonia, distill, absorb the distilled ammonia
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
in a specific amount of sulfuric acid (25 mmollZ), and obtain the amount equivalent
t o the nitrite ion and the nitrate ion by determination by the acid-base titrimetric
method. Separately determine the nitrite ion, subtract, and calculate the amount of
the nitrate ion.
Determination range: NOS- 1 to 140mg
Sulfuric acid (25 mmol/Z) As described in 36.1.2 (1)(b).
8951.
50 mmoW sodium hydroxide solution As described in 36.3 (1) (d).
Sodium hydroxide solution (40 g l l ) As described in 19 (1) ( g ) .
Sodium hydroxide solution (300 glZ) As described in 37.2.1 (1) (e).
Devarda’s alloy As described in 37.2.1 (1)(f).
Mixed solution of Methyl Red-Bromocresol Green As described in
13.1 (1) (a).
(a) Glassware Sufficiently wash with water prior to use.
(b) Distilling apparatus As described in 36.1.2 (2) (a). Wash sufficiently
with water prior t o use.
(a) Take a suitable amount of a sample (containing 1mg or over as NOS-, and
140 mg o r under as Nos- for the sum of NOZ- and NOS-). When the sample is
not neutral, adjust pH to approx. 7 with sodium hydroxide solution (40 g/Z) or
sulfuric acid (1+35).
(b) Transfer it t o a distillation flask by washing with water, add 10 ml of so-
dium hydroxide solution (300glZ) and several pieces of boiling tips (grain
diameter 2 t o 3 mm), add water t o make approx. 350 ml, remove ammonia
by carrying out the distilling operation of 36.1.2 (3)( c ) to (e)(6),detach the
condenser and the back-flow stopper, and wash sufficiently the inner tube
of the condenser and the in- and outside of the back-flow stopper with water.
217
K O101 : 1998
(c) Allow the residual solution in the distillation flask t o stand t o cool.
(d) Use a 500 ml Erlenmeyer flask as the receiver, add accurately 50 ml of sulfuric
acid (25 mmol/Z) thereto, and add 5 t o 7 drops of the mixed solution of Methyl
Red-Bromocresol Green as indicator.
(e) Quickly add 3 g of Devarda’s alloy t o the residual solution in the distilla-
tion flask, and add water t o make approx. 350 ml. Thereafter, assemble
the apparatus.
(0 Carry out the operation of 36.1.2 (3)(d)to (f)(7).
(g) Use the whole quantity of the distillate solution, and carry out the titrat-
ing operation of 36.3 (2)(a).
(h) Take approx. 100 ml of water into the distillation flask as a blank test, and
add 10 ml of sodium hydroxide solution (300 g/Z). Thereafter, carry out the
operation of (d) t o (0,and perform the titrating operation on the obtained
distillate solution in the same way as in (g).
(i) Separately, obtain the concentration of nitrite ion in the sample (mgN02-/Z)
in accordance with 37.1.1 or 37.1.2.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
6 ) Calculate the concentration of nitrate ion in the sample (mgNOs-/Z) accord-

where, N : nitrate ion (mgNOs-/Z)

titration (ml)
b : 50 mmol/l sodium hydroxide solution required for
titration in blank test (mi)
f : factor of 50mmol/Z sodium hydroxide solution
3.1 : nitrate ion equivalent of 1ml of 50 mmol/Z sodium
hydroxide solution (mg)
V : sample (mi)
C : nitrite ion in the sample obtained in (i) (mgNOz-lZ)
1.348 : coefficient when nitrite ion is converted t o nitrate
ion equivalent -
(2.3
37.2.3 Copper and cadmium column reduction-naphthylethylenediamine
absorptiometry Reduce nitrate ion in a sample with a copper and cadmium col-
umn to make nitrite ion, determine by the naphthylethylenediamine absorptiometry,
and obtain the concentration of nitrate ion.
Determination range: NOS- 0.8 t o 8 pg

218
K O101 : 1998
Hydrochloric acid (1+11) Prepare by using hydrochloric acid specified

in JIS K 8180.
Ammonium chloride-ammonia solution Dissolve 100 g of ammonium
chloride specified in JIS K 8116 in approx. 700 ml of water. Thereafter,
add 50ml of aqueous ammonia specified in JIS K 8085 and further add
water t o make 1 E.
Column activation solution Dissolve 38 g of disodium dihydrogen
ethylenediaminetetraacetate dihydrate specified in JIS K 8107 and 12.5 g
of copper (II) sulfate pentahydrate specified in JIS K 8983 in approx. 700 ml
of water joined by 70 ml of sodium hydroxide solution (80 gll) and further
drip sodium hydroxide solution (80glZ) to adjust pH of the solution to 7.
Thereafter, add water t o make 11.
Copper and cadmium column packing Take approx. 40 g of granular
cadmium (grain diameter 0.5 t o 2 mm) into a 300 ml Erlenmeyer flask, add
approx. 50 ml of hydrochloric acid (1+5),mix by shaking, clean the surface
of cadmium, discard washings, and wash five times with each approx. 100 ml
of water. Then add approx. 50 ml of nitric acid (1+39), mix by shaking,
clean the surface of cadmium, discard washings. After repeating this op-
eration twice, wash 5 times with each approx. 100 ml of water. Then, add
200 ml of column activation solution, allow to stand for 24 h, and form a
copper film on the surface of cadmium. This copper cadmium column packing
can be tightly stoppered as i t is t o be preserved.
Further, the copper and cadmium packing on the market may be used
instead of that prepared by this method.
Column packing liquid Dilute ammonium chloride-ammonia solution
ten times with water.
4-Aminobenzenesulfonamidesolution As described in 37.1.1 (1) (b).
N-1-Naphthylethylenediammoniumdihydrochloride solution As de-
scribed in 37.1.1 (1) (c).
Nitrate ion standard solution (1 mgNOs-/ml) Preliminarily heat potas-
sium nitrate specified in JIS K 8548 a t 105 to 110 O C for approx. 3 h, and
allow to stand t o cool in a desiccator. Take its 1.63 g, dissolve in a small
amount of water, transfer t o a 1O00 ml volumetric flask, and add water to
the marked line. Preserve in a dark place a t O t o 10 O C . Otherwise, use
nitrate ion standard solution Nos- 1O00 Specified in JIS K 0031.
Nitrate ion standard solution (10 pgNOS-/ml) Take 10 ml of nitrate ion
standard solution (1mgNOs-lml) into a 1 O00 ml volumetric flask, and add
water to the marked line. Prepare at the time of use.
(a) Copper and cadmium column Fill the bottom part of the glass tube as
given in Fig. 37.1 (A) with glass wool specified in JIS K 8251. After filling
it with column packing solution, make copper and cadmium column pack-
ing flow into so as not to come into contact with the air. Fill the upper part
with glass wool, and attach a cylindrical separating funnel thereto. Then,
make 100 ml of column packing solution, 200 ml of the solution obtained by
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
219
K O101 : 1998
diluting nitrate ion standard solution (1mgNOs-/ml) 100 times with the
column packing solution and further 100 ml of the column packing solution
in this order flow down at a flow rate of approx. 10 mumin from the cylin-
drical separating funnel. At that time, make the surface of the solution in
the column slightly above the packing.
Further, when the copper and cadmium column is not used, put the col-
umn packing solution up to the upper part so that the copper and cadmium
column packing does not contact the air. Since the copper and cadmium
column is deteriorated as it is used and the reduction rate of nitrate ion
decreases, reproduce by use of column activation solution as occasion de-
mands. For reproduction, fill the copper and cadmium column with the
column activation solution, and allow to stand for 2 to 3h. Thereafter,
wash with the column packing(8).
Note (8) Make approx. 20 ml of column activation solution flow to the cop-
per and cadmium column every time the copper and cadmium
column is used 15 to 20 times for the sample and then if it is washed
with approx. 100 ml of the column packing, decrease of the reducing
rate of nitrate ion can be prevented.
U n i t : mm
\/815,25
One way
cock
\ ,
i
,
L
- - I_
(A) Copper and cadmium column (B) Cylindrical separating funnel

bi, bz: Glass wool
C : Copper a n d cadmium packing
D : Column packing solution
Fig. 37.1 An example of copper and cadmium column
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
220
K O101 : 1998

Filter a sample with class 5C (or 6) filter paper, and discard approx. 50 ml
of the initial filtrate(9)(10). Take a suitable amount of the filtrate thereaf-
ter (containing 8 pg o r over as Nos-, and 80 pg or under as the NOa- of the
sum of NOZ- and NOS-) into a 100 ml volumetric flask.
Add 10 ml of ammonium chloride-ammonia solution thereto, and further
add water t o the marked line t o make reducing solution.
Put the reducing solution into the upper part cylindrical separating fun-
nel, make the reducing solution flow down a t approx. 10ml/min keeping
the solution surface in the copper and cadmium column slightly above the
packing, and discard approx. 30 ml of the effluent. Add the reducing solu-
tion, make it flow down in the same way, and collect 30 ml thereafter in a
50 ml measuring cylinder.
Take 10 ml from this effluent into a stoppered test tube, and carry out the
operation of 37.1.1 (3)(b) and (c).
Take water into a 100ml volumetric flask as a blank test, measure the
absorbance by performing the operation of (b)t o (d),and correct the ab-
sorbance obtained on the sample.
Obtain the amount of nitrate ion in 10 ml of the effluent in (d)from the
working curve, and calculate the concentration of nitrate ion in the sample
[the concentration of the sum of nitrite ion and nitrate ion (nitrate ion
converted amount)] (mgNOs-lZ).
Separately, obtain the concentration of nitrite ion in the sample (mgNOZ-lZ)
in accordance with 37.1.1.
Calculate the concentration of nitrate ion in the sample (mgNOs-lZ) accord-
N=a-bx1.348
where, N : nitrate ion (rngNOa-/Z)
a : the sum of nitrite ion and nitrate ion in the sample
which is calculated in (f) (mgNO3-/Z)
b : nitrite ion in the sample obtained in ( g ) (mgNOZ-/Z)
1.348 : coefficient when nitrite ion is converted t o nitrate
(-)
ion 62.00
46.01
Working curve Deal out step by step 0.8 t o 8 ml of nitrate ion standard
solution (10 pgNOs-/ml) into a 100 ml volumetric flask, carry out the op-
eration of (b)t o (e),and prepare the relation curve between the amount of
nitrate ion (Nos-) and the absorbance.
Notes (9) As described in Note (2). However, in the case where the colour
remains even though aggregate precipitation treatment is car-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ried out, this method can not be applied. In such a case, the
sample is determined in accordance with 37.2.1 or 37.2.2.
221
K O101 : 1998
(10) Since oxidizing matter and reducing matter interfere, they shall
be preliminarily removed. When oxidizing matter such as re-
sidual chlorine or the like coexists, after adding equivalent so-
dium sulfite solution (6.3 g/2) or sodium arsenite solution [after
dissolving 0.5 g of diarsenic trioxide specified in JIS K 8044 in
5 ml of sodium hydroxide solution (40 g l ) , add 6 ml of hydrochloric
acid (1+11)to make 100ml with water], the sample is tested.
Further, when reducing matter such as sulfite ion o r the like
coexists, the sample is made slightly alkaline and is joined by
equivalent hydrogen peroxide water (1+100). Thereafter, it is
tested.
37.2.4 Brucine absorptiometry The absorbance of the yellow compound to be

generated by the reaction of nitrate ion with brucine under strong acidity with sul-
furic acid, is measured t o determine nitrate ion.
Determination range: NO3- 5 to 1OOpg
(b) Sulfuric acid (20+3) Take 75 ml of water into a beaker, cool, add gradu-
ally 500 ml of sulfuric acid specified in JIS K 8951 with stirring, cool, stopper
tightly and preserve.
(c) Brucine and 4-aminobenzenesulfonic acid solution Dissolve 1g of
brucine n-hydrate (2,3-dimethoxystrychnizine-lO-on * n-hydrate) specified in
JIS K 8832 and 0.1 g of sulfanilic acid (4-aminobenzenesulfonicacid) specified
in JIS K 8586 in 3 ml of hydrochloric acid specified in JIS K 8180,and
add water to make 100ml.
(d) Nitrate ion standard solution (1 mgNOs-/ml) As described in 37.2.3
(1) (i).
(e) Nitrate ion standard solution (0.1 mgNOa/ml) Take 20 ml of nitrate
ion standard solution (1mgNOs-/ml) into a 200 ml volumetric flask, and add
water to the marked line. Prepare a t the time of use.
(a) Safety pipette 1ml
Filter a sample with class 5C (or 6) filter paper, and discard approx. 50 ml
of the initial fïltrate(9)(10) (11). Take each 2 ml from the filtrate thereafter
(containing 5 to 100 pg as NO3-1 into two 50 ml beakers(12) (Ad, (BI).
Add 1ml of brucine and 4-aminobenzenesulfonic acid solution t o (Ad by
use of a safety pipette and add 1ml of water t o (Bi) instead of the brucine
and 4-aminobenzenesulfonic acid solution.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
222
K O101 : 1998
Separately, take each 10 ml of sulfuric acid (20+3)(13)into two 50 ml bea-

kers (Ad, (Bz), carefully transfer the solution of the beaker (Ai) t o the bea-
ker (Ad, and sufficiently mix by shaking. Then, transfer the solution of
the beaker (Ad to the beaker (AI), and sufficiently mix by shaking again.
After repeating this operation 5 times, allow t o stand in a dark place('*)
for approx. 10 min.
Carefully transfer the solution of the beaker (BI) to the beaker (Bz), and
sufficiently mix by shaking. Then, transfer the solution of the beaker (B2)
to the beaker (BI),and sufficiently mix by shaking again. After repeating
this operation 5 times, allow t o stand in a dark place for approx. 10 min.
Add respectively each 10 ml of water t o beakers (Az) and (Bz), mix by shak-
ing, and wash the wall of the apparatus, join respectively washings t o the
solutions of (AI) and (BI), mix by shaking. Allow to stand in a cold dark
place for approx. 30 min.
Take a portion of the solution in the beaker (Ad into an absorption cell,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
use the solution in the beaker (BI) as reference solution, and measure the
absorbance near 410 nm in wavelength.
Take each 2 m l of water into two 50ml beakers(l2) (Ai), (BI) as a blank
test, obtain the absorbance by carrying out the operation of (b) t o (f),and
correct the absorbance obtained on the sample.
Obtain the amount of nitrate ion from the working curve, and calculate
the concentration of nitrate ion in the sample (mgNOa-lZ).
Working curve Deal out step by step 2.5 t o 50ml of nitrate ion stan-
dard solution (0.1 mgNOa-lml) into a 100 ml volumetric flask(15) and add
water t o the marked line. Take each 2 m l therefrom, perform the opera-
tion of (b)to (g), and prepare the relation curve between the amount of
nitrate ion (Nûa-) and the absorbance.
Notes (11) In the case where the alkalinity is strong, add sulfuric acid (1+5),
adjust pH t o approx. 7, and test.
(12) For this test, 4 pieces of the same shaped 50 ml beakers as those
in preparation of the working curve shall be used.
(13) If a 10 ml automatic burette is used, the operation becomes easy.
(14) I t is preferable that the beaker is covered by a cardboard case
or a wooden box.
(15) Since the yellow generated by the reaction of nitrate ion with
brucine does not strictly follow Lambert-Beer law, it is neces-
sary that the stage of nitrate ion standard solution is made small.
Remarks 7 When iron (II), iron (III) and manganese (IV) coexist, they cause
positive errors. However, if their concentration 1 mg/Z or under,
they are allowed t o coexist.
8 The reaction of nitrate ion with brucine becomes quick as tem-
perature rises. Therefore, the operation is performed a t the
same temperature as in preparation of the working curve.
223
K 0101 : 1998
37.2.5 Ion chromatography Nitrate ion in a sample is determined by the ion

chromatography. In the case where this method applies, the sample shall be tested
just after sampling without performing the preservation treatment of 3.3.
Determination range: Nos- 0.5 to 40 mg/E (16)
Repeatability: 2 to 10 % in coefficient of variation (different according to the
apparatus and measuring conditions)
Note (16) In the case of the combination system with a suppressor, the de-
termination range is 0.1 to 40 mgNO3-ll.
(a) Water Water A2 o r A3 specified in JIS K 0557.
(b) Eluent As described in 32.5 (1) (b).
(c) Reclaiming solution As described in 32.5 (1) (c).
(d) Nitrate ion standard solution (5 mgNOs-/ml) Preliminarily heat potas-
sium nitrate specified in JIS K 8548 at (105f2) "C for approx. 2 h, and al-
low t o stand to cool in a desiccator. Take its 0.815g, dissolve in a small
amount of water, transfer t o a 100 ml volumetric flask, and add water t o
the marked line. Preserve in a dark place at O t o 10 "C.
(e) Nitrate ion standard solution (0.5 mgNOa-fml) Take 10 ml of nitrate

ion standard solution (5 mgNOa-lml) into a 100 ml volumetric flask, and add
water t o the marked line. Prepare at the time of use,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(f) Anion mixed standard solution [(0.1mgCl-, 0.5 mgNOz-, 0.5 mgBr-,
0.5 mgNOa-, 1 mgS042-)/mll As described in 32.5 (1) (f).
(2) Apparatus The apparatus shall be as described in 32.5 (2). However, for the
detector, an ultraviolet absorption detector may be used.
(3) Preparatory operation Carry o u t as described in 32.5 (3).

(a) Carry o u t the operation of 32.5 (4)(a) and (b).
(b) Read the indicated value(l7) on the peak corresponding t o nitrate ion on
the chromatogram.
(c) If the sample is diluted, carry out the operation (a) and (b) for the same
amount of water as the sample, as a blank test, and correct the indicated
value(l7) obtained on the sample.
(d) Obtain the concentration of nitrate ion from the working curve, and calcu-
late the concentration of nitrate ion in the sample (mgNOs-/Z).
Working curve Deal out step by step 0.1 t o 8 ml of nitrate ion standard
solution (0.5 rngNOs-/ml)(1R) into a 100 ml volumetric flask, add water to
the marked line, carry o u t the operation of (a) and (b),and read the indi-
cated value (17) corresponding t o each nitrate ion. Separately, carry out the
operation of (a) and (b) on water as a blank test, and correct the indicated
224
K O 1 0 1 : 1998
value corresponding t o each nitrate ion. Thereafter, prepare the relation

curve between the amount of nitrate ion (NOa-) and the indicated value.
Notes (17) The indicated value means peak height o r peak area.
(18) In the case where anion other than nitrate ion is simultane-
ously tested, the anion mixed standard solution I(O.1 mgCl-,
0.5 mgNOz-, 0.5 mgBr-, 0.5 mgNOa-, 1mgS042-)/ml]shall be used.
Remarks 9 If bromide ion is 200 mglZ or under and sulfate ion is 500 mgll
or under when the concentration of nitrate ion is 1mgll, they do
not interfere.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
225
K O 1 0 1 : 1998
38 Organic nitrogen Decompose organic matter by pretreating a sample, con-

vert organic nitrogen t o ammonium ion, and separate by distillation. Thereafter,
determine the ammonium ion by the Indophenol Blue absorptiometry o r the acid-
base titrimetric method. Separately, determine ammonium ion in the sample before
the treatment, subtract, and obtain the organic nitrogen.
Since the organic nitrogen is liable to vary, carry out the test immediately. When
the test can not be immediately carried out, preserve in accordance with 3.3, and
carry out the test as soon as possible.
38.1 Pretreatment (Kjeldahl method) Add copper sulfate, sulfuric acid, and po-
tassium sulfate to a sample, and decompose organic matter by heating. Then, make
it alkaline by adding sodium hydroxide. Thereafter, distill, and make the distilled
ammonia be absorbed in sulfuric acid (25 mmol/Z).
Remarks 1 Though amino acid, polypeptide, protein, etc. are easily decomposed
by this method, nitro, nitroso, azo, heterocyclic compound (espe-
cially, a compound having a pyridine ring), etc. can not completely
decompose.
Sulfuric acid (25 mmol/Z) As described in 36.1.2 (1)(b).
8951.
Sodium hydroxide solution (40 g/Z) As described in 19 (1)(g).
Sodium hydroxide solution (500 gll) Dissolve 50 g of sodium hydrox-
ide specified in JIS K 8576 in water t o make 100 ml. Prepare a t the time
of use.
Potassium sulfate As specified in JIS K 8962.
Copper (II) sulfate pentahydrate As specified in JIS K 8983. Make
powdery and use.
(a) Glassware Wash sufficiently with water prior to use.
(b) Kjeldahl flask 200 ml Wash sufficiently with water prior to use.
(c) Distilling apparatus As described in 36.1.2 (2)(a). Wash sufficiently
with water prior to use.
(a) Take a suitable amount(1) of a sample into a 500 ml beaker, add sulfuric
acid (1+35)to make slightly acidic, and concentrate t o approx. 30 ml by
heating.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
226
K O101 : 1998
After standing t o cool, transfer contents into a 200ml Kjeldahl flask by

washing with a small amount of water.
Add 10 ml of sulfuric acid, 5 g of potassium sulfate and 2 g of copper (II)
sulfate pentahydrate, generate white fume of sulfuric acid by heating, and
successively decompose (2) organic matter by igniting for approx. 30 min.
After standing to cool, add a small amount of water, dissolve by heating,
and transfer to a distillation flask by washing with water to make approx.
300 ml.
Connect the distillation flask as shown in Fig. 36.1, use a 200 ml measuring
cylinder (with stopper) as the receiver, and put 50 ml of sulfuric acid (25 mmoll
1 ) therein (3).
After adding 40 ml of sodium hydroxide solution (500 gll) from the injec-
tion funnel on the upper part of the distillation flask, carry out the opera-
tion of 36.1.2 (3)(d) t o (f).
As a blank test, take 30 ml of water, and carry out the operation of (c)t o
(f) .
Notes (1) Take organic nitrogen to contain 32 pg or over as N for determi-
nation by Indophenol Blue absorptiometry, and take organic ni-
trogen to contain 0.23 mg or over as N and the sum of organic
nitrogen and ammonium ion to contain 30 mg or under as N for
determination by acid-base titrimetric method.
(2) The solution in the flask becomes colourless or light yellow.
(3) I n the case of the acid-base titrimetric method, use a 500ml
Erlenmeyer flask instead of the 200 ml measuring cylinder (with
stopper), add accurately 50ml of sulfuric acid (25mmol/Z) of
36.1.2 (i)(b),and add 5 to 7 drops of the mixed solution of Methyl
Red and Bromocresol Green of 13.1 (i)(a) as indicator.
Remarks 2 Nitrate ion and nitrite ion do not interfere the determination
of organic nitrogen by this method.
3 In the case where organic nitrogen is determined after pre-
liminarily removing ammonium ion in the sample, carry out
the operation of 36.1.2(3)(a)t o ( e ) ,and carry out the pre-
treatment of (3)(a) t o (f) on the said residual solution.
38.2 Indophenol Blue absorptiometry Determine ammonium ion by the Indophe-

nol Blue absorptiometry on the distillate solution by the pretreatment (Kjeldahl
method), obtain the sum of ammonium ion contained in the sample and ammonium
ion generated from organic nitrogen, separately determine ammonium ion in the sample,
subtract, and calculate organic nitrogen.
Determination range: N 4 t o 80pg
(b) Sodium phenoxide solution As described in 36.2 (i)(d).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
227
K O101 : 1998
(c) Sodium hypochlorite solution (effective chlorine 10 gll) As described

in 36.2 (1) (e).
(d) Ammonium ion standard solution (1 mgNH*+/ml) As described in 36.2
(1) (f).
( e ) Ammonium ion standard solution (10 pgNH4+/ml) As described in 36.2
(1)(g).
(a) Glassware Sufficiently wash with water prior t o use.
(a) Take a suitable amount of the distillate solution obtained in 38.1 (3)(f)(con-
taining 4 to 80 pg as N) into a 50 ml volumetric flask, and add water to
make approx. 25 ml. Thereafter, carry out the operation of 36.2 (3)(b)to
(e) to measure the absorbance.
(b) Partially take the same amount as in (a) from the distillate solution ob-
tained in 38.1 (3)(g) as a blank test, measure the absorbance by carrying
out the operation of (a),and correct the absorbance obtained on the sample.
(c) Obtain the amount (mg) of ammonium ion in the aliquot distillate solution
from the working curve of 36.2 (3).
(d) Separately, obtain the concentration of ammonium ion in the sample
(rngNHr+/Z)in accordance with 36.2.
(e) Calculate the concentration of organic nitrogen in the sample (mgN/Z) ac-
cording t o the following formula.
[
N = ax-x--
'Ooo
VI
2oo
v
2
A
where, N : organic nitrogen (mgN/Z)

a : amount of ammonium ion in the distillate solution
of (cl (rngNH4')
Vi : sample used in 38.1 (3)(a) (ml)
VZ: distillate solution partially taken in (a) (ml)
A : ammonium ion in the sample obtained in (d)
(mgNH4+/Z)
0.776 6 : coefficient when ammonium ion is converted t o
nitrogen equivalent -
(:23
38.3 Acid-base titrimetric method Determine ammonia by the acid-base titri-
metric method on the distillate solution by the pretreatment (Kjeldahl method), obtain
the sum of ammonium ion contained in the sample and ammonium ion generated
from organic nitrogen, separately determine ammonium ion in the sample, subtract,
and calculate organic nitrogen.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
228
K O 1 0 1 : 1998
Determination range: N 0.23 to 30 mg

(b) 50 mmol/Z Sodium hydroxide solution As described in 36.3 (1) (d).
(a) Use the total amount of the distillate solution obtained in 38.1 (3) (f), and
carry out the titrating operation of 36.3 (2) (a).
(b) As a blank test, use the total amount of the distillate solution obtained in
38.1 (3) ( g ) , and carry out the same titrating operation as in (a).
(c) Separately, obtain the concentration of ammonium ion in the sample
(mgNH4+/Z)in accordance with 36.3.
(d) Calculate the concentration of organic nitrogen in the sample (mgNlZ) ac-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
cording t o the following formula.
N = ( b - a)x f x 1000x0.700 -Ax0.776 6

V
where, N : organic nitrogen (mgNlZ)
b : 50 mmol/Z sodium hydroxide solution required for
titration in blank test (ml)
a : 50 mmolll sodium hydroxide solution required for
titration (ml)
f : factor of 50 mmolll sodium hydroxide solution
0.700 : nitrogen equivalent of 1ml of 50 mmolll sodium
hydroxide solution (mg)
V : sample used in 38.1 (3)(a) (ml)
A : ammonium ion in the sample obtained in ( c )
(mgNH4+/1)
0.776 6 : coefficient when ammonium ion is converted t o
nitrogen equivalent -
(:s4:04)
Remarks 4 Boric acid solution (saturated) may be used instead of sulfu-
ric acid (25 mmolll) of 38.1 (3)(e). In that case, the operation
of Remarks 5 of 36 is carried out. However, for the blank test,
the value obtained by titrating the distillate solution obtained
by the operation of 38.1 (3) (cl t o (f) by using 30 ml of water
in the same way as for the sample, is used.
229
K O 1 0 1 : 1998
39 Total nitrogen The following methods shall be applied: sum total method by
which a t first the nitrogen corresponding t o nitrite ion and nitrate ion and the ni-
trogen corresponding t o ammonium ion and organic nitrogen are tested, and they
are totalled; ultraviolet absorptiometry by which all nitrogen compounds are con-
verted into nitrate ion, and then determined; hydrazinium sulfate reducing method;
copper cadmium column reducing method; and thermal decomposition method.
Because nitrogen compounds easily decompose, test shall be carried out immedi-
ately after sampling. When immediate test is impossible, it should be kept accord-
ing to 3.3, and then tested a s soon as possible.
39.1 Sum total method Carry out distillation after adding sodium hydroxide t o
a sample, eliminate ammonia produced by decomposition of ammonium ion and of
partial organic nitrogen compound, add Devarda’s metal, reduce nitrite ion and ni-
trate ion into ammonia, separate it by distillation, and determine the amount of the
nitrogen by Indophenol Blue absorptiometry. Separately, add copper sulfate, potas-
sium sulfate, sulfuric acid into sample, decompose it thermally, change organic ni-
trogen to ammonium ion, distill it after making it alkaline, separate it together with
the ammonium ion that has been contained already in the sample, and determine
the quantity of nitrogen owing to Indophenol Blue absorptiometry. Calculate con-
centration of total nitrogen after making i t together with the nitrogen correspond-
ing t o nitrite ion and nitrate ion obtained in advance.
Determination range: N 8 t o 160yg
(1) Reagents Reagents shall be as follows.
Sulfuric acid (25 mmol/Z) Follow 36.1.2 (1) (b).
Sulfuric acid (1+35) Follow 36.1.2 (1)(c).
Sodium hydroxide solution (40 glZ) Follow 19 (1)(g).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Sodium hydroxide solution (300 g/Z) Follow 37.2.1 (1)(e).
Devarda’s metal Follow 37.2.1 (1)(f).
Potassium sulfate Specified in JIS K 8962.
Copper (II) sulfate pentahydrate Specified in JIS K 8983. Use it un-
der powdered condition.
Sodium phenoxide solution Follow 36.2 (1)(d).
Sodium hypochlorite solution (effective chlorine 10 glZ) Follow 36.2
(1)( e ) .
Phenolphthalein solution (5 g/Z) Follow 13.2 (1)(a).
Ammonium ion standard solution (1 mgNH4+/ml) Follow 36.2 (1)(f).
(m) Ammonium ion standard solution (10 pgNH4+/ml) Follow 36.2 (1)(g).
230
K O101 : 1998
(2) Apparatus Apparatus shall be as follows.

(a) Glassware Wash sufficiently with water prior t o use.
(b) Kjeldahl flask 200ml Wash sufficiently with water prior t o use.
(c) Distilling apparatus Follow 36.1.2 (2) (a). Wash sufficiently with wa-
ter prior t o use.
(d) Photometer Spectrophotometer or photoelectric photometer
(3) Operation Operations shall be as follows.
Take 50 ml(1) of sample, and when it is not neutral adjust it to pH about
7 using sodium hydroxide solution (40g/Z) o r sulfuric acid (1+35).
Carry out the operations shown in 37.2.1 (3)(b)t o (f).
Pipet 25 m1P) out of the distillate solution of the sample, into a 50 ml volu-
metric flask.
Carry out the operations shown in 36.2 (3)(b)t o (e) t o measure the absor-
bance.
Carry out a blank test as follows: Take about 50 ml of water, after adding
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
10 ml of sodium hydroxide solution (300 g/Z) carry out the operations shown
in 37.2.1 (3)(d) t o (f), measure the absorbance owing t o the operations of
( e ) and (d) on the distillate solution, and correct the absorbance obtained
a t (d).
Making use of the working curve prepared in 36.2 (31,found the amount of
ammonium ion (mg) contained in a 25 ml aliquot taken from the distillate
solution obtained in ( e ) .
Separately, take 50ml(3) of sample in a 200ml Kjeldahl flask, and carry
out the operations in 38.1 (3)( e ) to (f).
Pipet 25 ml(2) of this distillate solution into a 50 ml volumetric flask, and
measure the absorbance after operations of 36.2 (3)(b)t o (e).
Carry out a blank test as follows: Take 50 ml of water, carry out the op-
erations shown in ( g ) and (h), and correct the absorbance obtained at (h).
Making use of the working curve prepared in 36.2 (3),find the amount of
ammonium ion (mg) contained in a 25 ml aliquot taken at (h).
Calculate the concentration (mgN/Z) of total nitrogen in the sample in ac-
cordance with the following formula (4).
1000 200 1000 200
N=ax- X-x 0.776 6 + b X- x -x 0.776 6
50 25 50 25
where, N : total nitrogen (mgN/Z)
a : ammonium ion obtained at operation (f) (mg)
b : ammonium ion obtained a t operation 0’) (mg)
i-)
0.776 6 : coefficient to convert ammonium ion t o equivalent
nitrogen 14.01
18.04
231
K O101 : 1998
Notes (1) In the case of determining total nitrogen with low concentration,
increase the amount of sample. In this case, however, use a x - 1O00
V
instead of ax- 'Ooo in the formula shown in (k).
50
where, V : sample (mi)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(2) When 200 ml of distillate solution contains 0.8 mg o r more of
ammonium ion, take a suitable amount of distillate solution (con-
taining less than 0.4 mg of NHs+)into a 100 ml volumetric flask
in which 25 ml of sulfuric acid (25 mmolíl) has been put, add water
up to the marked line, and then pipet 25 ml out of this solution.
(3) When the sample containing a low concentration of total nitro-
gen shall be tested, increase amount of the sample and carry out
the operations shown in 38.1 (3)(a)and (b). In this case, however,
use bx- 'Ooo instead of bx- 'Ooo in the formula shown in (k).
V 50
where, V : sample (mi)
(4) When the operations in ( c ) or (h) are carried out according t o
Note (2), use ax- 100 o r b x - 100 instead of a o r b in the formula
C d
shown in (k)respectively. c or d represents the amount of solu-
tion taken in a 100 ml volumetric flask (mi) when carrying out
the operation in Note (2) respectively.
39.2 Ultraviolet absorptiometry Add alkaline potassium peroxodisulfate solu-

tion in a sample, change nitrogen compounds to nitrate ion by heating it at about
120 O C , and dissolve organic compounds. Make pH of this solution 2 t o 3, and mea-
sure the absorbance by nitrate ion using 220 nm wavelength for determination. This
method can be applied t o the sample where organic substance is easily decomposed
and its content is rather small, and which does not contain bromide ion or chro-
mium enough to give hindrance t o determination.
Determination range: N 5 t o 50pg
Hydrochloric acid (1+16) Prepare using the hydrochloric acid specified
in JIS K 8180.
Hydrochloric acid (1+500) Prepare using the hydrochloric acid speci-
fied in JIS K 8180.
Sodium hydroxide-potassium peroxodisulfate solution Add 20 g of
sodium hydroxide for nitrogen compounds analysis specified in JIS K 8826
in 500 ml of water, and dissolve 15 g of potassium peroxodisulfate for ni-
trogen and phosphorus analysis specified in JIS K 8253. Prepare each time .
it is needed(5).
Nitrogen standard solution (0.1 mgN/ml) Heat potassium nitrate speci-
fied in JIS K 8548 at 105 to 110 "C for about 3 h, and let it cool in a des-
iccator, Dissolve 0.722 g of this in a little amount of water, transfer it in
a 1O00 ml volumetric flask, and add water up t o the marked line. Store it
in a dark place a t O t o 10 O C .
232
K O101 : 1998
(f) Nitrogen standard solution (20 pgN/ml) Pipet 50 ml of nitrogen stan-

dard solution (0.1 mgNíml) in a 250 ml volumetric flask, and add water up
t o the marked line. Prepare each time it is needed.
Note (5) Nitrogen content of this solution should be 0.4 mglZ or less.
Decomposing bottle This should be ethylene tetrafluoride resin bottle
or heat-resisting and pressure-resisting glass bottle (capacity about 100 ml),
and be used in a high pressure steam sterilizer (about 120 "C)(6).
High pressure steam sterilizer Capable of heating at about 120 "C speci-
fied in JIS T 7322 or JIS T 7324.
Photometer Spectrophotometer
Absorption cell Made of quartz glass
Note (6) Instead, a glass ampule (capacity about 100 mi), which can be used
in a high pressure steam sterilizer (about 120 OC), may be em-
ployed.
Pipet 50 ml of the sample(') in a decomposing bottle.
Add 10 ml of sodium hydroxide-potassium peroxodisulfate solution, quickly
stopper it closely, and mix them.
Heat it in a high pressure steam sterilizer, and after it's temperature reaches
about 120 O C , heat it for 30 min t o decompose.
Take out the bottle from the sterilizer, and let it cool,
Pipet 25 ml of supernatant(8) in a 50 ml beaker.
Add 5 ml of hydrochloric acid (1+16)(9),and adjust its pH to 2 t o 3.
Put a part of the solution in an absorption cell, and measure the absor-
bance at 220nm wavelength.
Place 50 ml of water in a decomposing bottle to make a blank test, mea-
sure its absorbance after the operations from (b)to ( g ) , and correct the
Find the quantity of total nitrogen in the solution pipetted at (e) making
use of the working curve, and calculate the concentration (mgNIZ) of total
nitrogen in the sample according to the following formula (10).
60 1000
N=ax25x50
a : total nitrogen in 25 ml solution pipetted at ( e )(mg)
Working curve Pipet step by step 1 to 10 ml nitrogen standard solution
(20 pgN/ml) in as many 100 ml volumetric flasks, and dilute them with water
up to the marked line. Respectively, pipet its 25 ml into a 50 ml beaker,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
233
K O101 : 1998
add 5 ml hydrochloric acid (1+500), transfer a part of it in an absorption

cell, and measure its absorbance with 220 nm wavelength. Separately, take
25 ml of water in a 50 ml beaker for blank test, add 5 ml of hydrochloric
acid (1+500), measure its absorbance a t 220 nm wavelength, and correct
the absorbance obtained on the nitrogen standard solution. Draw the re-
lation curve between the absorbances and the quantities of nitrogen (N) in
pipetted 25 ml solutions.
Notes (7) When total nitrogen in 50 ml sample is 0.1 mg or more, take a
suitable amount of sample (contains less than 0.2 mg as N ) in a
100ml volumetric flask, add water up t o the marked line, and
then use it. Provided that, when total nitrogen in 50 ml sample
is 0.1 mg or more and its pH is out of 5 to 9, take a suitable
amount of the sample (contains less than 0.2 mg as N), neutral-
ize it with hydrochloric acid (1+11)o r sodium hydroxide solu-
tion (40gíZ), transfer into a 100 ml volumetric flask, and add water
(8) Be careful not t o contain precipitation of hydroxide. If neces-
sary, filtrate through glass fiber filter paper with l p m or less
pore diameter, and use the filtrate after discarding first 5 t o 10 ml
filtrate.
(9) When the precipitation of hydroxide takes place in the solution
obtained after decomposition, take 25 ml of filtrate at Note ( S ) ,
add 5 ml of low-concentrated hydrochloric acid answering t o the
quantity of hydroxide, and then adjust its pH to 2 t o 3.
(10) When the sample prepared by the operation in Note (7) is employed
at the operation in (a),calculate the concentration (mgNIZ) of to-
tal nitrogen in the sample according to the following formula.
60 1000 100
N=ax-x- -
25 50 V
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
a : total nitrogen in 25 ml solution taken at (e)

(mg)
V : sample (mi)
Remarks When total nitrogen in a 50 ml sample is less than 0.1 mg and
its pH is out of 5 to 9, take a suitable amount of sample (50
to 100ml), neutralize it with hydrochloric acid (1+11)or so-
dium hydroxide solution (40glZ) (Record the amount of solu-
tion required for neutralization.), take 50 ml from this solution
into a decomposing bottle, and carry out the operations in (3)(b)
t o ( e ) . Provided that in this case the following formula should
be used.
60 1000 V + b
N=ax-x-x-
25 50 V
where, N : total nitrogen (mgN1Z)
a : total nitrogen in 25 ml sample pipetted af-
ter pretreatment (mg)
234
K O101 : 1998
b : hydrochloric acid (1+11)and sodium hydrox-

ide solution (40gll) needed for neutraliza-
tion (mi)
V : sample (ml)
2 When the concentration of total nitrogen in the solution moved
into an absorption cell at (3)(g)is less than 0.4mgl2, use a
50 mm absorption cell. Provided that a 50 mm cell should be
used for both a blank test and the preparation of working curve.
For the preparation of working curve, take step by step 1 to
10 ml of the nitrogen standard solution (4 pgN/ml), which has
been prepared by diluting five times the nitrogen standard
solution (20 pgN/ml), and prepare the working curve similarly
to the working curve drawn a t (3).
3 In this method, even the presence of 10 mgll of bromide ion
or 0.1 mgll of chromium may disturb the test. Such a sample
shall not be treated with this method.
39.3 Hydrazinium sulfate reducing method Add alkaline solution of potassium

peroxodisulfate in the sample, heat it at about 120 "C to change nitrogen compound
into nitrate ion and decompose organic substance. Reduce the nitrate ion in this
solution t o nitrite ion by hydrazinium sulfate with copper catalyst, and determine it
by an absorptiometry with naphthylethylenediamine to find the concentration of total
nitrogen. This method can be applied t o the case where organic substance in the
sample is easily decomposed and the sample is a little in quantity.
Determination range: N 0.33 t o 3.3yg
Repeatability: 3 to 10 % by coefficient of variation
(b) Sodium hydroxide-potassium peroxodisulfate solution Follow 39.2
(1)(d).
(c) Copper-zinc solution Dissolve 0.08 g of copper (II) sulfate pentahydrate
specified in JIS K 8983 and 1.76g of zinc sulfate heptahydrate specified
in JIS K 8953 in water up t o 200 ml, and dilute its 5 ml with water up t o
250 ml.
(d) Hydrazinium sulfate solution (7 glZ) Dissolve 3.5 g of hydrazinium sulfate
specified in JIS K 8992 in water up t o 500 ml.
(e) Hydrazinium sulfate solution (0.7 gll) Dilute ten times the hydrazinium
sulfate solution (7 g l l ) w i t h water. Prepare each time it is needed.
(0 4-aminobenzenesulfonamidesolution Follow 37.1.1 (1) (b).
(g) N -1-naphthylethylenediammoniumdichloride solution Follow 37.1.1
(1) (cl.
(h) Nitrogen standard solution (20 pgN/ml) Follow 39.2 (1) (f).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
235
K O101 : 1998
(i) Nitrogen standard solution (4 pgN/ml) Pipet 20 ml of nitrogen stan-

dard solution (20pgNlml) in a 100ml volumetric flask, and dilute it with
water up t o the marked line. Prepare each time it is needed.
(a) Decomposing bottle Follow 39.2 (2) (a).
(b) Test tube with ground stopper Use it with the same material and shape.
(c) Water bath Capable of controlling its temperature at (35+1) "C
(d) High pressure steam sterilizer Follow 39.2 (2) (b).
(e) Photometer Spectrophotometer or photoelectric photometer

(a) Place 50 ml of sample(11)(12) in a decomposing bottle.
(b) Carry out the operations in 39.2 (3)(b)t o (d).
(c) Pipet 10 ml of supernatant(*)(13j in a test tube with ground stopper.
(d) Add 1 ml of copper-zinc solution, agitate it, add 1 ml of hydrazinium sul-
fate solution (0.7 gl0, agitate it, and immerse it in a water bath at (35+1) "C.
(e) Take it out from the water bath 2 h later, and cool it t o room temperature.
(f) Carry out the operations in 37.1.1 (3)(b) and ( c ) .
(g) Take 50 ml of water for a blank test in a decomposing bottle, carry out the
operations in (b)t o (0,
measure its absorbance, and correct the absorbance
obtained about the sample.
(h) Find the quantity of total nitrogen in 50 ml solution taken in a decompos-
ing bottle making use of the working curve ( I d ) , calculate the concentration
(mgNlZj of total nitrogen in the sample according to the following formula.
1 O00
N = ~ x -
50
a : total nitrogen in 50 ml solution taken in a
decomposing bottle (mg)
Working curve Pipet step by step 1 to 10 ml of nitrogen standard solu-
tion (4 ygNlml) in as many 100 ml volumetric flasks, and add water up to
the marked line respectively. Carry out the operations in (a)to (g) on this
solution, and prepare the relation curve between absorbances and the quan-
tities of nitrogen (N) in 50ml solution taken in a decomposing bottle.
Notes (11) Follow Note (7). Provided that the following formula should be
used t o calculate the concentration (mgNIZ) of total nitrogen in
the sample solution.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
236
K O101 : 1998
1000 100
N=~x-x-
50 V
V : sample (mi)
Follow Remarks 1. Provided that the following formula should
be used t o calculate the concentration (mgN/Z) of total nitrogen.
1000 V + b
N=~x-x-
50 V
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
b : hydrochloric acid (1+11) and sodium hydrox-
ide solution (40 glZ) required for neutraliza-
tion (mi)
V : sample (mi)
When quantity of nitrogen in 50 ml solution taken in a decom-
posing bottle is 20 pg or more, take a suitable amount (less than
30 pg as N) of supernatant into a 100 ml volumetric flask, add
5 ml of sodium hydroxide solution (40 g/Z), and add water up t o
the marked line.
When Note (13) is adopted a t the operation in (cl, obtain the
quantity of total nitrogen (mg) in 50ml sample taken in a de-
composing bottle owing to multiplication of the amount (mg) of
100
nitrogen found on the working curve by c. However, c means
the amount (ml) of supernatant taken in the 100 ml volumetric
flask.
Remarks 4 In case where the sample is sea water or the like, because
inorganic substance affects the reducing ratio of nitrate ion,
the following standard addition method shall be adopted.
Place 40 ml of sample in a decomposing bottle, and add 10 ml
of water. Hereafter, carry out the operation in (3)(b)to (g),
measure absorbance, and find the quantity (mg) of total ni-
trogen in 40ml sample making use of the below-mentioned
working curve. Separately, take 50 ml of water for a blank
test in a decomposing bottle, carry out the operation in (3)(b)
to (g), measure absorbance, and find the equivalent quantity
of nitrogen (mg) making use of the working curve in (3). Then,
calculate the concentration (mgN/Z) of total nitrogen in the
sample according to the following formula.
1O00
N = (a - b ) x -
40
237
K O101 : 1998
a : quantity of total nitrogen in 40 ml sample

(mg)
b : nitrogen obtained by blank test (mg)
Working curve Pipet step by step 1 t o 8 ml of nitrogen standard solu-
tion (4 pgNlml) in as many decomposing bottles, add respectively 40 ml of
sample, add water up to 50 ml, carry out the operations in 3 (b)to (f),measure
its absorbance, and correct the measured value by subtracting the absor-
bance given by 40ml sample from the above value.
In case of the sample, as sea water, containing a lot of magnesium ion,
it lowers pH of the solution after decomposing operation, and decreases the
reducing ratio owing to mingling a part of magnesium in supernatant o r
filtrate. Therefore the supernatant, whose pH has been adjusted to 12.6
to 12.8 by adding sodium hydroxide solution (40gll) into the solution after
decomposing, shall be used. When this operation is carried out, the calcu-
lating formula for total nitrogen shall be corrected because of dilution.
When the quantity of nitrogen in 40 ml sample is 10 pg o r more, take a
suitable amount of sample (less than 25 pg as N), carry out the operation
according t o Note ('), and pipet 40 ml from this solution in a decomposing
bottle.
When the quantity of nitrogen in 40ml sample is less than 1Opg and
its pH is out of 5 t o 9, carry out the neutralization according to Remarks 1,
and take 40 ml from this solution into a decomposing bottle. When these
operations are carried out, use this solution when preparing working curve,
and when calculating total nitrogen with formula, correct it because of
dilution, respectively.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
39.4 Copper and cadmium column reducing method Add alkaline solution
of potassium peroxodisulfate into a sample, and heat it about 120 "C t o make nitro-
gen compounds to nitrate ion and to decompose organic substance. Reduce the ni-
trate ion in this solution owing to copper and cadmium column to nitrite ion, determine
it using an absorptiometry with naphthylethylenediamine, and obtain the concen-
tration of total nitrogen. This method can be applied t o the case where organic sub-
stance in sample is small and easily decomposed.
Determination range: N 0.2 t o 2 y g
Repeatability: 3 t o 10 % by coefficient of variation
(b) Hydrochloric acid (1+11) Prepare using the hydrochloric acid specified
in JIS K 8180.
(c) Ammonium chloride-ammonia solution Follow 37.2.3 (1) (e).
(d) Sodium hydroxide-potassium peroxodisulfate solution Follow 39.2
(1) (d).
(e) Column activation solution Follow 37.2.3 (1) (d).
(0 Copper and cadmium column packing Follow 37.2.3 (1) (e).
238
K O101 : 1998
Column packing solution Follow 37.2.3 (i)(f).

4-aminobenzenesulfonamidesolution Follow 37.1.1 ( i )(b).
N-1-naphthylethylenediammoniumdichloride solution Follow 37.1.1
(1)(cl.
Nitrogen standard solution (0.1 mgN/ml) Follow 39.2 (i)(e).
Nitrogen standard solution (2 pgN/ml) Pipet 10 ml of nitrogen stan-
dard solution (0.1 mgN/ml) in a 500 ml volumetric flask, and add water up
t o the marked line. Prepare each time it is needed.
(a) Decomposing bottle Follow 39.2 (2) (a).
(b) High pressure steam sterilizer Follow 39.2 (2)(b).
( c ) Copper and cadmium column Follow 37.2.3 (2) (a).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

Place 50 ml of sample(l5)(16) in a decomposing bottle.
Carry out the operations in 39.2 (3)(b) to (d).
Add 10 ml of hydrochloric acid (1+11)in the decomposing bottle, agitate it,
and transfer the solution in a 100 ml volumetric flask(l7).
Wash inside wall of the bottle several times with a little water, and put
the washings into the flask of (e).
Add 10 ml of ammonium chloride-ammonia solution, and add water up to
marked line t o make the solution for reduction.
Carry out the operations in 37.2.3 (3)( c ) and (d).
Take 50 ml of water for a blank test in a decomposing bottle, carry out the
operations in (b) to (f), measure its absorbance, and correct the absorbance
obtained about the sample.
Find the quantity of total nitrogen in the solution for reduction on the working
curve, calculate (18) the concentration (mgNIZ) of total nitrogen in the sample
according to the following formula.
1O00
N=ax-
50
a : total nitrogen in 100 ml solution for reduction (mg)
Working curve Pipet step by step 1 to 10 ml of nitrogen standard solu-
tion (2pgN/ml) in as many 100 ml volumetric flasks, carry out respectively
the operations in (e) and (f),and measure its absorbance. Separately, take
about 50 ml of water in a 100 ml volumetric flask, carry out the operations
(e) and (f), measure absorbance, correct the absorbance obtained on the
239
K O101 : 1998
nitrogen standard solution (2 pgN/ml), and prepare the relation curve be-
tween absorbances and quantities of nitrogen (N).
Notes (15) Follow Note ( 7 ) . Provided that the following formula should be
used to calculate the concentration (mgN/Z) of total nitrogen in
the sample.
1000 100
N=ax-x-
50 V
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
a : total nitrogen in 100 ml of solution for re-

duction (mg)
V : sample (ml)
(16) Follow Remarks 1. Provided that the following formula should
be used to calculate the concentration (mgN/Z) of total nitrogen
in the sample.
1000 V+b
N=ax-x-
50 V
a : total nitrogen in 100 ml of solution for re-
duction (mg)
b : hydrochloric acid ( 1+11)and sodium hydrox-
ide solution (40 glZ) needed for neutraliza-
tion (mi)
V : sample (ml)
(17) When the total nitrogen in 50 ml sample taken in a decompos-
ing bottle at (a) is 20 pg or more, use a 200 to 500 ml volumet-
ric flask, add ammonium chloride-ammonia solution at operation
(e)so as to make its volume 10 ml per 100 ml of final liquid vol-
ume, and add water up to the marked line to prepare the solu-
tion for reduction.
(18) When Note (17) is carried out in the operation of (c),correct owing
to multiplying the formula by &, where c is the volume of the
volumetric flask employed (mi).
39.5 Thermal decomposition method Decompose thermally nitrogen compound

in the sample to produce ammonia or nitrogen, and determine it. Otherwise, after
converting them into nitrogen monoxide, determine nitrogen by chemiluminescence.
Finally obtain total nitrogen by either way.
Determination range: N 1 to 200mgA
Repeatability: 3 to 10 % by coefficient of variation (depending on apparatus and
measuring condition)
(i) Reagents Reagents shall be as follows.
240
K O101 : 1998
(b) Total nitrogen standard solution (0.2 mgN/ml) Heat potassium nitrate
specified in JIS K 8548 at 105 to 110 "C for about 3 h, and let it be cooled
in a desiccator. Dissolve its 1.444 g in a little water, transfer it into a 1 O00 ml
volumetric flask, and add water up to the marked line. Keep it in a dark
place at O t o 10°C.
(a) Microsyringe 20 t o 1 5 0 ~ 1
(b) Homogenizer or mixer
(c) Total-nitrogen analyzer
(3) Preparatory operation Preparatory operations shall be as follows.
Get ready for running a total-nitrogen analyzer.
Inject a definite amount (for instance: 20 pl) of total-nitrogen standard solution
(0.2 mgN/ml) into a sample injection port of the total-nitrogen analyzer with
a microsyringe, and control the sensitivity of the analyzer so as t o make
an indicated value (peak height) about 80 % of the maximum graduated
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
scale.
Repeat the operation in (b), and confirm the indicated value shows con-
stant.
Agitate the sample(l9) enough t o make it uniform, inject its definite amount
[for instance, the same amount as in (b)] with a microsyringe at sample
injection port, read an indicated value, and find the rough concentration of
total nitrogen in the sample comparing with (c).
Note (19) When the sample gives 200 mgN/Z or more concentration of total
nitrogen, dilute it suitably with water to submit to be tested.
(4) Preparation of working curve Working curve shall be prepared as follows.

Pipet step by step total-nitrogen standard solution (0.2 mgN/ml) in as many
100ml volumetric flasks so as to make the rough concentration of total
nitrogen in the sample found in preparatory operation (3)(d)to position
centrally, add water up to the marked line, and prepare total-nitrogen stan-
dard solutions with various concentration.
Inject a definite amount [for instance, the same amount as in (3)(b)]of
the highest concentrated one among total-nitrogen standard solutions pre-
pared in (a) with a microsyringe at the sample injection port, and control
the sensitivity of the total-nitrogen analyzer so as t o make the indicated
value show 80 % of the maximum scale.
Inject successively a definite amount [specified in (b)]of variously concen-
trated total-nitrogen standard solution prepared in (a)with a microsyringe
a t a sample injection port, and read indicted values,
Take the same amount of water as in (cl with a microsyringe for a blank
test, find an indicated value similarly to (cl, correct the indicated values
a t ( c ) , and draw a relation curve between indicated values and the concen-
trations of nitrogen (N).
241
K O 1 0 1 : 1998
( 5 ) Operation Operation shall be as follows.

When sample contains suspension, agitate it using a homogenizer or mixer
t o make it disperse uniformly.
Inject a definite amount [for instance, the same amount as in (4)(b)]of
the sample with a microsyringe at the sample injection port, and read an
indicated value.
Take the same amount of water as in (b)for a blank test with a microsyringe,
carry out similarly to (b) to read an indicated value, and correct the indi-
cated value in (b).
Making use of the working curve previously prepared, find the concentra-
tion of total nitrogen in the injected sample, and calculate the concentra-
tion (mgNII) of total nitrogen in the sample.
Remarks 5 There are following types in total nitrogen analyzer. The
ammonia generating and determining method is as follows: de-
compose thermally the sample in hydrogen gas flow, convert
total nitrogen into ammonia through catalyst, and determine
it by coulometric titration method or electric-conductivity
measuring method.
The nitrogen generating and determining method is as fol-
lows: decompose thermally the sample in helium gas flow,
convert total nitrogen into nitrogen gas through catalyst, and
then determine using a electric conductivity measuring method.
The determining method by a chemiluminescence type is
as follows: decompose thermally the sample in oxygen gas flow
in order t o make total nitrogen into nitrogen monoxide, react
it with ozone, and measure the chemiluminescence (650 t o
900 nm wavelength) which can be produced with it is oxidized
into nitrogen dioxide.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
242
K O101 : 1998
40 Sulfide ion (S2-) For the determination of sulfide ion, methylene-blue absorp-
tiometry or iodometry shall be adopted.
Because sulfide ion is so unstable that it is easily oxidized or dispersed in air as
hydrogen sulfide, it is necessary t o carry out testing immediately after sampling. If
immediate testing after sampling is impossible, it shall be kept in accordance with
3.3 and be tested as soon as possible.
40.1 Methylene-blue absorptiometry Measure the absorbance of methylene blue

[3,7-bis(dimethylamino)phenothiazine-5-iuml, which is produced by the reaction be-
tween sulfide ion and N,N’-dimethyl-p-phenylenediamine under the presence of iron
(III) chloride, and determine sulfide ion.
Determination range: S2- 5 t o 40 pg

Sulfuric acid ( l + l ) Described as 4.4 (1)(b).
N,N’-dimethyl-p-phenylenediammonium solution Add 0.8 g of N ,N y -
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
dimethyl-p-phenylenediammoniumdichloride in sulfuric acid (1+1)to make
total 100ml solution. Prepare this each time i t is needed.
Iron (III) chloride solution Dissolve 10 g of iron (III) chloride hexahy-
drate specified in JIS K 8142 in water t o make total 100ml.
Diammonium hydrogenphosphate solution (400 g/Z) Dissolve 40 g of
diammonium hydrogenphosphate specified in JIS K 9016 in water to make
total 100ml.
Sulfide ion standard solution (1mgS2-/ml) Take 7.6 g of sodium sul-
fide nonahydrate crystal specified in JIS K 8949, wash its surface with a
little water, put it on filter paper t o remove moisture, dissolve it in oxy-
gen-free water of 2 (12) (a) to make 1I, and transfer into an airtight ves-
sel. Standardize this each time it is needed.
Standardization Pipet 20 ml of iodine solution (50 mmol/Z)(l)into a 300 ml
Erlenmeyer flask with ground stopper, and add 0.5 ml of hydrochloric acid
(l+l). Then pipet 20 ml of the sulfide ion standard solution with a whole
pipet, and add this into the iodine solution with touching its tip in the iodine
solution(2). Immediately stopper it tightly, swirl it, and let it stand for
few minutes. Titrate it with 0.1 mol/Z sodium thiosulfate solution(3), when
yellow colour of the solution becomes faint add 1ml of starch solution as
indicator (10 g/Z) (4), and titrate until the blue colour by starch solution dis-
appears. Separately, take 20 ml of iodine solution (50 mmol/Z) into a 300 ml
Erlenmeyer flask with ground stopper, add 0.2 ml of hydrochloric acid (l+l),
and titrate it with 0.1 mol/Z sodium thiosulfate solution. Calculate the
quantity of sulfide ion in 1 ml of sulfide ion standard solution in accordance
with the following formula.
1
S = ( b - U ) x f x -x 1.603
20
where, S : sulfide ion standard solution (mgS2-/ml)
243
K O101 : 1998
a : 0.1 mol/Z sodium thiosulfate solution needed for

titration (mi)
b : 0.1 moVZ sodium thiosulfate solution equivalent t o
20 ml of iodine solution (50 mmol/Z) (ml)
f : factor of 0.1 mol/Z sodium thiosulfate solution
1.603: quantity of sulfide ion equivalent t o 1 ml of
0.1 mol/Z sodium thiosulfate solution (mg)
(0 Sulfide ion standard solution (10pgS2-/ml) When it is needed, take
10 ml of sulfide ion standard solution (i mgS2-/ml)of (e) in a 1O00 ml volu-
metric flask, and add oxygen-free water of 2 (12)(a)up to marked line. Cal-
culate the concentration of this solution on the base of the concentration of
sulfide ion standard solution ( i mgS2-/ml) of (e).
Notes (1) Described as 24.1 (i)(d).
(2) Add sulfide ion standard solution (imgS2-/ml)into iodine solu-
tion acidified with hydrochloric acid previously.
(3) Follow 22.1.2 (i)(d).
(4) Follow 22.1.2 ( i )(i),
Take a suitable amount of sample(5) ( 6 ) (containing 5 to 40 pg as S2-) in a
50 ml measuring cylinder (with a stopper), add oxygen-free water of 2 (12)(a)
to make total about 40 ml, add 1ml(7) of sulfuric acid (l+l), add the water
of the same kind up to 50 ml marked line.
Add 0.5 ml of N,W-dimethyl-p-phenylenediammonium solution, mix by shak-
ing it, add 1ml of iron (III) chloride solution, agitate it, and let it stand for
approx. 1 min.
Add 1.5 ml diammonium hydrogenphosphate solution (400 g/Z), mix by shaking
it, and let it stand for approx. 5 min.
Separately, take 1 ml of sulfuric acid ( l + l )in a 50 ml of measuring cylin-
der (with a stopper), add water up t o 50 ml marked line, and carry out the
operations in (b)and ( c ) .
Put the solution of (c) in an absorption cell, and measure its absorbance in
the vicinity of 670nm wavelength with making the solution of (d)refer-
ence solution.
Find the quantity of sulfide ion on the working curve, and calculate the
concentration (mgS2-/Z)of sulfide ion in the sample.
Working curve Pipet step by step 0.5 t o 4 ml of sulfide ion standard so-
lution (10 pgS2-/ml)into as many 50 ml measuring cylinders (with a stop-
per), carry out the operations of (a) t o (e),and prepare the relation curve
between absorbances and the quantities of sulfide ion (S2-).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
244
K O101 : 1998
Notes (5) In case of determining dissolved sulfide ion, filtrate it through

filter paper 5 grade C or 6 grade immediately after sampling,
and employ the filtrate after discarding about first 50 ml of fil-
trate.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
When the test cannot be carried out immediately after sampling,

either treat it by storing according t o 3.3 o r treat to fix as zinc
sulfide according t o Remarks 2. When treated according t o Re-
marks 2, separate sulfur as hydrogen sulfide from zinc sulfide
according t o 40.2 (3), and then carry out operations. In this case,
however, in order to absorb hydrogen sulfide, use sodium hydroxide
solution (20 mmol/Z) instead of zinc acetate solution.
('1 The colouring can be strongest when pH is 0.4 to 1.0. If the sample
is alkaline, neutralize with sulfuric acid (l+l),
and then add further
1ml of sulfuric acid (l+l).
Remarks 1 This method has considerably few disturbing coexisting ma-
terials, but oxidizing materials and reducing materials disturb.
Sulfite ion and thiosulfate ion disturb if 10mglZ o r more ex-
ists in it. Thiocyanate ion gives disturbance even if a very
small amount.
2 The fixing treatment of sulfide ion as zinc sulfide is as fol-
lows.
Prepare solution of dissolving 20 g of zinc sulfate hepta-
hydrate specified in JIS K 8953 in 100 ml of water and solu-
tion of sodium carbonate (100 g/Z), mix them in equal volume
each time it is needed, and prepare the suspensoid of basic
zinc carbonate. Employ an incubation bottle of 19 (2) (a)as a
sample container, take sample in it with care not t o leave
bubbles in, add the suspensoid of basic zinc carbonate by about
2 ml per 100 ml of sample, stopper tightly with no bubble in-
side, and then mix them by tumbling. Suspensoid 10ml of
basic zinc carbonate can fix about 50 mg of sulfide ion.
Successively, separate the precipitation through filter paper
5 grade C or a centrifuge, and carry out the test on this pre-
cipitation. Owing t o these operations, sulfite ion and thiosul-
fate ion coexisting in the sample can be separately determined.
The amount of sample for calculation of sulfide-ion concen-
tration in sample shall be the value given by subtracting added
volume (ml) of suspensoid of basic zinc carbonate from the ca-
pacity (ml) of the incubation bottle.
40.2 Iodometry Add a definite excess amount of iodine solution and hydrochloric
acid into the solution containing sulfide ion or sulfide, titrate remained iodine with
sodium thiosulfate solution with starch solution as indicator, and determine sulfide ion.
Determination range: S2- 0.2 mg o r more
Remarks 3 If a sample is directly titrated, the reducing material as sulfite ion
or thiosulfate ion is determined as sulfide ion because of similar
reaction, therefore the previous separation of sulfide ion is necessary.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`--- 245
K O101 : 1998

Hydrochloric acid Specified in JIS K 8180.
Sulfuric acid ( l + l )Follow 4.4 ( i )(b).
Hydroxylammonium chloride solution (100gll) Dissolve 10 g of hydro-
xylammonium chloride specified in JIS K 8201 in water to make total 100 ml,
Zinc acetate solution Dissolve 24 g of zinc acetate dihydrate specified
in JIS K 8356 in water to make total 100ml.
Starch solution (10 g l l ) Follow 22.1.2 ( i )(i).
Iodine solution (5 mmol/Z) Dissolve 50 ml of iodine solution (50 mmol/Z)
of 24.1 (i)(b)in water to make 500 ml. Preserve it in a coloured glass bottle.
10 mmol/Z sodium thiosulfate solution Follow 28.3 ( i )(e).
Nitrogen High purity grade 2 nitrogen specified in JIS K 1107.
(a) Generating and absorbing apparatus for hydrogen sulfide The ex-
ample of this apparatus shall be as shown in Fig. 40.1(8).
246
K O101 : 1998
Unit: mm
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
A: Round-bottom flask with interchange- F: Trapping sphere (Kjeldahl

able ground stopper 300ml or type)
1O00 ml GI, Gz : 200 ml Erlenmeyer flask with
B: Introducing pipe with connector interchangeable ground stop-
C: Side-arm connector Per
D: Funnel for pouring H: Rubber t u b e
E: Ground cock I: Interchangeable ground p a r t
Note (8) In case of directly generating hydrogen sulfide from a sample, flask
(A) should be 1 O00 ml, and in case of generating from fixed zinc
sulfide, it should be 300 ml.
Fig. 40.1 Example of generating and absorbing
apparatus for hydrogen sulfide
(3) Separating operation Separating operations shall be as follows.
(a) Dilute 5 ml of zinc acetate solution with water up t o 100 ml, and put 50 ml
each in Erlenmeyer flasks (GI) and (G2).
247
K O101 : 1998
Place a suitable amount of sample(5) (9) (usually 500 ml) in a flask (A), and
if iron (III) is contained add either 1ml of hydroxylammonium chloride
solution (100 gll) or 0.1 g of L(+)-ascorbicacid specified in JIS K 9502. Then
pour 100 ml of sulfuric acid (l+l) from an above fixed pouring funnel,
Heat the flask (A) at about 50 O C , flow nitrogen (or carbon dioxide) slowly
for about 20 min, expel hydrogen sulfide, and let it be absorbed in zinc acetate
solution.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Note (9) When hydrogen sulfide is generated from the separated precipi-
tation which is fixed as zinc sulfide as shown in Remarks 2, transfer
the precipitation with water into the flask (A) together with fil-
ter paper, and add about 50 ml of water. Then add 50 ml of hy-
drochloric acid (l+l)from a pouring funnel and hereafter carry
out as shown in (c).
Remarks 4 If bubbling is violent, add preferably defoaming agent as diphe-
nyl ether.
5 Passing carbon dioxide for about 1h can expel hydrogen sul-
fide without heating.
6 The method to generate hydrogen sulfide directly from sample
can separate sulfide ion from metal element as iron or zinc
besides thiosulfate ion, but the separation from sulfite ion is
not complete. The method shown in Note (9) gives previous
separation of sulfite ion and thiosulfate ion.
Add a definite amount of iodine solution (5 mmollt) in Erlenmeyer flasks

(GI) and (G2) in Fig. 40.1, in which hydrogen sulfide has been absorbed, so
as t o make amount of iodine solution be excess. Because amount of hydro-
gen sulfide is almost absorbed in the Erlenmeyer flask (GI), nearly all io-
dine solution (5 mmol/t) should be added t o the Erlenmeyer flask (GI).
Add respectively 2.5 ml of hydrochloric acid(10) into the Erlenmeyer flasks
(GI) and (Gd, agitate sufficiently, and transfer the contents into a 500 ml
Erlenmeyer flask. Wash the Erlenmeyer flasks (GI) and (Gd with water,
and put these washings together with the solution.
Titrate it with 10 mmollE sodium thiosulfate solution, when iodine colour
of yellow becomes faint, add about 1ml of starch solution (10 glt) as indi-
cator, and titrate until blue colour by starch solution disappears.
Take 100 ml of water in a 500 ml Erlenmeyer flask for a blank test, add
5 ml of zinc acetate solution and iodine solution (5 mmoVZ) by the same amount
as the test, add further 5 ml of hydrochloric acid, and carry out the opera-
tion of (cl.
Calculate the concentration (mgS2-lZ)of sulfide ion in the sample in accor-
dance with the following formula.
S = ( b - a )x f XEX
V 0.160 3
where, S : sulfide ion (rngS2-lZ)
248
K O101 : 1998
a : 10 mmol/E sodium thiosulfate solution needed for

titration (mi)
b : 10 mmol/Z sodium thiosulfate solution needed for
titration of blank test (ml)
f : factor of 10 mmol/l sodium thiosulfate solution
V : sample (mi)
0.160 3 : sulfide ion equivalent t o 1ml of 10 mmolll sodium
Note (10) First add iodine solution ( 5 mmol/Z), and add hydrochloric acid.
If the reverse is carried out, there is a fear to lose as hydrogen
sulfide.
Remarks 7 When the sample, containing a little disturbing substance, is
fixed as zinc sulfide according t o Remarks 2, the titration
without the separation treatment in (3)is permissible. In this
case, filtrate the precipitation resulted from fixation through
filter paper 5 grade C, wash it with water, transfer it into a
300 ml Erlenmeyer flask together with filter paper, and add
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
about 100 ml of water. Add a definite amount of iodine solu-
tion ( 5 mmol/Z), add 5 ml of hydrochloric acid, agitate sufficiently
to react them, hereafter carry out the operations in (e) t o (e),
and find the concentration of sulfide ion in the sample.
If fixed zinc sulfide precipitates in coloured condition, co-
existence of metal elements is thought, which may disturb the
titration, therefore they must be eliminated as hydrogen sul-
fide by the separation treatment in Note (9).
249
K O101 : 1998
41 Sulfite ion (S032-) For the determination of sulfite ion, iodometry shall be
adopted.
41.1 Iodometry After adding acetic acid-sodium acetate buffer solution into a
definite amount of iodine solution, add a sample, and titrate excess iodine with so-
dium thiosulfate solution with starch solution as indicator. Separately, take the same
amount of sample, acidify it, boil it to expel sulfite ion as sulfur dioxide, hereafter
carry out the same titration operations, and making this a blank test value, correct
the influence by reducing material as thiosulfate ion and so on. Because sulfite ion
is easily oxidized by air, carry out the test immediately after sampling.
Determination range: s0s2-0.2 mg or more
(a) Sulfuric acid (1+35) Prepare using the sulfuric acid specified in JIS K
8951.
(b) Sodium hydroxide solution (40 gll) Follow 19 ( i )(g).
(c) Acetic acid-sodium acetate buffer solution (pH 3.9) Dissolve 75 g of
sodium acetate trihydrate specified in JIS K 8371 in 500 ml of acetic acid
(1+2).
(d) Starch solution (10 g/Z) Follow 22.1.2 ( i >
(i).
(e) Phenolphthalein solution (5 gll) Follow 13.2 ( i )(a).
(f) Iodine solution (5 mmol/Z) Follow 40.2 ( i )(f).
(9) 10 mmolll sodium thiosulfate solution Follow 40.2 (i)(g).
(h) Nitrogen High purity grade 2 nitrogen specified in JIS K 1107.
Pipet 20 ml of iodine solution (5 mmol/Z) in an Erlenmeyer flask, and add
10 ml of acetic acid-sodium acetate buffer solution (pH 3.9).
Pipet a suitable amount of sample (containing 0.2 t o 10 mg as so32-),and
pour gently it into the flask with touching the tip of the pipet to the solu-
tion t o mix them well.
Titrate it with 10 mmol/2 sodium thiosulfate solution, add about 1ml of starch
solution (10 g/Z) as indicator when yellow colour of the solution becomes
faint, and titrate until the blue colour of the starch solution disappears.
Take the same amount of the sample as used for the test in an Erlenmeyer
flask for a blank test, add 6 t o 7 ml of sulfuric acid (1+35), boil gently for
several minutes while passing nitrogen gas(1) on the surface of the liquid
to expel sulfur dioxide, and then cool it while passing nitrogen as well.
After cooling, neutralize it with sodium hydroxide solution (40g/Z) using
phenolphthalein solution (5 g l l ) as indicator. Add 20 ml of iodine solution
(5 mmol/Z) and 10 ml of acetic acid-sodium acetate buffer solution (pH 3.9),
and titrate it with 10 mmol/Z sodium thiosulfate solution according t o the
operation in (cl.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
250
K O101 : 1998
(0 Calculate the concentration of sulfite ion (mgS032-/Z)in the sample accord-

S = ( b- a)x f x Ex 0.4003
V
where, S : sulfite ion (mgS032-/Z)
a : 10 mmol/Z sodium thiosulfate solution needed for
titration Cml)
b : 10 mmol/Z sodium thiosulfate solution needed for
f : factor of 10 mmol/Z sodium thiosulfate solution
V : sample (ml)
0.400 3 : sulfite ion equivalent t o 1ml of 10 mrnol/Z sodium
Note (1) In order to remove oxygen in nitrogen gas, pass the gas through
a gas washing bottle in which 75 ml of sodium hydroxide solution
(600 gil) and 15 ml of water containing previously 5 g of pyrogallol
(1,2,3-benzenetriol) specified in JIS K 8780 have been put,
Remarks 1 Sulfide ion makes disturbance because of consuming iodine,
and it cannot be corrected even by the blank test in (2)(d).
To remove the disturbance by sulfide ion, the following is ef-
fective; fix sulfide ion as zinc sulfide according t o Remarks 2
in 40, filtrate it through filter paper 5 grade C (or by centrifugal
separation), and determine sulfite ion using its filtrate.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
2 Iron (III) ion and copper (II) ion oxidize iodide ion t o disturb.
3 To sample from piping o r an apparatus? the sampler shown
in Fig. 41.1 is convenient. In case of using this sampler, sam-
pling for test shall be carried out as follows.
Connect lower ends of two samplers with the piping for sam-
pling via soft vinyl chloride tube and Y-type tube. Direct the
outlet of the sampler to upward, and no connection there. Con-
trol the flow rate of sample at the outlet of the sampler. When
temperature of sample is high, adjust it lower than room tem-
perature by 1 or 2 degrees.
Adjust flow rate t o fill both 2 samplers during 8 t o 12 s at
the same time, and flow the sample sufficiently through pip-
ing to wash piping and samplers and t o replace contents with
sample.
Next, shut the cock located a t upper end of 2 samplers, si-
multaneously shut the cock at lower end, disconnect connect-
ing tube, and confirm there remains no bubble owing t o the
reversing of the samplers. If there remain bubbles, discard
the sample from both samplers, and retry sampling. Use one
of them for test, and another for a blank test. In advance,
place 20 ml of iodine solution (5 mmoVZ) in a beaker, add 10 ml
of acetic acid-sodium acetate buffer solution (pH 3.9), discard
the sample kept in the legs located at both sides of the sampler
251
K O101 : 1998
for test, wash them with water, and fill them with 2 m l of
ethanol (95) specified in JIS K 8102. With care not to lose
the ethanol kept in the lower side as far as possible, immerse
the leg of the sampler in the sample in a beaker, and pour
the sample with quietly opening upper cock and lower cock.
Hereafter, carry out the test according to (2)(c) and the fol-
lowing items.
Unit: mm
Inside
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
_ x
A, B : Ground cock
Fig. 41.1 Example of sampler
252
K O101 : 1998
42 Sulfate ion (Sod2-) For the determination of sulfate ion, barium chromate-
diphenylcarbazide absorptiometry, barium chromate absorptiometry, gravimetry, or
ion chromatography shall be applied.
42.1 Barium chromate-diphenylcarbazideabsorptiometry Precipitate barium

sulfate by adding acidic suspensoid of barium chromate into a sample, add aqueous
ammonia containing calcium ion and ethanol, let the excess barium chromate pre-
cipitate, and then separate it by a centrifuge. Change chromate ion resulted from
displacement of sulfate ion into dichromate ion, react it with diphenylcarbazide (1,5-
diphenylcarbonohydrazide), and measure the absorbance of reddish violet generated
t o determine sulfate ion.
Determination range: Sod2- 2 to 50pg
(a) Hydrochloric acid (l+l> Prepare using hydrochloric acid specified in JIS
K 8180,
(b) Acidic suspensoid of barium chromate (A) Mix 100 ml of acetic acid
(1+15) and 100 ml of hydrochloric acid (1+500), add 0.5 g of barium chro-
mate, agitate sufficiently t o make it suspensoid, and store in a polyethyl-
ene bottle.
Prepare barium chromate as follows. Dissolve 8 g of potassium chro-
mate specified in JIS K 8312 in about 800 ml of water, add 10 ml of acetic
acid (6 mol/,!) (Dissolve 35 ml of acetic acid specified in JI$ K 8355 in wa-
ter t o make total 100 ml.), and heat it to about 70 "C. While stirring the
solution violently, pour drop by drop 100 ml of barium chloride solution (Dis-
solve l o g of barium chloride dihydrate specified in JIS K 8155 in water
to make total 100 ml.) heated at about 70 "C t o precipitate barium chro-
mate, and let it stand. Discard supernatant, and repeat decantation 4 times
using about 500 ml warm water respectively. Transfer the precipitation
into a precipitation tube for a centrifuge, and wash it 2 or 3 times with
cold water by centrifugal separation. Remove this precipitation in a glass
filter, filtrate with suction, heat it at 105 to 110 "C for about 1h, let it cool
in a desiccator, and grind it in an agate mortar.
(c) Aqueous ammonia containing calcium Dissolve 1.85 g of calcium chlo-
ride dihydrate specified in JI$ K 8122 in 500 ml of aqueous ammonia (3+4),
put it in a polyethylene bottle, and store it t o prevent it from invasion of
carbon dioxide in air. It is convenient to store as shown in Fig. 42.1.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
253
K O101 : 1998
/
i
/
A: Polyethylene bottle
500 ml
B: Buret (with side tube)
5 ml
rc c: One-way cock
A''' D: Absorption tube for
ol u'' carbon dioxide (packed

with soda lime)
E: Rubber stopper
~ I"
_. I __ - ___- F: Rubber tube
Fig. 42.1 Example of storing aqueous
ammonia containing calcium
Ethanol (95) Specified in JIS K 8102.
Diphenylcarbazide solution Dissolve 1g of 1,5-diphenylcarbonohydrazide
(diphenylcarbazide) specified in JIS K 8488 in 100 ml of ethanol (95). Prepare
this solution each time it is needed.
Sulfate ion standard solution (1 mgS042-/ml) Heat potassium sulfate
specified in JIS K 8962 at about 700 "C for about 30 min, and let it cool in
a desiccator. Weigh its 1.815 g, dissolve in a little amount of water, trans-
fer it into a 1 O00 ml volumetric flask, and add water up to marked line.
Otherwise, use sulfate ion standard solution S042-1 O00 specified in JIS K
0028.
Sulfate ion standard solution (10 ~gS04~-/ml) Pipet 10 ml of sulfate
ion standard solution (i mgSOP/ml) in a 1 O00 ml volumetric flask, and
add water up to marked line. Prepare this solution each time it is needed.
(a) Glass filter Büchner funnel type 3G4
(b) Centrifugal separator
(c) Precipitation tube for centrifuge 20 t o 30 ml with ground stopper
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
254
K 0101 : 1998
(d) P h o t o m e t e r Spectrophotometer or photoelectric photometer
(3) Operation Operation shall be as follows.

Put 10 ml of sample (containing 2 t o 50 pg a s Sod2-)
in a precipitation tube
for a centrifuge, and keep it at 20 t o 30 OC('). Add 4 ml of acidic suspen-
soid of barium chromate (A) kept 20 t o 30 O C , agitate it, and let it stand
for 2 t o 3 min.
Add gently about 1ml of supernatant of aqueous ammonia containing cal-
cium using a buret or pipet shown in Fig. 42.1, mix them, add 10 ml of
ethanol (95), and after agitating for 1 min, let it stand for about 1 0 min.
Centrifugalize it, move supernatant in a glass filter, and filtrate i t with
weak suction.
Add 2 ml of diphenylcarbazide solution and 1.4 ml of hydrochloric acid (1+1)
in the filtrate, agitate it, and let it stand for 5 min t o colour it.
Separately, take 10ml of water for a blank test, and carry out the opera-
tions in (a) t o (d)a t the same time as the sample.
Transfer a part of the solution obtained a t (d) in an absorption cell, and
measure its absorbance in the vicinity of 540 nm wavelength with making
the solution of (e) as reference solution.
Find the quantity of sulfate ion on the working curve, and calculate the
concentration of sulfate ion (mgS042-lZ)in the sample.
Working c u r v e Pipet stepwise 0.2 t o 5 ml of sulfate ion standard solu-
tion (10 pgS042-/ml)into as many precipitation tubes for a centrifuge, di-
lute respectively with water up t o 10 ml, carry out the operations in (a) t o
(f), and prepare the relation curve between absorbances and the quantities
of sulfate ion (SO4*-).
Note (1) The same working curve can be obtained within temperature range
from 20 t o 30 " C during reaction.
42.2 B a r i u m c h r o m a t e absorptiometry Add acidic suspensoid of barium chro-

mate in sample t o precipitate barium sulfate, add aqueous ammonia containing cal-
cium ion and ethanol in order to precipitate excess barium chromate, and centrifugalize
them. Measure the absorbance of yellow caused by chromate ion produced by the
replacement with sulfate ion, and determine sulfate ion.
Determination range: SO.,--
50 t o 500pg
( i ) Reagents Reagents shall be as follÓws.

(a) Acidic suspensoid of b a r i u m chromate (B) Add 2.5 g of barium chro-
mate(?)in 200 ml mixture of 100 ml of acetic acid (1+15) and 100 ml of hy-
drochloric acid (1+500), agitate well t o make suspensoid, and store it in a
polyethylene bottle.
Note (2) This is the barium chromate prepared in 42.1 (i)(b).
(b) Aqueous ammonia containing calcium Follow 42.1 (1)( c ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
255
K 0101 : 1998
(c) Ethanol (95) Specified in 31s K 8102.
(d) Sulfate ion s t a n d a r d solution (0.1 mgS042-/ml) Pipet 20 ml .of sulfate

ion standard solution ( imgS042-/ml)stated in 42.1 (1)(0 in a 200 ml volu-
metric flask, and add water up t o marked line.

(a) Centrifugal separator
(b) Precipitation tube for centrifuge 20 t o 30ml with ground stopper

Take 10 ml of sample (containing 50 t o 500 pg as S042-)in a precipitation
tube for a centrifuge, and keep it a t 20 t o 30 O C ( ’ ) . Add 4 ml of acidic sus-
pensoid of barium chromate (B)kept a t 20 t o 30 “C into it, agitate it, and
let it stand for 2 t o 3 min.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Add gently about 1ml of supernatant of aqueous ammonia containing cal-

cium using a buret or pipet shown in Fig. 42.1, mix them, add 10 ml of ethanol
(951, and after agitating for 1 min, stand it for about 1 0 min.
Centrifugalize it, take supernatant in an absorption cell, and measure its
absorbance in the vicinity of 370 nm wavelength.
Take 10 ml of water for a blank test, carry out the operations in (a)t o (c),
measure its absorbance, and correct the absorbance obtained on the sample.
concentration of sulfate ion (mgSO4”/Z) in the sample.
Working curve Pipet stepwise 0.5 t o 5 ml sulfate ion standard solution
(0.1 mgS042-/ml) into a s many precipitation tubes for a centrifuge, dilute
respectively with water up t o 10 ml, carry out the operations in (a)t o (d),
and draw the relation curve between absorbances and the quantities of sulfate
ion (sod2-).
Remarks 1 Any ion of nitrate, carbonate, and hydrogencarbonate, which
coexists by 50 mg/Z or more, disturbs. Any ion of phosphate,
arsenate, selenate, vanadate, and metal lead disturbs even if
very small amount, therefore they must be removed by pre-
treatment.
For removal of carbonate ion and hydrogencarbonate ion,
boil with adding hydrochloric acid. [In advance, take a part
of sample, neutralize with hydrochloric acid using mixture of
Methyl Red-Bromocresol Green ( * > as indicator, and calculate
amount of hydrochloric acid (1+100)t o neutralize it. Add the
same amount of hydrochloric acid (1+100)into the sample. Be
careful not t o add excess hydrochloric acid.]
In case of phosphate ion, add 2 ml of calcium chloride solu-
tion ( i l g/2) and 1 ml of equivolume mixture of sodium hydroxide
solution (10 g/Z) and sodium carbonate solution (13 gll) into 10 ml
256
K O101 : 1998
of sample. After 10min standing, centrifugalize it, take a

definite amount of supernatant, neutralize with hydrochloric
acid (1+100), heat it in a water bath for about 10min t o re-
move carbon dioxide. After cooling it, dilute with water up t o
20 ml, take 10 ml out of it, and determine sulfate ion accord-
ing to the operation in (3). The working curve for the case
where removal treatment of phosphate ion is carried out should
be made by using the sulfate ion standard solution that has
been treated similarly.
Note (*) Follow 13.1 (i)(a).
42.3 Gravimetry Precipitate sulfate ion as barium sulfate, and determine it by

means of weighing its mass.
Determination range: S042- 10mg o r more
Repeatability: 2 % by coefficient of variation
(a) Hydrochloric acid Specified in JIS K 8180.
(b) Hydrochloric acid (1+50) Prepare using hydrochloric acid specified in
JIS K 8180.
( c ) Barium chloride solution (100 g/Z> Dissolve 11.7 g of barium chloride
dihydrate specified in JIS K 8155 in water t o make total 100ml.
(d) Silver nitrate solution (10 gll) Dissolve 1g of silver nitrate specified
in JIS K 8550 in water to make total 100 ml.

Place a suitable amount (containing 10 mg or more as S 0 2 - ) of sample in
a porcelain evaporation dish, add 3 ml of hydrochloric acid, evaporate to
dryness on a boiling water bath, and heat it for about 20 min more.
After cooling, moisten it with 2 ml of hydrochloric acid, add 20 to 30 ml of
warm water, heat for several min, filtrate it through filter paper 5 grade
B, and wash it several times with hydrochloric acid (1+50).
Add water into the filtrate t o make total 100m1, warm on a water bath,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
pour drop by drop warmed barium chloride solution (100 g/Z) with constant
stirring, and when n o precipitation is generated, add excessively 20 t o 50 %
of added amount.
Warm for 20 to 30 min on a boiling water bath, and let it stand for 3 or
4 h.
Filtrate it through filter paper 6 grade or 5 grade C, and wash with water
until no reaction by chloride ion is found in the Filtrate [confirm by silver
nitrate solution (10 g/Z)I.
Put the precipitation together with the filter paper in a porcelain crucible
which has been made constant at 800 O C , dry it, heat gradually t o carbon-
ize the filter paper first, and then t o ash it.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
257
K 0101 : 1998
(g) Successively, heat a t 800 "C for about 30 min, let it cool in a desiccator,
and weigh its mass.
(h) Repeat the operation in (g) until constant weight is achieved.
(i) Calculate the concentration of sulfate ion (mgS042-/Z)in the sample in ac-
cordance with the following formula.
S=ax- 'Ooo ~0.4116

V
where, S: sulfate ion (mgS042-/1)
a: mass of barium sulfate (mg)
V: sample (mi)
0.411 6 : sulfate ion equivalent t o 1mg of barium sulfate
(mg)
42.4 Ion chromatography Sulfate ion in sample shall be determined using an

ion chromatography.
Determination range: sod2- 1 t o 100 mglZ(3)
Note (3) When combined with a suppressor, it becomes 0.2 t o 100 mgS042-/Z.
( i ) Reagents Reagents shall be as follows.
(a) Water Water A3 or A4 specified in JIS K 0557.
(b) Eluent Follow 32.5 ( i )(b).
(c) Reclaiming solution Follow 32.5 (i)( c ) .
(d) Sulfate ion standard solution (10 mgS042-/ml) Heat potassium sulfate
specified in JIS K 8962 at about 700 "C for about 30 min, and cool it in a
desiccator. Dissolve its 1.815 g in a little amount of water, transfer it in a
100 ml volumetric flask, and add water up t o marked line.
(e) Sulfate ion standard solution (i mgS042-/ml) Pipet 10 ml of sulfate ion
standard solution (10 mgS042-/ml) in a 100 ml volumetric flask, and add
water up t o marked line. Prepare this each time it is needed.
(f) Anion-mixed standard solution L(O.1 mgCl-, 0.5 mgNOz-, 0.5 mgBr-,
0.5 mgNOs-, 1 mgS042-)/ml] Follow 32.5 ( i )(f).
(2) Apparatus Apparatus shall conform t o 32.5 (2).
(3) Preparatory operation Preparatory operations shall be as described in 32.5 (3).

(a) Carry out the operations in 32.5 (4) (a) and (b).
(b) Read the indicated value(4) on the peak corresponding t o sulfate ion on the
chromatogram.
258
K O101 : 1998
(c) When the sample is diluted, carry out the operations in (a)and (b)on water
as the same amount of the sample, as a blank test, and correct the indi-
cated value(4) obtained on the sample.
(d) Find the concentration of sulfate ion on the working curve, and calculate
the concentration of sulfate ion (mgS042-/Z}in sample.
Working curve Pipet step by step 0.1 to 10 ml of sulfate ion standard
solution (1 mgS042-/m1)15)in as many 100 ml volumetric flasks, respectively
add water up t o marked line, carry out the operations in (a)and (b),and
read the indicated value (4) corresponding to each sulfate ion. Separately,
take water for a blank test, carry out the operations in (a)and (b),and
correct the indicated value corresponding to each sulfate ion, and draw the
relation curve between quantities of sulfate ion (S042-) and the indicated
value. The working curve shall be prepared when sample is measured.
(6) When anions other than sulfate ion are simultaneously tested, an-
ion-mixed standard solution [0.1 mgCl-, 0.5 mgNo,-, 0.5 mgBr-,
0.5 mgNOB-, 1mgS042-)/ml]shall be used.
Remarks 2 When concentration of sulfate ion is 1mglE, if bromide ion is
200 mg/Z or less and nitrate ion is 400 mg/Z or less, they do
not disturb the test.
3 Follow Remarks 11 of 32.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
259
K O101 : 1998
43 Phosphorus compound and total phosphorus Phosphorus compound means

the phosphorus contained in the phosphorus compounds existing in water, such as
phosphoric acid, polyphosphoric acid, animal matter, plant matter, and so on, and
classified into phosphate ion, hydrolytic phosphorus, and total phosphorus.
When testing the sample filtrated, it is classified into dissolved one and suspended
one.
Because phosphorus compounds are easily changed, test shall be carried out im-
mediately after sampling. If immediate testing is impossible, preserve it according
to 3.3, and test it as soon as possible.
43.1 Phosphate ion (Pod3-)
43.1.1 Molybdenum blue (ascorbic acid reduction) absorptiometry Carry out

the reaction of phosphate ion to hexaammonium heptamolybdate and potassium
tartoratoantimonate (III) to produce hetero-poly compound, reduce the hetero-poly
compound by L(+)-ascorbic acid, and measure the absorbance by molybdenum blue
to determine the phosphate ion.
Determination range: P043- 2.5 to 75 pg
Ascorbic acid solution (72 gll) Dissolve 7.2 g of L(+)-ascorbicacid specified
in JIS K 9502 in water to make total 100 ml. Store in a dark place a t O t o
10°C. Don't use the coloured solution.
Ammonium molybdate solution Dissolve 6 g of hexaammonium
heptamolybdate tetrahydrate specified in JIS K 8905 and 0.24 g of bis[(+)-
tartrato] diantimonate (III) dipotassium trihydrate specified in JIS K 8533
in about 300 ml of water, add 120 ml of sulfuric acid (2+1), dissolve 5 g of
ammonium amidosulfate(1) specified in JIS K 8588 in it, and add water
up t o 500ml.
Ammonium molybdate-ascorbic acid mixed solution Mix ammonium
molybdate solution and ascorbic acid solution (72 g l l ) t o make their volume
ratio 5:l. Prepare this mixed solution each time it is needed.
Phosphate ion standard solution (0.1 mgP043-/ml) Heat potassium
dihydrogenphosphate (for pH standard solution) Specified in JIS K 9007
a t (105I2)"C for about 2 h, and let it cool in a desiccator. Take its O. 143 3 g ,
dissolve it in a little water, transfer it in a 1O00 ml volumetric flask, and
add water up to marked line. Store it in dark place a t O t o 10 OC. Other-
wise, use Pod3-100 of reference material-standard solution-phosphate ion
Phosphate ion standard solution (5 ~gP04~-/ml) Pipet 10 ml of phos-
phate ion standard solution (0.1 mgP043-/ml) in a 200 ml volumetric flask,
and add water up to marked line. Prepare it each time it is needed.
Note (1) When sample does not coexist with nitrate ion or nitrite ion, ad-
dition of ammonium amidosulfate can be eliminated. (See Remarks
3.)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
260
K O101 : 1998
( 2 ) Apparatus Apparatus shall be as follows.

Take a suitable amount (containing 2.5 t o 75 pg as Pod3-) of sample(2) (3)
in a 25 ml measuring cylinder (with stopper), and add water up to the marked
line of 25 ml.
Add 2 ml of ammonium molybdate-ascorbic acid mixed solution, agitate it,
let it stand a t 20 t o 40 O C ( 4 ) for about 15 min.
Put a part of the solution in a n absorption cell, and measure the absor-
bance in the vicinity of 880 nm wavelength(5) (6).
Take 25 ml of water for a blank test, carry out the operations in (b)and (c),
Find the quantity of phosphate ion on the working curve, and calculate the
concentration of phosphate ion (mgP0d3-/Z)in the sample (7).
Working curve Pipet step by step 0.5 t o 15ml of phosphate ion stan-
dard solution ( 5 ~..tgPO~~-/ml)
in as many 25 ml measuring cylinders (with
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
stopper), carry out respectively the operations in (a) t o (d),and draw the
relation curve between quantities of phosphate ion (Po43-)and absorbances.
Notes (2) When dissolved phosphate ion is determined, use the sample which
was filtrated by 3.3.
(3) If sample is acidic, add 2 or 3 drops of p-nitrophenol solution
(1g/Z) [follow 43.2 (1) ( g ) ] as indicator, and neutralize it with
sodium hydroxide solution (40gll) until getting faint yellow,
Provided, if hydroxide precipitation by such as aluminium is pro-
duced neutralization should be stopped just before precipitating.
(4) This shall be the same colouring temperature as that at work-
ing curve preparing.
(5) When the photometer cannot measure the absorbance near 880 nm
wavelength, measure the absorbance near 7 10 nm wavelength.
(6) When sample has turbidity or colour, take the same amount of a
sample as in (a), use 2 ml of ammonium molybdate solution in-
stead of 2 ml of ammonium molybdate-ascorbic acid mixed solu-
tion, carry out the operations in (a) and (b),and measure
absorbance making this solution reference solution. Otherwise,
measure the absorbance of this solution, and correct the absor-
bance obtained on the sample. In these cases, however, the sample
does not need the correction by a blank test carried out in (d).
When adopting this method, the sample with serious turbid-
ity gives a large error,
(7) In case where concentration of phosphorus is expressed by
(mgP/Z), the following conversion formula is used.
Concentration of phosphorus (mgPIZ)
= concentration of phosphate ion (mgP043-/Z)x 0.326 1
261
K 0101 : 1998
Remarks 1 When arsenic (V)is contained in sample a s it colours just as

phosphate ion does, eliminate the disturbance by the follow-
ing operations.
Add 2.5 ml of sulfuric-acidic sodium disulfite-sodium thio-
sulfate solution into a suitable amount of sample, let it stand
for 1 t o 2 min, neutralize its acidity according t o Note (9, and
carry out the operations in (a) t o ( e ) .
The preparation of sulfuric-acidic sodium disulfite-sodium
thiosulfate solution shall be a s follows.
Dissolve 1 4 g of sodium disulfite specified i n JXS K 8501
in 100 ml of water, add 20 ml of sulfuric acid (l-tl)into 40 ml
of the above solution, add 40 ml of solution in which 0.56 g of
sodium thiosulfate pentahydrate specified in JIS K 8637 is
dissolved in water (prepare this solution when it is needed).
2 A lot of coexisting chloride ion and sulfate ion (about 4 %'o> does
not disturb test. Therefore, this is suitable for the sample
containing a lot of salts and the sample in which a lot of salts
has been produced by pretreatment. The coexistence of much
ammonium ion and potassium ion, however, gives turbidity,
which leads t o disturbance.
3 In case of no addition of ammonium amidosulfate in ammo-
nium molybdate solution, existing of several gram of nitrate
ion or 0.25 mg of nitrite ion gives serious fading of molybde-
num blue about 15 min later since adding reagents. Further
coexisting of a lot of these ions disturb its highest colouring.
When ammonium molybdate solution mixed with ammonium
amidosulfate is used, nitrate ion gives no disturbance. Nitrite
ion also gives no disturbance until about 20 mg. If amount of
nitrite ion is more than this, supplement ammonium amido-
sulfate i n proportion t o its amount. Ammonium amidosulfate
can be supplemented by making another solution.
4 Existence of iron (III) by 30mg or more fades molybdenum
blue. If adding of ascorbic acid solution is increased, the dis-
turbance can be lessened.
5 When phosphate is contained a s suspension, the absorbance
may increase gradually even about 15 min later since addi-
tion of reagent.
6 When sample has low concentration of phosphate ion, the fol-
lowing is possible; increase the amount of a sample and am-
monium molybdate-ascorbic acid mixed solution, colour
molybdenum blue, extract i t with 2,6-dimethyl-4-heptanone
[diisobutyl ketone (DIBK)], and determine it.
7 Using the following operations, colouring and determination
is possible.
Take a suitable amount of sample (containing 5 t o 150pg
as POq3-)in a 50 ml volumetric flask, dilute with water t o make
total about 40 ml, add 3.5 ml of ammonium molybdate-ascor-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
bic acid mixed solution, add water up t o marked line and agitate
262
K O101 : 1998
it, let it stand for about 15 min at 20 t o 40 "C for colouring,

and measure its absorbance in the vicinity of 880nm wave-
length (or near 710nm wavelength). Carry out a blank test
using water, and correct the absorbance.
Prepare working curve by pipetting 1 t o 30 ml of phosphate
ion standard solution (5 pgP0d3-/ml) and operating similarly
t o the sample side.
43.1.2 Molybdenum blue [tin (II) chloride reduction] absorptiometry Carry

out the reaction of phosphate ion to hexaammonium heptamolybdate (ammonium mo-
lybdate) to produce hetero-poly compound, reduce this hetero-poly compound by tin
(II) chloride, measure the absorbance of produced molybdenum blue, and determine
phosphate ion.
Determination range: P O P 5 t o 150pg

Ammonium molybdate solution Dissolve 15 g of hexaammonium
heptamolybdate tetrahydrate specified in JIS K 8905 in water, add this
solution into sulfuric acid [Pour gently stirringly 182ml of sulfuric acid
specified in JIS K 8951 into 600ml water and let it cool.] with stirring,
add 10 g of ammonium amidosulfate(1) specified in JIS K 8588, dissolve
it, and add water t o make total 1E.
Tin (II) chloride solution Dissolve 1 g of tin (II) chloride dihydrate speci-
fied in JIS K 8136 in 5 ml of hydrochloric acid specified in JIS K 8180 (If
necessary, heat it.), add water t o make total 50 ml, add 2 or 3 granules of
tin specified in JIS K 8580,and store it in a coloured glass bottle. If tur-
bidity takes place, don't use it.
Phosphate ion standard solution (10 pgP0d3-/ml) Pipet 20 ml of phos-
phate ion standard solution (0.1 mgP043-/ml) of 43.1.1 (1)(e) in a 200 ml
volumetric flask, and dilute with water up to marked line.

Take a suitable amount (containing 5 to 150 pg as P043-) of sample(2)(3) in
a 50 ml volumetric flask, and add water t o make about 40 ml.
Add 5 m l of ammonium molybdate solution, agitate it, add 0.25ml of tin
(II) chloride solution, add water up t o marked line, agitate it, and let it
stand for about 15 min.
Put a part of the solution in an absorption cell, and measure its absorbance
in the vicinity of 700 nm wavelength(8).
Take 40 ml of water for a blank test, carry out the operations in (b) and (e),
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
263
K O101 : 1998
(e) Find quantity of phosphate ion on the working curve, and calculate the con-
centration of phosphate ion (mgP0PlZ) in the sample (7).
Working curve Pipet step by step 0.5 t o 15 ml of phosphate ion standard
solution (10 pgP043-/rnl)in as many 50 ml volumetric flasks, respectively carry
out the operations in (a)to (d), measure absorbances, and draw the relation
curve between quantities of phosphate ion (P043-)and absorbances.
Note (8) When sample has turbidity or colour, take the same amount of
sample as in (a),carry out the operations in (a)and (b) except
addition of tin (II) chloride solution, and measure the absorbance
making this solution reference solution. Otherwise, measure the
absorbance of this solution, and correct the absorbance obtained
on the sample. In these cases, however, the sample does not need
the correction by a blank test carried out in (d).
In this operation, the error will increase for samples with seri-
ous turbidity.
Remarks 8 When arsenic (VI is contained in the sample, prepare sulfu-
ric-acidic sodium disulfite-sodium thiosulfate solution according
to Remarks 1, add a suitable amount of a sample into 5 ml of
this solution, let it stand for 1 t o 2 min, neutralize its acidity
according t o Note (9, add water up t o about 40 ml, and carry
out the operations in tb) to (e).
9 Existence of chloride ion, iodide ion, and bromide ion some-
what weakens the colouring. The existence of C1- 75 mg,
I- 6 mg, and Br- 25 mg decreases absorbance by about 5 %.
Therefore, if sample contains a lot of halide ion, when pre-
paring working curve, either add halide ion t o get the same
amount as contained in the sample or adopt the method in
43.1.1 in which there is no influence by halide.
10 Coexistence of a lot of sulfate ion increases somewhat colouring.
Coexistence of 500 mg of sulfate ion other than that in am-
monium molybdate solution increases about 3 % of absorbance,
and 1 g does about 5 %. Accordingly, when a lot of sulfate ion
is contained, like the case of halide ion in Remarks 9, add sulfate
ion t o make the same coexistence as the sample, and prepare
the working curve. Otherwise use the method in 43.1.1.
11 Coexistence of 150 mg of sulfite ion gives about 5 % positive
error.
12 Follow Remarks 3.
13 Coexistence of 2 mg of iron (III) makes molybdenum blue begin
t o fade about 15 min after adding reagent. In order t o pre-
vent this disturbance, adjust pH to be about 2 by aqueous am-
monia (1t50) [Just before precipitating iron (III) hydroxide,
if iron (III) hydroxide is produced, dissolve it by dripping ni-
tric acid (1+25), and be careful the excess of nitric acid (1+25)
does not become 1 ml or more.], and add 1ml of potassium
iodide-sodium sulfite solution. [Dissolve 2 g of potassium iodide
specified in JIS K 8913 and l o g of sodium sulfite specified
in JIS K 8061 in water t o make total 100ml.I In this case
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
264
K O101 : 1998
the solution shows strong reddish brown, but let it stand until
the colour disappears. [This will disappear within 5 to 10 min,
but if a lot of iron (III) is contained, it may be effective to im-
merse it in boiling water for 30 t o 60 s.] Carry out the opera-
tions in and after (a)on this solution, and determine amount
of phosphate ion.
Adding potassium iodide-sodium sulfite solution can remove
the influence by nitrate ion and nitrite ion.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
14 The influence by silica is +5 % error even if it exists 500 times
more than phosphate ion.
16 In case of the sample having low concentration of phosphate
ion, coloured molybdenum blue can be determined after it is
extracted by 1-butanol.
Take a suitable amount of sample (containing 2 t o 40 pg as
P043-)in a 100 ml separatory funnel, and make liquid amount
50 ml. (Mark previously the line for 50 ml on the separatory
funnel.) Add 1ml of sulfuric acid (1+50) and 15 ml of 1-bu-
tanol specified in JIS K 8810, agitate it, and transfer water
layer t o another 100 ml separatory funnel. Add 6.5 ml of
ammonium molybdate solution in water layer, agitate it, then
add 0.25 ml of tin (II) chloride solution, agitate it, and let it
stand for 10 t o 15 min. Add 10 ml 1-butanol and agitate t o
extract molybdenum blue, after standing discard water layer,
put a part of 1-butanol layer in an absorption cell and mea-
sure its absorbance in the vicinity of 730 nm wavelength making
1-butanol reference solution.
To prepare working curve, take step by step 2 to 40ml of
phosphate ion standard solution (1~gP04~-/ml), which has been
prepared by diluting five times phosphate ion standard solu-
tion (5 ygP043-lml), and carry out similarly t o the operation
for the sample.
In this method, during the first extraction by 1-butanol, water
layer is saturated with 1-butanol, and simultaneously disturbing
material for colouring in sample is extracted. When arsenic
(V) coexists, follow Remarks 8. Provided that, after adding
5 ml of sulfuric-acidic sodium disulfite-sodium thiosulfate so-
lution, standing for more than about 2 h should be necessary.
Concerning other disturbing materials, refer Remarks 3, Re-
marks 13, Remarks 14, and Remarks 15.
43.2 Hydrolytic phosphorus This means the phosphorus which changes to phos-
phate ion owing to hydrolysis when a sample is boiled in acidic condition.
Add mixed acid of sulfuric acid-nitric acid in a sample, boil it, change to phos-
phate ion, determine it, subtract phosphate ion given before hydrolysis from the value,
and represent converted value t o phosphate ion.
Determination range: Po43-2.5 to 75 pg by 43.1.1 and P043-5 t o 150 yg by 43.1.2
265
K O101 : 1998

(b) Mixed acid of sulfuric acid-nitric acid Pour stirringly and carefully
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
300 ml of sulfuric acid specified in JIS K 8951 in about 600 ml of water,

mix them, and let it cool. Add 4 ml nitric acid specified in JIS K 8641 and
water to make it total 11.
(cl Sodium hydroxide solution (40g/Z) Follow 19 (1)(g).
(d) Ascorbic acid solution (72gll) Follow 43.1.1 (1)(b)(9).
(e) Ammonium molybdate solution Follow 43.1.1 (1)(c) (10).
(0 Ammonium molybdate-ascorbic acid mixed solution Follow 43.1.1
(1)(d)(9).
(g) p-nitrophenol solution (1gil) Dissolve 0.1 g of p-nitrophenol specified
in JIS K 8721 in water to make total 100ml.
(h) Phosphate ion standard solution ( 6 ~ g P 0 4 ~ - / r n lFollow
) 43.1.1 (1)(0.
Notes (9) When using method in 43.1.2,prepare the solution of tin (II) chlo-
ride in 43.1.2 (1)( c ) instead of this solution.
(10) When using method in 43.1.2, prepare the solution of ammonium
molybdate in 43.1.2 (1)(b) instead of this solution.
Take a suitable amount (containing 1 mg or less as P O P I of sample(3) (11)
in a 200 ml beaker, add water to make up to 50 t o 100 ml, and add 1ml of
mixed acid of sulfuric acid-nitric acid.
Boil it gently. When amount of liquid becomes 25 ml or less, add water to
keep liquid level at 25 to 50 ml, and boil for about 90 min(l2).
After cooling, filtrate it through filter paper 5 grade B, and wash with warm
water 3 to 4 times.
Put the filtrate and washings together, drip (3) sodium hydroxide solution
(40 g/Z), with several drops of p-nitrophenol solution (1gll) as indicator, until
the solution turns faint yellow, and transfer it in a 100 ml volumetric flask,
and dilute with water up to marked line.
Take a suitable amount of this solution, obtain quantity of phosphate ion
according to 43.1.1 (or 43.1.2)(13), convert it to the concentration of phos-
phate ion in the sample (mgP043-lZ), subtract the concentration of phosphate
ion in the sample (mgP043-lZ) determined by 43.1.1 (or 43.1.2) from this
value to find the hydrolytic phosphorus, and express it with concentration
of phosphate ion (mgP043-/Z)(7).
Notes (11) When determining dissolved hydrolytic phosphorus, use filtrated
sample according to Note (2).
266
K O101 : 1998
(12) Diphosphate ion, tripolyphosphate ion, and so on change t o phos-

phate ion within about 1h.
(13) When a lot of salts are contained in the solution that has coloured,
the method in 43.1.1, giving tough resistance to their influences,
is preferable. In case of adopting 43.1.2, refer to Remarks 9
and Remarks 10.
43.3 Total phosphorus Owing to such as potassium peroxodisulfate decomposi-

tion, nitric acid-perchloric acid decomposition, or nitric acid-sulfuric acid decompo-
sition, decompose the organic substance contained in sample, determine phosphate ion
contained in this solution, and express it as the concentration of total phosphorus.
43.3.1 Potassium peroxodisulfate decomposition Add potassium peroxodisulfate

in a sample, heat it in a high-pressure steam sterilizer t o decompose organic sub-
stances, and determine phosphate ion in this solution to find the concentration of
total phosphorus.
Determination range: P 1.25 t o 25pg
Ascorbic acid solution (72 g/Z) Follow 43.1.1 (1) (b).
Potassium peroxodisulfate solution (40g/Z) Dissolve 4 g of potassium
peroxodisulfate (for nitrogen and phosphorus measurement) specified in JIS
K 8263 in water to make total 100 ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Ammonium molybdate solution Follow 43.1.1 (1) (c). Provided that am-
monium amidosulfate is not added(l4).
Ammonium molybdate-ascorbic acid mixed solution Follow 43.1.1
(1) (d).
Phosphorus standard solution (50 p gP/ml) Heat potassium dihydrogen-
phosphate (for pH standard solution) specified in JIS K 9007 at (105f2)"C
for about 2 h, and let it cool in a desiccator. Take its 0.220 g, dissolve in a
little amount of water, transfer it in a 1O00 ml volumetric flask, and add
water up t o the marked line. Store it in a dark place at 1 to 10 O C .
Phosphorus standard solution ( 5 pgP/ml) Pipet 20 ml of phosphorus
standard solution (50 pgP/ml) in a 200 ml volumetric flask, and add water
up to the marked line. Prepare this solution each time it is needed.
Note (14) In total phosphorus measurement, because nitrite ion has been
eliminated by decomposition of the pretreatment, ammonium
amidosulfate is not necessarily added.
(a) Decomposing bottle This is a heat-resisting and pressure-resisting glass
bottle (capacity about 100ml), and can be used in high-pressure steam
sterilizer (about 120 "C)(15).
267
K 0101 : 1998
(b) High-pressure steam sterilizer One specified in JIS T 7322 or JIS T

7324 capable of heating at about 120 O C .
Note (15) A glass ampule (capacity about 100 mi), which can be used in high-
pressure steam sterilizer (about 120 O C ) may be used.
(a) Take 50 ml of sample(16) in a decomposing bottle.
(b) Add 10 ml of potassium peroxodisulfate solution (40 g/Z), and keep it with
close stoppering, followed by mixing.
(c) Heat it in a high-pressure steam sterilizer, and heat for 30 min more after
it reaches about 120°C.
(d) Take out the decomposing bottle, and after cooling(l7) pipet 25 ml of super-
natant(l8) ( 1 9 ) in a test tube with a ground stopper.
(e) Carry out the operations in 43.1.1 (3)(b)and ( c ) , and measure absorbance(20).
(0 Take 50 ml of water in a decomposing bottle for a blank test, carry out the
operations in (b) t o (e), measure absorbance, and correct the absorbance
obtained on the sample.
(g) Find the quantity of phosphorus in 25 ml of solution pipetted at (d) on the
working curve, and calculate the concentration of total phosphorus (mgPIZ)
in the sample in accordance with the following formula(21).
60 1000
P=ax-x-
25 50
where, P : total phosphorus (mgPíZ)
a : total phosphorus in 25 ml solution pipetted in (d)
(mg)
Working curve Pipet step by step 1 to 20 ml of phosphorus standard so-
lution ( 5 pgPlml) in as many 100 ml volumetric flasks, respectively add water
up t o marked line. Take each 25 ml in a test tube with a ground stopper,
carry out the operations in 43.1.1 (3)(b) and (cl, and measure its absor-
bance. Separately, take 25 ml of water in a test tube with a ground stop-
per for a blank test, carry out the operations in 43.1.1 (3)(b) and (c), measure
its absorbance, and correct the absorbance obtained on phosphorus stan-
dard solution (5 pgP/ml). Draw the relation curve between the quantities
of phosphorus (Pl in 25 ml aliquot solution and absorbances.
Notes (16) If 50 ml of sample contains 60 pg o r more total phosphorus, take
a suitable amount of sample (containing less than 0.12 mg as
P) in a 100ml volumetric flask, and add water up to marked
line. However, if 50 ml of sample contains 60 pg or more total
phosphorus and its pH is out of 5 t o 9, the following is neces-
sary: take a suitable amount of a sample (containing less than
0.12 mg as P), neutralize it using sulfuric acid (1+35) o r sodium
hydroxide solution (40 gL), transfer it in a 100 ml volumetric flask,
and add water up t o marked line.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
268
K 0101 : 1998
When sample contains chloride ion, add 1m of sodium hydrogen-

sulfite solution (50 g/¿) in the solution after decomposing in or-
der t o prevent the disturbance of colouring by molybdenum blue
owing to the generation of chlorine.
In case of turbidity in supernatant, filtrate it through filter pa-
per 5 grade C or a glass fiber filter with pore diameter of l pm
o r less, and use the filtrate after discarding initial 5 t o 1 0 m l
filtrate.
When metal hydroxide precipitates in the solution after decom-
posing, add sulfuric acid (1+35)or, if necessary, sodium hydrox-
ide solution (40 gil) until the precipitation will dissolve. (Record
the amount of these added solutions.)
If there is turbidity in the solution after dissolving precipi-
tation of metal hydroxide, additionally carry out the operation
in Note ('8).
When arsenic (VIis contained in supernatant a t the operation
of (d),add 5 ml of sulfuric-acidic sodium disulfite-sodium thio-
sulfate solution prepared according t o Remarks l into the solu-
tion after decomposing, let it stand for 1 t o 2 min, neutralize its
acidity according t o Note ( 3 ) , take 25 ml of the supernatant, and
60+c
carry out the operation in ( e ) . When this is adopted, use -
60 25
instead of 25 in the formula ( g ) . Provided that c means the
sum total (mi) of sulfuric-acidic sodium disulfite-sodium thio-
sulfate solution, sodium hydroxide solution (40 g/Z) and sulfuric
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
acid (1+35) which are used at Note (20).

If the operation in Note (16) is adopted a t the operation (a), cal-
culate the concentration of total phosphorus (mgP/Z) in the sample
according t o the following formula.
60 1000 100
P=ax-x-x-
25 50 V
where, P : total phosphorus (mgP/Z)
a : total phosphorus in 25 ml aliquot solution
taken a t (d) (mg)
V : sample (ml)
When, a t the operation of (d), the operation in Note ( 1 7 ) o r
Note (19) is carried out or the operations in both Note ( ' 7 ) and
Note (19) are carried out, use respectively - 61 -(60+b) (61+b)
60 25' 25
, o r ____
25
instead of 25 in the formula in ( g ) or the above-mentioned for-
mula. Provided that b means the amount of sulfuric acid (1+35)
o r sodium hydroxide solution ( 4 0 g l Z ) (mi) which is added a t
Note ( 1 9 ) .
Remarks 17 When the pH of sample is out of 5 t o 9, carry out the decom-
posing operation after neutralization a t Note (16), however, in
case of less than 60 pg of total phosphorus in 50 ml sample, take
a suitable amount (50 t o 100 ml) of sample, neutralize it with
sulfuric acid (1+35)and sodium hydroxide solution (40 glZ), place
269
K 0101 : 1998
50 ml out of this solution into a decomposing bottle. In this

case, however, record the volume (mi) of both solutions required
for neutralization, and use the following formula instead of the
one in (3)( g ) for the calculation of the concentration of total
phosphorus (mgP/Z) in the sample.
- y - 1000
P = a x - x60 V+b
25 50 " v
where, P : total phosphorus (mgPIZ)
a : total phosphorus in 25 ml aliquot so-
lution taken at (d)(mg)
b : sulfuric acid (1+35) and sodium hy-
droxide solution (40 gll) required for
neutralization (mi)
V : sample (ml)
18 When the concentration of phosphorus in a sample, which has
been taken in a decomposing bottle at (3)(a),is poor, for ex-
ample less than 0.1 mg/Z, use an absorption cell with 50 mm
length optical path for measuring absorbance at (3)(e).
When measuring absorbance for the preparation of work-
ing curve and blank test, use a n absorption cell with 50 mm
length optical path.
In this case, use the phosphorus standard solution (0.5 pgP/
mi), prepared from the phosphorus standard solution ( 5 FgP/
mi) by diluting 10 times, instead of the phosphorus standard
solution ( 5 yP/ml) in (1)(g).
19 When handling sample with poor concentration of phospho-
rus and with no prospect to get precise determination, either
carry out the following heating and concentrating process, o r
extract molybdenum blue by solvent in Remarks 20.
Take 100 to 250 ml of sample in a beaker of 200 t o 500 ml,
add 1 t o 2 drops of sulfuric acid (2+1), and heat it o n a hot
plate to concentrate until it becomes 50 ml or less. Neutral-
ize it with sodium hydroxide solution (40g/Z), put it in a de-
composing bottle (which has been marked at 50 ml level), make
it 50 ml by adding water, and carry out the operation in (3)(b)
and the following items. Provided that, for calculation of the
concentration of total phosphorus in a sample, use -'Ooo instead
V
'Ooo in the formula of (3)( g ) , where V means the amount
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
of -
50
of the sample (mi).
20 If the molybdenum blue, which has been coloured according
t o (3),is extracted by 2,6-dimethyl-4-heptanone[diisobutyl
ketone (DIBK)], a trace of phosphorus can be determined.
After the operations in (3)(a)to ( c ) ,transfer the solution(16)
in the decomposing bottle into a 100 ml separatory funnel, wash
the decomposing bottle with 10 ml of water, and put together
270
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the washings in the separatory funnel. Add 5.5 ml of ammo-

nium molybdate-ascorbic acid mixed solution, and let it stand
for about 15 min at 20 to 40 O C ( 4 ) . Add 5 ml of 2,6-dimethyl-
4-heptanone in the separatory funnel and agitate it for about
5 min. After standing, discard water layer, and place a part of
Z76-dirnethyl-4-heptanone layer (If there is turbidity by water
drops and so on, filtrate quickly through dried filter paper.) in
an absorption cell, and measure the absorbance in the vicinity
of 640 nm wavelength.
Take 50 ml of water for a blank test in a decomposing bottle,
carry out the same operations as carried out on the sample,
measure the absorbance, and correct the absorbance obtained
on the sample. Find the quantity of total phosphorus in the
sample on the working curve, and calculate the concentration
of total phosphorus in the sample (mgPIZ) in accordance with
the following formula.
1O00
P=ax-
V
a : total phosphorus measured (mg)
V : sample (mi)
Working curve Prepare phosphorus standard solution
(0.5pgP/ml) by diluting 10 times phosphorus standard solu-
tion (5 pgP/ml) in (1) (g), pipet step by step 1 t o 25 ml of the
standard solution into as many 100 ml volumetric flasks, and
add water up to marked line. Respectively, take its 50 ml into
a 100 ml separatory funnel, add 20 ml of water, carry out the
same operations as carried on the sample, and measure its
absorbance. Take 70 ml of water in a 100 ml separatory fun-
nel for a blank test, after similar operations are carried out,
measure absorbance, correct the absorbance obtained on phos-
phorus standard solution (0.5 pgP/ml), and draw the relation
curve between absorbances and the quantities of phosphorus
(P) in an aliquot 50 ml solution.
43.3.2 Nitric acid-perchloric acid decomposition method Add nitric acid in

a sample, concentrate it with heating, add nitric acid and perchloric acid, heat on it,
decompose organic substance, determine phosphate ion in this solution, and obtain
the concentration of total phosphorus. This method shall be applied t o the sample
containing a lot of organic substances and organic phosphorus compounds which are
hardly decomposed.
Determination range: P 1.25 t o 25 pg
(b) Nitric acid Specified in JIS K 8541.
27 1
K O101 : 1998
(c) Perchloric acid Specified in JIS K 8223.

(d) Ascorbic acid solution (72gil) Follow 43.1.1 (i)(b).
(e) Sodium hydroxide solution (40gll) Follow 19 (i)(g).
(0 Sodium hydroxide solution (200 gll) Follow 35.1.1.1 ( i )(c).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(9) Ammonium molybdate solution Follow 43.1.1 (i)( c ) .

(h) Ammonium molybdate-ascorbic acid mixed solution Follow 43.1.1
(1)(d).
(i) p-nitrophenol solution (ig/Z) Follow 43.2 (i)(g).
(j) Phosphorus standard solution (5 pgP/ml) Follow 43.3.1 (i)(g).


(a) Take 50 ml(22) of sample in a beaker.
(b) Add nitric acid t o make it weak acidic, and heat gently on a hot plate to
concentrate it 15 t o 20ml.
(c) Add 2 t o 5 ml of nitric acid, heat it again t o concentrate it t o about 10 ml,
add 2 m l of nitric acid, and heat it again t o concentrate t o about 10m1,
followed by letting it cool.
(d) Add little by little 5ml(23) of perchloric acid. Heat on a hot plate, when
white fume of perchloric acid is generated cover the beaker with a watch
glass, and keep the white fume from perchloric acid to circulate inside wall
of the beaker(24) (25).
(e) After letting it cool, add about 30 ml of water(26).
(f) Add several drops ofp-nitrophenol solution (1g/Z) for indicator, at first add
sodium hydroxide solution (200 g/Z) and then sodium hydroxide solution
(40 gll) for neutralization, and have the colour of the solution faint yellow (27).
(g) Transfer this solution into a 50ml volumetric flask, and add water up t o
marked line.
(h) Pipet 25 ml of this solution in a test tube with a ground stopper(28), carry
out the operations in 43.1.1 (3)(b) and ( c ) , and measure absorbance.
(i) Take the same amount of water as that of the sample in (a) for a blank
test in a beaker, carry out the operations in (b)t o (h),measure absorbance,
and correct the absorbance obtained on the sample.
(j) Find the quantity of phosphorus in 25 ml solution pipetted at (h)on the
working curve, and calculate the concentration of total phosphorus (mgPl0
in the sample in accordance with the following formula(29).
50 1000
P=ax-x-
25 V
272
K 0101 : 1998
a : total phosphorus in 25 ml aliquot solution at (h)

(mg)
V : sample (mi)
Working curve Prepare in accordance with the operations of working curve
in 43.3.1 (3)(g).
When the concentration of total phosphorus in sample is poor,
50ml o r more is permissible. When sample contains a lot of
chloride ion and concentration of total phosphorus is high, less
than 50 ml is permissible.
When sample contains a lot of chloride ion, add more than equiva-
lent amount to chloride ion.
Because thermally decomposing operation using perchloric acid
may bring about explosion depending on the kind of the sample,
the following cautions should be attended to.
(i) Easily oxidized organic substance should be completely
decomposed according t o the operations in (b)and ( c ) be-
fore adding perchloric acid.
(i;) Adding perchloric acid should be carried out after concen-
trated liquid is cooled without fail.
(iii) Thermal decomposition should be carried out under the
condition of coexisting of perchloric acid and nitric acid.
(iv) Never let dry up the concentrated liquid.
When this operation cannot decompose organic substance and
yellow remains in the solution, repeat the operation as adding
2 ml of nitric acid and then heating.
If necessary, dissolve soluble salts by heating, and in case where
unsoluble remainder is found even by heating, filter through filter
paper 5 grade C o r glass fiber filter with 1ym o r smaller pore
diameter, wash the filter paper with a little water and put to-
gether filtrate and washings.
If the precipitation of metal hydroxide is found during neutral-
ization, addition of sodium hydroxide solution (40gll) should be
stopped just before precipitation is about t o appear. If neces-
sary, control by sulfuric acid (1+35).
When there is 25 pg or more of total phosphorus in pipetted 25 ml
solution, take a suitable aliquot (Finally 25 yg or less phospho-
rus is t o be obtained.) out of this solution in a 50 ml measuring
cylinder (with a stopper), and dilute with water t o make total
25 ml.
50
When the operation in Note (28) is carried out in (h),use - in-
b
stead of 50 in the formula in ci). Provided that b means amount
25
of solution pipetted in a measuring cylinder (with a stopper) (ml).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
273
K O101 : 1998
43.3.3 Nitric acid-sulfuric acid decomposition method Add nitric acid in a

sample, heat it t o concentrate, add nitric acid and sulfuric acid, heat again to de-
compose organic substance, determine the phosphorus ion contained in this solution,
and obtain the concentration of total phosphorus. This method shall be applied to
the sample containing a lot of organic substances and organic phosphorus compounds
which are hardly decomposed.
Determination range: P 1.25 t o 25 pg
Nitric acid Specified in JIS K 8541.
Sulfuric acid Specified in JIS K 8951.
Ascorbic acid solution (72gll) Follow 43.1.1 (1)(b).
Sodium hydroxide solution (40gll) Follow 19 (1)( g ) .
Sodium hydroxide solution (200g l l ) Follow 35.1.1.1 (1)( c ) .
Ammonium molybdate solution Follow 43.1.1 (1)( c ) .
Ammonium molybdate-ascorbic acid mixed solution Follow 43.1.1
(1) (d).
p-nitrophenol solution (1g/Z) Follow 43.2 (1)(g).
Phosphorus standard solution (5 ygP/ml) Follow 43.3.1 (1) (g).
Carry out the operations in 43.3.2 (3)(a)and (b).
Add 2 ml(23) of sulfuric acid (l+l)and 2 t o 5 ml of nitric acid into the solu-
tion of which operation (a)has been finished, heat t o concentrate it until
white fume of sulfuric acid is generated, heat strongly on, and stop it after
violent white fume of sulfuric acid is generated a little while, followed by
letting it cool.
Add 5 ml of nitric acid in this solution, and heat it again until white fume
is again generated (25).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
After cooling it, add about 30ml of water, and boil it gently for about
10 min(26).
Carry out the operations in 43.3.2 (3)(f) t o (h).
Take the same amount of water as that of the sample taken in (a) for a
blank test in a beaker, carry out the operations in 43.3.2 (3)(b),carry out
the operations in (b)to (e),measure absorbance, and correct the absorbance
on the sample.
274
K O101 : 1998
( g ) Find the quantity of phosphorus in 25 ml aliquot solution taken at ( e )on the

working curve, and calculate the concentration of total phosphorus (mgPIZ)
in the sample in accordance with the formula in 43.3.2 (3)u’).
Working curve Prepare in accordance with the operations of working curve
in 43.3.1 (3)(g).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
275
K O101 : 1998
44 Silica (Sioz) Silica in water is classified to ionic silica (ionic silicic acid), dis-
solved and colloidal silica, and total silica, and they are expressed as silica oxide
(IV) (Sioz).
44.1 Ionic silica Ionic silica means the silica which produces yellow heteropoly
compound as a result of reaction with hexaammonium heptamolybdate.
44.1.1 Molybdenum yellow absorptiometry Measure the absorbance by yellow

of heteropoly compound produced by the reaction of ionic silica and hexaammonium
heptamolybdate, and determine silica.
Determination range: Si02 0.1 to 1mg
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(1) Reagents Reagents shall be as follows, and stored in a polyethylene bottle.

Water Water A3 specified in JIS K 0557. For preparation of reagents
and operation, always use this water.
Hydrochloric acid (1+1) Prepare using hydrochloric acid specified in JIS
K 8180.
Ammonium molybdate solution (100 g/Z) Dissolve 21.2 g of hexa-
ammonium heptamolybdate tetrahydrate specified in JIS K 8905 in water
to make total 200ml.
Oxalic acid solution Dissolve 20 g of oxalic acid dihydrate specified in
JIS K 8519 in water to take total 200 ml.
Silica standard solution (1 mgSiOdml) Grind sand-like quartz (99.9 %
or more) in an agate mortar, heat at 700 to 800 "C for about 1h, and let it
cool in a desiccator. Take its 0.500g in a platinum crucible, add 4 g of
sodium carbonate, reference material for volumetric analysis, specified in
JIS K 8005, mix them well, and heat them for about 40min for fusing.
After cooling it, dissolve the fused material in water, transfer in a 500 ml
volumetric flask, and add water up to marked line.
Silica standard solution (0.1 rngSiOdm1) Pipet 20 ml of silica standard
solution (1mgSi0dml) in a 200 ml volumetric flask, and add water up to
marked line.
(3) Operation Operations shall be as follows,
(a) Filtrate sample(1), take its 50 ml(2) (containing 0.1 to 1mg as Sioz) in a
50 ml measuring cylinder (with a stopper), control its temperature a t about
20 "C.
(b) Add 1ml of hydrochloric acid ( l + l ) and 2 ml of ammonium molybdate so-
lution (100 g/Z), agitate it(3), and let it stand for 5 min.
(c) Add 1.5 ml(4) of oxalic acid solution, agitate it, and let it stand for 1min(5).
276
K O101 : 1998
(d) Immediately, transfer a part of the solution in an absorption cell, and measure
its absorbance with 410 t o 450 nm wavelength,
(e) Take about 50 ml water for a blank test, carry out the operations in (a) t o
(d), measure the absorbance, and correct the absorbance obtained on the
sample.
(f) Find the quantity of silica on the working curve, and calculate the concen-
tration (mgSi0dZ) of silica in the sample.
Working curve Pipet step by step 1 to 10 ml of silica standard solution
(0.1mgSiOz/ml) in as many 50 ml measuring cylinders (with a stopper),
respectively, add water up t o 50 ml marked line, control its temperature t o
about 20"C, carry out the operations in (b) to (e), and draw the relation
curve between quantities of silica (Si02) and absorbances.
Notes (1) Filtrate through filter paper 5 grade C (or filter paper 6) o r fil-
ter media with 0.45 t o l km pore diameter. Discard about ini-
tial 50 ml of filtrate, and use the filtrate obtained thereafter.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(2) If sample gives high concentration of silica, take a suitable amount

of sample and dilute it with water up to 50ml.
(3) At this time, pH becomes 1.1 t o 1.6.
(4) If there is no coexistence of phosphate ion, don't add oxalic acid
solution. Provided that the working curve should be prepared
similarly.
(6) When oxalic acid solution is added, the time of standing should
be strictly kept. Long standing time results in the fading of yellow
colour of heteropolycompound by silica.
44.1.2 Molybdenum blue absorptiometry Carry out the reaction between ionic
silica and hexaammonium heptamolybdate to produce heteropolycompound, reduce
the compound by L(+)-ascorbic acid to produce molybdenum blue, and measure its
absorbance t o determine silica.
Determination range: Sioz 10 t o 100 pg
(1) Reagents Reagents shall be as follows, and stored in a polyethylene bottle.

(a) Water Follow 44.1.1 (i)(a).
(b) Hydrochloric acid (l+l) Follow 44.1.1 (i)(b).
( c ) Sulfuric acid (1+5) Take five volume water in a beaker, cool it, and add
one volume sulfuric acid specified in JIS K 8951 gradually with stirring.
(d) Ammonium molybdate solution (100 gll) Follow 41.1.1 (i)(c).
(e) Oxalic acid solution Follow 44.1.1 (i)(d).
(f) Ascorbic acid solution (100gll) Dissolve 10 g of L(+)-ascorbicacid speci-
fied in JIS K 9502 in water up to 100 ml. Store in a dark place a t 10 "C
or lower. Don't use the coloured solution.
277
K O101 : 1998
(g) Silica standard solution (10 pgSiOzlm1) Pipet 20 ml of silica standard

solution (0.1 mgSiOz/ml) stated in 44.1.1 (1)(f) in a 200 ml volumetric flask,
add water up to the marked line. Prepare this solution each time it is needed.
(a) Photometer Spectrophotometer o r photoelectric photometer

Filtrate sample(l), take its 50 ml(2) (containing 10 to 100 pg as SiOd in a
50 ml measuring cylinder (with a stopper), and keep its temperature at about
20 OC.
[or 1ml of sulfuric acid (1+5)]and 2 ml
Add 1ml of hydrochloric acid (l+l)
of ammonium molybdate solution (lOOgll), agitate i t ( 3 ) , and let it stand
for 5 min.
Add 1.5 ml of oxalic acid solution, agitate it, and let it stand for 1 min(%
Add 1ml of ascorbic acid solution (100 g/Z), agitate it, and let it stand for
about 10min.
Transfer a part of the solution in an absorption cell, and measure its ab-
sorbance in the vicinity of 815 nm wavelength.
Take 50 ml of water for a blank test, control its temperature at about 20 "C,
carry out the operations in (b) to (e),measure absorbance, and correct the
Find the quantity of silica on the working curve, and calculate the concen-
tration of silica (mgSi0dZ) in sample.
Working curve Pipet stepwise 1 to 1 0 m l of silica standard solution
(10 pgSiOz/ml) in as many 50 ml measuring cylinders (with a stopper), re-
spectively, add water up to marked line of 50 ml, control its temperature
at about 20 OC, carry out the operations in (b) to (f),measure absorbance,
and draw the relation curve between quantities of silica (Si02) and absor-
bances.
Remarks 1 When the colouring of molybdenum blue is weak because of
low concentration of silica, an absorption cell with 20 mm o r
50mm optical length may be used for absorbance measure-
ment. The value for a blank test becomes large. In this case,
44.1.3 shall be recommendable.
Molybdenum blue extraction absorptiometry Carry out the reaction

between ionic silica and hexaammonium heptamolybdate to produce hetero-
polycompound, reduce the compound by L (+)-ascorbic acid to produce molybdenum
blue, extract this by 1-butanol, and measure the absorbance of organic layer t o de-
termine silica. This method can be applied to the sample containing a low concen-
tration of silica.
Determination range: Si02 0.5 to 1Opg
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
278
K O101 : 1998
(1) Reagents Reagents shall be as follows, and be kept in a polyethylene bottle.

Water Water A3 specified in JIS K 0007 (distill using distilling appara-
tus made of stainless steel o r copper). Use this water for the preparation
of reagents to be used in this test and for the operation of this test.
Sulfuric acid (2.5 mol/Z)-ammonium molybdate (188 g/Z) mixed solu-
tion Mix, while stirring and cooling, 140 ml of sulfuric acid specified in
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
JIS K 8951 into about 300 ml of water. Add into this the solution which
has been prepared by dissolving 200 g of hexaammonium heptamolybdate
tetrahydrate specified in JIS K 8905 in about 500ml water, transfer it
into a 1O00 ml volumetric flask, and add water up t o marked line.
Sulfuric acid (2+1) Take 1 volume of water in a beaker, cool, and add
gradually, while stirring, 2 volume of sulfuric acid specified in JIS K 8951
in the above water.
Ascorbic acid solution (100 g/Z) Follow 44.1.2 (1)(f).
Sodium sulfate Specified in JIS K 8987.
1-butanol Specified in JIS K 8810.
Silica standard solution (1 mgSiO2/ml) Follow 44.1.1 (1)(e). Provided
that water specified in (a) shall be used.
Silica standard solution (50 pgSiOdm1) Pipet 25 ml of silica standard
solution (1 mgSiOdm1) into a 500 ml volumetric flask, and add water up t o
marked line. Prepare this solution each time it is needed.
Silica standard solution (1 pgSiOdm1) Pipet 10 ml of silica standard
solution (50ygSiOz/ml) into a 500ml volumetric flask, and add water up
t o marked line. Prepare this solution each time it is needed.
(2) Tool and apparatus Tools and apparatus shall be as follows.
(a) Separatory funnel 300 ml one made of plastics.
Place 200ml (containing 0.5 t o 1Oyg as SiOd of sample in a separatory
funnel.
Add 4 ml of sulfuric acid (2.5 mol/Z)-ammonium molybdate (188 g/Z) mixed
solution, agitate it, and then let it stand for 20 min while keeping its tem-
perature at about 25 "C.
Add 25 ml of sulfuric acid (2+1),agitate it, immediately add 2 ml of ascor-
bic acid solution (100 g/Z), agitate it(61, and let it stand for 10 min.
Add 25 ml of 1-butanol, and agitate it for about 2 min t o extract molybde-
num blue.
After standing, put 1-butanol layer in a 10 ml test tube with ground stop-
per, and add sodium sulfate for dehydration.
Place this into a 20 mm absorption cell(7), measure the absorbance in the
vicinity of 800 nm wavelength with making 1-butanol reference solution.
279
K O101 : 1998
(g) Following the next operations, obtain a blank test value based on sulfuric
acid (2.5 mol/Z)-ammonium molybdate (188 g/Z) mixed solution, and correct
the absorbance obtained on the sample.
Take respectively 200ml of water into separatory funnels (A) and (B),
add 4 ml of sulfuric acid (2.5 mol/Z)-ammonium molybdate (188 gll) mixed
solution in (A) and 8 ml in (B), and agitate them. Let them stand for 20 min
while keeping solution temperature at about 25 "C. Then, carry out the
operations in ( c ) to (f), and measure respectively absorbances of (A) and
(B), followed by making them a and b. Calculate the blank test value c
based on sulfuric acid (2.5 mol/Z)-ammonium molybdate (188 g/Z) mixed
solution in accordance with the following formula.
c=b-a
(h) Find the quantity of silica on the working curve, and calculate the concen-
tration (pgSi0dZ) of silica in the sample.
Working curve Pipet step by step 0.5 to 10 ml of silica standard solution
(1pgSiOJm1) into as many separatory funnels, respectively, add water(8) up
to 200 ml, carry out the operations in (b)to (0.Separately, take the water
used in this operation by 200 ml, carry out the operations in (b) t o (0,
cor-
rect the absorbance obtained on silica standard solution, and draw the rela-
tion curve between the quantities of silica (Sioz) and absorbances.
Notes (6) Immediately after adding sulfuric acid (2+1) and agitating them,
add ascorbic acid solution (100 g/Z) and agitate them.
(7) When the concentration of silica in sample is 10 pgSiOd2 or more,
an absorption cell 10mm long can be used. In this case, how-
ever, a 10mm absorption cell should be used when measuring
absorbance of a blank test and working curve drawing. When
drawing working curve, instead of 0.5 to 10 ml of silica standard
solution (1 pgSiOdml), employ 1 t o 10 ml of silica standard solu-
tion (2 pgSiOdm1) which has been prepared by diluting 25 times
silica standard solution (50 pgSiOJm1).
(8) Use the same water as the water used when preparing silica
standard solution.
Remarks 2 When the high concentration (20 t o 400 pgSiOd2) of silica in
sample gives strong colouring in the solution which was treated
with the operation in (a)to (cl, the absorbance of the solution
may be measured using a 50mm absorption cell and wave-
length of 815 nm. In case of blank test, carry out the opera-
tion in (g) excepting the extraction by 1-butanol, measure
absorbance at wavelength 815 nm using a 50 mm absorption
cell, and calculate the blank test value similarly to (g).
The working curve should be prepared using 0.4 t o 8 ml of
silica standard solution (10 ygSiOz/ml) which has been prepared
by diluting 5 times silica standard solution (50 pgSiOdm1). In
this case, 50ml of the sample should be used and carry out
the operation similarly t o this.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
280
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
K O 1 0 1 : 1998
44.2 Dissolved and colloidal silica Dissolved and colloidal silica means the silica
contained in the solution obtained after filtration of sample. After filtration of sample,
add sodium hydrogencarbonate, boil it to vary silica into ionic state, and then deter-
mine it owing to molybdenum yellow absorptiometry o r molybdenum blue absorp-
tiometry.
(b) Hydrochloric acid (l+l) Follow 44.1.1 (i)(b).
(c) Sodium hydrogencarbonate Specified in JIS K 8622 and containing
0.002% or less of SiOs.

(a) Beaker made of tetrafluoroethylene resin 200 ml
(b) Platinum dish
Filtrate sample (11, take a suitable amount (containing 2 mg or less dissolved
and colloidal silica) in a 200 ml tetrafluoroethylene resin beaker (or plati-
num dish), and add water to make total 50 to 100ml.
Add 0.20 g of sodium hydrogencarbonate per 100 ml of the sample, and heat
it in a boiling water bath for about 20 min.
After cooling, neutralize it with hydrochloric acid (l+l) to make its pH5,
transfer it into a 100 ml volumetric flask, and add water up to marked line.
Pipet a suitable amount(9) of this solution, colour it by molybdenum yel-
low owing to 44.1.1 or by molybdenum blue owing to 44.1.2,and measure
its absorbance.
Take the same amount of water as the sample for a blank test, carry out
the operations in (b) to (d) to measure absorbance, and correct the absor-
Conforming to 44.1.1 (3)(f) or 44.1.2 (3)(g),calculate the concentration of
dissolved and colloidal silica (mgSiOdZ) in the sample,
Note (9) When using molybdenum yellow absorptiometry, make the amount
of Sioz in dissolved and colloidal silica 0.1 to 1 mg, and when using
molybdenum blue, make it 10 to 100 pg.
44.3 Total silica For testing total silica, after changing all silica in water into
ionic state, apply molybdenum yellow absorptiometry, molybdenum blue absorptiometry,
o r gravimetry.
44.3.1 Fusion by sodium carbonate Add sodium carbonate in a sample, evapo-

rate it to dryness, fuse it to vary silica to ionic state, and determine it by either of
molybdenum yellow absorptiometry or molybdenum blue absorptiometry.
281
K O101 : 1998

(b) Hydrochloric acid (l+l> Follow 44.1.1 (1)(b).
(cl Sodium carbonate Specified in JIS K 8005, containing 0.001 % or less
of Sion.

--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Beaker made of tetrafluoroethylene resin 200 ml (or platinum dish)

(b) Platinum crucible
(c) Photometer Spectrophotometer o r photoelectric photometer
Place a suitable amount (containing 2 mg or less of total silica) of sample
in a 200 ml tetrafluoroethylene resin beaker (or platinum dish), add 0.20 g
of sodium carbonate, and heat t o concentrate it to about 5 ml.
Remove the concentrated sample into a platinum crucible with a little water,
and again evaporate it to dryness.
Heat gently to carbonize organic substance, ash it, heat strongly t o fuse,
and let it cool.
Add water t o dissolve the fused sample by heating, let it cool, transfer it t o
a beaker, and neutralize it with hydrochloric acid (1+1)to make its pH about 5.
When there is turbidity, filtrate it, wash it with water, place washings and
filtrate into a 100 ml volumetric flask, and add water t o marked line.
Take a suitable amount(l0) of this solution, colour it by molybdenum yel-
low owing t o 44.1.1 or by molybdenum blue owing to 44.1.2,and measure
its absorbance.
the operations in (a) t o (f),measure its absorbance, and correct the absor-
Calculate the concentration of total silica (mgSiOslE) in the sample owing
to either 44.1.1 (3)(f) o r 44.1.2 (3)( g ) .
Note (10) When using molybdenum yellow absorptiometry, make the amount
of Si02 in total silica 0.1 t o 1mg, and when using molybdenum
blue, make it 10 to 1OObg.
44.3.2 Gravimetry Add hydrochloric acid and perchloric acid in a sample, heat
it, generate white fume of perchloric acid, and make silica undissolved state by de-
hydration. Add water t o dissolve salts, separate silica by filtration, heat t o get con-
stant weight, dispel silica adding sulfuric acid and hydrofluoric acid, and determine
the silica making use of its decreased weight.
Determination range: Si02 5 mg or more
282
K O101 : 1998

(b) Hydrochloric acid (l+l) Follow 44.1.1 (1)(b).
(c) Hydrochloric acid (1+50) Prepare using hydrochloric acid specified in
JIS K 8180.
(d) Sulfuric acid (1+2) Take 2 volume of water in a beaker, cool, and add
gradually, while stirring, 1 volume of sulfuric acid specified in JIS K 8951.
(e) Perchloric acid Specified in JIS K 8223.
(0 Hydrofluoric acid Specified in JIS K 8819.
(2) Tool Tools shall be as follows.
(a) Platinum crucible
(a) Take a suitable amount (containing 5 m g o r more of Si02) of sample in a
beaker, and add 10 ml of hydrochloric acid ( l + l )and 15 ml of perchloric
acid.
(b) Heat it t o evaporate, when deep white fume from perchloric acid is gener-
ated, cover the beaker with a watch glass, heat it for about 15 min succes-
sively, and let it cool.
(c) Add 100ml of warmed water t o dissolve soluble salts, filtrate it through
filter paper 5 grade B, wash several times residue and filter paper with
warmed hydrochloric acid (1+50), and then wash them with warmed water
several times.
(d) Put the residue and filter paper together in a platinum crucible which has
been made to be constant weight at 1O00 O C , dry it at first, then heat gradu-
ally, carbonize the filter paper, ash it, and heat at about 1O00 "C t o make
it constant weight.
(e) Moisten the residue with a few drops of sulfuric acid (1+2), add about 5 ml
of hydrofluoric acid, dispel silica with careful heating, continuously heat it
t o dry up, and heat at about 1O00 "C to make it constant weight.
(f) For a blank test, put a few drops of sulfuric acid (1+2) and 5 ml of hydro-
fluoric acid in a platinum crucible which has been made constant weight
a t about 1O00 O C , carry out the operation in (e), obtain the ignition resi-
due of hydrofluoric acid, and make it a blank test value of hydrofluoric acid.
(g) Calculate the concentration of silica (mgSiO2lZ) in the sample in accordance
with the following formula.
1 O00
s = [wl
- (wz - w3)]
x7
where, S : total silica (mgSiOdZ)

W1 : mass of residue obtained at (d) (mg)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
283
K O 1 0 1 : 1998
W z: mass of residue obtained at ( e ) (mg)

W3 : blank test value of hydrofluoric acid (mg)
V : sample (mi)
Remarks 3 When a lot of organic substance is contained in sample, add
nitric acid specified in JIS K 8541 to acidify it, heat it t o evapo-
rate. When the amount of liquid becomes small, add 10 to
20 ml of nitric acid and heat, let it cool, add 5 ml of perchloric
acid, carry out the operations on and after (3)(b),and deter-
mine it.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
284
K 0101 : 1998
45 Boron (B) For the determination of boron, methylene blue absorptiometry,

azomethine H absorptiometry or ICP atomic emission spectrometry shall be applied.
46.1 Methylene blue absorptiometry Change boron compounds into tetra-

fluoroborate ion by adding sulfuric acid and hydrofluoric acid, add methylene blue
[3,7-bis(dimethylamino)phenothiazine-5-ium chloride], extract produced ion association
with 1,Z-dichloroethane, and measure its absorbance to determine the boron con-
tained.
Determination range: B 0.1 to 1pg
(i) Reagents Reagents shall be as follows, and they shall be stored in polyethyl-
ene bottles.
(a) Water Water A3 specified in JIS K 0557 (prepared using distilling ap-
paratus made of quartz glass or metal).
(b) Sulfuric acid (3+97) Prepare using sulfuric acid specified in JIS K 8951.
(c) Hydrofluoric acid (1+9) Prepare using hydrofluoric acid specified in JIS
K 8819.
(d) Silver sulfate solution (0.3 g/Z) Dissolve 0.15 g of silver sulfate speci-
fied in JIS K 8965 in water t o make up the volume t o 500 ml.
(e) Methylene blue solution (0.4 g/Z) Dissolve 0.48 g of methylene blue (usu-
ally trihydrate) specified in JIS K 8897 in water to make up the volume t o
100 ml. Pipet 10 ml of this solution in a 100 ml volumetric flask, add wa-
(0 1,2-dichloroethane Specified in JIS K 8465.
(g) Boron standard solution (0.1 mgB/ml) Dissolve 0.572 g of boric acid
(orthoboric acid) specified in JIS K 8863 in water, transfer it in a 1O00 ml
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
volumetric flask, and add water up t o the marked line.

(h) Boron standard solution ( i pgB/ml) Pipet 10 ml of boron standard so-
lution (0.1 mgB/ml) in a l O00 ml volumetric flask, and add water up t o the
marked line.
(i) Boron standard solution (0.1 pgB/ml) Pipet 20 ml of boron standard
solution (i pgB/ml) in a 200 ml volumetric flask, and add water up to the
marked line. Prepare this solution just before use.

(a) Glassware Made of quartz glass or soda lime glass.
(b) Separatory funnel Made of polyethylene 50 ml
(cl Photometer Spectrophotometer or photoelectric photometer
(a) Take a suitable amount (containing 0.1 t o 1pg as B) of sample(1) ( 2 ) in a
separatory funnel, add water up t o 15 ml, add 3 ml of sulfuric acid (3+97),
285
K O101 : 1998
3 ml of methylene blue solution (0.4g/Z), and 10 ml of 1,2-dichloroethane,

then shake for about 1min, and let it stand(3)(4).
(b) Discard 1,S-dichloroethane layer(5), add 3 ml of hydrofluoric acid (1+9) in
water layer, and let it stand for about 1h.
(c) Add 10 ml of 1,2-dichloroethane, shake violently for about 1min, and let it
stand.
(d) Transfer 1,Z-dichloroethane layer in another separatory funnel, add 5 ml
of silver sulfate solution (0.3 g/Z), agitate it for about 1 min, wash 1,2-
dichloroethane layer, and let it stand.
(e) Place a part of 1,2-dichloroethane layer in an absorption cell, and measure
its absorbance in the vicinity of 660nm wavelength with making 1,2-
dichloroethane a reference solution.
(0 Take 15 ml of water for a blank test, carry out the operation in (a)t o ( e ) ,
and correct the absorbance obtained on the sample.
(g) Find the quantity of boron on the working curve, and calculate the concen-
tration (mgB/Z) of boron in the sample.
Working curve Pipet in stages 1 t o 10ml of boron standard solution
(0.1 pgB/ml) in as many 50 ml separatory funnels as the stages, respectively
carry out the operations in (a)to (f), and draw the relation curve between
the quantities of boron (B) and absorbances.
Notes (1) When there coexist a lot of organic substances in sample, take a
definite amount of sample in a platinum dish, add 0.1 g of so-
dium carbonate specified in JIS K 8625, evaporate it t o dryness,
and fuse it. After letting it cool, add water, heat it t o dissolve
the melt, add sulfuric acid (3+97) to neutralize, and make the
volume of liquid definite.
Take a suitable amount (containing 0.1 to 1pg of boron) of
this solution in a separatory funnel, add water t o make up the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
volume t o 15 ml, add 3 ml of sulfuric acid (3+97) and 3 ml of hy-
drofluoric acid (1+9), shake it, and let it stand for about 1 h. Add
3 ml of methylene blue solution (0.4g/Z), after shaking add 10 ml
of 1,2-dichloroethane, shake violently for about 1min, and ex-
tract boron ion association. Hereafter, carry out the operations
in and after (d).
(2) When sample is not neutral, neutralize it with sulfuric acid (3+97)
o r sodium hydroxide solution (40g/Z).
(3) When fluoride ion coexists, the operation in (a)will make boron
lose owing to the extraction of boron, so the operation in Note
(1) is needed.
(4) The separation of 1,Z-dichloroethanelayer from water layer takes

considerable duration.
(5) This extraction can eliminate anion surface-active agent and the
like in sample.
286
K O101 : 1998
Remarks 1 Though chromate ion gives a disturbance, the operation that

a few drops of hydrogen peroxide (1+100)is added and then
excess of hydrogen peroxide is decomposed by boiling, can elimi-
nate this disturbance.
2 When methylene blue is attached on a separatory funnel or
absorption cell, it should be removed by rinsing with ethanol.
46.2 Azomethine H absorptiometry The absorbance of yellow complex gener-

ated by reaction of boric acid to azomethine H [8-N-(2-hydroxybenzylidene)-amino-
l-hydroxy-3,6-naphthalenedisulfonatelat about pH 6 is measured to determine boron.
Determination range: B 5 t o 25 pg
Repeatability: 3 t o 10 % of coefficient of variation
Remarks 3 This applies t o the sample of little turbidity.
(i) Reagents The following reagents shall be used. Preserve in a polyethylene bottle.
Water Follow 45.1 (1)(a).
Azomethine H solution Dissolve 1.0 g of azomethine H-sodium salt [8-
N-hydroxybenzylidene)-amino-l-hydroxy-3,6-napht halenedisulfonate-sodium
salt] and 3.0 g of L(+)-ascorbicacid specified in JIS K 9502 in a little amount
of water, transfer it into a 100 ml volumetric flask and add water to the
marked line. Preserve in a polyethylene bottle. This solution is stable for
one week if preserved in a dark place at 4 t o 6 O C .
Buffer solution (pH 5.9) Add 250 g of ammonium acetate specified in JIS
K 8359, 15 ml of sulfuric acid specified in JIS K 8951,5 ml of phosphoric
acid specified in JIS K 9005, 1.0 g of citric acid monohydrate specified in
JIS K 8283 and 1.0g of disodium dihydrogen ethylenediamine tetraacetate
dihydrate specified in JIS K 8107 in 250 ml of water, and dissolve by heating.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Azomethine H mixed solution Mix equal volume of azomethine H so-
lution and buffer solution (pH 5.9). Prepare when it is used.
Boron standard solution ( i pgB/ml) Follow 45.1 (i)(h).
(2) Tool and apparatus Tool and apparatus shall be as follows.
(a) Glassware Follow 45.1 (2) (a).
(a) Take a suitable amount (containing 5 t o 25 pg as B) of sample(6) (7) in a
100 ml polyethylene beaker and add water to make 25 ml.
(b) Add 10 ml of azomethine H mixed solution and allow to stand for about 2 h
in a dark place at 20 O C .
(c) Transfer a part of the solution into a absorption cell(') and measure its
absorbance in the vicinity of 410 nm wavelength.
(d) Take 25 ml of water, for a blank test, in a 100 ml polyethylene beaker, carry
out the operation of (b) and ( c ) t o measure the absorbance, and correct the
287
K O101 : 1998
(e) Find the quantity of boron on the working curve, and calculate the concen-
tration (mgBIZ) of boron in the sample,
Working curve Pipet in stages 5 to 25 ml of boron standard solution (1pgB/
mi) in as many 100ml polyethylene beakers, respectively carry out the
operation in (a) t o (d),and draw the relation curve between the quantities
of boron (B) and absorbances.
Notes (6) If suspended solid is contained, remove it by means of filtration
or centrifugation.
(7) 1 to 5 pg of boron can be determined if a 50 mm absorption cell
is employed.
Remarks 4 In this method, sodium, potassium, calcium, magnesium, zinc,
phosphate, sulfate or nitrate does not interfere,
Iron, manganese, aluminium, copper chromium, beryllium,
titanium, vanadium or zirconium gives positive error.
45.3 ICP atomic emission spectrometry Spray sample in inductively coupled

plasma, measure the emission by boron at 249.773 nm wavelength, and determine
boron.
Determination range: B 20 t o 8 O00 kg/2
Repeatability: 2 t o 10 % of coefficient of variation (depending on apparatus and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(i) Reagents Reagents shall be as follows, and stored in polyethylene bottles.

(a) Water Follow 45.1 (i)(a).
(b) Boron standard solution (20 pgB/ml) Pipet 50 ml of boron standard so-
lution (0.1 mgB/ml) stated in 45.1 (i)(g) in a 250 ml volumetric flask, and
add water up to the marked line. Prepare this solution just before use.

(a) Glassware Follow 45.1 (2) (a).
(b) ICP emission spectrometer

(a) Spray the sample(6) in a plasma through the sample introducing part in
accordance with 5.8 of JIS K 0116, and measure emission strength at
249.773 nm wavelength(8) (9) (lo).
(b) Take water for a blank test, carry out the operation in (a),and correct the
emission strength obtained on sample.
(c) Find the quantity of boron on the working curve, and calculate the concen-
tration of boron (pgBIZ) in the sample.
Working curve Pipet in stages 0.1 t o 40 ml of boron standard solution
(20pgBíml) in as many 100ml volumetric flasks as the stages, and add
respectively water up t o the marked line. Carry out the operations in (a)
respectively. Separately, take water for a blank test, and carry out the
operation in (a), correct the emission strength obtained on the standard
288
K O101 : 1998
solution, and draw the relation curve between the quantities of boron (B)
and emission strengths. Prepare the working curve when sample is mea-
sured.
Notes (8) The apparatus capable of simultaneous measuring of 2 or more
spectra with different wavelength can use an internal standard
method. The procedure for the internal standard method shall
be as follows. Take a suitable amount of sample in a 100ml
volumetric flask, add 10 ml of yttrium solution (50 p.gY/ml) [Dis-
solve 0.318 g of yttrium (III) oxide in 5 ml of high purified nitric
acid specified in JIS K 9901 with heating, dispel nitrogen ox-
ide, cool it, transfer it in a 250 ml volumetric flask, and add water
up t o the marked line. Pipet 10 ml out of this solution in a 200 ml
volumetric flask, and add water up to the marked line.], and add
water up t o the marked line. Carry out spraying of this solu-
tion as described in (3)(a),measure its emission strength at
249.773 nm and 371.029 nm (yttrium) simultaneously, and ob-
tain the emission-strength ratio of boron and yttrium.
Separately, take in stages 0.1 to 40 ml of boron standard so-
lution (20 pgB/ml) in as many 100 volumetric flasks as the stages,
add respectively 10 ml of yttrium solution (50 pgY/ml), and re-
spectively add water up t o the marked line. Carry out spraying
of these solutions as stated in (3)(a),measure respectively its
emission strength a t 249.773 nm and 371.029 nm wavelength si-
multaneously, draw the relation curve between the concentration
of boron and the emission-strength ratio of boron to yttrium, and
make this the working curve. On this working curve, find the
quantity of boron that is corresponding to emission-strength ra-
tio obtained from the sample, and calculate the concentration of
boron (ygB/Z) in the sample.
(9) When the sample with high salts concentration interferes applying
working curve method, it would be preferable t o employ the stan-
dard addition method described in 5.8.3 (2) of JI$ K 0116. In
this case, background correction should be done irrespective of
kind of the sample.
(10) In case of the apparatus capable of using emitting higher order
spectrum, the measurement may be made using higher order
spectrum.
Another wavelength may be adopted, if its accuracy and pre-
ciseness have been previously confirmed.
Remarks 5 When the solution containing boron is sprayed in a plasma
torch, its memory effect is longer than other elements, there-
fore prior t o spraying next solution, water spraying of suffi-
cient duration is needed for elimination of memory effect caused
by the last sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
289
K O101 : 1998
46 Arsenic (As) For the determination of arsenic, silver diethyldithiocarbamate

absorptiometry, hydride-generation atomic absorption method or hydride-generation
ICP atomic emission spectrometry shall be applied.
46.1 Silver diethyldithiocarbamate absorptiometry After oxidizing arsenic

by potassium permanganate, coprecipitate it with iron (III) hydroxide, separate the
arsenic, and concentrate. Dissolve the precipitation with sulfuric acid and hydro-
chloric acid, generate arsenic hydride by adding potassium iodide, tin (II) chloride,
and zinc, make this be absorbed in the chloroform solution of silver diethyldithio-
carbamate (silver&, IV-diethylcarbamo-dithioicacid), measure the absorbance of reddish
violet produced in the solution, and determine arsenic.
Determination range: As 2 t o 1Oyg
Repeatability: 2 t o 10 % by coefficient of variation.
Hydrochloric acid For arsenic analysis specified in JIS K 8180. Use
this one for the preparation of reagents and operations.
Hydrochloric acid (l+l) Prepare using the hydrochloric acid described
in (a).
Sulfuric acid (1+5) Prepare using the sulfuric acid specified in JIS K
8951.
Aqueous ammonia (1+2) Prepare using aqueous ammonia specified in
JIS K 8085.
Potassium permanganate solution (3gil) Dissolve 0.3g of potassium
permanganate specified in JIS K 8247 in water t o make total 100ml.
Hydrogen peroxide solution (1+30) Prepare using hydrogen peroxide
Iron (III) solution (10mgFe/ml) Dissolve 5 g of iron (III) chloride hexahy-
drate specified in JIS K 8142 or 9 g of ammonium iron (III) sulfate 12-
water specified in JIS K 8982 and 5 ml of hydrochloric acid stated in (a)
in water to make total 100ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Potassium iodide solution (200 g/Z) Dissolve 20 g of potassium iodide

Specified in JIS K 8913 in water t o make total 100 ml.
Tin (II) chloride solution Dissolve 40 g of tin (II) chloride dihydrate speci-
fied in JIS K 8136 in hydrochloric acid stated in (a) t o make total 100 ml.
Add 2 or 3 pieces of tin specified in JIS K 8580 in the solution and pre-
serve in a coloured glass bottle. When using, take out a suitable amount,
and dilute it 10 times with water.
Lead (II) acetate solution (100gll) Dissolve 1 2 g of lead (II) acetate
trihydrate specified in JIS K 8374 in 1 or 2 drops of acetic acid specified
in JIS K 8355 and water t o make total 100ml.
290
K O 1 0 1 : 1998
Zinc Screen zinc for arsenic analysis specified in JIS K 8012 through the
test sieve specified in JIS Z 8801, and gather the zinc which has passed
1400 pm sieve opening and stopped on 1 O00 pm opening.
Silver diethyldithiocarbamate solution Dissolve 0.25g of silver N , N -
diethyldithiocarbamate specified in JIS K 9512 and 0.1g of brucine n-hy-
drate (2,3-dimethoxystrychinizine-lO-onen-hydrate) specified in JIS K 8832
in chloroform specified in JIS K 8322 to make total 100ml.
Metacresol purple solution (1g/Z) Dissolve 0.1 g of metacresol purple
specified in JIS K 8889 in 50 ml of ethanol (95) specified in JIS K 8102,
and add water to make total 100ml.
Chloroform Specified in JIS K 8322.
Arsenic standard solution (0.1 mgAs/ml) Heat arsenic oxide (III), ref-
erence material for volumetric analysis, (diarsenic trioxide) specified in JIS
K 8005 at 105 "C for about 2 h, and let it cool in a desiccator. Weigh its
0.132 g as 100 % of As.203, dissolve in 2 ml of sodium hydroxide solution
(40 g/Z), add water t o make total 500 ml, add sulfuric acid (1+10) to make
it very slightly acidic, transfer it in a 1O00 ml volumetric flask, and add
water up to the marked line. Otherwise, use arsenic reference material,
standard solution, As 100, specified in JIS K 0026.
Arsenic standard solution (i pgAs/ml) Pipet 10 ml of arsenic standard
solution (0.1 mgAs/ml) in a 1O00 ml volumetric flask, and add water up to
the marked line. Prepare this solution each time it is needed.

(a> Arsenic hydride generator Fig.46.1 and Fig. 46.2 show its example.
Unit: mm
A: Arsenic hydride generat-

ing bottle 100 ml
B: Introducing tube
b: Glass wool moistened
with lead (II) acetate
solution (100 g/Z)
C: Absorbing tube for ar-
senic hydride (with
ground stopper)
__
H D: Rubber stopper
46.1 --&ample of arsenic hydride generator
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
.
29 1
K O101 : 1998
Unit: mm
A: Arsenic hydride generat-
ing bottle 100ml
B: Introducing tube
b: Glass wool moistened
with lead (II) acetate
solution (100gíl)
C: Absorbing tube for ar-
senic hydride (with
ground stopper)
b
/--
D: Tube for zinc putting in
E: Ground flat surface
F: Spring for pressing
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
i
0
-1
1
I
L- Ii
Fig. 46.2 Example of arsenic hydride generator
(3) Concentrating operation Concentrating operations shall be as follows.

Take a suitable amount (containing 2 t o 10 pg as As) of sample, add 3 ml
of nitric acid per 1I of the sample, and drip potassium permanganate solu-
tion (3gll) for colouring the solution.
Boil this solution, and when the colour by permanganate disappears drip
further potassium permanganate solution ( 3 g/Z), followed by boiling on (1).
Decompose the excess permanganate by dripping of as little hydrogen per-
oxide (1+30) as possible.
Add 2 ml of iron (III) solution (10 mgFe/ml) and 2 or 3 drops of metacresol
purple solution (1g/Z) as indicator, and while keeping the solution at about
80 "C neutralize stirring it with aqueous ammonia (1+2) until the colour of
the solution turns purple(2).
292
K O101 : 1998
(e) After settling down the precipitation, filtrate it through small size filter
paper 5 grade A.
Notes (1) When decomposing is difficult because of organic substance dis-
turbing coprecipitation, decompose through the operation in 4.3
o r 4.4, and then carry out the operation in (d).
(2) The coprecipitation of arsenic brought by iron (III) hydroxide is
suitably carried out a t pH 9 to 10.

Transfer the precipitation obtained in (3)( e )in an arsenic hydride gener-
ating bottle with as little water as possible by rinsing. Dissolve the pre-
cipitation attached t o the filter paper with 18 ml of warm sulfuric acid (1+5)
and 2 ml of hydrochloric acid (l+l), wash the filter paper, put this solution
in the arsenic hydride generating bottle to dissolve the above precipitation,
and add water t o make total about 40ml.
Add 15 ml of potassium iodide solution (200 g/Z) and 5 ml of tin (II) chlo-
ride solution, shake it, and let it stand for about 10 min.
Connect an arsenic hydride generating bottle, introducing tube, and arsenic-
hydride absorbing tube which contains 5 ml of silver diethyldithiocarbamate
solution, and then put in swiftly about 3 g of zinc into the arsenic hydride
generating bottle(3). Place the bottle in a water bath at 25 O C , let it stand
for about 1h, make the arsenic hydride be absorbed in silver diethyl-
dithiocarbamate solution, and colour it.
Add chloroform in this solution to make total 5 ml.
Transfer a part of this solution in an absorption cell, and measure its ab-
sorbance in the vicinity of 510nm wavelength with making chloroform a
reference solution.
Take 2 ml of iron (III) solution (10 mgFe/ml) for a blank test in the arsenic
hydride generating bottle of the arsenic hydride generator, add 18ml of
sulfuric acid (1+5)and 2 ml of hydrochloric acid (l+l),add water up to about
40m1, carry out the operations in (b)t o (d), measure its absorbance, and
correct the absorbance obtained on the sample.
Find the quantity of arsenic on the working curve(4), and calculate the con-
centration (pgAs/Z) of arsenic in the sample.
Working curve Pipet step by step 2 t o 10ml of arsenic standard solu-
tion (ipgAs/ml) in as many arsenic hydride generating bottles, respectively
add 2 ml of iron (III) solution (10 mgFe/ml), 18 ml of sulfuric acid (1+5) and
2 ml of hydrochloric acid (l+l), and add water to make it total 40 ml. Carry
out the operations in (b) t o (f) simultaneously with that on the sample,
measure each absorbance, and draw the relation curve between the quan-
tities of arsenic (As) and absorbances.
Notes (3) When using the arsenic hydride generator shown in Fig. 46.2,
put about 3 g of zinc in the tube for zinc putting in, and after
connecting the absorbing tube, add the zinc in the sample by
revolving the tube for zinc putting in.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
293
K O101 : 1998
(4) Because this working curve changes its inclination, it shall be

prepared when test is carried out.
46.2 Hydride-generation atomic absorption method Pretreat the sample to

change arsenic into arsenic hydride, introduce it into hydrogen-argon flame, mea-
sure an atomic absorption by arsenic at 193.7nm wavelength, and determine ar-
senic.
Determination range: As 5 to 50pg/Z
Hydrochloric acid Follow 46.1 (i)(a).
Hydrochloric acid (l+l) Prepare using the hydrochloric acid described
in (a).
Sulfuric acid (l+l) Follow 4.4 ( i )(b).
Potassium permanganate solution (3 gll) Follow 46.1 (i)(f).
Potassium iodide solution (200 g / l ) Follow 46.1 (i) (i).
Tin (II)chloride solution Dissolve 10 g of tin (II) chloride dihydrate speci-
fied in JIS K 8136 in 100 ml of hydrochloric acid described in (a).
Zinc powder For arsenic analysis, specified in JIS K 8013.
Iron (III) solution (10 mgFe/ml) Follow 46.1 (1) (h).
Argon Argon grade 2 specified in JIS K 1105.
Arsenic standard solution (0.1 pgAs/ml) Pipet 10 ml of arsenic stan-
dard solution (i pgAs/ml) stated in 46.1 (i)(9)into a 100 ml volumetric flask,
add 2 ml of hydrochloric acid (l+i) in (b), and add water up to the marked
line.

(a) Arsenic hydride generator Fig. 46.3 shows its example.
(b) Atomic absorption analyzer One capable of correcting background.
(c) Arsenic hollow cathode lamp
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
294
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
- I f ,
I 11 c- 8
E - - __
--- __ _
_ A: Cock
-
Argon
I
,-
1-1
- _ __
emri-1,
€3:Reaction vessel
C: Reservoir
Fig. 46.3 Example of construction of arsenic hydride generator

Take a suitable amount (containing 0.1 to 1 pg as As) of sample(5)in a 100 ml
beaker(", add 1 ml of sulfuric acid (1+1) and 2 ml of nitric acid, and drip
potassium permanganate solution (3 g / l ) until the solution colours.
Heat it on a hot plate(? until white fume of sulfuric acid is generated(*).
Let it cool t o room temperature, add 4 ml of hydrochloric acid (l+l), heat
gently to dissolve residue, let it cool, transfer it in a reaction vessel of the
arsenic hydride generator, and add water to make total 20ml.
Add 2 ml of potassium iodide solution (200 g/Z), 2 ml of tin (II) chloride solution
and 1 ml of iron (III) solution (10 mgFe/ml), shake them and let it stand
for about 15min.
Connect the arsenic hydride generator and atomic absorption analyzer, replace
air in this system with argon, add promptly(9) (10) 1.0 g of zinc powder in
the reaction vessel, run a magnetic stirrer, and generate arsenic hydride (11).
Rotate the cock t o introduce arsenic hydride into hydrogen-argon (12) flame,
and read the indicated value(l3) at 193.7 nm wavelength.
Take the same amount of water as that of the sample for a blank test, carry
out the operations in (a) t o (f), read indicated value, and correct the value
obtained on the sample.
Find the quantity of arsenic o n the working curve, and calculate the con-
centration of arsenic (pgAsll) in the sample.
Working curve Pipet step by step 1 t o 10ml of arsenic standard solu-
tion (0.1 pgAs/ml), carry out the operations in (c) t o tg), and draw the re-
lation curve between the quantities of arsenic (As) and indicted values.
Prepare this working curve when sample is tested.
Notes (5) When testing the sample containing no organic substance, nitrate,
nor nitrite, the following procedures are effective instead of (a)
to (cl.
295
K O101 : 1998
Take a suitable amount (containing 0.1 to 1 pg as As) of sample

in a 100 ml beaker, add 4 ml of hydrochloric acid (l+l),heat it
for several minutes a t near boiling, and let it cool. Transfer it
in the reaction vessel, and add water up to 20 ml.
When a lot of organic substance is contained, the following
procedures may be available instead of (a)and (b).
Add 1 ml of sulfuric acid ( l + l )2, ml of nitric acid, and 3 ml
of perchloric acid specified in JIS K 8223,into the suitable amount
of sample, and heat it until white fume is generated in order t o
decompose the organic substance.
Be careful about borosilicate glass because sometimes it contains
arsenic. Beakers made of tetrafluoroethylene resin should be
preferably used.
If colour by permanganate disappears while heating, supplement
potassium permanganate solution ( 3 g/Z).
The existence of nitric acid interferes the generation of arsenic
hydride, therefore the sufficient generation of white fume of sulfuric
acid is necessary to eliminate nitric acid.
Arsenic hydride tends t o be abruptly generated immediately af-
ter adding powdered zinc, it should be attended not t o let it es-
cape.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Because zinc powder contains a trace of arsenic, added amount

of zinc power should be kept constant. For this purpose the fol-
lowing methods should be taken: (i)adding a zinc tablet made
by being shaped with binder, (2) adding dense water suspensoid
of zinc powder using a fountain pen filler, and (3) adding a defi-
nite amount of zinc powder that is wafered.
Instead of zinc powder, sodium tetrahydroborate can be used.
In this case, addition of tin (II) chloride solution and iron (III)
solution is not needed. (The generating condition of arsenic hy-
dride by sodium tetrahydroborate is dependent on a n arsenic
hydride generator.)
Instead of hydrogen-argon flame a thermal absorption cell can
be used.
When adding sodium tetrahydroborate, there is such a way
to add bit by bit its solution or using tablet. The acid concen-
tration during addition varies according t o a generator, so it should
be attended. Generally it is done under about 1mold, but the
adding amount of hydrochloric acid ( l + l )a t ( c ) should be con-
trolled t o small side according to the remnant of sulfuric acid a t
operation (b) in (3).
The amount of gas collected is known by either pressure o r vol-
ume.
Instead of argon, high purity nitrogen grade 2 specified in JIS
K 1107 may be used.
(13) Absorbance or its proportional value is valid.
296
K O101 : 1998
Remarks 1 For concentrating arsenic hydride only, fix arsenic hydride using
a cold trap where a U-tube filled with glass beads is soaked
in liquid nitrogen. After pulling up the U-tube, recover it to
room temperature as its both ends are closed, and send gas-
ified arsenic hydride into flame using argon.
2 Instead of Fig. 46.3, a continuous-type hydride generator,
employing sodium tetrahydroborate, can be used. Fig. 46.4
shows its example. The procedures needed in this case are as
follows.
c A: Pump
Argon 4-
1 BI, Bz:
C:
D:
Mixing joint
Reaction tube
Pressure gauge
Sodium tetra- 52 E: Flowmeter
hydroborate
solution
2
hm
C
j-
Bi ;--II
Sample :
A . 1 WI I
!i I
Hydrochloric acid : f’
I
l
(Potassiumiodide - -i-- ---------J
solution)
Fig. 46.4 Example of continuous-type hydride generator

After letting cool the sample carried out with operations in
46.2 (3)(a) and (b),transfer it in a 20 ml volumetric flask, and
add water up to the marked line. While flowing argon in the
apparatus, introduce continuously this solution, hydrochloric
acid (i to 6 mol/Z), and sodium tetrahydroborate solution [Dis-
solve 5 g of sodium tetrahydroborate in 500 ml of sodium hy-
droxide solution (0.1 mol/Z).] into the apparatus with the flow
rate of 1t o 10 mlímin respectively (Such factors as flow rate
of sample, hydrochloric acid, sodium tetrahydroborate solution,
and concentration of hydrochloric acid and sodium tetra-
hydroborate solution are different according to type of a n
apparatus.) using a constant delivery pump in order t o pro-
duce arsenic hydride.
After separating generated arsenic hydride from solution,
introduce the gas containing arsenic hydride into hydrogen-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
argon flame or a thermal absorption cell, and read the indi-

cated value at 193.7 nm wavelength. For a blank test, carry
out the operations in 46.2(3)(a)and (b)on water, read the
indicated value similarly t o the sample side, and correct the
value obtained on the sample. On the working curve, find the
quantity of arsenic, and calculate the concentration of arsenic
(pgAs/Z) in the sample.
297
K O101 : 1998
Iron, nickel, o r cobalt, if each coexists over the degree as 5,

10, or 80 times or over as amount of arsenic respectively, dis-
turbs the generation of arsenic hydride. If either 4 ml potas-
sium iodide solution (100 g/Z) is added when the sample is made
20 ml by introducing in a volumetric flask after pretreatment,
or potassium iodide solution (20 t o 100 gll) is introduced into
an arsenic hydride generator together with other solution a t
the rate of 1 t o 10mlímin (Concentration and flow rate de-
pend on an apparatus.), the disturbance by iron can be elimi-
nated even when iron coexists as 1O00 times as the existence
of arsenic.
Working curve Take step by step 1to 10 ml of arsenic stan-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
dard solution (0.1 pgAs/ml)(*) in as many 20 ml volumetric
flasks, and add water up to the marked line. Hereafter treat
respectively similarly t o the sample side, and draw the rela-
tion curve between the quantities of arsenic (As) and indicated
values. Prepare the working curve when the sample is mea-
sured.
Note (*) For thermal absorption cell, the sensitivity is supe-
rior 10 to 50 times compared with hydrogen-argon
flame (depending on apparatus and operating con-
ditions), therefore, reduce the amount of arsenic stan-
dard solution ( O . 1 pg/ml) suitably.
46.3 Hydride-generation ICP atomic emission spectrometry Pretreat sample

t o make arsenic arsenic hydride, spray it, through a sample introducing part, into
an inductively coupled plasma, and measure the emission of arsenic a t 193.696 nm
wavelength to determine arsenic.
Determination range: As 1 t o 10pglZ
Repeatability: 3 to 10 % by coefficient variation (depending on apparatus and
operating conditions)
(1) Reagents Use reagents below.

Hydrochloric acid Follow 46.1 (1) (a).
Nitric acid As specified in J I S K 8541.
Sulfuric acid ( l + l >Follow 4.4 ( i )(b).
Potassium bromide solution ( i moUZ) Dissolve 11.9 g of potassium bro-
mide specified in J I S K 8506 in water t o make 100 ml.
Sodium tetrahydroborate solution (10 g/Z) Dissolve 5 g of sodium
tetrahydroborate in sodium hydroxide solution (O. 1mol/,!) t o make 500 ml.
Prepare when it is used.
Argon Follow 46.2 ( i )ci).
Arsenic standard solution (0.1 pgAs/ml) Follow 46.2 ( i )(k).
298
K O 1 0 1 : 1998

(a) Arsenic hydride generator One with continuous type.
(b) ICP atomic emission spectrometer
Take suitable amount (containing 0.05 t o 0.5 pg as As) of sample(14) in a
beaker, add 1 ml of sulfuric acid (i+l) and 2 ml of nitric acid, heat it on a
hot plate t o generate white fume of sulfuric acid.
Cool it t o room temperature, add 8 ml of water, 8 ml of hydrochloric acid
and 8 ml of potassium bromide solution (i mol/Z), and heat for about 50 min
at about 50°C.
Cool it to room temperature, transfer this solution into a 50 ml volumetric
flask and add water up t o the marked line.
Connect the arsenic hydride generator with the ICP atomic emission spec-
trometer, and introduce the solution of (cl,sodium tetrahydroborate solu-
tion (10 g/Z) (15) and hydrochloric acid (1to 6 mol/Z) (15) continuously with a
constant pump while flowing argon to generate arsenic hydride.
Spray arsenic hydride through a sample introducing part into the plasma
and measure the emission strength at 193.696 nm wavelength.
the operation (a) to (e), and correct the emission strength obtained on the
sample.
Find the amount on the working curve and calculate the concentration
(pgAs/Z) of the arsenic in the sample.
Working curve Take 0.5 t o 5 ml of arsenic standard solution (0.1 pgAs/
mi) step by step in as many 50 ml volumetric flasks, add acid and potas-
sium bromide solution (i mol/Z) t o make the same concentration as that for
the sample pretreated, and add water up t o the marked line. Carry out
the operation (d) to (f) and draw a relation curve between the amount of
arsenic and the emission strength. Prepare the working curve when the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
sample is measured.
Notes (14) When sample contains a lot of organic matter, add 1 ml of sul-
furic acid (l+l), 2 ml of nitric acid and 3 ml of perchloric acid
specified in JIS K 82.23 in the operation (a) and decompose the
organic matter by generating white fume.
(15) The concentration of hydrochloric acid and sodium tetrahydro-
borate solution depends on the apparatus.
(16) The flowing amount of sample, hydrochloric acid o r sodium
tetrahydroborate solution depends on the apparatus.
Remarks 3 Plasma becomes unstable sometimes because hydrogen which
is by-produced at hydride generation is introduced into plasma,
therefore, be careful that the amount of hydrogen is not too
much in the initial introduction.
299
K O101 : 1998
4 It may be influenced by acid and salt if they coexist. When it

is influenced, use an interference retardant in accordance with
hydride-generation atomic absorption method, as appropriate.
5 If a working curve method cannot be applied because the salts
concentration of sample is dense, it is preferable to use the
standard addition method specified in 5.8.3 (2) of JIS K 0116.
In this case, the background correction must be carried out
irrespective of type of sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
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K O101 : 1998
47 Sodium (Na) For the determination of sodium, flame emission photometry,

flame atomic absorption method, ion-selective electrode method, or ion chromatog-
raphy shall be applied.
47.1 Flame emission photometry Spray sample in acetylene-air flame, hydro-

gen-oxygen flame or the like, and measure the strength of emission at 589.0 nm wave-
length t o determine sodium.
Determination range: Na 30 t o 300 pg/Z, 0.3 to 3 mgll, 3 to 30 mgll
(a) Sodium standard solution (i O00 mgNalE) Heat sodium chloride, stan-
dard reagent for quantitative analysis, specified in JIS K 8005 a t 600 "C
for about 1h, and let it cool in a desiccator. Take its 2.542 g as NaC1 100 %,
dissolve it in a little of water, transfer it in a 1O00 ml volumetric flask,
and add water up t o the marked line. Store it in a polyethylene bottle.
(b) Sodium standard solution (3 to 30 mgNalZ) Take step by step several
aliquots from sodium standard solution (i O00 mgNaíZ), and dilute them with
water to prepare standard solutions of 3 t o 30 mgNalZ (l).
Note (1) When measuring a low concentration, prepare the standard solu-
tion with 30 t o 300 pgNalZ or 0.3 to 3 mgNalZ.
(a) Flame photometer
(a) Spray sodium standard solution (30 mgNalZ)(2)in the flame of a flame pho-
tometer, and adjust its indicated value at 589.0nm wavelength to be 100.
(b) Spray water, and adjust the indicated value t o be O .
(c) Successively spray sodium standard solution (3 t o 30 mgNa/Z)(l),and draw
the relation curve between the concentrations of sodium (Na) and indicted --`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
values t o prepare the working curve.

(d) Spray sample(3) (4) (In case of sodium concentration of 30 mglZ o r more, it
should be diluted.), read indicated value, and find the concentration of sodium
(mglZ) in the sample on the working curve.
Notes (2) When measuring a low concentration, use the standard solution
with 3 mgNall or 0.3 mgNdZ.
(3) When suspensoid is contained, eliminate it by filtration o r cen-
trifugal separation.
(4) When interfering substances are contained, either of the follow-
ing shall be adopted: measure after diluting sample to the level
of neglecting their influence; or prepare sodium standard solu-
tions (3 to 30 m g N d ) containing interfering substance as the same
level as that in sample and then draw the working curve.
301
K O101 : 1998
Remarks 1 Coexistence of potassium and calcium gives a positive error.

Coexistence of lithium, barium, free acid, phosphate, borate,
oxalate, silica, glucose, gelatine, and so on gives a negative
error. Magnesium and sulfate ion give almost no influence.
When a lot of silicate coexists, take a suitable amount of
sample in a quartz glass beaker or platinum dish, add hydro-
chloric acid (l+l) to make it acidic, and evaporate to dryness.
After cooling it, add 5 drops of hydrochloric acid (l+l) and a
little of water, heat it to dissolve, filtrate and make it defi-
nite volume with water.
47.2 Flame atomic absorption method Spray sample into a flame such as acety-
lene-air flame, and measure the atomic absorption by sodium at 589.0 nm wavelength
to determine sodium.
Determination range: Na 0.05 t o 4mglZ

(a) Sodium standard solution (0.1 mgNa/ml) Pipet 10 ml of sodium stan-
dard solution (1O00 mgNalZ) stated in 47.1 (1)(a) in a 100 ml volumetric
flask, and add water up t o the marked line. Prepare this solution each
time it is needed.
(b) Sodium standard solution (10 pgNa/ml) Pipet 20 ml of sodium stan-
dard solution (0.1 mgNa/ml) in a 200 ml volumetric flask, and add water
up to the marked line. Prepare this solution each time it is needed.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Flame atomic absorption analyzer
(b) Sodium hollow cathode lamp
Spray the sample(3) in flame according t o 6 of JIS K 0121,and read indi-
cated value (5) a t 589.0 nm wavelength.
Take water for a blank test, carry out the operation in (a),and correct the
indicated value obtained on the sample.
Find the quantity of sodium on the working curve, and calculate the con-
centration of sodium (pgNalZ) in the sample.
Working curve Pipet stepwise 0.5 t o 40 ml of sodium standard solution
(10 pgNdml) in as many 100 ml volumetric flasks, and respectively add water
up to the marked line. Carry out the operation in (a) on this solution.
Separately carry out the operation in (a) on water as a blank test, correct
the indicated value obtained on the standard solution, and draw the rela-
tion curve between the quantities of sodium (Na) and indicated values.
Note ( 5 ) Absorbance o r its proportional value is valid.
302
K O 1 0 1 : 1998
47.3 Ion-selective electrode method Control pH of sample to be 10.2 to 10.6,

and measure electric potential using sodium ion-selective electrode as an indicator
electrode in order to determine sodium.
Determination range: Na 1 t o 100mglZ

(a) Tris buffer solution Take 60 g of 2-amino-2-hydroxymethyl-1,3-propanediol
[tris(hydroxymethyl)aminomethanel specified in JIS K 9704,and dissolve
in water to make total 11.
(b) Sodium standard solution (100mgNalZ) Pipet 20 ml of sodium stan-
dard solution (1O00 mgNalZ) stated in 47.1 (1)(a) in a 200 ml volumetric
flask, and add water up to the marked line.
(c) Sodium standard solution (10 mgNalE) Pipet 20 ml of sodium standard
solution (100 mgNalZ) in a 200 ml volumetric flask, and add water up to
the marked line.
(d) Sodium standard solution (1 mgNalZ) Pipet 20 ml of sodium standard
solution (10 mgNalZ) in a 200 ml volumetric flask, and add water up to the
marked line.
( 2 ) Apparatus Apparatus shall be as follows.
(a) Potentiometer Follow 31.2 (2) (a).
(b) Indicator electrode Sodium electrode [Glass membrane (6) electrode]
(c) Reference electrode Follow 31.2 (2)( c ) . Provide that the liquid kept
in outer cylinder should be ammonium nitrate solution (100 glZ) or potas-
sium nitrate solution (100 g/Z).
(d) Magnetic stirrer Follow 31.2 (2)(d).
Note (6) When a sodium electrode made of liquid membrane is used, fol-
low Remarks 4.
(3) Preparation of working curve The preparation of working curve shall be

as follows.
Take 100 ml of sodium standard solution (1mgNalZ) in a 200 ml beaker,
and add 10 ml(7) of tris buffer solution.
Immerse an indicator electrode ( 8 ) (9) and a reference electrode (10) (111, strongly
stir i t ( 1 3 ) with a magnetic stirrer(l2) to the degree not to let bubbles touch
the electrode.
Measure temperature of the solution, and measure electric potential (14) with
a potentiometer.
Take each 100 ml of sodium standard solution (10 mgNalZ) and sodium stan-
dard solution (100 mgNaA) in two 200 ml beakers, and add respectively 10 ml
of tris buffer solution(7). Control the temperature of each solution to be
within f 1"C of the temperature a t ( c ) , carry out the operations in (b)and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
303
K O101 : 1998
( c ) , and measure the electric potential of sodium standard solution (10 t o

100 mgNalZ )( 14).
(e) Graduate the concentration of sodium (mgNalZ) on logarithm axis of semi-
logarithm section paper and the electric potential (mV) on uniform axis,
and draw the relation curve between the concentrations of sodium and electric
potentials (15).
Tris buffer solution is for adjustment of pH as 10.2 t o 10.6 while
measuring and t o make the ionic strength constant.
Immerse a sodium ion-selective electrode in sodium standard
solution (1mgNalZ) when it is used, and after a point gives Sta-
bility, begin to measure electric potential.
Follow Note (12) in 31.
The response time of sodium ion-selective electrode is 2 to 3 min
when solution temperature is 10 t o 3 0 ° C .
The difference of potential between sodium standard solution
1mgNa4 and 100 mgNaL shall fall within 100 to 120 mV (25 OC).
And the working curve in the range of 1t o 100 mgNa/Z of so-
dium concentration becomes straight.

(a) Take 100 ml(16) of sample in a 200 ml beaker, and add 10 mlP) of tris buffer
solution. Adjust solution temperature within k1 "C of the solution tem-
perature at (3)(c).
(b) Carry out the operations in (3)(b)and ( c ) , find the concentration of sodium
on the working curve, and calculate the concentration of sodium (mgNalZ)in
the sample.
Note (16) When tris buffer solution cannot give pH 10.2 t o 10.6 because of
acidity o r strong alkalinity of sample, in advance place sample
in a 200 ml measuring flask, adjust its pH t o be about 10 using
aqueous ammonia (l+l) or hydrochloric acid (1+2), and add wa-
ter up t o the marked line. Take 100 ml from this solution, carry
out the operations in and after (a),correct the measured concen-
tration of sodium (mgNalZ}answering to the rate of dilution, and
obtain the concentration of sodium (mgNalZ) in the sample.
Remarks 2 In case of an ion densitometer, carry out the operations in (3)(a)
t o ( c ) on both sodium standard solution (1 mgNalZ) and sodium
standard solution (100 mgNalZ), and adjust the pointer of ion
densitometer t o show respectively 1 mgNalZ and 100 mgNalZ.
Further, confirm the pointing of the ion densitometer making
use of sodium standard solution (10mgNal2).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
304
K O101 : 1998
3 Allowance limits of main coexisting materials t o sodium ion-

selective electrode (glass membrane) shall be as follows by the
maximum ratio.
Li+: 4.5 x 10, K+: 4 x lo2, "4:' 1.3x lo3, Ag+: 0.2
4 When using sodium ion-selective electrode made of liquid
membrane, use lithium acetate solution (1 mol/Z) or lithium
acetate buffer solution (Dissolve 102 g of lithium acetate
dihydrate in about 500 ml of water, add 2 ml of acetic acid
specified in JIS K 8355,and add water up to 12. This makes
its pH 6 to 7.) as outer-cylinder liquid of a reference electrode.
To prepare working curve, add 10 ml of lithium acetate buffer
solution per 100ml sodium standard solution, control pH as
6 t o 7, and carry out the operations in and after (3)(b). Carry
out measurements under the same condition as when prepar-
ing working curve.
Allowance limits of main coexisting materials in this case
shall be as follows by the maximum ratio.
K+: 1.25 x lo2, Li+: 6.25 x io3, NH$: 1.4 x lo3, Ca2+:1.5 x lo4
Coexisting of surfactant causes drift, so be careful.
47.4 Ion chromatography Sodium in sample shall be determined by ion chro-

matography.
Determination range: Na 0.1 to 30mglZ
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Eluent Follow 36.5 (1) (b).
Reclaiming solution Follow 36.5 (1) (cl.
Sodium standard solution (1 mgNa/ml) Follow 47.1 (1) (a).
Sodium standard solution (0.1 mgNdml1 Pipet 10 ml of sodium stan-
dard solution (1mgNa/ml) in a 100 ml volumetric flask, and add water up
to the marked line.
Alkali metals-ammonium ion mixed standard solution [(0.1mgNH$,
0.1 mgNa, 0.1 mgK)/ml] Follow 36.5 (1)(f).
(2) Tool and apparatus Follow 36.6 (2).

(3) Preparatory operation Preparatory operations shall be carried out accord-
ing to 36.5(3).
(a) Carry out the operations in 36.5 (4) (a) and (b).
305
K O101 : 1998
(b) Read the indicated value(17) of the peak corresponding to sodium o n chro-
matogram.
(c) When the sample is diluted, carry out the operations of (a)and (b)as a
blank test on the same amount of water as the sample, and correct the
(d) On the working curve, obtain the concentration of sodium, and calculate
the concentration of sodium (mgNalZ) in the sample.
Working curve Pipet stepwise 0.1 to 30 ml of sodium standard solution
(0.1 mgNa/ml)(l8)in as many 100 ml volumetric flasks, add respectively water
up to the marked line, carry out the operations in (a)and (b),and read the
indicated value corresponding to each sodium. Separately, carry out the
operations in (a)and (b>on water for a blank test, correct the indicated
value corresponding to each sodium, and draw the relation curve between
the quantities of sodium (Na) and indicated values. Prepare the working
curve when sample is measured.
(18) When ammonium ion and potassium are tested a t the same time,
use alkali metals-ammonium ion mixed standard solution
[(0.1mgNHc+, 0.1 mgNa, 0.1 mgK)/mll.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 5 When sodium concentration is 1 mgll, if ammonium ion and
potassium are respectively 100 mglZ or less, they will not dis-
turb determination.
6 Follow Remarks 9 in 36.
7 Follow Remarks 10 in 36.
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K O101 : 1998
48 Potassium (K) For the determination of potassium, flame emission photom-

etry, fiame atomic absorption method, or ion chromatography shall be applied.
48.1 Flame emission photometry Spray sample into acetylene-air flame, oxy-
gen-hydrogen flame, etc., and measure the strength of emission at 766.5 nm or 769.9 nm
wavelength to determine potassium.
Determination range: K 40 to 400 pglZ, 0.4 t o 4 mglZ, 4 t o 40 mgll
Repeatability: 3 t o 10 % of coefficient of variation
(a) Potassium standard solution (1 O 0 0 mgW1) Heat potassium chloride
specified in JIS K 8121 at 500 to 600 "C for about 1h, and let it cool in a
desiccator. Take its 1.907 g, dissolve in a little of water, transfer it into a
1O00 ml volumetric flask, and add water up to the marked line. Store in
a polyethylene bottle.
(b) Potassium standard solution (4 to 40 mgW1) Take step by step potas-
sium standard solution (lOOOmgWZ), dilute them with water to prepare
standard solution of 4 to 40 mgWZ (1).
Note (1) When measuring a low concentration, prepare the standard solu-
tion with 40 t o 400 pgWZ or 0.4 to 4 mgWZ.
(a) Flame photometer
Spray potassium standard solution (40mgWZ)(2) in the flame of the flame
photometer, and adjust its indicated value at 766.5 nm or 769.9 nm wave-
length t o be 100.
Spray water, and adjust the indicated value to be O .
Successively spray potassium standard solution (4to 40 mgWI)(1),and draw
the relation curve between the concentrations of potassium (K) and indi-
cated values t o prepare the working curve.
Spray sample(3) (4) (In case of potassium concentration of 40 mglZ o r more,
it should be diluted.), read indicated value, and find the concentration of
potassium (mgWZ) in the sample on the working curve.
Notes (2) When measuring a low concentration, use the standard solution
with 4mgWZ or 0.4mgWZ.
(3) When suspensoid is contained, eliminate it by filtration or cen-
trifugal separation.
(4) When interfering substances are contained, either of the follow-
ing shall be adopted: measure after diluting sample t o the level
of neglecting their influence; o r prepare potassium standard so-
lutions (4 to 40 mgWZ) containing interfering substances as the
same level as that in sample, and draw the working curve.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
307
K 0101 : 1998
Remarks 1 Coexistence of sodium gives a positive error. Concerning the

influences by other coexisting substances, follow Remarks 1
of 47.
48.2 Flame atomic absorption method Spray sample in a flame such as acety-
lene-air, and measure the atomic absorption by potassium at 766.5 nm wavelength
t o determine potassium.
Determination range: K 0.05 t o 5 mg/Z
(a) Potassium standard solution (0.1mgWml) Take 10 ml of potassium
standard solution (1 O00 mgWZ) stated in 48.1 (1)(a) in a 100 ml volumet-
ric flask, and add water up t o the marked line. Prepare this solution each
time it is needed.
(b) Potassium standard solution (10pgWm1) Take 20 ml of potassium stan-
dard solution (0.1 mgWml) in a 200 ml volumetric flask, and add water up
to the marked line. Prepare this solution each time it is needed.

(a) Flame atomic absorption analyzer
(b) Potassium hollow cathode lamp
(a) Spray sample(3) in flame according to 6 of JIS K 0121, and read the indi-
cated value a t 766.5 nm wavelength(5).
(b) Carry out the operation in (a>for a blank test using water, and correct the
indicated value in the sample.
(c) Find the quantity of potassium on the working curve, and calculate the
concentration of potassium (mgWZ) in the sample.
Working curve Pipet step by step 0.5 to 50ml of potassium standard
solution (10 pgWml) into as many 100 ml volumetric flasks, and add water
up t o the marked line. Carry out the operation in (a)on this solution. Sepa-
rately, carry out the operation a t (a) as a blank test on water, correct the
indicated value obtained on the standard solution, and draw the relation
curve between the quantities of potassium (K) and indicated values, Pre-
pare the working curve when the sample is measured.
Note (5) Absorbance o r its proportional value is valid.
48.3 Ion chromatography Potassium in a sample shall be determined by ion chro-

matography.
Determination range: K 0.1 to 30mglZ
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
308
K O101 : 1998

(a) Water Water A2 or A3 specified in JIS K 0557.
(b) Eluent Follow 36.5 (1) (b).
(c) Reclaiming solution Follow 36.5 (1) (e).
(d) Potassium standard solution (1 mgWml) Follow 48.1 (1) (a).
(e) Potassium standard solution (0.1 mgK/ml) Pipet 10 ml of potassium
standard solution (1mgWml) into a 100 ml volumetric flask, and add wa-
(0 Alkali metals-ammonium ion mixed standard solution [(0.1 mgNHd+,
0.1 mgNa, 0.1 mgK)/ml] Follow 36.5 (1) (f).
(2) Tool and apparatus Follow 36.5 (2).

(3) Preparatory operation Preparatory operations shall be carried out accord-
ing to 36.5(3).
(a) Carry out the operations in 36.5 (4)(a) and (b).
(b) Read the indicated value(6) of the peak corresponding to potassium on chro-
matogram.
(c) When the sample is diluted, carry out the operations of (a) and (b)as a
blank test on the same amount of water as the sample, and correct the
(d) On the working curve, obtain the concentration of potassium, and calcu-
late the concentration of potassium (mgWZ) in the sample.
Working curve Pipet step by step 0.1 t o 30ml of potassium standard
solution (0.1 mgWml) (7) into as many 100 ml volumetric flasks, respectively
add water up to the marked line, carry out the operations in (a) and (b),
and read the indicated value corresponding to each potassium. Separately,
carry out the operations in (a) and (b) on water for a blank test, correct
the indicated value corresponding t o each potassium, and draw the rela-
tion curve between the quantities of potassium (KI and indicated values.
Prepare the working curve when sample is measured.
Notes (6) It means peak height or peak area.
(7) When ammonium ion and sodium are tested a t the same time,
use alkali metals-ammonium ion mixed standard solution
[(0.1 mgNH4+, 0.1 mgNa, 0.1 mgK)/ml].
Remarks 2' When potassium concentration is 1mgll, if ammonium ion and
sodium are respectively 100 mgll or less, they will not disturb
determination.
When methane amine (monomethylamine) among amines co-
exists in sample, it overlaps with the peak of potassium to dis-
turb.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
309
K O101 : 1998
49 Calcium (Ca) For the determination of calcium, chelatometric titration, flame

atomic absorption method, o r ICP atomic emission spectrometry shall be applied.
49.1 Chelatometric titration Make pH of sample 12 or more, add HSNN [2-

hydroxy-1- I( 2’-hydroxy-4’-sulfo-1’-naphthalenyl)azoI- 3-naphthalenecarboxyl acid (no-
menclature by IUPAC)] as indicator, and titrate the solution with disodium dihydrogen
ethylenediaminetetraacetate solution to determine calcium.
Determination range: Ca 0.2 t o 5 m g

Potassium hydroxide solution Dissolve 250 g of potassium hydroxide
specified in JIS K 8574 in water t o make total 500 ml, store it in a poly-
ethylene bottle.
Potassium cyanide solution (100 g/Z) Dissolve 10 g of potassium cya-
nide specified in JIS K 8443 in water to make total 100 ml. Store it in a
polyethylene bottle.
Hydroxylammonium chloride solution (100 gll) Follow 40.2 (1) ( c ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
HSNN solution Dissolve 0.5 g of 2-hydroxy-l-(2 Hydroxy-4-sulfo-l-

naphthylazo)-3-naphthoic acid specified in JIS K 8776 in 100 ml of metha-
nol specified in JIS K 8891, and add 0.5 g of hydroxylammonium chloride
specified in JIS K 8201. Store in a coloured glass bottle.
10 mmol/Z EDTA solution Heat disodium dihydrogen ethylenediamine-
tetraacetate dihydrate specified in JIS K 8107 at 80 “C for about 5 h, and
let it cool in a desiccator. Dissolve its 3.722 g in a little of water, transfer
it into a 1O00 ml volumetric flask, and add water up t o the marked line.
One milliliter of this solution is equivalent t o 0.400 8 mg of Ca.

(a) Take a suitable amount (containing 5 mg o r less as Ca) of sample(1) in a
beaker, and add water to make about 50ml.
(b) Add 4 ml of potassium hydroxide solution, stir sufficiently, and let it stand
for about 5 min(?.
(c) Add 0.5 ml of potassium cyanide solution (100 glZ) and 0.5 ml of hydroxyl-
ammonium chloride solution (100 g l ) , followed by stirring.
(d) Add 5 o r 6 drops(*) of HSNN solution(3) as indicator, and titrate it with
10 mmol/Z EDTA solution until the solution turns blue from reddish violet.
(e) With the following formula, calculate the concentration (mgCalZ) of calcium
in the sample.
C=ax- ‘Oo x 0.400 8

V
where, C : calcium (mgCa/Z)
a : 10 mmol/Z EDTA solution needed for titration (ml)
310
K O101 : 1998
V : sample (ml)
0.4008: calcium equivalent t o 1 m l of 10mmolA EDTA
solution (mg)
Notes (1) When suspensoid is contained, separate it through filtration or
centrifugal operation.
When sample contains the organic substances and coloured
substances which affect the tests, neutralize it after the opera-
tion in 4. Provided that the method in 4.4 does not apply.
(2) Large amount of precipitation after standing makes the end point
obscure. In this case, the following is preferable: estimate rough
amount of titrant at the first titration, take the same amount of
sample in another beaker, add 10 mmoVZ EDTA solution by the
volume 1ml less than that needed at the first titration, add 4 ml
of potassium hydroxide solution, agitate sufficiently, and let it
stand for about 5 min. Then, add 0.5 ml of potassium cyanide so-
lution (100 gll) and 0.5 ml of hydroxylammonium chloride solution
(100 glZ), and agitate it. Add 5 or 6 drops of HSNN solution as in-
dicator, titrate again it with 10 mmoVZ EDTA solution, and make
it an end point when the solution turns blue.
(3) Instead of HSNN solution, addition of about 0.1 g powder, which
is the ground mixture t o uniform of both 0.5 g of 2-hydroxy-142-
hydroxy-4-sulfo-l-naphthylazo)-3-naphthoic acid specified in JIS
K 8776 and 50g of potassium sulfate specified in JIS K 8962,
can be available.
(4) If this indicator stands a long time, its end point may become
not clear because of it being oxidized.
49.2 F l a m e atomic absorption m e t h o d Spray sample in a flame such as acety-

lene-air, and measure atomic absorption by calcium at 422.7 nm wavelength t o de-
termine calcium.
Determination range: Ca 0.2 t o 4mg/Z
Repeatability: 2 t o 10 % by coefficient of variation (depending on apparatus and
(a) Hydrochloric acid (l+l) Prepare using hydrochloric acid specified in JIS
K 8180.
(b) L a n t h a n u m (III) solution (50gLalZ) Dissolve bit by bit 29 g of lantha-
num (III) oxide in 500 ml of hydrochloric acid (l+l).
(c) Calcium standard solution (0.5 mgCa/ml) Heat calcium carbonate speci-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
fied in JIS K 8617 a t 180 "C for about 1h, and let it cool in desiccator.
Take its 1.249 g, disperse it in about 50 ml of water, and dissolve it by adding
20 ml of hydrochloric aid (l+l).Heat it at almost boiling for several min-
utes to eliminate carbon dioxide. After cooling it, transfer it into a 1 O00 ml
311
K O101 : 1998
(d) Calcium standard solution (20 pgCa/ml) Pipet 10 ml of calcium stan-

dard solution (0.5 mgCalm1) into a 250 ml volumetric flask, add 5 ml of hy-
drochloric acid ( l + l )and
, add water up t o the marked line.

(a) Flame atomic absorption analyzer Capable of correcting background.
(b) Calcium hollow cathode lamp
Take a suitable amount (containing 20 t o 400 pg as Ca) of sample(5) in a
100 ml volumetric flask, add 2 ml of hydrochloric acid (l+l),and add wa-
ter to the marked line.
Pipet 10ml of this solution into a dried beaker, and add 1 m l of lantha-
num (III) solution (50 gLalZ).
Spray the solution stated in (b)into the flame according to 6 of JIS K 0121,
and read the indicated value(6) at 422.7 nm wavelength.
Take the same amount of water as that of the sample for a blank test, carry
out the operations in (a)to ( c ) , and correct the indicated value on the sample.
Find the quantity of calcium on the working curve, and calculate the con-
centration of calcium (mgCa/Z) in the sample.
Working curve Pipet step by step 1 to 20ml of calcium standard solu-
tion (20 pgCa/ml) into as many 100 ml volumetric flasks, respectively add
hydrochloric acid (l+l)to get the same concentration as the sample side,
and add water up to the marked line. Carry out the operations in (b) and
( c ) on this solution. Separately, add hydrochloric acid (l+l) in water to get
the same concentration as the sample, as a blank test, carry out the opera-
tions in (b) and ( c ) , correct the indicated value obtained on the standard
solution, and draw the relation curve between the quantities of calcium (Ca)
and indicated values. Prepare the working curve when the sample is tested.
Notes (5) When suspensoid is contained, eliminate it through filtration or
centrifugal separation.
(6) Absorbance o r its proportional value shall be valid.
Remarks 1 Such as phosphate, sulfate, o r aluminum disturbs determina-
tion, but it can be restrained by adding lanthanum (III) solu-
tion (50 gLalZ).
2 The coexistence of a lot of magnesium (1 O00 mg/Z or more) gives
a negative error.
49.3 ICP atomic emission spectrometry Spray sample into inductively coupled
plasma through the sample introducing part, and measure the emission by calcium
at 393.367 nm wavelength to determine calcium.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Determination range: Ca 10 t o 200 pg/Z, 0.2 t o 5 mglZ

312
K O101 : 1998

(a) Hydrochloric acid ( l + l ) Follow 49.2 (1) (a).
(b) Calcium standard solution (20 pgCa/ml) Follow 49.2 (1) (d).
(c) Mixed standard solution [(20 pgCa, 10 pgMg, 20 pgAl)/mll Place 20 ml
of calcium standard solution (0.5 mgCa/ml) in 49.2 (1) ( c ) , 10 ml of magne-
sium standard solution (0.5 mgMg/ml) in 50.2 (1) ( c ) , and 20 ml of alumi-
num standard solution (0.5 mgAl/ml) in 59.2 (1) (b)into a 500 ml volumetric
flask, add 10 ml of hydrochloric acid (1+i), and add water up to the marked
line. Prepare when it is needed.
(a) ICP atomic emission spectrometer
Take a suitable amount (containing 1 t o 500 yg as Ca) of sample(5) into a
100 ml volumetric flask, add hydrochloric acid (1+1)to make the concen-
tration of hydrochloric acid about O . 1molll, and add water up to the marked
line.
Spray the solution in (a)into a plasma through the sample introducing part
in accordance with 5.8 of JIS K 0116, and measure emission strength at
393.367nm wavelength(') (8) (9).
For a blank test, prepare the solution whose concentration of hydrochloric
acid is about 0.1 mol/Z by diluting hydrochloric acid (l+l)
with water, carry
out the operation of (b), and correct the emission strength obtained on the
sample.
Find the quantity of calcium on the working curve, and calculate the con-
centration (mgCa/Z) of calcium in the sample.
Working curve Pipet step by step 0.05 t o 1ml (or 1 t o 25 ml)(l*)of cal-
cium standard solution (20 pgCaím1) into as many 100 ml volumetric flasks,
add hydrochloric acid (l+l) to make final concentration of acid the same
as that of sample, and add water up t o the marked line. Carry out the
operations in (b)on these solutions. Separately, take water for a blank
test, add hydrochloric acid (l+l) to make the solution the same acid con-
centration as that in the sample side, carry out the operation in (b),cor-
rect the emission strength obtained on the standard solution, and draw the
relation curve between the quantities of calcium (Ca)and emission strengths.
Notes (7) When the apparatus capable of simultaneously measuring two spec-
tra with different wavelength is used, an internal standard method
can be applicable. When the internal standard method is applied
the procedures are as follows: take a suitable amount of sample
into a 100 ml volumetric flask, add 10 ml of yttrium solution
(50 pgY/ml) [Follow Note (8) in 45.1, add hydrochloric acid (l+l)
to make final concentration of hydrochloric acid about 0.1 moVZ,
add water up t o the marked line. Carry out spraying operation
in (3)(b)on this solution, and measure emission strengths at both
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
313
K O101 : 1998
393.367 nm and 371.029 nm wavelength, and obtain the emission-

strength ratio of calcium and yttrium.
Separately, pipet step by step 0.05 t o 1ml (or 1 to 25 ml) of
calcium standard solution (20 pgCa/ml) into as many 100 ml volu-
metric flasks, add respectively 10 ml of yttrium solution (50 pgY/
mi), add hydrochloric acid (l+l) t o make the concentration of hy-
drochloric acid about 0.1 mol/Z, and add water up t o the marked
line. Carry out the spraying operation in (3)(b)on these solu-
tions, measure each emission strength at both 393.367nm and
371.029 nm (yttrium) wavelength, and draw the relation curve
between the emission-strength ratio of calcium to yttrium and the
concentration of calcium. Make this curve the working curve. On
this working curve, find the quantity of calcium corresponding to
emission-strength ratio obtained on the sample, and calculate the
concentration (mgCaí2) of calcium in the sample.
(8) When working curve method cannot be applied because of high
concentration of salts in sample, the standard addition method
described in 5.8.3 (2) of JIS K 0116 is preferably applicable. In
this case, however, the correction of background is necessary
irrespective of sample type.
(9) In case of the apparatus capable of using high-order spectrum
lines, these lines can be used.
Another wavelength can be used if its exactness and accuracy
have been confirmed.
(10) When magnesium and aluminum are simultaneously to be tested,
use mixed standard solution C(20 pgCa, 10 pgMg, 20 pgAl)/mll,
and prepare the working curve at the testing condition for each
metal element.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
314
K O101 : 1998
50 Magnesium (Mg) For the determination of magnesium, chelatometric titration,

flame atomic absorption method, or ICP atomic emission spectrometry shall be ap-
plied.
50.1 Chelatometric titration Add buffer solution in a sample to control its pH

to be 10,add Eriochrome black T [sodium 3-hydroxy-4- I(
1-hydroxy-2-naphthaleny1)azol-
7-nitro-1-naphthalenesulfonate(nomenclature by IUPAC)] as indicator, titrate this
solution with disodium dihydrogen ethylenediaminetetraacetate solution, find the titrant
needed for the total of calcium and magnesium, and subtract the titrant for calcium
to determine magnesium.
Determination range: 0.15 to 5 mg as Ca on the total of Mg and Ca

Potassium cyanide solution (100 g/Z) Follow 49.1 (1)(b).
Hydroxylammonium chloride solution (100 g/Z) Follow 40.2 (1) (cl.
Ammonium chloride-ammonia buffer solution (pH 10) Follow 22.1.2
(1) (b).
Eriochrome black T solution ( 6 g/Z) Dissolve 0.5 g of Eriochrome black
T [sodium i-(l-hydroxy-2-naphthylazo)-6-nitro-2-naphthol-4-sulfonate]
speci-
fied in JIS K 8736 in 100 ml of methanol specified in JIS K 8891, and add
0.5g of hydroxylammonium chloride specified in JIS K 8201. Store this
solution in a coloured glass bottle.
10 mmoW EDTA solution Follow 49.1 (1) (e). One milliliter of this so-
lution corresponds to 0.243 l mg of Mg.

Take a suitable amount (containing 5 mg or less as Ca on the total of Mg
and Ca) of sample(1) in a beaker, add water to make total about 50 ml.
Add 0.5 ml of potassium cyanide (100g/Z), a few drops of hydroxylammonium
chloride solution (100g/Z), and 1 ml of ammonium chloride ammonia buffer
solution (pH 10).
Add 2 or 3 drops of Eriochrome black T solution (5 g/Z) as indicator,
Titrate it with 10 mmol/Z EDTA solution until the solution turns blue from
red (2).
Separately, take the same amount of the sample, carry out the operations
in 49.1 (2) (a) to (d), and obtain titrant (ml) of 10 mmol/Z EDTA solution
corresponding t o the amount of calcium.
Calculate the concentration of magnesium (mgMg/Z) in the sample in ac-
cordance with the following formula.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
315
K O101 : 1998
v vca
M,=(a_bj,1000xO.2431
MZ=
where,
(; vo,)
MI : magnesium (mgMg/Z)
X1OOOX1.001
a : 10 mmol/Z EDTA solution needed for titration

(mi)
b : 10 mmol/Z EDTA solution needed for titration in
the operation in 49.1 (2)(mi)
V : sample (mi)
v c a : sample a t 49.1(2) (mi)
0.243 1: magnesium equivalent to 1ml of 10 mmol/Z
EDTA solution (mg)
Mz : calcium carbonate equivalent to magnesium
(mgCaCO&
1.001 : calcium carbonate equivalent to 1ml of
10 mmol/Z EDTA solution (mg)
When suspensoid is contained, remove it through filtration or
When sample contains the organic substances and coloured
substances which affect the tests, neutralize it after the opera-
tion in 4. Provided that the method in 4.4 does not apply.
Because Eriochrome black T slowly changes its colour, titrate
gradually with full stirring near colour changing point.
50.2 Flame atomic absorption method Spray sample into a flame such as acety-
lene-air, and measure the atomic absorption by magnesium a t 285.2 nm wavelength
to determine magnesium.
Determination range: Mg 20 to 400 pg/Z
Hydrochloric acid (l+l) Follow 49.2 (1)(a).
Lanthanum (III) solution (50 gLa/Z) Follow 49.2 (1)(b).
Magnesium standard solution (0.5 mgMg/ml) Heat magnesium oxide
specified in JIS K 8432 at 700 to 800 "C for about 30 min, and let it cool in
a desiccator. Dissolve its 0.829 g in 20 ml of hydrochloric acid (l+l), trans-
fer it into a 1O00 ml volumetric flask, and add water up t o the marked line.
Magnesium standard solution (2pgMg/ml) Pipet 10 ml of magnesium
standard solution (0.5 mgMg/ml) into a 100 volumetric flask, and add water
up t o the marked line. Take 10 ml of this solution into a 250 ml volumetric
flask, add 5 ml of hydrochloric acid (l+l),and add water up to the marked
line.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
316
K O101 : 1998

(a) Flame atomic absorption method Capable of correcting background.
(b) Magnesium hollow cathode lamp
(a) Take a suitable amount (containing 2 to 40 pg as Mg) of sample(3)in a 100 ml
volumetric flask, add 2 ml of hydrochloric acid (l+l)? add water up t o the
marked line.
(b) Take 10 ml of this solution in a dried beaker, and add 1 ml of lanthanum
(III) solution (50 gLalZ).
(c) Spray the solution of (b) into the flame in accordance with 6 of JIS K 0121,
and read the indicated value(4) at 285.2 nm wavelength.
(d) Take the same amount of water for a blank test as that of the sample, carry
out the operations in (a)t o (cl,and correct the indicated value obtained on
the sample.
(e) Find the quantity of magnesium on the working curve, and calculate the
concentration (ygMglZ) of magnesium in the sample.
Working curve Pipet step by step from 1 to 20ml of magnesium stan-
dard solution (2 pgMglm1) into as many 100 ml volumetric flasks, add re-
spectively hydrochloric acid (l+l) t o make them the same acid concentration
as the sample side, and add water up to the marked line. Carry out the
operations in (b)and ( c ) on this solution. Separately, add hydrochloric acid
(l+l) in water, as a blank test, t o make the same acid concentration as the
sample, carry out the operations in (b) and ( c ) t o correct the indicated value
obtained on the standard solution, and draw the relation curve between
the quantities of magnesium (Mg) and indicated values. Prepare the working
curve when the sample is measured.
Notes (3) When suspensoid is contained, remove it through filtration or
(4) Absorbance or its proportional value shall be valid.
Remarks 1 Even small amount of aluminum (2 mglZ) disturbs the test, but
the addition of lanthanum (III) solution (50 gLa/Z) can restrain
it.
50.3 ICP atomic emission spectrometry Spray sample in inductively coupled

plasma through the sample introducing part, and measure the emission by magne-
sium at 279.553 nm wavelength t o determine magnesium.
Determination range: Mg 5 to 100 pg/Z, 0.1 t o 3 mglZ
(a) Hydrochloric acid (l+l) Follow 49.2 (i)(a).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
317
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Magnesium standard solution (10 pgMg/ml) Pipet 5 ml of magnesium

standard solution (0.5 mgMg/ml) stated in 50.2 (1) (cl into a 250 ml volu-
metric flask, add 5 ml of hydrochloric acid (l+l), and add water up t o the
marked line.
(c) Mixed standard solution [ ( 2 0 pgCa, 10 pgMg, 20 pgAl)/mll Follow
49.3 (1)(cl.
(a) Take a suitable amount (containing 0.5 t o 300 pg as Mg) of sample(3) in a
100ml volumetric flask, add hydrochloric acid (l+l) to make the concen-
tration of hydrochloric acid about 0.1 mol/Z, and add water up to the marked
line.
(b) Spray the solution in (a) into the plasma through the sample introducing
part according t o 5.8 of JIS K 0116, and measure the emission strength a t
279.553 nm wavelength(5) (9
(c) Take water for a blank test, add hydrochloric acid (l+l) t o make the con-
centration of hydrochloric acid about 0.1 mol/Z, carry out the operation in
(b),and correct the emission strength obtained on the sample.
(d) Find the quantity of magnesium on the working curve, and calculate the
concentration (pgMglZ) of magnesium in the sample.
Working curve Pipet step by step 0.05 to 1ml (or 1 to 30 ml)(8) of mag-
nesium standard solution (10 pgMg/ml) into as many 100 ml volumetric flasks,
respectively add hydrochloric acid (l+l) to make the same acid concentra-
tion as that of the sample and add water up t o the marked line. Carry out
the operation in (b)on this solution. Separately, take water for a blank
test, add hydrochloric acid (l+l) t o make the same acid concentration as
that of the sample, carry out the operation in (b),correct the emission strength
the quantities of magnesium (Mg) and emission strengths. Prepare the
working curve when sample is measured.
the procedures are as follows: take a suitable amount of sample
into a 100 ml volumetric flask, add 10 ml of yttrium solution
(50 pgY/ml>[follow Note (8) in 451, add hydrochloric acid (l+l) to
make final concentration of hydrochloric acid about 0.1 mol/,!, add
water up to the marked line. Carry out the spraying operation
in (3)(b) on this solution, and measure emission strengths at both
279.553 nm and 371.029 nm (yttrium) wavelength, and obtain the
emission-strengths ratio of magnesium and yttrium.
Separately, pipet step by step 0.05 to 1ml (or 1 t o 30 ml) of
magnesium standard solution (10 pgMg/ml) in as many 100 ml
volumetric flasks, respectively add 10 ml of yttrium solution
318
K O101 : 1998
(50 pgY/ml), add hydrochloric acid (l+l) to make the concentra-

tion of hydrochloric acid about 0.1 moVZ, and add water up t o the
marked line. Carry out the spraying operation in (3)(b)on these
solutions, measure each emission strength at both 279.553 nm and
371.029 nm wavelength, and draw the relation curve between the
emission-strength ratio of magnesium to yttrium and the concen-
tration of magnesium. Make this curve the working curve. On
this working curve, find the quantity of magnesium correspond-
ing to emission-strength ratio obtained on the sample, and calcu-
late the concentration (pgMg/Z) of magnesium in the sample.
(6) When the working curve method cannot be applied because of
high concentration of salts in sample, the standard addition method
described in 5.8.3 (2) of JIS K 0116 is preferably applicable. I n
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

has been confirmed.
(8) When calcium and aluminum are simultaneously t o be tested,
use mixed standard solution [(20 pgCa, 10 pgMg, 20 pgAl)/ml],
and prepare the working curve at the testing condition for each
metal element.
319
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`--- K O101 : 1998
51 Copper (Cu) For the determination of copper, diethyldithiocarbamic acid

absorptiometry, flame atomic absorption method, electric heating atomic absorption
method, ICP atomic emission spectrometry or ICP mass spectrometry shall be ap-
plied.
51.1 Diethyldithiocarbamic acid absorptiometry Add both citric acid salts and
disodium dihydrogen ethylenediaminetetraacetate (EDTA) as a masking agent for
coexisting metals in the sample, adjust its pH t o be about 9 using aqueous ammo-
nia, add sodium N,N-diethyldithiocarbamate(sodium diethylcarbamodithio acid), ex-
tract the yellowish brown copper complex using butyl acetate, and measure its
absorbance to determine copper.
Determination range: Cu 2 to 30pg
Aqueous ammonia (l+l) Prepare using aqueous ammonia specified in
JIS K 8085.
Diammonium hydrogencitrate solution (100 g l l ) Dissolve 10 g of
diammonium hydrogencitrate specified in JIS K 8284 in water to make
total 100ml.
Diammonium hydrogencitrate containing copper should be purified as
follows.
Dissolve 10 g of diammonium hydrogencitrate in 80 ml of water, make
its pH about 9 by adding aqueous ammonia ( l + l )add , water to make total
100 ml. Place it in a separatory funnel, add 2 ml of sodium diethyldithio-
carbamate solution (10 gll) stated in (e) and 10 ml of butyl acetate stated
in ( g ) into the funnel, agitate violently, and let it stand. Filtrate water
layer through dried filter paper, and use the filtrate from which small drops
of butyl acetate were removed.
EDTA solution Dissolve 2 g of disodium dihydrogen ethylenediamine-
tetraacetate dihydrate specified in JIS K 8107 in water up t o 100 ml.
Sodium diethyldithiocarbamate solution (10 gll) Dissolve 1.3 g of
sodium N , N-diethyldithiocarbamate trihydrate (diethyldithiocarbamodithio
acid sodium trihydrate) specified in JIS K 8454 in water to make total
100 ml. Store in a coloured bottle, and never use after two weeks or longer.
Metacresol purple solution (1 gll) Follow 46.1 (1)(n).
Butyl acetate Specified in JIS K 8377.
Copper standard solution (0.1mgCulm1) Wash the copper, standard
reagent for quantitative analysis, specified in JIS K 8005 with hydrochlo-
ric acid (1+3), wash with water, wash with ethanol (99.5) specified in JIS
K 8101,then wash with diethyl ether specified in JIS K 8103, place im-
mediately it in a desiccator, and let it stand for 12 h or longer. Weigh 0.100 g
Cu on the base of 100 % Cu, put it in 20 ml of nitric acid (l+l), boil it to
dissolve copper and dispel nitrogen oxide, let it cool, introduce into a 1 O00 ml
320
K O101 : 1998
volumetric flask, and add water up to the marked line. Otherwise, take
0.393g of copper (II) sulfate pentahydrate specified in JIS K 8983, dis-
solve it in 20 ml of nitric acid (l+l),transfer it in a 1 O00 ml volumetric
flask, and add water up to the marked line. Or use copper, reference material,
standard solution, Cu 100, specified in JIS K 0010.
(i) Copper standard solution (1 pgCu/ml) Pipet 10 ml of copper standard
solution (0.1 mgculml) into a 1 O00 ml volumetric flask, add 20 ml of nitric
acid (l+l), and add water up to the marked line.
(a) Separatory funnel With 100 ml or 300 ml capacity.
Take a suitable amount(1) (containing 2 t o 30 pg of Cu) of the sample which
has been treated with 4 in a separatory funnel, add 2 or 3 drops of metacresol
purple solution (1 glE) as indicator, and add 5 ml of diammonium hydrogen-
citrate solution (100 gll) and 1 ml of EDTA solution.
Add aqueous ammonia ( l + l )t o neutralize until solution turns into a faint
violet(2) (pH about 9), and add water up to 50 ml(3).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Add 2 ml of sodium diethyldithiocarbamate solution (10 gl0, and mix them,

add 10 ml of butyl acetate(*),agitate violently for about 3 min, and let it
stand.
Discard water layer, remove butyl acetate layer into a ground-stoppered
test tube in which about l g of sodium sulfate (anhydrate) has been put,
and shake well(5).
Place a part of the solution in an absorption cell, and measure its absor-
bance in the vicinity of 440nm wavelength with making butyl acetate as
reference solution.
Take about 20 ml of water for a blank test, carry out the operations in (a)
to (e),measure its absorbance, and correct the absorbance obtained on the
sample.
Find the quantity of copper on the working curve, and calculate the con-
centration (mgCulZ) of copper in the sample.
Working curve Take stepwise from 2 t o 30ml of copper standard solu-
tion (1 pgCu/ml) in as many separatory funnels, carry out respectively the
operations in (a) to (f),and draw the relation curve between the quantities
of copper (Cu) and absorbances.
Notes (1) If sample, with low copper concentration, has no organic substance
and turbidity, the following is permissible: take a suitable amount
of sample that is up t o 250m1, and carry out according t o the
operations in 4.1 and then in (a)t o (f). I n this case, use the
whole amount pretreated, and use the same amount of reagents
as those used in (a)t o (d). Prepare the working curve similarly
to the sample side.
32 1
K O101 : 1998
(2) Instead of metacresol purple solution (1glZ) a t (a), a pH meter

or pH test paper can be available.
(3) In advance, put a mark on a separatory funnel.
(4) Such as chloroform, or benzene may be used as solvent for ex-
traction. When sample contains a sort of anion surfactant (for
example, sulfonic acid type) o r tannin, the extraction of copper
becomes incomplete.
(5) Either way may be useful; filtrate through dried filter paper, or
use a funnel of which the stem was packed with dried absorbent
cotton.
Remarks 1 When no addition of EDTA solution, sodium diethyldithio-
carbamate reacts with many metal elements. However, the
complexes of almost all metals such as mercury, arsenic, lead,
tin, antimony, and so on are colourless. The complexes of iron,
nickel, cobalt, o r others have colour, but in this method, they
are masked owing to addition of EDTA solution.
2 Bismuth, extracted together with copper, shows yellow, but
when its amount is 2 times or less than that of copper, it gives
no influence.
When its amount is more than 2 times, the following pro-
cedures are necessary: make the absorbance measured a t (3)
AI, separately take the same amount of sample as used for
copper test, carry out the operation in (a),add 3 ml of potas-
sium cyanide (50 gll) to make copper cyanocomplex, carry out
the operations in (b)to (f) and extract only bismuth complex,
measure its absorbance, and make it A2. The absorbance by
copper is AI -Az.
3 When sample is pretreated according to 4.1, if it contains cya-
nide compound, sufficient heating is necessary.
51.2 Flame atomic absorption method Spray sample, which has been pretreated,
into acetylene-air flame, and measure atomic absorption by copper a t 324.8 nm
wavelength to determine copper.
Determination range: Cu 0.2 to 4mglZ
(1) Reagent Reagent shall be as follows.
(a) Copper standard solution (10 pgCu/ml) Take 50 ml of copper standard
solution (0.1 mgCdml) stated in 51.1 (1) (h)into a 500 ml volumetric flask,
add 10 ml of nitric acid (l+l), and add water up t o the marked line,

(b) Copper hollow cathode lamp
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
322
K O 1 0 1 : 1998

(a) Treat sample according t o 4.5.
Remarks 4 The preparatory operations for the sample with low concentra-
tion of copper and without substances disturbing extracting
operation shall be either as follows o r according t o Remarks 5.
Take 500 ml (or definite volume from 100 to 500 mi) of sample
in a beaker, add 10 ml of hydrochloric acid specified in JIS K
8180 and boil for about 5 min. After cooling, transfer into a
1 0 0 0 m l separatory funnel (or 200 to 500ml), add 10ml of
diammonium hydrogencitrate solution (100 g/Z) and 2 or 3 drops
of metacresol purple solution (1gll) as indicator, and add aque-
ous ammonia (l+l) until the colour of solution turns faint purple.
Add 5 ml of sodium diethyldithiocarbamate (10 gA), mix by
shaking, add 10 t o 20 ml of butyl acetate specified in JIS K
8377, shake violently for about 1min, and allow to stand.
Separate butyl acetate layer and put it in a 100ml beaker.
Add 5 ml of butyl acetate in water layer and repeat extrac-
tion operation. Combine the extracted butyl acetate layer in
the said beaker. Volatilize butyl acetate by heating, add 2 ml
of nitric acid specified in JIS K 8541 and 2 ml of perchloric
acid specified in JIS K 8223,heat, and decompose organic sub-
stances. After almost dryness, allow t o cool. Dissolve resi-
due in 10 ml of nitric acid (1+15), and use i t for determination
of copper (it may also be used for determination of zinc, cad-
mium, nickel, lead, etc.).
The liquid which was made definite volume by adding bu-
tyl acetate into the extracted butyl acetate layer, o r the butyl
acetate layer which was prepared by once extraction under speci-
fied extracting condition, can be directly used for atomic ab-
sorption spectrometry owing to the spraying as it is. Instead
of butyl acetate specified in JI$ K 8377,4-methy1-2-pentanone
specified in JI$ K 8903 or 2,6-dimethyl-4-heptanone (diisobutyl
ketone DIBK) can be serviceable.
When using 2,6-dimethyl-4-heptanone, the addition is enough
little because the mutual solubility between water and 2,6-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
dimethyl-4-heptanone is almost zero.

5 Take 200 ml of sample, acid-treat similarly to Remarks 4, and
adjust its pH to be 3.5 to 4.0. Add 20 ml of ammonium sulfate
solution (saturated). Add 5 ml of 1-pyrrolidinecarbodithioicacid
ammonium salt (ammonium pyrrolidine-N-dithiocarbamate)
(APDC) solution (10 g/Z), shake it gently, and let it stand for
about 3 min. Then add 10 ml of 4-methyl-2-pentanone speci-
fied in JIS K 8903,agitate violently for about 3 min, let it stand.
Separate organic layer and put it in a 100 ml beaker. Add 5 ml
of 4-methyl-2-pentanone in the water layer, and repeat the ex-
tracting operation. Put the extracted organic layer into the
said beaker. Treat this organic layer similarly to Remarks 4
and use it copper determination (it can also be used for deter-
mination of zinc, cadmium, nickel, lead, etc.).
323
K O101 : 1998
In addition, a definite amount which the extracted 4-methyl-

2-pentanone layer is added the same solvent, or the 4-methyl-
2-pentanone layer carried out restore/extraction once with
constant extracting condition may be sprayed for determination.
2,6-dimethyl-4-pentanone may be used instead of 4-methyl-
2-pentanone.
Spray the sample, which has been carried out by preparatory treatment in
(3),into flame according to 6 of JIS K 0121,and read the indicated value (6)
a t 324,8 nm wavelength.
Take the same amount of water as that of the sample for the preparatory
treatment in (3)for a blank test, carry out the operations in (3)and (4)(a)
similarly to the sample, and correct the indicated value obtained on the
sample.
centration (mgCu/Z) of copper in the sample.
Working curve Pipet step by step from 2 t o 40 ml(7) of copper standard
solution (10 pgCu/ml) into as many 100 ml volumetric flasks, respectively
add acid t o make the same acid concentration as the sample carried out
the operation in (3)(a),and add water up to the marked line(8). Carry out
the operation in (a) on this solution. Separately, for a blank test, add acid
in water t o make the same acid concentration as the sample carried out
the operation in (3)(a),carry out the operation in (a) to correct the indi-
cated value obtained on the standard solution, and draw the relation curve
between the quantities of copper (Cu) and the indicated values. Prepare
the working curve when sample is measured.
Notes (6) Absorbance or its proportional value is valid.
(7) When solvent extraction is carried out as preparatory operation,
the amount of copper standard solution (10 pgCu/ml) shall be
suitably decreased.
(8) When such as butyl acetate layer, 4-methyl-2-pentanonelayer, or
2,6-dimethyl-4-heptanonelayer is directly sprayed after prepara-
tory operation in Remarks 4 and Remarks 5, working curve is pre-
pared as follows: dilute copper standard solution (10 pgCu/ml) into
suitable concentration (0.1 to 1kgCu/ml), pipet step by step from
2 to 40 ml of the solution, make them 500 ml (or a definite amount
from 100 to 500 mi), carry out the operations in Remarks 4 and
(4)(a)and (b)similarly to the sample, and draw the relation curve
between the quantities of copper (Cu) and the indicated values.
51.3 Electric heating atomic absorption method After pretreatment of a sample,

atomize it in an electric furnace, and measure the atomic absorption by copper a t
324.8 nm wavelength t o determine copper.
Determination range: Cu 5 t o lOOpg/Z
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
324
K O101 : 1998
Remarks 6 Because this method is easily affected by coexisting acids or salts

or their concentrations, the sample which is less affected shall be
adopted.

(a) Water Water A3 specified in JIS K 0557. In advance, carry out a blank
test on the element to be determined t o verify there is no interference.
(b) Nitric acid (l+l)Prepare using highly purified nitric acid specified in
JIS K 9901.
(c) Copper standard solution (1 pgCu/ml) Follow 51.1 (1)(i).
(2) Tool and apparatus Tools and apparatuses shall be as follows.
(a) Electric heating atomic absorption analyzer Electrically heating type
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
and capable of correcting background.
(b) Exothermic body Made of graphite or heat-resisting metal.
(c) Copper hollow cathode lamp
(d) Flow gas Argon grade 2 specified in JIS K 1105.
(e) Micropipet Push-button type micro-volume meter for liquid specified in
JIS K 0970,5 t o 5 0 ~ 1o, r automatic injection device.
(3) Preparatory operation Preparatory operation shall be as follows.
(a) Treat sample in accordance with 4.5.
Inject a definite amount (for instance, 10 to 5 0 ~ 1 of ) sample, which has
been pretreated as in (31, into an exothermic body using a micropipet, dry
it according t o the operations in 6 of JIS K O121 (at 100 t o 120 "C for 30
to 40 s), ash it (at 600 to 1O00 "C for 30 t o 40 s), then atomize i t ( 9 ) (2 200
t o 2 700 "C for 3 to 6 s), and read(l0) the indicated value(6) at 324.8 nm wave-
length.
Take the same amount of water as that of the sample at preparatory op-
erations in (3) for a blank test, carry out the operations in (3) and (4) (a)
similarly t o the sample, and correct the indicated value on the sample.
centration (pgCu/Z) of copper in the sample.
Working curve Pipet step by step from 0.5 to 10 ml of copper standard so-
lution (1 pgCu/ml) into as many 100 ml volumetric flasks, add acid to make
the same acid concentration as the sample carried out the operation in (3)(a),
and add water up to the marked line. Carry out the operation in (a) on this
solution. Separately, for blank test, take water, add acid to make the same
acid concentration as the sample carried out the operation in (3)(a),carry
out the operation in (a),correct the indicated value obtained on the standard
solution, and draw the relation curve between the quantities of copper (Cu)
and the indicted values. Prepare the working curve when sample is tested.
325
K O101 : 1998
Notes (9) The condition for drying, ashing, or atomizing varies depending
upon apparatus, and they may be affected by such as injected
volume of sample and concentration of coexisting salts.
(10) Repeat successively a t least 3 times the operations in (a), and
confirm the indicated values sufficiently agree.
51.4 ICP atomic emission spectrometry After a sample is pretreated, spray

into inductively coupled plasma, and measure the emission by copper at 324.754nm
wavelength t o determine copper.
Determination range: Cu 20 t o 5000pg/l
(a) Copper standard solution (10 ygculml) Follow 51.2 (1) (a).
(b) Mixed standard solution [(loygCu, 10 pgZn, 8 ygCd, 10 pgNi, 10 pgPb,
10 pgMn, 10 ygFe)/mll Place 50 ml of copper standard solution (0.1 mgCd
ml) stated in 51.1 (1) (h),50 ml of zinc standard solution (0.1 mgZn/ml) stated
in 52.1 (1)(a), 40 ml of cadmium standard solution (0.1 mgCd/ml) stated
in 53.1 (1)(a), 50 ml of nickel standard solution (0.1 mgNi/ml) stated in
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
54.1 (1)(i),50 ml of lead standard solution (0.1 mgPb/ml) stated in 56.1 (1) (a),
50 ml of manganese standard solution (0.1 mgMn/ml) stated in 58.1 (1)(a),
and 5 ml of iron standard solution (1mgFe/ml) stated in 60.1 (1)(g) into
500 ml volumetric flasks, respectively, add 3 ml of nitric acid (l+l), and
add water up to the marked line. Prepare this solution when it is used.
(a) Treat sample according to 4.5.
Remarks 7 When the sample, which has been preparatorily operated (pre-
treatment), has a rich concentration of sodium, potassium,
magnesium, and calcium, and poor in copper concentration,
the following procedures shall be carried out.
Take 500 ml (or a definite amount of 100 to 500 mi) of sample
in a beaker, add 5 ml of hydrochloric acid specified in JIS K
8180, and boil it for about 5 min. After cooling it, add 10 ml
of acetic acid-sodium acetate buffer solution (pH 5) [Dissolve
19.2g of sodium acetate trihydrate specified in JIS K 8371
and 3.4 ml of acetic acid specified in JIS K 8355 in water t o
make total 12.1, and adjust its pH to 5.2 using aqueous am-
monia (l+l) or nitric acid (1+10). Transfer this solution into
a 1O00 ml (or 200 to 500 ml) separatory funnel, add 2 ml of
1-pyrrolidinecarbodithioacid ammonium salt solution (20 g/l)
and 2 ml of methanol solution (20 g/E) of hexamethylene-
ammonium-hexamethylenecarbamodithio acid (hexamethylene-
ammonium-hexamethylenedithiocarbamic acid), then mix them,
326
K O101 : 1998
add a definite amount (5 t o 20 ml) of xylene specified in JIS

K 8271, and agitate violently t o mix them for about 5min,
followed by settling down. Discard water layer, and put xy-
lene layer in a ground-stoppered test tube.
This solution can be used for determination of cadmium,
nickel, lead, manganese, iron, vanadium respectively, and also
used for the simultaneous determination of copper with them.
The acetic acid-sodium acetate buffer solution (pH 5) to be
used in this operation should be purified, prior to its use, by
mixing with such as 1-pyrrolidinecarbodithioacid ammonium
salt solution, methanol solution of hexamethyleneammonium-
hexamethylenecarbamodithio acid, and xylene.
Spray the sample, which has been pretreated as in (3),into plasma through
the sample introducing part according to 5.8 of JIS K 0116, and measure
emission strength at 324.754 nm wavelength (11) (12) (13).
Take the same amount of water as that of the sample a t preparatory treat-
ment in (3)for a blank test, carry out the operations in (3) and (4) (a)similarly
to the sample, and correct the emission strength obtained on the sample.
centration (pgCul2) of copper in the sample.
Working curve Pipet step by step from 0.2 to 50 ml(14) (15) of copper stan-
dard solution (10 pgCu/ml) into as many 100 ml volumetric flasks, add re-
spectively acid t o make the same acid concentration as the sample carried
out the operation (3)(a),and add water up to marked line. Carry out the
operation in (a)on this solution. Separately take water for a blank test,
and add acid t o make the same acid concentration as the sample carried
out the operation (3)(a),carry out the operation in (a),correct the emis-
sion strength obtained o n the standard solution, and draw the relation curve
between the quantities of copper (Cu) and emission strengths. Prepare this
Notes (11) When the apparatus capable of simultaneously measuring two
spectra with different wavelength is used, an internal standard
method can be applicable. When the internal standard method
is applied the procedures are as follows: Take a suitable amount
of sample, which has been treated in (3)(a),into a 100 ml volu-
metric flask, add 10 ml of yttrium solution (50 pgY/ml) [Follow
Note (8) of 451, add acid t o make the same acid concentration as
the sample in (4)(a), add water up t o the marked line. Carry
out the spraying operations in (4) (a>on this solution, measure
emission strengths a t both 324.754 nm and 371.029 nm (yttrium)
wavelength, and obtain the emission-strengths ratio of copper
and yttrium.
Separately, pipet step by step from 0.2 to 50 ml of copper stan-
dard solution (10pgCu/ml) into as many 100 ml volumetric flasks,
add respectively 10 ml of yttrium solution (50 ygY/ml), add acid
to make the same acid concentration as the sample of (4)(a),
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
327
K O101 : 1998
and add water up t o the marked line. Carry out the spraying
operations in (4)(a)on these solutions, measure emission strengths
a t both 324.754 nm and 371.029 nm wavelength, draw the rela-
tion curve between emission-strength ratio of copper t o yttrium
and the concentration of copper, and make it the working curve.
On this working curve, find the quantity of copper correspond-
ing to the emission-strength ratio obtained on the sample, and
calculate the concentration (pgCulZ) of copper in the sample.
high concentration of salts in sample, the standard addition
method described in 5.8.3 (2) of JIS K 0116 is preferably appli-
cable. In this case, however, the correction of background is
necessary irrespective of sample type,
(13) I n the case of the apparatus capable of using high-order spec-
trum lines, these lines can be used.
Another wavelength can be used if its exactness and accu-
racy has been confirmed.
(14) When, after making preparatory operations according to Remarks
7, xylene layer is directly sprayed, the working curve shall be
prepared as follows: dilute copper standard solution (10 pgCu/
mi) t o suitable concentration (0.1 t o 1 pgCulml), take stepwise
from 0.2 to 50 ml of the solution, make them 500 ml (or a defi-
nite amount of 100 t o 500ml), carry out the operations in Re-
marks 7 and (4)(a)and (b) similarly t o the sample side, and
draw the relation curve between the quantities of copper (Cu)
and the emission strengths.
(15) When zinc, cadmium, nickel, lead, manganese, iron are simul-
taneously tested, use mixed standard solution [(10 pgCu, 10 pgzn,
8 pgCd, 10 pgNi, 10 pgPb, 10 pgMn, 10 pgFe)/mll, and prepare
preferably each working curve under the test condition of each
metal element.
51.5 ICP mass spectrometry After sample was pretreated, add internal standard --`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
substance, spray into an inductively coupled plasma through the sample introducing
part, measure the ionic current in number of masses/electrical charges of both copper
and the internal standard substance and obtain the ratio between the ionic current
by copper and that by internal standard substance t o determine copper.
Determination range: Cu 0.5 to 25 pgll, 10 t o 500 pgll

(a) Water Follow 51.3 (1)(a).
(b) Nitric acid (l+l)Follow 51.3 (1) (b).
(c) Yttrium solution (1 pgY/ml)(l6) Put 20 ml of yttrium solution (50 pgY/
mi) of Note (8) in 45 in a 1 O00 ml volumetric flask, add 1.5 ml of nitric
acid (1+1)and add water up t o the marked line. Prepare when it is used.
328
K O101 : 1998
Copper standard solution (1 pgCu/ml) Follow 51.1 (1)(i).

Copper standard solution (50 ngCu/ml) Put 50 ml of copper standard
solution (1pgCu/ml) of 51.1 (1)(i) in a 1O00 ml volumetric flask, add 3 ml
and add water up to the marked line. Prepare when it
of nitric acid (l+l)
is used.
Mixed standard solution [(ipgCu, 1 pgZn, 1 pgCd, 1 pgPb, 1 pgMn,
1 pgCr)/ml] Take each 10 ml of copper standard solution (0.1mgCu/ml)
of 51.1 (1) (h),zinc standard solution (0.1 mgZn/ml) of 52.1 (1)(a),cadmium
standard solution (0.1 mgCd/ml) of 53.1 (1)(a), lead standard solution
(0.1mgPb/ml) of 56.1 (1)(a), manganese standard solution (0.1 mgMn/ml)
of 58.1 (1)(d) and chromium standard solution (0.1 mgCr/ml) of 61.1 (1)(f)
in a 1O00 ml volumetric flask, add 1.5 ml of nitric acid (l+l) and add wa-
ter up t o the marked line. Prepare when it is used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Mixed standard solution [ ( 5 0 ngCu, 50 ngZn, 50 ngCd, 50 ngPb,
50 ngMn, 50 ngCr)/ml] Take each 5 ml of copper standard solution
(10 pgCu/ml) of 51.2 (1)(a),zinc standard solution (10 pgZním1) of 52.1 (1)(b),
cadmium standard solution (10 pgCd/ml) of 53.1 (1)(b),lead standard so-
lution (10 pgPb/ml) of 56.3 (1)(a),manganese standard solution (10 pgMn/
ml) of 58.2 (1)(a) and chromium standard solution (10 pgCr/ml) of
61.1.2 (1)(a)in a 1O00 ml volumetric flask, add 1.5ml of nitric acid (l+l)
and add water up t o the marked line. Prepare when it is used.
Note (16) This solution is used as internal standard substance. Indium
(In), bismuth (Bi), etc. other than yttrium may be used as in-
ternal standard substance. Their preparation methods are as
follows.
Indium solution (1 pgIn/ml) Add 10 ml of highly purified ni-
tric acid specified in JIS K 9901 in 0.250g of indium, dissolve by
heating, expel nitrogen oxide, cool, transfer into a 250 ml volumet-
ric flask and add water up to the marked line. When it is used,
take 1ml of this solution in a 1O00 ml volumetric flask, add 3 ml
of nitric acid (l+l) and add water up t o the marked line.
Bismuth solution (1pgBi/ml) Add 10 ml of nitric acid (l+l)
in 0.279 g of bismuth trioxide, dissolve by heating, cool, trans-
fer into a 250 ml volumetric flask and add water up to the marked
line. When it is used, take 1ml of this solution in a 1O00 ml
volumetric flask, add 20ml of nitric acid (l+l) and add water
Apparatus Apparatus is as follows.
(a) ICP mass spectrograph
Remarks 8 Ones having equivalent performance t o inductively coupled
plasma as the ion source may be used.
9 For sample spraying, an ultrasonic wave nebulizer o r those
having equivalent performance thereto may be used. In this
case, the lower limit value of determination may be lowered
some one figure, but washing shall be carried out sufficiently
with care for memory effect.
329
K O101 : 1998
10 Verify that there is no contamination from the materials of

sampling cone and skimmer cone.
(3) Preparatory operation Preparatory operation shall be carried out as follows (17).
(a) The sample shall be treated according to 4.6.
(b) Take a suitable amount (including 0.05 to 50 pg as Cu) of the sample treated
in (a)in a 100 ml volumetric flask, add 1 ml of yttrium solution (1pgY/ml),
add nitric acid (l+l) to make final nitric concentration 0.1 t o 0.5 mol/Z, and
Note (17) Be careful not t o contaminate the sample from the tester. Sur-
gical rubber gloves (not adhering powder) specified in JIS T 9107
should be used.
(4) Operation Operation shall be carried out as follows(18).

Make the ICP mass spectrograph ready to run, spray the solution in (3)(b)
into a inductively coupled plasma through the sample introducing part, read
the indicated value ( 2 0 ) in the number of masses/electrical charges (19) of copper
and yttrium, and obtain the ratio between the indicated value of copper
and that of yttrium.
Take water, for a blank test, the same amount as the sample in (3)(b),
carry out the operations in (3) and (4)(a)similarly to the sample, obtain
the ratio between the indicated value of copper and that of yttrium, and
correct the ratio of indicated values of copper and yttrium obtained on the
sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Obtain the amount of copper on the working curve and calculate the con-
centration of copper (pgCu/Z) in the sample.
Working curve Pipet step by step from 1 to 50 rnl(21) of copper standard
solution (50 ngCu/ml or 1 pgCu/ml) into as many 100 ml volumetric flasks,
add 1 ml of yttrium solution (1pgY/ml), add nitric acid (l+l)t o make acid
concentration the same as that of the sample in (3)(b),and add water up
to the marked line. Carry out the operation in (a)on this solution. Sepa-
rately put l ml of yttrium solution (1 pgY/ml) as a blank test in a 100ml
volumetric flask, add nitric acid (l+l) to make the same acid concentra-
tion as the sample of (3)(b),and add water up to the marked line. Carry
out the operation in (a), correct the ratio of indicated values obtained on
the standard solution, and draw a relation curve of the ratio between the
indicated value t o the amount of copper (Cu) and the indicated value of
yttrium. Prepare the working curve when the sample is measured,
Notes (18) If the existence of interfering substance is not clear, carry out
qualitative analysis by an ICP mass spectrometer before deter-
mination t o estimate interference against number of measuring
masses of target element and internal standard substance. When
interference is found, change of internal standard substance, di-
lution of sample o r carrying out of pretreatment is carried out
to decrease interference.
(19) To set the number of masses, refer to Table 51.1. When there
are stable isotopes, measure using number of masses/electrical
330
K O101 : 1998
charges of plural isotopes t o estimate the interference by spec-

tra. If the interference by spectra cannot be ignored, further dilute
the sample and measure. If influenced still, remove interfering
matrix using a suitable separation method and measure.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Table 51.1 Example of measuring masses
Element name Number of masses
Copper 63, 65
Zinc 66, 68, 67
Cadmium 111, 114
Lead 208, 206, 207
Manganese 55
Chromium 53, 52, 50
Yttrium 89
Indium 115
Bismuth 209
(20) The indicated value means ionic current in number of masses/

electrical charges of the target elements.
(21) When copper, zinc, cadmium, manganese and chromium are de-
termined simultaneously, use the mixed standard solution
[(i pgCu, 1pgZn, 1pgCd, 1pgPb, 1pgMn, 1pgCr)/ml] or the mixed
standard solution [(50 ngCu, 50 ngZn, 50 ngCd, 50 ngPb, 50 ngMn,
50ngCr)/ml], and draw a working curve under the test condi-
tions of each metal element.
Remarks 11 For the sample able t o ignore influence by interfering sub-
stance in the operation of Note (181, the addition of internal
standard substance may be omitted and determine by a work-
ing curve method.
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K O101 : 1998
52 Zinc (Zn) For the determination of zinc, flame atomic absorption method, electric
heating atomic absorption method, ICP atomic emission spectrometry or ICP mass
spectrometry shall be applied.
52.1 Flame atomic absorption method Spray the sample which has been pre-
treated in a acetylene-air flame, and measure the atomic absorption by zinc at 213.9 nm
wavelength to determine zinc.
Determination range: Zn 0.05 t o 2mgll
(1) Reagents Reagent shall be as follows.
(a) Zinc standard solution (0.1mgZdm1) Wash zinc, reference material for
volumetric analysis, specified in JIS K 8005 with hydrochloric acid (1+3),
wash with water, wash with ethanol (99.5) specified in JIS K 8101, then
wash with diethyl ether specified in JIS K 8103, put promptly into a des-
iccator and allow t o stand for 12 h o r longer. Take 0.100 g in respect t o
Zn 100 %, add it in 20 ml of nitric acid (l+l>,dissolve by boiling and expel
nitrogen oxide. After cooling, transfer into a 1O00 ml volumetric flask and
add water up to the marked line. Otherwise, use zinc standard solution
Zn 100 specified in JIS K 0011.
(b) Zinc standard solution (10pgZn/ml) Pipet 50 ml of zinc standard so-
lution (0.1 mgZn/ml) in a 500 ml volumetric flask, add 10 ml of nitric acid
(l+l), and add water up to marked line.
(2) Tool and apparatus Tools and apparatuses shall be as follows.
(b) Zinc hollow cathode lamp
Remarks 1 When sample, with low zinc concentration, does not contain
the substance disturbing extraction operation, preparatory
operation shall be carried out according to Remarks 4 o r 5 of
51.
(a) Spray the sample which has been pretreated in (3)into flame according to
6 of JIS K 0121, and read the indicated value(1) at 213.9 nm wavelength.
(b) Take the same amount of water as that of the sample at preparatory op-
eration in (3)for a blank test, carry out the operations in (3) and (4)fa)
similarly to the sample side, and correct the indicated value obtained on
the sample.
(c) Find the quantity of zinc on the working curve and calculate the concen-
tration (mgZdZ) of zinc in the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
332
K O101 : 1998
Working curve Pipet step by step from 0.5 to 20 ml(2) of zinc standard
solution (10 pgZn/ml) into as many 100 ml volumetric flasks, respectively
add acid t o make the same acid concentration as the sample carried out
the operation in (3)(a),and add water up to the marked line(3). Carry out
the operations in (a) on this solution. Separately, to the water for a blank
test add acid t o make the acid concentration the same as that of the sample
carried out the operation in (3)(a), correct the indicated value obtained on
the standard solution by carrying out the operation in (a), and draw the
relation curve between the quantities of zinc (Zn) and indicated values.
Notes (1) Absorbance or its proportional value shall be valid.
(2) When extraction solvent is applied for preparatory operation, the
amount of zinc standard solution (10 pgZn/ml) can be lessened
according t o circumstances.
(3) When such as butyl acetate layer, 4-metyl-Z-pentanone layer, or
Z76-dimethyl-4-heptanone layer is directly sprayed after the pre-
paratory operation in Remarks 1, the working curve shall be
prepared as follows.
Dilute zinc standard solution (10 pgZn/ml) into suitable con-
centration (0.1 t o 1pgZn/ml), pipet step by step from 0.5 t o 20 ml
of its solution, make them about 100 ml, carry out the operations
in Remarks 1, and (4) (a) and (b)similarly t o the sample, then
draw the relation curve between the quantities of zinc (Zn) and
indicated values.
52.2 Electric heating atomic absorption method After pretreating sample, at-
omize by an electric heating furnace, and measure the atomic absorbance by zinc at
213.9 nm wavelength t o determine zinc.
Determination range: Zn 1 t o 2OpglZ
Remarks 2 This method is easily influenced by type and concentration of co-
existing acid and salt, therefore, apply to samples having less in-
fluence.
(i) Reagents Reagents shall be used as follow.
(a) Water Water A3 specified in JIS K 0557. Carry out a blank test on the
elements t o be determined in advance t o verify that there is no interference.
(b) Nitric acid ( l + l )Prepare using highly purified nitric acid specified in
JIS K 9901.
(c) Zinc standard solution ( i pgzdml) Take 10 ml of zinc standard solu-
tion (10 pgZn/ml) of 52.1 ( i )(b) in a 100 ml volumetric flask, add 2 ml of
of (b) and add water up t o the marked line. Prepare when
nitric acid (l+l)
it is used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
333
K O101 : 1998
(2) Tools and apparatus Tools and apparatus shall be as follows.

(a) Electric heating atomic absorption analyzer Electric heating type ca-
pable of correcting background.
(b) Exothermic body Made of graphite or heat resisting metal
(c) Zinc hollow cathode lamp
(d) Flow gas Argon class 2 specified in JIS K 1105.
(e) Micropipet Piston operated micro-volumetric apparatus 5 to 50 pl speci-
fied in JIS K 0970, or automatic injection apparatus.
(a) Treat the sample according to 4.5.
(4) Operation Operation shall be carried out as follows.
Inject a definite amount (for example, 10 t o 50 p1) of the sample which was
carried out the preparatory operation in (3)to the exothermic body with
the micropipet, dry it (at 100 to 120°C for 30 t o 40s) according t o the
operation in 6 of JIS K 0121, incinerate it (at 600 to 1 0 0 0 ° C for 30 t o
40 s), then atomize it(*)(at 2 200 t o 2 700 "C for 3 t o 6 s), and read the
indicated value(1) at 213.9 nm wavelength(5).
For a blank test, take the same amount of water as that of the sample carried
out in (3)preparatory operation, carry out the operations in (3) and (4)(a)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
similarly t o the sample, and correct the indicated value obtained on the
sample.
Find the amount of zinc on a working curve, and calculate the concentra-
tion (pgZn/Z) of zinc in the sample.
Working curve Pipet step by step 0.1 to 2 ml of zinc standard solution
(1 pgZn/ml) into as many 100 ml volumetric flasks, add respectively acid
to make the acid concentration the same as that of the sample carried out
the operation in (3)(a),and add water up t o the marked line. Carry out
the operation in (a) on this solution. Separately, for a blank test, add acid
in water t o make the acid concentration the same as that of the sample
carried out the operation in (3)(a),carry out the operation in (a)to correct
the indicated value obtained on the standard solution, and draw a relation
curve between the quantities of zinc (Zn) and the indicated values. Pre-
pare a working curve when the sample is measured.
Notes (4) Conditions of drying, incinerating and atomizing depend on ap-
paratus, also may depend on injecting amount of sample and con-
centration of coexisting salts.
(5) Repeat at least 3 times the operation in (a) successively, and
confirm that the indicated values are fit.
52.3 ICP atomic emission spectrometry Spray the sample which has been pre-
treated into inductively coupled plasma through the sample introducing part, and
measure emission by zinc at 213.856 nm wavelength t o determine zinc.
334
K O101 : 1998
Determination range: Zn 10 t o 6 O00 pg/Z


(a) Zinc standard solution (10 pgZn/ml) Follow 52.1 ( i )(b).
(b) Mixed standard solution [(lopgCu, 10 pgzn, 8 pgCd, 10 pgNi, 10 pgPb,
10 pgMn, 10 pgFe)/ml] Follow 51.4 ( i )(b).
Remarks 3 When the sample which has been pretreated has high concen-
tration of sodium, potassium, magnesium, calcium, and low
of zinc, it is allowable t o carry out the operation according t o
Remarks 7 in 51 to determine zinc.
Spray the sample which has been pretreated as in (3) into plasma through
emission strength a t 213.856 nm wavelength(6) (7) (*l.
Take the same amount of water as that of the sample at preparatory op-
eration stated in (3)for a blank test, carry out the operations in (3) and
(4)(a)similarly t o the sample side, and correct the emission strength ob-
Find the quantity of zinc on the working curve, and calculate the concen-
tration (pgZníE) of zinc in the sample.
Working curve Pipet step by step from 0.1 t o 60 ml of zinc standard so-
lution (10 pgZníml)(9)(10) into as many 100 ml volumetric flasks, add re-
spectively acid t o make acid concentration the same as the sample carried
out the operation in (3)(a), and add water up to the marked line. Carry
out the operation in (a) on this solution. Separately, take water for a blank
test, add acid t o make acid concentration the same as the sample carried
out the operation in (3)(a),carry out the operation in (a),correct the emis-
sion strength obtained on the standard solution, and draw the relation curve
between the quantities of zinc (Zn) and emission strengths. Prepare the
Notes ( 6 ) When the apparatus capable of simultaneously measuring two spec-
tra or more with different wavelengths is used, an internal stan-
dard method can be applicable. When the internal standard
method is applied t o the procedures are as follows: take a suitable
amount of sample, which has been treated as in (3)(a),into a
100 ml volumetric flask, add 10 ml of yttrium solution (50 pgY/ml)
[Follow Note (8) in 451, add acid to make the same acid concentra-
tion as the sample in (4)(a),and add water up t o the marked line.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
335
K O101 : 1998
Carry out the spraying operations in (4) (a)on this solution, mea-
sure emission strengths at both 213.856 nm and 371.029 nm (yt-
trium) wavelength, and obtain the emission-strength ratio of zinc
and yttrium.
Separately, pipet step by step from 0.1 to 60 ml of zinc stan-
dard solution (10 pgZníml) into as many 100 ml volumetric flasks,
add respectively 10 ml of yttrium solution (50 pgY/ml), add acid
to make the same acid concentration as the sample in (4) (a),and
add water up to the marked line. Carry out the spraying opera-
tions in (4) (a) on these solutions, measure emission strengths
at both 213.856 nm and 371.029 nm wavelength, draw the rela-
tion curve between emission-strength ratio of zinc to yttrium and
concentration of zinc, and make it working curve. On this working
curve, find the quantity of zinc corresponding to the emission-
strength ratio obtained on the sample, and calculate the concen-
tration (pgZdZ) of zinc in the sample.
(8) I n case of the apparatus capable of using high-order spectrum
has been confirmed.
(9) When, after making preparatory operations according t o Remarks
prepared as follows: dilute zinc standard solution (10 pgZn/ml)
t o suitable concentration (0.1 t o 1 pgZn/ml), pipet step by step
from 0.1 to 60 ml of the solution, make them 500 ml (or a defi-
nite amount of 100 to 500ml), carry out the operations in Re-
marks 3 and (4) (a)and (b) similarly to the sample side, and draw
the relation curve between the quantities of zinc (Zn) and the
emission strengths. --`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(10) When copper, cadmium, nickel, lead, manganese, and iron are
simultaneously tested, use mixed standard solution [( 10 pgCu,
10 pgZn, 8 pgCd, 10 pgNi, 10 pgPb, 10 ygMn, 10 ygFe)/mll, and
prepare preferably the working curve under the test condition
of each metal element.
52.4 ICP mass spectrometry Pretreat a sample, add an internal standard sub-
stance, spray it into an inductively coupled plasma through the sample introducing
part, measure the ionic current in each number of masses/electric charges of zinc
and internal standard substance, and find the ratio between ionic current of zinc
and that of internal standard substance t o determine zinc.
Determination range: Zn 0.5 to 25pg/Z, 10 to 500pglZ
336
K O101 : 1998

Water Follow 52.2 (1) (a).
Nitric acid ( l + l ) Follow 52.2 (1) (b).
Yttrium solution (1 pgY/ml)('l) Follow 51.5 (1) ( c ) .
Zinc standard solution (1 pgZn/ml) Follow 52.2 (1)( c ) .
Zinc standard solution (50 ngZn/ml) Take 50 ml of zinc standard so-
lution ( i pgZním1) in a l O00 ml volumetric flask, add 3 ml of nitric acid
(l+l) and add water up t o the marked line. Prepare when it is used.
1 pgCr)/mll Follow 51.5 (1) (f).
50 ngMn, 50 ngCr)/ml] Follow 51.5 (1) (g).
Note (11) Follow Note (16) of 51.
Remarks 4 Follow Remarks 8 of 51.
(3) Preparatory operation Preparatory operation shall be as follows (12).
(a) Treat a sample according to 4.5.
(b) Take a suitable amount (containing 0.05 t o 50 pg as Zn>of sample treated
in (3)(a) in a 100 ml volumetric flask, add 1 ml of yttrium solution (i pgY/
mi), add nitric acid (l+l) to make final concentration of nitric acid 0.1 to
0.5 mol/Z and add water up to the marked line.
(4) Operation Operation shall be carried out as follows(l3).
Make the ICP mass spectrograph ready t o run, spray the solution in (3)(b)
into the inductively coupled plasma through the sample introducing part,
read the indicated value (15) in the number of masses/electric charges (14) of
zinc and yttrium, and obtain the ratio between the indicated value of zinc
and that of yttrium.
Take the same amount of water for a blank test as that of the sample treated
in (3)(a),carry out the operations in (3)and (4) (a)similarly t o the sample,
obtain the ratio between the indicated value of zinc and that of yttrium,
and correct the ratio of the indicated values between zinc and yttrium ob-
Find the amount of zinc on a working curve, and calculate the concentra-
tion (pgZn/Z) of zinc in the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
337
K O101 : 1998
Working curve Pipet step by step 1 t o 50 ml of the zinc standard solu-

tion (50 pgZn/ml or 1pgZn/ml)(16) in as many 100 ml volumetric flasks, add
1ml of yttrium solution (i pgY/ml), add nitric acid (l+l) t o make the same
acid concentration as that of the sample carried out the operation in (3)(b)
and add water up t o the marked line. Carry out the operation in (a)on
this solution. Separately put 1ml of yttrium solution (i pgY/ml) as a blank
test in a 100 ml volumetric flask, add nitric acid (l+i) to make the same
acid concentration as that of the sample of (3)(b),and add water up to the
marked line. Carry out the operation in (a),correct the ratio of indicated
values obtained on the standard solution, and draw a relation curve of the
ratio between the indicated value t o the amount of zinc (Zn) and the indi-
cated value of yttrium. Prepare the working curve when the sample is
measured.
Notes (13) Follow Note ('8) of 51.
(14) Follow Note (I9) of 51.
(15) Follow Note (20) of 51.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
338
K O101 : 1998
53 Cadmium (Ca) For the determination of cadmium, flame atomic absorption

method, electric heating atomic absorption method, ICP atomic emission spectrom-
etry or ICP mass spectrometry shall be applied.
treated into acetylene-air flame, and measure atomic absorption by cadmium a t
228.8 nm wavelength t o determine cadmium.
Determination range: Cd 50 t o 2 O00 pg/Z
(i) Reagents Reagent shall be as follows.
(a) Cadmium standard solution (0.1 mgCd/ml) Dissolve 0.100 g of cadmium

(99.9 % or more) in 20 ml of nitric acid (l+i>.Expel nitrogen oxide by boil-
ing, cool, transfer into a 1O00 ml volumetric flask and add water up to the
marked line. Otherwise use cadmium standard solution Cd 100 specified
in JIS K 0012.
(b) Cadmium standard solution (10 pgCd/ml) Pipet 50 ml of cadmium stan-

dard solution (0.1 mgCdlm1) into a 500 ml volumetric flask, add 10 ml of
nitric acid (l+l),

(b) Cadmium hollow cathode lamp
(a) Treat sample according to 4.5.

Remarks 1 When sample, with a low concentration of cadmium, has no sub-
stance disturbing extraction operation, the preparatory opera-
tion shall be the following or conform to Remarks 4 o r 5 of 51.
2 When sample contains a lot of iron or manganese, separate
and concentrate cadmium by the following method.
Separation by ion exchange resin
(a) Add hydrochloric acid specified in JIS K 8180 in a suit-
able amount of sample to make about 2 mol/Z acidic hy-
drochloric acid. Flow it to an ion exchange column [I type
strong basic anion-exchange resin which is prepared into
chloride ion form has been filled in a column (for example,
inside diameter lOmm, 200mm in length)] a t a rate of
about 3 ml/min t o adsorb cadmium as chromium complex,
and wash i t with hydrochloric acid ( 1+9).
(b) Change the receiver, elute with nitric acid (1+12) and make
the eluent a definite amount.
339
K O 1 0 1 : 1998
3 When sample contains a lot of zinc, copper, and so on, add a

proper amount of hydrobromic acid into a suitable amount
sample t o make the concentration of hydrobromic acid solu-
tion about 0.5 moVZ, add 4-methyl-2-pentanonesolution (1vol %)
of trioctylamine (N,N-dioctyl-1-octaneamine) by 10 ml per 50 ml
of the above solution, shake them, and extract cadmium. Spray
directly the extracted 4-methyl-2-pentanone layer for atomic
absorption analysis.

Spray the sample which has been pretreated in (3)into flame in accordance
with 6 of JIS K 0121,and read the indicated value(1) a t 228.8 nm wave-
length.
Take the same amount of water as that of the sample a t preparatory op-
eration in (3)for a blank test, carry out the operations in (3) and (4)(a)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
similarly to the sample side, and correct the indicated value obtained on
the sample.
Find the quantity of cadmium on the working curve, and calculate the con-
centration (mgCd/Z) of cadmium in the sample.
Working curve Pipet step by step 0.5 to 20 ml(2) of cadmium standard
solution (10 ygCd/ml) into as many 100 ml volumetric flasks, add respec-
tively acid to make them the same acid concentration as the sample car-
ried out the operation (3)(a),and add water up to the marked line(% Carry
out the operation in (a) on this solution. Separately take water for a blank
test, add acid t o make the acid concentration the same as that of the sample
carried out the operation (3)(a),carry out the operation in (a),correct the
curve between the quantities of cadmium (Cd) and indicated values. Pre-
pare the working curve when sample is measured.
Notes (1) Absorbance o r its proportional value shall be valid.
(2) When solvent extraction is applied as preparatory operations, the
amount of cadmium standard solution (10 ygCd/ml) shall be suit-
ably lessened.
(3) When butyl acetate layer, 4-methyl-2-pentanone layer or 2,6-dim-
ethyl-4-heptanone layer is directly sprayed after the preparatory
operations in Remarks 1, the working curve shall be prepared
as follows.
Dilute cadmium standard solution (10 ygCd/ml) into suitable
concentration (0.1 t o 1pgCd/ml), pipet step by step 0.5 t o 20 ml
out of the solution, make them up t o about 100 ml, carry out the
operations in Remarks 1 and (4) (a)and (b) similarly to the sample
side, and draw the relation curve between the quantities of cad-
mium (Cd) and indicated values.
Remarks 4 Existence of a lot of halide of alkali metals gives a positive
error owing to molecular absorption, light scattering, and so
on, In this case, either shall be carried out, correcting back-
ground o r advance separation of cadmium.
340
K O101 : 1998
53.2 Electric heating atomic absorption method Atomize the sample which
has been pretreated and was added with palladium (II) nitrate as a matrix modifier
in an electric furnace, and measure atomic absorption by cadmium a t 228.8 nm wave-
length to determine cadmium.
Determination range: Cd 0.5 t o lOyg/Z
Remarks 5 This method is easily affected by kind or concentration of coexisting
acid o r salts, therefore this can be applied to the sample which is
less affected.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
element to be determined to verify that there is no interference in use.
(b) Nitric acid (l+l) Prepare using highly purified nitric acid specified in
JIS K 9901.
(c) Palladium (II) nitrate solution (10 pgPd/ml) Dissolve 0.108 g of palla-
dium (II) nitrate in 10 ml of nitric acid (l+l),
transfer it into a 500 ml volu-
metric flask, and add water up to the marked line. Take 20 ml of this solution
into a 200 ml volumetric flask, and add water up to the marked line.
(d) Cadmium standard solution (1 pgCd/ml) Take 10 ml of cadmium stan-
dard solution (10 pgCd/ml) of 53.1 (1) (b)in a 100 ml volumetric flask, add
2 ml of nitric acid (l+l)
(e) Cadmium standard solution (0.1 pgcdml) Pipet 10 ml of cadmium stan-
dard solution (ipgCd/ml) into a 100 ml volumetric flask, add 2 ml of nitric
acid (l+l),and add water up to the marked line.

(a) Electric heating atomic absorption analyzer Electric heating type and
capable of correcting background.
(c) Cadmium hollow cathode lamp
(e) Micropipet Piston operated micro-volumetric apparatus specified in JIS
K 0970, 10 t o 500 p1, or automatic injection device.

(a) Take 15 ml of sample preparatorily operated in (3)in several 20 ml volu-
metric flasks, prepare solutions with adding 0.1 t o 2 ml of cadmium stan-
dard solution (0.1 ygCd/ml) in 3 stages o r more and the one without adding
341
K O101 : 1998
the standard solution, and add nitric acid ( l + l so

) that the acid concentra-
tion of each solution attains the same, then add water up t o the marked
line.
Take a definite amount, not less than 100 p1, of sample which has been car-
ried out (a) into a small vessel using a micropipet, add the same volume of
palladium (II) nitrate solution (10 pgPdml), and mix them sufficiently.
Inject a definite amount (for instance, 10 to 50 p1) of the sample which was
treated in (b) into an exothermic body using a micropipet, dry it (at 100 to
120 O C , for 30 t o 40 s ) according t o 6 of JIS K 0121, ash it (at 500 t o 800 "C,
for 30 to OS), then atomize it(") (at 1600 t o 2200°C, for 3 to 6 s ) , and
read indicated value(1) a t 228.8 nm wavelength(5).
Carry out a blank test as follows. Take water of the same amount as that
of the sample preparatorily operated in (3),carry out the operation in (3)
similarly t o the sample, and put its 15 ml into a 20 ml volumetric flask.
Add nitric acid ( l + l )t o make the acid concentration the same as that of
the sample of (4)(a), and add water up t o the marked line. Carry out the
operations of (b)and ( c ) to correct the indicated value obtained on the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Draw a relation curve between adding amount of cadmium and the indi-
cated value to find the quantity of cadmium, and calculate the concentra-
tion (pgCdlZ) of cadmium in the sample.
Notes (4) The conditions for drying, ashing, and atomizing are different
according t o apparatus. They are often influenced by an injected
amount of sample and concentration of coexisting salts.
(5) Repeat successively at least three times the operation in ( c ) , and
confirm that indicated values sufficiently agree.
53.3 ICP atomic emission spectrometry Spray the sample which has been pre-
treated into inductively coupled plasma through the sample introducing part and
measure emission by cadmium at 214.438 nm wavelength to determine cadmium.
Determination range: Cd 8 t o 2 O00 pg/Z
(a) Cadmium standard solution (8 pgCd/ml) Pipet 40 ml of cadmium stan-
dard solution (0.1 mgCd/ml) stated in 53.1 ( i )(a) in a 500 ml volumetric
flask, add 10 ml of nitric acid (l+l),
(b) Mixed standard solution [(lopgCu, 10 pgZn, 8 pgCd, 10 pgNi, 10 pgPb,
10 pgMn, 10 pgFe)/ml] Follow 51.4 ( i )(b).

342
K O101 : 1998
Remarks 6 When the solution which has been preparatorily operated con-
tains sodium, potassium, magnesium, calcium of high concen-
tration, and low concentrated cadmium, cadmium may be
determined after operations according to Remarks 7 in 51.
(a) Spray the sample which has been preparatorily operated in (3)into plasma
through the sample introducing part according t o 5.8 of JIS K 0116, and
measure emission strength a t 214.438 nm wavelength(6) ( 7 ) (8).
(b) Take the same amount of water as that of the sample at the preparatory op-
eration in (3)for a blank test, carry out the operations in (3)and (4) (a) simi-
larly to the sample, and correct the emission strength obtained on the sample.
(c) Find the quantity of cadmium on the working curve, and calculate the con-
centration (pgCd4) of cadmium in the sample.
Working curve Pipet step by step 0.1 t o 25 ml(9) (10) of cadmium stan-
dard solution (8 pgCd/ml) into as many 100 ml volumetric flasks, add re-
spectively acid t o make the same acid concentration as the sample which
has been carried out in (3)(a),and add water up t o the marked line. Carry
out the operation in (a) on this solution. Separately, take water for a blank
test, add acid t o make the same acid concentration as the solution carried
out in (3)(a),carry out the operation in (a),correct emission strength ob-
tained on the standard solution, and draw the relation curve between the
quantities of cadmium (Cd) and emission strengths. Prepare the working
curve when sample is measured.
or more spectra with different wavelength is used, an internal
standard method can be applicable. When the internal standard
method is applied the procedures are as follows: take a suitable
amount of the sample which has been treated in (3)(a)into a
100 ml volumetric flask, add 10 ml of yttrium solution (50 pgY/
mi) [Follow Note (8) in 451, add acid to make the same acid con-
centration as that of the sample in (4) (a), and add water up t o
the marked line. Carry out the spraying operation in (4) (a),and
measure the emission strengths a t both 214.438nm and
371.029 nm wavelength (yttrium), and obtain the emission-strength
ratio of cadmium and yttrium.
Separately, pipet step by step 0.1 t o 25 ml of cadmium stan-
dard solution (8pgCd/ml) into as many 100 volumetric flasks,
add respectively 10 ml of yttrium solution (50 pgY/ml), add acid
to make the same acid concentration as that of the sample in
(4)(a), and add water up to the marked line. Carry out the
spraying operation in (4)(a) on these solutions, measure the
emission strengths at both 214.438 nm and 371.029 nm wave-
length, draw relation curve between emission-strength ratio of
cadmium t o yttrium and concentration of cadmium, and make it
working curve. On this working curve, find the quantity of cad-
mium corresponding to the emission-strength ratio obtained on
the sample, and calculate the concentration (pgCd/Z) of cadmium
in the sample.
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K O101 : 1998
When working curve method cannot be applied because of high

whatever sample may be used.
I n case of the apparatus capable of using high-order spectrum
has been confirmed.
When, after making preparatory operations according t o Remarks
prepared as follows: dilute cadmium standard solution (8 pgCd/
mi) to suitable concentration (0.1 to 0.8 pgCd/ml), pipet step by
step 0.1 t o 25 ml of the solution, make them 500 ml (or a defi-
nite amount of 100 to 500ml), carry out the operations in Re-
marks 6 and (4)(a)and (b)similarly to the sample side, and draw
the relation curve between the quantities of cadmium (Cd) and
emission strengths.
When copper, zinc, nickel, lead, manganese, and iron are simul-
taneously tested, use mixed standard solution [( 10 pgCu, 10 pgZn,
preferably the working curve under the test condition of each metal
element.
part, measure the ionic current in each number of masses/electric charges of cad-
mium and internal standard substance, and find the ratio between ionic current of
cadmium and that of internal standard substance to determine cadmium.
Determination range: Cd 0.5 t o 25 pg/Z, 10 t o 500 pg/Z

Water Follow 53.2 (i)(a).
Yttrium solution (1 pgY/ml)(ll) Follow 51.5 (1) (cl.
Cadmium standard solution ( i pgCd/ml) Follow 53.2 ( i )(d).
Cadmium standard solution (50 ngCd/ml) Take 50 ml of cadmium stan-
dard solution (ipgCd/ml) in a 1O00 ml volumetric flask, add 3 ml of nitric
acid (l+l) and add water up t o the marked line. Prepare when it is used.
1 pgCr)/ml] Follow 51.5 (1) (f).
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K O 1 0 1 : 1998
(g) Mixed standard solution [50 ngCu, 50 ngZn, 50 ngCd, 50 ngPb,

50 ngMn, 50 ngCr)/mll Follow 51.5 (i) (g).
(3) Preparatory operation Preparatory operation shall be as follows(12).
(b) Take a suitable amount (containing 0.05 t o 50 pg as Cd) of sample treated
in (3)(a) in a 100 ml volumetric flask, add 1ml of yttrium solution (ipgY/
ml), add nitric acid (l+l)t o make final concentration of nitric acid 0.1 to
0.5mol/l and add water up to the marked line.
(4) Operation Operation shall be carried out as follow(13).
Make the ICP mass spectrograph ready to run, spray the solution in (3)(b)
cadmium and yttrium, and obtain the ratio between the indicated value of
cadmium and that of yttrium.
in (3) (a),carry out the operations in (3) and (4)(a) similarly to the sample,
obtain the ratio between the indicated value of cadmium and that of yt-
trium, and correct the ratio of the indicated values between cadmium and
yttrium obtained on the sample.
Find the amount of cadmium on a working curve, and calculate the con-
centration (pgCdlZ) of cadmium in the sample.
Working curve Pipet step by step 1 t o 50ml of the cadmium standard
solution (50 ngCd/ml o r 1pgCd/ml)(16) in as many 100 ml volumetric flasks,
add 1ml of yttrium solution (i pgY/ml), add nitric acid (1+1)to make the
same acid concentration as the sample carried out the operation in (3)(b)
and add water up to the marked line. Carry out the operation in (a)on this
solution. Separately put 1ml of yttrium solution (i pgY/ml) as a blank test
in a 100 ml volumetric flask, add nitric acid (l+l) to make the same acid
concentration as the sample of (3)(b),and add water up t o the marked line.
Carry out the operation in (a),correct the ratio of indicated values obtained
on the standard solution, and draw a relation curve of the ratio between
the indicated value to the amount of cadmium (Cd) and the indicated value
of yttrium. Prepare the working curve when the sample is measured.
Notes (13) Follow Note (18) of 51.
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345
K O101 : 1998

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346
K O 1 0 1 : 1998
54 Nickel (Ni) For the determination of nickel, dimethylglyoxime absorptiometry,

flame atomic absorption method, o r ICP atomic emission spectrometry shall be ap-
plied.
64.1 Dimethylglyoxime absorptiometry Add citrate into a sample, make it weak

alkaline with aqueous ammonia, add 2,3-butanedionedioxime (dimethylglyoxime) t o
produce nickel complex, extract it with chloroform, and back-extract with diluted
hydrochloric acid. Add bromine and aqueous ammonia into the extract to oxidize
nickel, add again Z73-butanedionedioximeto make reddish brown nickel complex, and
measure its absorbance t o determine nickel.
Determination range: Ni 2 to 50pg
Hydrochloric acid (1+20) Prepare using hydrochloric acid specified in
JIS K 8180.
Aqueous ammonia (l+l)
and (1+5) Prepare using aqueous ammonia speci-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
fied in JIS K 8086.

Bromine water (saturated) Add 3 to 4 ml of bromine specified in JIS
K 8529 in 100 ml of water, agitate violently, and use supernatant after stand-
ing for a while.
Diammonium hydrogencitrate solution (100 g/Z) Dissolve 10 g of
diammonium hydrogen citrate specified in JIS K 8284 in about 80ml of
to adjust pH t o about 7, and add water
water, drip aqueous ammonia (l+l)
t o make 100ml.
Phenolphthalein solution (5 g/Z) Follow 13.2 ( i )(a).
Dimethylglyoxime solution in ethanol (10 g/Z) Dissolve 1 g of dimethyl-
glyoxime specified in JIS K 8498 in ethanol (95) specified in JIS K 8102
to make total 100ml.
Dimethylglyoxime solution in sodium hydroxide solution (10 g/Z)
Dissolve l g of dimethylglyoxime specified in JIS K 8498 in sodium hy-
droxide solution (10 g/Z), add sodium hydroxide solution (10 g/Z) to make total
100 ml. If there is undissolved matter, filtrate it.
Nickel standard solution (0.1mgNi/ml) Take 0.100g of nickel (99.9 %
or more) specified in JIS K 9062, dissolve in 20 ml of nitric acid (l+l), heat
it t o expel nitrogen oxide, let i t cool, transfer i t into a 1O00 ml volumetric
flask, and add water up to the marked line. Otherwise, take 0.673g of
ammonium nickel (II) sulfate hexahydrate specified in JIS K 8990, dis-
solve it in mixture of water and 10 ml of nitric acid (l+l), transfer into a
1O00 ml volumetric flask, and add water up to the marked line. Or, use
nickel standard solution, Ni 100, specified in JIS K 0013.
Nickel standard solution (5 ygNi/ml) Pipet 50 ml of nickel standard
solution (0.1 mgNi/ml) into a 1O00 ml volumetric flask, add 20 ml of nitric
acid (l+l), and add water up t o the marked line.
347
K O101 : 1998

(a) Separatory funnel
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(3) Operation Operations shall be as follow.
Take a suitable amount(') (containing 2 t o 50 pg as Ni) of the sample, which
has been treated in 4, into a separatory funnel, add 5 ml of diammonium
hydrogencitrate solution (100 g/l) and 2 or 3 drops of phenolphthalein so-
lution(2) (5 gll) as indicator, and drip aqueous ammonia (1+5) until solu-
tion turns faint red. Drip 2 o r 3 drops of aqueous ammonia (1+5), and add
water to make total about 100ml.
Add 2 ml of dimethylglyoxime ethanol solution (10 g / l ) and 10 ml of chloro-
form, agitate violently for about 1min, let it stand, and transfer chloro-
form layer in another separatory funnel. Add 5 ml of chloroform in water
layer, agitate violently for about l m i n for extraction, let it stand, take
chloroform layer, and put it into the above separatory funnel. Repeat this
operation once more.
Add 10 to 20ml of aqueous ammonia (1+50) into the separatory funnel
retaining chloroform layer, shake for about 30 s, and after standing trans-
fer chloroform layer into another separatory funnel.
Put 10 ml of hydrochloric acid (1+20)in the separatory funnel keeping chlo-
roform layer, agitate violently for about 1min and back-extract nickel. After
standing, transfer chloroform layer into another separatory funnel. Into
this chloroform layer, again add 5 ml of hydrochloric acid (1+20),and re-
peat the back-extraction. Discard chloroform layer, and put water layer
into the above water layer, and transfer it into a 25 ml volumetric flask.
Add 2 ml of bromine water (saturated), shake them, and let it stand for
about 1min. Add aqueous ammonia (l+l) t o neutralize, add more 2 ml of
aqueous ammonia (l+l), and cool it with running water to room tempera-
ture o r below.
Add 2 ml of dimethylglyoxime sodium hydroxide solution (10 g/O, shake t o
colour nickel, and add water up to the marked line.
Place a part of this solution in an absorption cell, and measure its absor-
bance in the vicinity of 450 nm wavelength.
Take about 50 ml of water for a blank test, carry out the operations in (a)
t o ( g ) t o measure absorbance, and correct the absorbance obtained on the
sample.
Find the quantity of nickel on the working curve, and calculate the con-
centration (pgNill) of nickel in the sample.
Working curve Pipet step by step 0.4 to 10 ml of nickel standard solu-
tion (5 pgNi/ml) in as many 25 ml volumetric flasks, respectively carry out
the operations in (e) t o (h),and draw the relation curve between the quan-
tities of nickel (Ni) and absorbances.
348
K O101 : 1998
Notes (1) When sample, with low concentration of nickel, has no organic
substance and turbidity, it is allowable to take a suitable amount
up t o 500 ml of sample and to determine according to the opera-
tions in 4.1 and then (a) to (h). In this case, use all amount of
the sample which has been pretreated, and use the same amount
of reagents as used in (a) to (f). Prepare working curve by simi-
lar operations t o the sample side.
Litmus paper may be available.
treated into an acetylene-air flame, and measure atomic absorption by nickel at
232.0 nm wavelength t o determine nickel.
Determination range: Ni 0.3 t o 6mglZ
(i) Reagent Reagent shall be as follows.
(a) Nickel standard solution (10 pgNi/ml) Pipet 50 ml of nickel standard
solution (0.1 mgNi/ml) stated in 54.1 (i)(i) into a 500 ml volumetric flask,
add 10 ml of nitric acid (l+l>,and add water up t o the marked line.

(a) Flame atomic absorption analyzer Capable of background correction
(b) Nickel hollow cathode lamp
(a) Treat a sample according t o 4.5.
Remarks 1 The preparatory operation for the sample which has low con-
centration of nickel and no substances disturbing extraction
shall be carried out either by the following or by Remarks 2
and Remarks 3.
Take 100ml of the sample into a beaker, add 5 m l of hy-
drochloric acid specified in JIS K 8180, boil for about 5 min,
and let it cool. Carry out the operations according to 54.1 (3) (a)
t o ( c ) , and extract nickel as dimethylglyoxime complex into
chloroform layer. Put together all chloroform layer, add 10 ml
of hydrochloric acid (1+20), shake it, and back-extract nickel.
After separating water layer, add 5 ml of hydrochloric acid
(1+20) into chloroform layer, and back-extract nickel. Put
together all back-extracts, and transfer it into a 25 ml volu-
metric flask, add water up t o the marked line, and use this
solution for determination of nickel.
2 Follow Remarks 4 in 5 1 as appropriate.
3 Follow Remarks 5 in 51 as appropriate.
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349
K O101 : 1998
(a) Spray the sample which has been pretreated in (3) into a flame according
to 6 of JIS K 0121, and read the indicated value(3) at 232.0 nm wavelength.
(b) Take the same amount of water as that of sample at pretreatment of (3)
for a blank test, carry out the operations in (3) and (4)(a) similarly to the
sample side, and correct the indicated value obtained on the sample.
(c) Find the quantity of nickel on the working curve, and calculate the con-
centration (mgNilZ) of nickel in the sample.
Working curve Pipet stepwise 3 to 60 ml(4) of nickel standard solution
(10 pgNilm1) into as many 100 ml volumetric flasks, respectively add acid
to make the acid concentration the same as that of the sample carried out
the operation in (3)(a),and add water up t o the marked line. Carry out
the operation of (a) on this solution. Separately take water for a blank
test, add acid t o make the acid concentration the same as that of the sample
carried out the operation of (3)(a), carry out the operation in (a), correct
the indicated value obtained on the standard solution, and draw a relation
curve between the amounts (Ni) of nickel and indicated values, Prepare
the working curve when sample is measured.
Notes (3) Absorbance o r its proportional value shall be valid.
(4) When solvent extraction method is applied as preparatory op-
erations, the amount of nickel standard solution shall be suit-
ably lessened.
54.3 ICP atomic emission spectrometry Spray a sample into an inductively

coupled plasma through the sample introducing part after pretreatment, measure
the emission by nickel a t 221.647 nm wavelength t o determine nickel.
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Determination range: Ni 40 to 2 O00 pglZ

(a) Nickel standard solution (10 pgNi/ml) Follow 54.2 ( i )(a).
10 pgMn, 10 pgFe)/mll Follow 51.4 (i)(b).

(a) Pretreat a sample according t o 4.5.
Remarks 4 When the solution which has been pretreated has high con-
centration of sodium, potassium, magnesium, calcium, and so
on, and low of nickel, it is allowable t o carry out the opera-
tions according t o Remarks 7 in 51 t o determine nickel.
350
K O101 : 1998

(a) Spray the sample which has been pretreated in (3)into a plasma through
emission strength at 221.647 nm wavelength(5) (6) (7).
(b) Take the same amount of water as that of sample at pretreatment of (3)
for a blank test, carry out the operations in (3) and (4)(a) similarly to the
sample side, and correct the emission strength obtained on the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(c) Find the quantity of nickel on the working curve, and calculate the con-
centration (pgNi/Z) of nickel in the sample.
Working curve Pipet step by step 0.4 t o 20 ml(8) (9) of nickel standard
solution (10 pgNi/ml) into as many 100 ml volumetric flasks, respectively
add acid t o make the same acid concentration as the sample which has been
pretreated in (3)(a),and add water up t o the marked line. Carry out the
operation in (a)on this solution. Separately take water for a blank test,
add acid to make the same acid concentration as the sample in (3)(a),carry
out the operation in (a),correct the emission strength obtained on the stan-
dard solution, and draw the relation curve between the quantities of nickel
(Ni) and emission strengths. Prepare the working curve when sample is
measured.
spectra with different wavelength is used, an internal standard
method can be applicable. When the internal standard method
is applied, the procedures are as follows: take a suitable amount
of sample which has been treated in (3)(a)into a 100 ml volu-
metric flask, add 10ml of yttrium solution (50pgY/ml) [follow
Note ( 8 ) in 451, and add acid to make the same acid concentra-
tion as the sample in (4)(a),and add water up t o the marked
line. Carry out the spraying operation in (4)(a) on this solu-
tion, measure emission strengths a t both 221.647 nm and
371.029 nm (yttrium) wavelength, and obtain emission-strength
ratio of nickel and yttrium.
Separately, pipet step by step 0.4 to 20 ml of nickel standard
solution (10 pgNi/ml) into as many 100 ml volumetric flasks, re-
spectively add 10 ml of yttrium solution (50 pgY/ml), add acid t o
make the same acid concentration as the sample in (4) (a), and
tions in (4)(a) on these solutions, measure emission strengths
a t both 221.647 nm and 371.029 nm wavelength, draw the rela-
tion curve between emission-strength ratio of nickel to yttrium
and concentration of nickel, and make it working curve. On this
working curve, find the quantity of nickel corresponding t o the
concentration (pgNil2) of nickel in the sample.
concentration of salts in a sample, the standard addition method
351
K O101 : 1998

has been confirmed.
prepared as follows: dilute nickel standard solution (10 pgNi/ml)
to suitable concentration (0.2 to 1 ygNi/ml), pipet step by step
0.4 t o 20 ml of the solution, make them 500 ml (or a definite
amount of 100 t o 500 mi), carry out the operations in Remarks 4
and (4)(a)and (b)similarly to the sample side, and draw the
relation curve between the quantities of nickel (Ni) and the
emission strengths.
(9) When copper, zinc, cadmium, lead, manganese, and iron are si-
multaneously tested, use mixed standard solution [( 10 pgCu,
10 pgZ, 8 pgCd, 10 pgNi, 10 ygPb, 10 pgMn, 10 ygFe)/ml], and
prepare preferably a working curve under the test condition of
each metal element.
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352
K O101 : 1998
55 Tin (Sn) For the determination of tin, phenylfluorone absorptiometry, querce-

tin absorptiometry or ICP atomic emission spectrometry shall be applied.
65.1 Phenylfluorone absorptiometry Make tin tin (IV) with potassium perman-
at existence of citric
ganate, add phenylfluorone (2,3,7-trihydroxy-9-phenyl-6-fluorone)
acid, polyvinyl alcohol t o generate yellow complex, and measure its absorbance.
Determination range: Sn 3 t o 40pg

Hydrochloric acid (1+11) Prepare using hydrochloric acid specified in
JIS K 8180.
K 8180.
Sulfuric acid (l+l) Follow 4.4 (i)(b).
L(+)-ascorbicacid Specified in JIS K 9502.
Citric acid solution (100 g/Z) Dissolve 11g of citric acid monohydrate
specified in JIS K 8283 in water t o make total 100 ml.
Aqueous ammonia (l+l)Prepare using aqueous ammonia specified in
JIS K 8085.
Potassium permanganate solution (3 glZ) Follow 46.1 ( i )(f).
Polyvinyl alcohol solution (5 g/Z) Dissolve 0.5 g of polyvinyl alcohol (about
80 mol% of saponification degree) specified in JIS K 9550 in water t o make
total 100ml.
Bromocresol green solution (0.5g/Z) Dissolve 0.05 g of bromocresol green
specified in JIS K 8840 in 20 ml of ethanol (95) specified in JIS K 8102,
and add water t o make total 100ml.
Phenylfluorone solution (0.1 g/Z) Dissolve 0.05 g of phenylfluorone speci-
fied in JIS K 9547 in the mixture of 100 ml ethanol (95) specified in JIS
K 8102 and 10ml hydrochloric acid (1+2), and add ethanol (95) t o make
total 500ml.
Tin standard solution (0.1 mgSn/ml) Put 0.100 g of tin specified in JIS
K 8580 in a beaker, add 10 ml of sulfuric acid, cover it with a watch glass,
and heat it t o dissolve. After cooling, add 200 ml of hydrochloric acid (1+4)
for dissolving, and let it cool t o room temperature. Transfer it into a 1O00 ml
volumetric flask, and add hydrochloric acid (1+4) up to the marked line.
Tin standard solution (2 pgSn/ml) Pipet 10 ml of tin standard solution
(0.1 mgSn/ml) in a 500 ml volumetric flask, and add hydrochloric acid (1+10)

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353
K 0101 : 1998

Take a suitable amount (containing 7.5 t o 100 pg as Sn) of the sample which
has been treated in 4, add 5 ml(1) of sulfuric acid (l+l),heat it to generate
white fume of sulfuric acid, and concentrate t o nearly dry.
After cooling, add gradually 10ml of hydrochloric acid (1+4), heat it t o
dissolve, cool it t o room temperature, transfer it into a 50ml volumetric
flask, and add water up t o the marked line.
Pipet 20 ml each into a 50 ml volumetric flask and a 100 ml beaker.
Add water into the beaker up to about 50 ml, add 2 to 3 drops of bromocresol
green solution (0.5 glZ) as indicator, and titrate it until the solution turns
green with aqueous ammonia (l+l), and record the volume of aqueous
ammonia (l+l) needed for neutralization.
Into the solution kept in the 50ml volumetric flask, drip potassium per-
manganate solution (3glZ) until the solution turns faint red, and let it stand
for about 5min t o oxidize tin.
Add about 0.1 g of L(+)-ascorbic acid, shake it, and reduce excessively added
permanganate.
Add 5 ml of citric acid solution (100 gll) and 1.5 ml of hydrochloric acid (l+ll),
and add the same amount of aqueous ammonia (l+l) as needed t o neutral-
ize a t (d)(pH becomes 1.5 to 2.0).
Add 5 ml of polyvinyl alcohol solution (5 gll) and 5 ml of phenylfluorone
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
solution (0.1 glZ), shake them, add water up t o the marked line, and let it
stand for about 20 min.
Pipet 10 ml of hydrochloric acid (1+4)in a 50 ml volumetric flask for a blank
test, add water up t o the marked line. Hereafter carry out the operations
in ( c ) to (h).
Place a part of the solution obtained a t (h)into an absorption cell, and
measure absorbance in the vicinity of 510 nm wavelength with making the
solution obtained at (i) a reference solution.
Find the quantity of tin on the working curve, and calculate the concentra-
tion (pgSníZ) of tin in the sample.
Working curve Pipet step by step 1.5 t o 20 ml of tin standard solution
(2pgSn/ml) in as many 50 ml volumetric flasks and as many 100 ml bea-
kers, carry out the operations in (d)to ci), and draw the relation curve between
the quantities of tin (Sn) and absorbances.
Note (1) When sulfuric acid is used a t pretreatment, don’t add here.
Remarks 1 When a sample, with low concentration of tin, has no organic
substance and turbidity, take a suitable amount of sample t o
500 ml or less, and concentrate it by means of the coprecipitation
method with manganese (IV) oxide (manganese dioxide) as fol-
lows.
Take 100 ml of the sample, add 3 ml of nitric acid and 2 ml
of manganese (II) nitrate solution [Dissolve 16 g of manganese
354
K O101 : 1998
(II) nitrate hexahydrate specified in JIS K 8568 in water up t o

100 mi.], and boil gently. Then, add 2 ml of potassium perman-
ganate solution (30 glZ) (dissolve 3 g of potassium permangan-
ate specified in JIS K 8247 in water to make 100 mi) per 100 ml
of solution, and boil gently for 5 to 10 min to precipitate man-
ganese (IV) oxide. After standing for about 20min, filtrate
through filter paper 5 grade B, and wash several times with
warm water. Transfer the precipitate into the original beaker,
dissolve the precipitate on the filter paper by washing the pa-
per with dripping alternately 10 ml of nitric acid (i+l)and hy-
drogen peroxide (1+30), and wash the paper with warm water.
Put these washings into the original beaker, heat t o dis-
solve manganese (IV) oxide, heat on t o decompose hydrogen
peroxide, and make about 200 ml by adding water. Add 1 ml
of manganese (II) nitrate solution in this solution, and heat
t o boil gently. Add 2 ml of potassium permanganate solution
(30 g/Z), boil it for 5 t o 10 min, and again precipitate manga-
nese (IV) oxide. Filtrate this, and wash it with warm water.
Put the precipitate into the original beaker, dissolve the pre-
cipitate on the filter paper by dripping mixture of 10 ml of warm
sulfuric acid (1+3) and 5 ml of hydrogen peroxide (1+30), and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
put the filtrate into the original beaker. Wash the paper with
water, and put washings together. Heat it t o dissolve the pre-
cipitate and t o generate white fume of sulfuric acid as well,
and concentrate it until near drying. After cooling, add 10 ml
of hydrochloric acid (1+4), and heat it t o dissolve residue. After
letting it cool, transfer this solution into a 50 ml volumetric
flask, add water t o the marked line, hereafter follow the op-
erations in (3)( c ) t o (j).
2 Disturbing elements are such as germanium, zirconium, anti-
mony, bismuth, iron, etc. As to iron, the reduction by L(+)-
ascorbic acid prevents its disturbance if its amount is about
10 mg o r less. As to antimony, after oxidizing it to 5 valences,
and as t o bismuth, after colouring it, then add 0.3 g of disodium
dihydrogen ethylenediaminetetraacetate dihydrate specified in
JIS K 8107 to change them individual complex, and they don’t
disturb if their amounts are 0.5 mg or less respectively.
55.2 Quercetin absorptiometry Make tin tin (IV) with potassium permanga-
nate and reduce the excess permanganate. Add quercetin [2-(3,4-dihydroxyphenyl)-
3,5,7-trihydroxy-4H-benzopyrane-4-one],generate yellow tin-quercetin complex, extract
4-methyl-2-pentanone, and measure its absorbance to determine tin.
Determination range: Sn 2 t o 20pg
K 8180.
(b) Sulfuric acid (l+l) Follow 4.4 ( i )(b).
355
K O101 : 1998
Sulfuric acid (1+19) Prepare using sulfuric acid specified in JIS K 8951.
L(+)-ascorbicacid One specified in JIS K 9502.
Potassium permanganate solution (3 gll) Follow 46.1 (1) (f).
Sodium sulfate One specified in JIS K 8987.
Thiourea solution (50 gld) Dissolve 10 g of thiourea specified in JIS K
8635 in water to make 200 ml.
4-methyl-2-pentanone One specified in JIS K 8903.
Quercetin solution ( i gll) Dissolve 0.2 g of quercetin in about 100 ml of
ethanol (99.5) specified in JIS K 8101, add 10 ml of hydrochloric acid specified
in JIS K 8180, and make it 200 ml with ethanol (99.5). Prepare when it is
used.
Tin standard solution (5 pgSn/ml) Take 10 ml of tin standard solution
(0.1 mgSdm1) of 55.1 ( i )(k)in a 200 ml volumetric flask and add hydro-
chloric acid (1+10) up t o the marked line.
(2) Tool and apparatus They shall be as follows.
(a) Separatory funnel 100 ml
(3) Operation Operations shall be carried out as follows.

Take a suitable amount (containing 2 to 20 pg as Sn) of the sample oper-
ated in 4, add 5 ml of sulfuric acid(l), heat to generate white fume of sul-
furic acid, concentrate t o near drying.
After cooling, add 5 ml of sulfuric acid, dissolve by heating, drip potassium
permanganate solution ( 3 g/Z) until the solution turns to faint red, and let
it stand for about 5 min t o oxidize tin.
Add 0.1 g of L(+)-ascorbicacid, mix by shaking and reduce the excess per-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
mangana te.
Transfer it together with a little amount of water into the 100 ml separatory
funnel, add 20 ml of thiourea solution (50 g/Z) and 15 ml of hydrochloric acid
(l+l)and mix by shaking. Add 20 ml of quercetin solution ( ig/Z), shake
again, and stand for about 15 min.
Add 15 ml of 4-methyl-2-pentanone7shake violently for about 1 min, and
extract tin-quercetin complex.
Discard a water layer, add 25 ml of sulfuric acid (1+19) in the organic layer
and mix by shaking for about 30 s.
Discard a water layer, add about 5 g of sodium sulfate in the organic layer
and mix by shaking.
For a blank test, take 5 ml of sulfuric acid (1+19) and carry out the opera-
tions of (b)t o (g).
356
K O101 : 1998
(i) Take a part of the organic layer of ( g ) in an absorbing cell, and measure
the absorbance in the vicinity of 440 wavelength with making the organic
layer of (h)reference solution.
Cj) Find the amount of tin on the working curve, and calculate the concentra-
tion (pgSdZ) of tin in the sample.
Working curve Pipet step by step 0.4 t o 4 m l of tin standard solution
(5 pgSn/ml), add 5 ml of sulfuric acid (1+19)respectively, heat to make volume
about 5 ml, and add potassium permanganate solution ( 3 g/Z) until the colour
of solution turns to faint red. Hereafter, carry out the operations ( c ) t o (i)
and draw a relation curve between the amount (Sn) of tin and the absor-
bances.
Remarks 3 For samples, not containing organic matter and turbidity, with
low concentration of tin, separate and concentrate them by
means of the coprecipitation method with manganese (IV) oxide
according t o Remarks 1, hereafter, carry out the operations
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(3)(b)and after t o determine tin.
55.3 ICP atomic emission spectrometry After the sample is pretreated, spray
the sample into an inductively coupled plasma through the sample introducing part,
and measure emission by tin at 189.989 nm wavelength to determine tin.
Determination range: Sn 0.4 t o 2 mg/Z
(a) Tin standard solution (10 pgSn/ml) Take 10 ml of tin standard solu-
tion (0.1 mgSn/ml) of 55.1 ( i )(k)in a 100 ml volumetric flask, and add hy-
drochloric acid (1+10) up t o the marked line.
(3) Preparatory operation The preparatory operation shall be as follows.
(4) Operation The operations shall be carried out as follows.
(a) Spray the sample operated in (3)into a plasma through the sample intro-
ducing part according to 5.8 of JIS K 0116, and measure the emission strength
at 189.989 wavelength(2)( 3 ) (4).
(b) Take the same amount of water as that of the sample at the preparatory
operation stated in (3)for a blank test, carry out the operations in (3) and
(4) (a) similarly t o the sample, and correct the emission strength obtained
on the sample.
(cl Find the quantity of tin on the working curve, and calculate the concentra-
tion (mgSn/Z) of tin in the sample.
357
K O 1 0 1 : 1998
Working curve Pipet step by step 4 t o 20ml of tin standard solution

(10 pgSn/ml) into as many 100 ml volumetric flasks, add respectively acid
to make the same acid concentration as that of the sample carried out the
operation in (3)(a)and add water up t o the marked line. Carry out the
operation in (a) on this solution. Separately, take water for a blank test,
add acid to make the same acid concentration as that of the sample carried
out the operation in (3)(a),carry out the operation in (a),correct the emis-
sion strength obtained on the standard solution, and draw the relation curve
between the quantities of tin (Sn) and emission strengths. Prepare the
When the apparatus capable of simultaneously measuring two
spectra o r more with different wavelengths is used, an internal
standard method can be applicable. When the internal standard
method is applied, take a suitable amount of sample treated in
(3)(a)in a 100 ml volumetric flask, add 10 ml of yttrium solu-
tion (50pgY/ml) [follow Note (8) of 451, add acid t o make the same
acid concentration as that of the sample of (4) (a), and add wa-
ter up t o the marked line. Carry out the spraying operation in
(4)(a) on this solution, measure emission strengths a t both
189.989 nm and 371.029 nm (yttrium) wavelength, and obtain the
ratio of emission strengths between tin and yttrium.
Separately, pipet step by step 4 to 20 ml of tin standard so-
lution (10 pgSn/ml) in as many 100 ml volumetric flasks, add re-
spectively 10 ml of yttrium solution (50 pgY/ml) t o make the same
acid concentration as that of the sample of (4)(a),and add wa-
ter up t o the marked line. Carry out the spraying operation of
(4)(a),measure emission strengths a t both 189.989 nm and
371.029 nm wavelength, draw the relation curve between emis-
sion-strength ratio of tin to yttrium and concentration of tin, and
make it working curve. On this working curve, find the quan-
tity of tin corresponding to the emission-strength ratio obtained
on the sample, and calculate the concentration (mgSn/Z) of tin
in the sample.
When working curve method cannot be applied because of high
specified in 5.8.3 (2)of JIS K 0116 is preferably applied. In this
case, the correction of background is necessary irrespective of
sample type.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
358
K O101 : 1998
56 Lead (Pb) For the determination of lead, flame atomic absorption method, electric
heating atomic absorption method, ICP atomic emission spectrometry or ICP mass
spectrometry shall be applied.
treated into an acetylene-air flame, and measure atomic absorption by lead at 283.3 nm
wavelength to determine lead.
Determination range: Pb 1 t o 20mglZ
(a) Lead standard solution (0.1 mgPb/ml) Take 0.100 g of lead (99.9 % o r
more) Specified in JIS K 8701, dissolve it by adding 40ml of nitric acid
(1+3),heat to expel nitrogen oxide, let it stand to cool, transfer into a 1O00 ml
volumetric flask and add water up to the marked line. Otherwise, take
0.160 g of lead (II) nitrate specified in JIS K 8563, dissolve it in 20 ml of
nitric acid (l+l) and a suitable amount of water, transfer into a 1O00 ml
volumetric flask and add water up to the marked line. Otherwise use lead
standard solution Pb 100 specified in JIS K 0015.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Flame atomic absorption analyzer One capable of background correc-
tion
(b) Lead hollow cathode lamp
(a) Treat a sample according t o 4.5(1).
Note (1) In case of direct spraying water solution, the pretreatment using
sulfuric acid should not be carried out because sulfate ion gives a
negative error on lead determination.
Remarks 1 The preparatory operation for the sample which has a low con-
centration of lead and has not a disturbing substance shall
be carried out according t o Remarks 4 or 5 of 51.
(a) Spray the sample which has been pretreated as in (3)into a flame according
to 6 of JIS K 0121, and read the indicated value(2) at 283.3 nm wavelength.
(b) Take the same amount of water as that of sample pretreated in (3)for a
blank test, carry out the operations in (3)and (4) (a) similarly to the sample
side, and correct the indicated value on the sample.
(c) Find the quantity of lead on the working curve, and calculate the concen-
tration (mgPblZ) of lead in the sample.
Working curve Pipet step by step 1t o 20 ml(3) of lead standard solution
(0.1 mgPblm1) into as many 100 ml volumetric flasks, respectively add acid
to make the same acid concentration as the sample carried out in (3)(a),
and add water up to the marked line(*). Carry out the operation in (a) on
359
K O101 : 1998
this solution. Separately, take water for a blank test, add acid t o make
the same acid concentration as that of the sample carried out in (3)(a),
carry out the operation in (a) t o correct the indicated value obtained on
the standard solution, and draw the relation curve between the quantities
of lead (Pb) and indicated values. Prepare the working curve when a sample
is measured.
Notes (2) Absorbance or its proportional value shall be valid.
(3) When solvent extraction is carried out as preparatory operation,
the amount of lead standard solution (0.1 mgPb/ml) shall be
suitably decreased.
(4) When such as butyl acetate layer, 4-methyl-2-pentanone layer,
or 2,6-dimethyl-4-heptanone layer is directly sprayed after pre-
paratory operation in Remarks 1, working curve is prepared as
follows: dilute lead standard solution ( O . 1mgPb/ml) into suitable
concentration (i t o 5 pgPb/ml), pipet step by step 1 to 20 ml of
the solution, make them about 500 ml (or definite amount from
100 t o 500 ml), carry out the pretreatment in (3)similarly t o the
sample, and draw the relation curve between the quantities of
lead (Pb) and indicated values.

add palladium (II) nitrate as a matrix modifier, atomize in an electric furnace, and
measure atomic absorption by lead at 283.3 nm wavelength t o determine lead.
Determination range: Pb 5 t o lOOpg/Z
Remarks 2 Because this method is easily affected by the kind of coexisting
salts or acids and their concentrations, the sample which is less
affected shall be adopted.

(a) Water Water A3 specified in JIS K 0557. Carry out a blank test on el-
ement to be determined, and verify that there is no interference for using.
JIS K 9901.
(c) Palladium (II) nitrate solution (10pgPb/ml) Follow 53.2 ( i )( c ) .
(d) Lead standard solution (ipgPb/ml) Pipet 10 ml of lead standard so-
lution (0.1 mgPb/ml) of 56.1 ( i )(a) into a l O00 volumetric flask, add 20 ml
of nitric acid (l+l),
(c) Lead hollow cathode lamp
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
360
K 0101 : 1998

K 0970, 10 to 500 pl. Or an automatic injection device.


Put 15 ml of sample carried out the preparatory operation in (3)in as many
20 ml volumetric flasks, prepare solutions by adding lead standard solution
(1 pgPb/ml) 3 steps or more in the range from 0.1 to 2 ml respectively and
one not added the standard solution, add nitric acid (l+i)to make each acid
concentration of the solutions the same, and add water up to the marked line.
Take a definite amount, 100 pl or more, of the sample which has been pre-
treated as in (a) into a small vessel using a micropipet, and add palladium
(II) nitrate solution (10 pgPd/ml) by the same volume as the sample, and
mix them sufficiently.
Inject a definite volume (for instance, 10 to 50 pl) of the sample which has
been treated in (b)into an exothermic body using a micropipet, carry out
the operation according t o 6 of JIS K 0121,dry it (100 t o 120 "C for 30 t o
40 s), ash it (500 to 800 "C for 30 to 40 SI, then atomize i t ( 5 ) ( i 800 t o 2 500 "C
for 3 t o 6 s), and read the indicated value(2) at 283.3 nm wavelength(6).
Take the same amount of water as that of the sample at pretreatment in (3)
for a blank test, carry out the operations in (3)similarly to the sample side,
and put its 15 ml in a 20 ml volumetric flask. Then, add nitric acid ( l + l )to
make the acid concentration the same as that of the solution of (4)(a) and
add water up to the marked line. Carry out the operations in (b)and (c) on
this solution, and correct the indicated value obtained on the sample.
Find the quantity of lead by preparing a relation curve between the amount
of read and the indicated value, and calculate the concentration (pgPb/Z) of
lead in the sample.
Notes (5) The condition for drying, ashing, o r atomizing varies depending
upon apparatus, and they may be affected by such as injected
volume of a sample and concentration of coexisting salts.
(6) Repeat successively at least 3 times the operations in (c), and
confirm the indicated values agree.
56.3 ICP atomic emission spectrometry After pretreatment of a sample, spray

it in an inductively coupled plasma through the sample introducing part, and mea-
sure emission by lead at 220.351 nm wavelength to determine lead.
Determination range: Pb 0.1 t o 2 mg/Z
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
361
K O101 : 1998

(a) Lead standard solution (10 pgPb/ml) Pipet 10 ml of lead standard so-
lution (0.1 mgPb/ml) stated in 56.1 (i)(a) into a 100 ml volumetric flask,
add 10ml of nitric acid (i+l), and add water up to the marked line.
10 pgMn, 10 pgFe)/mll Follow 51.4 ( i )(b).
Remarks 3 When the solution, which has been preparatorily operated, has
rich concentration of sodium, potassium, magnesium and cal-
cium, and poor in lead concentration, the operation in Remarks
7 of 51 should preferably be carried out.
the sample introducing part according to 5.8 of JIS K 0116, measure emission
strength a t 220.351 nm wavelength(7) ( 8 ) (9).
(b) Take the same amount of water as that of sample at preparatory operation
in (3)for a blank test, carry out the operations in (3) and (4) (a) similarly
to the sample side, and correct the emission strength obtained on the sample.
( c ) Find the quantity of lead on the working curve, and calculate the concen-
tration of lead (mgPb/Z) in the sample.
Working curve Pipet step by step 1to 20 ml(10) (11) of lead standard solution
(10 pgPb/ml) into as many 100 ml volumetric flasks, respectively add acid
to make the same acid concentration as that of the sample pretreated in
(3) (a), and add water up t o the marked line. Carry out the operation in
(a) on this solution. Separately, take water for a blank test, add acid t o
make the same acid concentration as that of the sample pretreated in (3) (a),
carry out the operation in (a), correct the emission strength obtained on
of lead (Pb) and emission strengths. Prepare the working curve when sample
is measured.
the procedures are as follows: Take a suitable amount of sample,
which has been treated in (3)(a),into a 100 ml volumetric flask,
add 10 ml yttrium solution (50 pgY/ml) [follow Note (8) of 451, add
acid to make the same acid concentration as the sample in (4)(a),
and add water up to the marked line. Carry out the spraying
operation in (4)(a) on this solution, measure emission strengths
at both 220.351 nm and 371.029 nm (yttrium) wavelength, and
obtain the emission-strength ratio of lead and yttrium.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
362
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Separately, pipet step by step 1 t o 20 ml of lead standard so-

lution (10 pgPb/ml) into as many 100 ml volumetric flasks, re-
spectively add 10 ml of yttrium solution (50 kgY/ml>,add acid to
make the same acid concentration as the sample of (4)(a),and
tion in (4)(a)on these solutions, measure emission strengths at
both 220.351 nm and 371.029 nm wavelength, draw the relation
curve between emission-strength ratio of lead to yttrium and the
concentration of lead, and make it the working curve. On this
working curve, find the quantity of lead corresponding to the
concentration (mgPb/Z) of lead in the sample.
When the working curve method cannot be applied because of
described in 5.8.3 (2) of JIS K 0116 is preferably applied. In
this case, however, the correction of background is necessary what-
ever sample may be used.
I n the case of the apparatus capable of using high-order spec-
trum lines, these lines can be used.
has been confirmed.
When, after making preparatory operations according to Remarks
prepared as follows: dilute lead standard solution (10 pgPb/ml)
t o suitable concentration (0.1 t o 1pgPb/ml), take step by step 1
t o 20 ml of the solution, make them 500 ml (or a definite amount
of 100 t o 500ml), carry out the operations in Remarks 3, and
(4)(a)and (b)similarly t o the sample side, and draw the rela-
tion curve between the quantities of lead (Pb) and emission
strengths.
When copper, zinc, cadmium, nickel, manganese, and iron are
simultaneously tested, use mixed standard solution [( 10 pgCu,
10 pgZn, 8 pgCd, 10 pgNi, 10 pgPb, 10 kgMn, 10 pgFe)/mll, and
prepare preferably each working curve under the test condition
56.4 ICP mass spectrometry After pretreated sample, add internal standard sub-
part, measure ionic current in each number of masses/electric charges of lead and
internal standard substance, and obtain the ratio between ionic current of lead and
that of internal standard substance t o determine lead.
Determination range: Pb 0.5 to 25pg/Z, 10 to 50OpglZ
363
K O101 : 1998

Yttrium solution (1 pgY/ml)(l2) Follow 51.5 (1)( c ) .
Lead standard solution (1pgPb/ml) Follow 56.2 (1)(d).
Lead standard solution (50 ngPb/ml) Take 50 ml of lead standard so-
lution (1pgPb/ml) in a l O00 ml volumetric flask, add 3 ml of nitric acid
(l+l) and add water up t o the marked line. Prepare it when used.
Mixed standard solution [(iygCu, 1 ygZn, 1pgCd, 1 pgPb, 1 ygMn,
1 pgCr)/ml] Follow 51.5 (1) (f).
50 ngMn, 50 ngCr)/ml] Follow 51.5 (1)(g).
(3) Preparatory operation Preparatory operations shall be as follows ( 1 3 ) .
(b) Put a suitable amount (containing 0.01 to 50 pg as Pb) of the sample treated
in (3) (a) in a 100 ml volumetric flask, add 1ml of yttrium solution (1pgY1
mi), add nitric acid (l+l) t o make the final concentration of nitric acid 0.1
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
to 0.5 mol/Z, and add water up to the marked line.
(4) Operation Operations shall be carried out as follows(l4).
Let the ICP mass spectrograph be ready to run, spray the solution of (3)(b)
into an inductively coupled plasma through the sample introducing part,
lead and yttrium, and obtain the ratio between the indicated values of lead
and yttrium.
Take the same amount of water as that of the'sample of (3) (a)for a blank
test, carry out the operations in (3) and (4)(a)similarly to the sample, obtain
the ratio between the indicated values of lead and yttrium, and correct the
ratio between indicated values of lead and yttrium obtained on the sample.
Find the amount of lead on the working curve, and calculate the concen-
tration (pgPb/Z) of lead in the sample.
Working curve Take step by step 1t o 50 ml(17) of lead standard solution
(50 ngPb/ml o r 1 pgPb/ml) in as many 100 ml volumetric flasks, add 1 ml of
yttrium solution (1pgY/ml), add nitric acid (l+l)t o make the same acid
concentration as that of the sample of (3)(b),and add water up to the marked
line. Carry out the operation in (a) on this solution. Separately, add 1ml
364
K O 1 0 1 : 1998
of yttrium solution (1pgY/ml) as a blank test in a 100 ml volumetric flask,

add acid to make the same acid concentration as that of the sample of (3)(b),
add water up to the marked line, carry out the operation of (a)to correct the
indicated value obtained on the standard solution, and draw a relation curve
of the ratio between the indicated value t o the amount of lead (Pb) and the
indicated value of yttrium. Prepare the working curve when the sample is
measured.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
365
K O101 : 1998
57 Mercury (Hg) For the determination of mercury, atomic absorption spectrometry

by reduction-vaporization or atomic absorption spectrometry by heating-vaporization
shall be applied.
57.1 Atomic absorption spectrometry by reduction-vaporization After a

sample is pretreated with potassium permanganate, reduce mercury (II) with tin (II)
chloride. Measure atomic absorption, caused by mercury vapor generated by aera-
tion, a t 253.7 nm wavelength t o determine mercury.
Determination range: Hg 0.5 t o 10pglZ
Nitric acid Specified in JIS K 8541 and containing mercury by 0.1pgll
o r less.
Sulfuric acid (l+l) Follow 4.4 (1)(b). However, use the one containing
mercury by 1 pglZ or less.
Potassium permanganate solution (50g l l ) Dissolve 50 g of potassium
permanganate(1) specified in JIS K 8247 in water, filtrate through a glass
filter, and add water t o make total 11. Store it in a coloured glass bottle.
Potassium peroxodisulfate solution (50 gll) Dissolve 50 g of potassium
peroxodisulfate specified in JIS K 8253 in water t o make total 1Z(2).
Hydroxylammonium chloride solution (80 gll) Dissolve 8 g of hydroxyl-
ammonium chloride specified in JIS K 8201 in water t o make total 100 ml.
This solution should contain 1pgll or less of mercury. When purification
is needed, transfer this solution into a separatory funnel, add a little amount
of dithizone-chloroform solution (50 mgll) (dissolve 5 mg of dithizone specified
in JIS K 8490 in 100ml of chloroform with mercury content 0.5pglZ or
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
less), shake it, let it stand and discard chloroform layer. Repeat this op-
eration until chloroform layer gives no discolouring. Filtrate water layer
through dried filter paper to remove small particles of chloroform.
Tin (II) chloride solution Add 60 ml of sulfuric acid (1+20) onto 10 g of
tin (II) chloride dihydrate specified in JIS K 8136, dissolve by stirring it
while heating. After cooling, add water t o make total 100ml. This solu-
tion should contain 1pglZ or leas mercury. When Purification is needed,
pass high purity nitrogen specified in JIS K 1107. The storing time limit
of this solution shall be one week.
Mercury standard solution (0.5 mgHg/ml) Dissolve 0,339 g of mercury
(II) chloride specified in JIS K 8139 in a little of water, transfer this into
a 500 ml volumetric flask, add 5 ml of nitric acid (l+l), and add water up
to the marked line. Store in a borosilicate glass bottle.
Mercury standard solution (10pgHglm1) Pipet 10 ml of mercury stan-
dard solution (0.5 mgHg/ml) into a 500 ml volumetric flask, add 5 ml of nitric
acid (l+l), and add water up to the marked line. Store in a borosilicate
glass bottle. The storing time limit of this solution shall be one month.
366
K O101 : 1998
(i) Mercury standard solution (0.1 pgHg/ml) Pipet 10 ml of mercury stan-

dard solution (10 pgHglml) into a 1O00 ml volumetric flask, add 10 ml of
nitric acid (l+l), and add water up t o the marked line. Prepare each time
it is needed.
Notes (1) Use such reagent as for atomic absorption analysis which con-
tains little mercury.
(2) Ammonium peroxodisulfate specified in JIS K 8252 is also avail-
able. In both cases, mercury in solution should be 1 pglZ or less.
(a) Atomic absorption analyzer or atomic absorption analyzer for mer-
cury
(b) Apparatus for reduction-vaporizationof mercury(3) Use together with
atomic absorption analyzer.
(c) Mercury hollow cathode lamp or mercury lamp
Note (3) This apparatus consists of a vessel for reduction, absorption cell,
air pump, flow meter, drying tube, and connecting pipe. Fig. 57.1
and Fig. 57.2 exemplify its construction.
The detail of each construction unit shall be as follows.
Vessel for reduction A glass bottle (or Erlenmeyer flask) with
300 t o 350 ml capacity (Mark at the level of 250 mi.), or separatory
funnel type vessel (300 mi) as shown in Fig. 57.3.
Absorption cell 100 to 300 mm long, made of quartz glass, glass,
o r plastic (absorbing no mercury vapor), of which both ends have
a quartz glass window.
Air pump Diaphragm pump capable of sending gas with 0.5 t o
3 Zlmin, o r the air pump with equivalent performance. If the part
exposed t o mercury vapor is metal, it should be covered with such
as collodion.
Flow meter Capable of measuring flow rate of 0.5 to 5 Zlmin.
Drying tube A drying tube or U-type tube. Either it is packed
with drying agent such as magnesium perchlorate (for drying) speci-
fied in JIS K 8228 or calcium chloride (for drying) specified in
JIS K 8124, o r a cold-trap can be effective, If keep the tempera-
ture inside of an absorption cell about 10 "C higher than surround-
ings by means of as lighting small lamp in the cell, the drying
tube is needless.
Connecting pipe Soft vinyl chloride pipe
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
367
K 0101 : 1998
c A: Vessel for reduction

B: Drying t u b e
C: Flow m e t e r
D: Absorption cell
E: Air p u m p
F: Recorder
G: Mercury hollow cathode l a m p
(mercury lamp)
H: Detector for atomic absorption
I: Mercury eliminating device
Fig. 57.1 Example of construction
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
for closed circulating type
A: Vessel for reduction

B: Drying t u b e
C: Flow m e t e r
D: Absorption cell
E: Air p u m p
F: Recorder
G: Mercury hollow cathode
l a m p (mercury l a m p )
H: Detector for atomic
absorption
I: Mercury eliminating
device
Fig. 67.2 Example of construction

for open aerating type
368
K O101 : 1998
' Unit : mrn
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
i
Bulb diameter 010,diameter
of small hole 01 to 1 5 x 6 holes
One-way cock K-17
Fig. 57.3 Example of vessel for reduction

Take a suitable amount (containing 0.1 to 2 pg as Hg) of the sample(4) in
a 300 ml Erlenmeyer flask(5), and add water t o make about 150 ml.
Add 20 ml of sulfuric acid (l+l),
5 ml of nitric acid, and 20 ml of potassium
permanganate solution (50 g/Z)(6), shake them, and let it stand for about
15 min.
When red colour in the solution disappears, add little by little potassium
permanganate solution (50 gll) until red colour lasts about for 15 min.
Add 10 ml of potassium peroxodisulfate solution (50 gll), and heat it for about
2 h by immersing the 300 ml Erlenmeyer flask in a water bath kept a t about
95 "C.
Cool it to room temperature, and add 10ml of hydroxylammonium chlo-
ride solution (80 gll) t o reduce excess permanganate.
Immediately transfer the solution in a vessel for reduction(7), and after water
is added up to make 250m1, assemble a circulating network for aeration.
Add 10 ml of tin (II) chloride solution quickly, and run an air pump with
the optimal flow rate(8), which has been beforehand set up, in order t o cir-
culate air (9).
369
K O101 : 1998
Read indicated value (10) at 253.7 nm wavelength.

Turn the bypass cock(11), and continue aeration until the indicated value
returns original point.
Take water the same amount as that of the sample for a blank test, add
reagents the same amount as that added in the operations (b)to (d),carry
out the operations in (e)t o (i), read indicated value, and correct the indi-
cated value obtained on the sample.
Find the quantity of mercury on the working curve, and calculate the con-
centration (pgHglZ) of mercury in the sample.
Working curve Pipet step by step 1 t o 20 ml of mercury standard solu-
tion (0.1 ygHglml) into as many 300 ml vessels for reduction, respectively
add 20 ml of sulfuric acid (l+l) and water to make total 200 ml, and carry
out the operations in (f)t o (i). Separately, take 200 ml of water in a 300 ml
vessel for reduction for a blank test, add 20 ml of sulfuric acid (l+l), carry
out the operations (f) t o (i), correct the indicated values obtained on mer-
cury standard solution, and draw the relation curve between the quanti-
ties of mercury (Hg) and indicated values. Prepare the working curve when
sample is measured.
In case of a sample containing no organic substance or other dis-
turbances, the following is allowable: omit the operations in (a)
to (e),take a sample directly into a vessel for reduction, add 20 ml
of sulfuric acid (l+l), and carry out the operations (f) to (k). In
this case, prepare the working curve similarly to this.
Instead, an Erlenmeyer flask or glass bottle for reduction can
be used.
In case of a sample with less organic substance, this can be suitably
lessened.
When a vessel for reduction is used for decomposition, connect
it directly.
Because the optimal flow rate depends on the apparatus employed,
the optimal condition has been found in advance,
In case of open aerating type, attach a cock to air line of a re-
duction vessel, after adding tin (II) chloride solution, agitate
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
violently for about 2 min, connect to the apparatus, and open the
cock at the same time running the pump. The optimal aerating
rate should be previously found, but it usually is 1 to 1.5 Zlmin.
Absorbance or its proportional value shall be valid. In case of
open aerating type, measure peak height or peak area.
This should be exhausted in air passing through a gas washing
bottle with sulfuric acid (1+4) dissolving potassium permangan-
ate solution (50 g/Z).
1 If a sample containing a lot of chloride ion is handled, potas-
sium permanganate oxidizes chloride ion t o produce chlorine,
and this results in absorbing the light of 253.7 nm wavelength
and finally gives a positive error. In this case, add excessively
370
K O101 : 1998
hydroxylammonium chloride solution (80 gll) in order to suf-

ficiently reduce chlorine. Chlorine which exists in a vessel
for reduction should be expelled owing t o sending of such as
nitrogen gas.
2 Such as benzene or acetone gives a positive error by absorb-
ing the light of 253.7 nm wavelength. When the sample con-
tains this kind of volatile organic substance, any one of the
following procedures shall be adopted after pretreatment by
potassium permanganate.
(1) Mix by shaking with a little of hexane t o eliminate volatile
organic substance by extraction. (2)Correct background us-
ing such as a deuterium lamp. (3) Obtain the difference of
indicated values using a mercury hollow cathode lamp and
deuterium lamp, then carry out the same measurement with-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
out addition of tin (II) chloride solution, and determine mer-
cury as the difference of both indicated values. (4) Extract
and separate mercury as dithizone complex, and measure it
by heating vaporization method.
57.2 Atomic absorption spectrometry by heatingvaporization After a sample

is pretreated with potassium permanganate, extract mercury into organic solvent as
dithizone complex from sulfuric-acidic solution. Vaporize organic solvent, heat resi-
due t o generate mercury vapor, and measure its atomic absorption a t 253.7nm
wavelength t o determine mercury.
Determination range: Hg 0.5 to 10pglZ
Nitric acid Follow 57.1 (1) (a).
Sulfuric acid (l+l)Follow 57.1 (1)(b).
Potassium permanganate solution (50 g/Z) Follow 57.1 (1) (c).
Potassium peroxodisulfate solution (50 gll) Follow 57.1 (1) (d).
Hydroxylammonium chloride solution (80glZ) Follow 57.1 (1) (e).
Dithizone-chloroform (0.1gll) Dissolve 10 mg of dithizone specified in
JIS K 8490 in 100ml of chloroform specified in JIS K 8322. However,
the mercury content in the solution shall be 1pglZ or less.
2,3-dimercapto-l-propanolsolution (0.1 vol %) Mix 0.1 ml of 2,3-
dimercapto-l-propanol with 10 ml of chloroform specified in JIS K 8322.
Dilute 10 times with chloroform when it is needed.
Mercury standard solution (0.1 pgHg/ml) Follow 57.1 (1)(i).
37 1
K 0101 : 1998
(b) Atomic absorption analyzer or atomic absorption analyzer for mer-

cury
(c) Apparatus for heatingvaporization of mercury Use jointly with atomic
absorption analyzer. Fig. 57.4 shows the example of construction of an ap-
paratus for heating-vaporization.
(d) Mercury hollow cathode lamp or mercury lamp
A: Heating tube
B: Drying tube
C: Flow meter
D: Absorption cell
E: Pump
F: Burner
G: Mercury hollow cathode
lamp (mercury lamp)
I n II
H: Detector
I: Mercury eliminating device
J: Recorder
K: Porcelain boat
Fig. 57.4 Example of construction of apparatus
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
for heatingvaporization
Take a suitable amount (containing 0.1 t o 2 p g as Hg) of sample(4) in a
300 ml Erlenmeyer flask, and add water to make total about 150 ml.
Carry out the operations in 57.1 (3) (b)t o (e).
Transfer this solution into a separatory funnel, add 5 ml of dithizone-chlo-
roform solution (0.1 g/Z), agitate violently for about 2 min, and let it stand.
Separate chloroform layer into a test tube.
Add again 5 ml of dithizone-chloroform solution (0.1gll) into water layer,
and agitate violently for about 2 min, followed by standing.
Put this chloroform layer into the above test tube, add 5 ml of water, shake
it, and let it stand.
Remove water layer with a fountain pen filler.
Put a part (2.5 ml or less) of this chloroform solution in a porcelain boat.
372
K O 1 0 1 : 1998
solution (0.1 vol %), and aerate inside

Add 0.1 ml of 2,3-dimercapto-l-propanol
to volatilize chloroform.
Insert a porcelain boat in a heating tube of a mercury heating-vaporizer,
and start simultaneously suction (12) and heating.
Read indicated value (10) a t 253.7 nm wavelength.
Take the same amount of water as that of the sample for a blank test, add
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
the same amount of reagents as added in (b),carry out the operations in
( c ) t o (k), and correct the indicated value obtained on the sample.
Find the quantity of mercury on the working curve, and calculate the con-
centration (pgHgl2) of mercury in the sample.
Working curve Pipet step by step 1 t o 20 ml of mercury standard solu-
tion (0.1 pgHglm1) into as many separatory funnels, respectively add 20 ml
of sulfuric acid (l+l), add water t o make total 200m1, and carry out the
operations in ( c ) to (k). Separately take 200 ml of water and 20 ml of sul-
furic acid (l+l) in a separatory funnel for a blank test, carry out the op-
erations in ( c ) t o (k),correct the indicated values on mercury standard
solution, and draw the relation curve between the quantities of mercury
(Hg) and indicated values. Prepare the working curve when the sample is
measured.
Notes (12) The optimal suction amount must be obtained beforehand. It is
generally 1 t o 1.5Zlmin.
373
K 0101 : 1998
58 Manganese (Mn) F o r the determination of manganese, periodic acid absorp-

tiometry, flame atomic absorption method, electric heating atomic absorption method,
ICP atomic emission spectrometry or ICP mass spectrometry shall be applied.
58.1 Periodic acid absorptiometry After a sample is acidified with sulfuric acid,
add potassium periodate, heat it t o produce reddish violet permanganate ion, and
measure its absorbance to determine manganese.
Determination range: Mn 40 t o 500pg
Sulfuric acid (l+l)Follow 4.4 (1) (b).
Phosphoric acid Specified in JIS K 9005.
Potassium periodate Specified in JIS K 8249.
Manganese standard solution (0.1 mgMn/ml) Dissolve 0.288 g of po-
tassium permanganate specified in JIS K 8247 in the mixture of 150 ml of
water and 10 ml of sulfuric acid (l+l).Drip sodium hydrogensulfite solu-
tion (100 g/Z) (dissolve 10 g of sodium hydrogensulfite specified in JIS K
8059 in water t o make 100 ml) to decolour it by stirring, and boil it to expel
excess sulfur dioxide. After cooling, transfer into a 1O00 ml volumetric
flask, and add water up to the marked line. Otherwise, take 0.1OOg of
manganese (99.9 % o r more), dissolve it in 20 ml of sulfuric acid (1+3)with
heating, let it cool, transfer it into a 1000ml volumetric flask, and add
water up t o the marked line. Otherwise, use reference material-standard
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
solution, manganese, Mn 100, specified in JIS K 0027.
Manganese standard solution (20 pgMn/ml) Pipet 50 ml manganese
standard solution (0.1 mgMn/ml) into a 250 ml volumetric flask, and add
water up t o the marked line.
(a) Photometer Spectrophotometer o r photoelectric photometer
Take a suitable amount(2) (containing 40 t o 500 pg as Mn) of the sample
which has been treated(1) in 4, add 10 ml of sulfuric acid (1+1)(3),heat it
t o make white fume of sulfuric acid, and remove halides.
After cooling it, add about 20 ml of water and 1 ml of phosphoric acid, and
heat it to dissolve the content. If undissolved is found, filtrate it, wash
both the filter paper and precipitate with warmed water, put the filtrate
and washings together, and add water to make total about 45 ml.
Add 0.5 g(4) of potassium periodate, and heat it in boiling water bath for
30min(5) to make it colour.
Cool it in running water, transfer in a 50 ml volumetric flask, and add water
374
K O101 : 1998
(e) Place a part of this solution into a n absorption cell, and measure its absor-
bance in the vicinity of 525 nm or 545 nm wavelength.
(f) Take about 30 ml of water for a blank test, add 10 ml of sulfuric acid (l+l)
and 1 m l of phosphoric acid, carry out the operations in (c) t o (e), read
absorbance, and correct the absorbance obtained on the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(g) Find the quantity of manganese on the working curve, and calculate the
concentration (pgMn/Z) of manganese in the sample.
Working curve Pipet step by step 2 t o 25 ml of manganese standard so-
lution (20 pgMn/ml) into as many 100 ml beakers, respectively add water
t o make each total about 30 ml, add 10 ml of sulfuric acid (l+l) and 1ml
of phosphoric acid, carry out the operations in (c) to (f), and draw the re-
lation curve between the quantities of manganese (Mn) and absorbances.
Notes (1) When sample contains no organic substances and disturbances
affecting results, this pretreatment may be omitted.
(2) This should be at most 500ml.
(3) When sulfuric acid is used in pretreatment, don’t add sulfuric
acid, Provided that sulfuric acid should be controlled t o be about
5 ml.
(4) Instead of adding potassium periodate, the followings are permis-
sible: add 2 ml of silver nitrate solution (5 g/E) (dissolve 0.5 g of
silver nitrate specified in JIS K 8550 in water to make 100 mi)
and 5 ml of ammonium peroxodisulfate solution (200 g/Z) (dissolve
20 g of ammonium peroxodisulfate specified in JIS K 8252 in water
to make 100 mi), and boil for about 1min to colour. The concen-
tration of sulfuric acid for colouring shall be about 0.5 mol/Z. If
this way gives precipitate of silver chloride, drip nitric-acidified
mercury (II) nitrate solution (50 g/Z) (dissolve 5 g of mercury (II)
nitrate n-hydrate specified in JIS K 8558 in 20 ml of nitric acid
(1+2) and make it 100 ml with water) until the precipitate disap-
pears and add several more drops of the solution to fix chloride
ion.
(5) Too long a time for heating may decompose the generated per-
manganate ion, and it is important to keep accurate heating time.
Remarks 1 When determining dissolved manganese, use a suitable amount
(containing 40 t o 500 pg as Mn) of the sample filtrated in 3.2
(use filter paper 5 grade C) and carry out the operation in (3)(a)
and after.
2 In case of low concentrated manganese, it can be determined
after being concentrated by iron-coprecipitation method a s
follows.
Take suitable amount up t o 500ml of sample and heat to
about 90 OC, add 5 ml of ammonium iron (III) sulfate solution
(2 mgFe/ml) [dissolve 1.8 g of ammonium iron (III) sulfate 12
hydrate specified in JIS K 8982 in 10 ml of nitric acid (1+6)
and water to make total 100 mi], add 5 to 10 ml of hydrogen
375
K O101 : 1998
peroxide specified in JIS K 8230,add, while agitating, aque-

ous ammonia ( l + l )or sodium hydroxide solution (1OOglZ) t o
generate the precipitate of iron (III) hydroxide. After settling
the precipitate, filtrate it through filter paper 5 grade A, and
wash with warm water. Move the precipitate as completely
as possible t o the original beaker, dissolve the precipitate at-
tached on the filter paper with 50 ml of sulfuric acid (1+9)con-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
taining a little of hydrogen peroxide solution (1+10), and wash

the filter paper with water. Put the filtrate and washings into
the original beaker, heat it t o dissolve the precipitate and
simultaneously decompose hydrogen peroxide, and concentrate
the solution up t o about 40 ml. After adding 1 ml of phospho-
ric acid, carry out the operations in (3)( c ) t o (g) t o determine
manganese.
58.2 Flame atomic absorption method After sample is pretreated, spray it into
acetylene-air flame, and measure atomic absorption by manganese at 279.5 nm
wavelength t o determine manganese.
Determination range: Mn 0.1 to 4mglZ
(a) Manganese standard solution (10 pgMn/ml) Pipet 50 ml of manganese
standard solution (0.1 mgMním1) stated in 58.1 (1)(d)into a 500 ml volumetric
flask, add 10 ml of nitric acid (l+l), and add water up to the marked line.

(b) Manganese hollow cathode lamp
Remarks 3 When dissolved manganese is determined, take a suitable
amount of sample filtrated in 3.2 (use filter paper 5 grade C)
and treat according t o 4.5.
4 When concentration of manganese is poor, treat it according to
Remarks 2, and separate and concentrate manganese. Dissolve
precipitate in a little amount of hydrochloric acid (1+2) con-
taining a little hydrogen peroxide (1+10), and wash filter pa-
per with warm water. Put together filtrate and washings, and
make it a definite volume with 0.1 to 1 mol/Z acidic solution of
hydrochloric acid. Otherwise, treat it according to Remarks 5
in 51. In this case, extract it at pH 4.5 to 5.0.
Because 1-pyrrolidinecarbodithioacid complex of manganese
(APDC complex) is easily transferred to water layer, carry out
extraction and separation swiftly.
376
K O101 : 1998

Spray the sample which has been preparatorily treated in (3)into a flame
according t o 6 of JIS K 0121, and read indicated value(6) a t 279.5nm
wavelength.
Take water as the same amount as that of sample a t pretreatment in (3)
for a blank test, carry out the operations in (3) and (4)(a)similarly to sample
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
side, correct the indicated value obtained on the sample.

Find the quantity of manganese on the working curve, and calculate the
concentration (mgMníZ) of manganese in the sample.
Working curve Pipet step by step 1 t o 40 ml of manganese standard so-
lution (10 pgMníml) into as many 100 ml volumetric flasks, respectively add
acid to make the same acid concentration as that of sample treated in (3) (a),
and add water up t o the marked line. Carry out the operation in (a)on
this solution. Separately, take water for a blank test, add acid t o make
the same acid concentration as that of the sample treated in (3)(a),carry
out the operation in (a), correct the indicated value obtained on the stan-
dard solution, and draw the relation curve between the quantities of man-
ganese (Mn) and indicated values. Prepare the working curve when the
sample is measured.
Note (6) Absorbance o r its proportional value is valid.
Remarks 5 In case of a lot of silica, it is recommendable t o add calcium
(or magnesium) by 200 mglZ as an interference inhibitor.

atomize it in an electric furnace, and measure atomic absorption by manganese at
279.5 nm wavelength t o determine manganese.
Determination range: Mn 1 t o 30pg/Z
Remarks 6 This method is easily affected by the kind and concentration of
coexisting acid and salt, therefore applicable to the sample which
is less affected.
element t o be determined, and verify that there is no interference t o use.
(b) Nitric acid (l+l) Highly purified nitric acid specified in JIS K 9901.
(c) Manganese standard solution (1pgMdml) Pipet 10 ml of manganese
standard solution (10 pgMním1) stated in 58.2 ( i )(a) into a 100 ml volumet-
ric flask,add 2 ml of nitric acid (l+l),

377
K O101 : 1998
(b) Exothermic body Made of graphite o r heat resisting metal.

(c) Manganese hollow cathode lamp
(e) Micropipet Piston operated micro-volumetric apparatus, 10 to 50 pl, speci-
fied in JIS K 0970. Or an automatic injection device.
Remarks 7 When dissolved manganese is determined, follow Remarks 3.

Inject a definite amount (for instance, 10 to 50 pl) of the sample which has
been preparatorily treated in (3) into an exothermic body using a micropi-
pet, hereafter according t o 6 of JIS K 0121, dry it (100 to 120 "C for 30 to
40 s), ash it (500 t o 800 "C about for 30 s), atomize i t ( 7 ) (2 O00 t o 2 700 "C
for 4 to 6 s), and read indicated value(6) at 279.5 nm wavelength(8).
Take water the same volume as that of the sample at preparatory treat-
ment in (3)for a blank test, carry out the operations in (3)and (4) (a)similarly
t o sample side, and correct the indicated value obtained on the sample.
Find the quantity of manganese on the working curve, and calculate the
concentration (pgMníE) of manganese in the sample.
Working curve Pipet step by step 0.1 t o 3 ml of manganese standard so-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
lution (1 pgMn/ml) into as many 100 ml volumetric flasks, respectively add
acid t o make the same acid concentration as that of the sample which has
been pretreated in (3) (a),and add water up to the marked line. Carry out
the operation in (a) on these solutions. Separately, take water for a blank
test, add acid t o make the same acid concentration as that of the sample
treated in (3)(a),carry out the operation in (a), correct indicated value
the quantities of manganese (Mn) and indicated values. Prepare the working
curve when a sample is measured.
Notes (7) The condition for drying, ashing, or atomizing varies depending
upon apparatus. They may be also affected by such as injected
(8) Repeat successively at least 3 times the operation in (a),and con-
firm the indicated values agree.
58.4 ICP atomic emission spectrometry After pretreatment of a sample, spray

it in an inductively coupled plasma through the sample introducing part, measure
the emission by manganese at 257.610 nm wavelength t o determine manganese.
Determination range: Mn 10 t o 200 pglZ, 0.2 t o 5 mglZ
378
K O101 : 1998

(a) Manganese standard solution (10 pgMdm1) Follow 58.2 (1) (a).
(b) Mixed standard solution [(lopgCu, 10 pgzn, 8 pgCd, 10 pgNi, 10 pgPb,
10 pgMn, 10 pgFe)/ml] Follow 51.4 (1)(b).
Remarks 8 When dissolved manganese is determined, follow Remarks 3.
9 When the sample, which has been preparatorily treated, has
rich concentration of sodium, potassium, magnesium, calcium,
etc., and poor concentration in manganese, the operation in
Remarks 7 in 51 is preferably carried out for determination
of manganese.
its emission strength a t 257.610 nm wavelength (9) (10) (11).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Take water the same amount as that of the sample pretreated in (3)for a
blank test, carry out the operations in (3) and (4)(a)similarly to the sample,
and correct the emission strength obtained on the sample.
(c) Find the quantity of manganese on the working curve, and calculate the
concentration (pgMn/Z) of manganese in the sample.
Working curve Pipet step by step 0.1 t o 2 ml (or 2 t o 50 ml)(l2)(13) of
manganese standard solution (10 pgMn/ml) into as many 100 ml volumet-
ric flasks, respectively add acid to make the same acid concentration as
that of the sample pretreated in (3)(a),and add water up to the marked
line. Carry out the operations in (a) on these solutions. Separately, take
water for a blank test, add acid t o make the same acid concentration as
that of the sample pretreated in (3) (a), carry out the operation in (a),cor-
rect the emission strength obtained on the standard solution, and draw the
relation curve between the quantities of manganese (Mn) and emission
strengths. Prepare the working curve when the sample is measured.
spectrums with different wavelength is used, an internal standard
method can be applicable. When the internal standard method is
applied the procedures are as follows: Take a suitable amount of
the sample, which has been treated in (3)(a),into a 100 ml volu-
metric flask, add 10 ml of yttrium solution (50 pgY/ml) [Follow Note
(8) of 451, add acid t o make the same acid concentration as the
sample in (4) (a),and add water up t o the marked line. Carry out
the spraying operations in (4)(a) on this solution, measure
emission strength at both 257.610 nm and 371.029 nm (yttrium)
379
K O101 : 1998
wavelength, and obtain the emission-strength ratio of manganese

and yttrium.
Separately, pipet step by step 0.1 t o 2 ml (or 2 t o 50 mi) of
manganese standard solution (10 pgMn/ml) into as many 100 ml
volumetric flasks, respectively add 10 ml of yttrium solution
(50 pgY/ml), add acid to make the same acid concentration as the
sample of (4) (a),and add water up t o the marked line. Carry
out the spraying operation in (4) (a) on these solutions, measure
emission strengths at both 257.610 nm and 371.029 nm wave-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
length, draw the relation curve between emission-strength ratio
of manganese t o yttrium and the concentration of manganese,
and make it the working curve. On this working curve, find the
quantity of manganese corresponding t o the emission-strength
ratio obtained on the sample, and calculate the concentration
(pgMn/Z) of manganese in the sample.
high concentration of salts in a sample, the standard addition
necessary whatever sample may be used.
In case of the apparatus capable of using high-order spectrum
has been confirmed.
When, after making preparatory operations according to Remarks
prepared as follows: dilute manganese standard solution (10 pgMd
mi) to suitable concentration (0.1 to 1ygMn/ml), take step by step
0.2 to 4 ml (or 4 t o 100 mi), make them 500 ml (or definite amount
of 100 t o 500 ml), carry out the operations in Remarks 9 and (4)(a)
and (b)similarly t o sample side, and draw the relation curve
between the quantities of manganese (Mn) and emission strengths.
When copper, zinc, cadmium, nickel, lead, and iron are simulta-
neously tested, use mixed standard solution [(lo pgCu, 10 pgZn,
preferably each working curve under the test condition of each
metal element.
part, measure the ionic current in each number of masses/electric charges of man-
ganese and internal standard substance, and find the ratio between ionic current of
manganese and that of internal standard substance to determine manganese.
Determination range: Mn 0.5 to 25 pg/Z, 10 to 500 pg/Z
380
K O101 : 1998

Water Follow 58.3 (1)(a).
Nitric acid (l+l)Prepare using highly purified nitric acid specified in
JIS K 9901.
Yttrium solution (1 ygY/ml)(14) Follow 51.5 (1)(c).
Manganese standard solution (1 pgMn/ml) Follow 58.3 (1)( e ) .
Manganese standard solution (50 ngMn/ml) Take 50 ml of manganese
standard solution (1ygMn/ml) in a 1O00 ml volumetric flask, add 1.5 ml of
nitric acid (1+1) and add water up to the marked line. Prepare when it is used.
Mixed standard solution [(ipgCu, 1 pgZn, 1 ygCd, 1 ygPb, 1 pgMn,
Mixed standard solution [(50ngCu, 50 ngZn, 50 ngCd, 50 ngPb,
50 ngMn, 50 ngCr)/ml] Follow 51.5 (1)(g).
(3) Preparatory operation Preparatory operations shall be as follows (15).
(b) Take a suitable amount (containing 0.05 t o 50 yg as Mn) of sample treated
in (3) (a) in a 100 ml volumetric flask, add 1ml of yttrium solution (1pgY/
mi), add nitric acid (l+l) t o make final concentration of nitric acid 0.1 to
0.5 molll and add water up to the marked line.
Remarks 13 When dissolved manganese is determined, treat it according t o Re-
marks 3, and carry out the operation in (b).
(4) Operation Operation shall be carried out as follows(l6).

(a) Make the ICP mass spectrograph ready t o run, spray the solution in (3)(b)
manganese and yttrium, and obtain the ratio between the indicated value
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
of manganese and that of yttrium.

(b) Take the same amount of water for a blank test as that of the sample treated
in (3) (a),carry out the operations in (3) and (4) (a) similarly to the sample,
obtain the ratio between the indicated value of manganese and that of yttrium,
and correct the indicated values between manganese and yttrium obtained
on the sample.
381
K O 1 0 1 : 1998
(c) Find the amount of manganese on a working curve, and calculate the con-
centration (PgMníZ) of manganese in the sample.
Working curve Pipet step by step 1 to 50 ml of the manganese standard
solution (50 ngMn/ml o r 1pgMn/ml)(lg) in as many 100 ml volumetric flasks,
add 1ml of yttrium solution (1pgY/ml), add nitric acid ( l + l ) to make the
same acid concentration as the sample carried out the operation in (3)(b),
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
and add water up to the marked line. Carry out the operation in (a)on
this solution. Separately, put 1ml of yttrium solution (1pgY/ml>as a blank
test in a 100 ml volumetric flask, add nitric acid ( l + l ) to make the same
acid concentration as the sample of (3)(b),and add water up to the marked
line. Carry out the operation in (a),correct the indicated values obtained
the indicated value to the amount of manganese (Mn) and the indicated
value of yttrium. Prepare the working curve when the sample is measured.
382
K O101 : 1998
59 Aluminum (Al) For the determination of aluminum, quinolinol absorptiometry,

flame atomic absorption method, electric heating atomic absorption method, o r ICP
atomic emission spectrometry shall be applied.
59.1 Quinolinol absorptiometry Add hydroxylammonium chloride and 1 , l O -

phenanthroline into a sample, which has been faintly acidified, in order to mask iron,
add 8-quinolinol and ammonium acetate, and extract produced complex with chloro-
form. Wash it with ammonium chloride solution containing potassium cyanide to
remove the copper, nickel, cobalt, and so on extracted together with aluminum, and
then measure absorbance by aluminum complex to determine aluminum.
Determination range: Al 5 to 50pg

K 8180.
Aqueous ammonia (1+2) Prepare using aqueous ammonia specified in
JIS K 8085.
Hydroxylammonium chloride solution (100 g l l ) Follow 40.2 (1) (e).
Ammonium acetate solution (150 gll) Dissolve 15 g of ammonium ac-
etate specified in JIS K 8359 in water to make total 100 ml. Transfer this
solution into a separatory funnel, add 5 ml of 8-quinolinol chloroform solu-
tion (dissolve 2 g of 8-quinolinol specified in JIS K 8775 in 100 ml of chlo-
roform specified in JIS K 8322), shake them violently, let it stand, then
discard chloroform layer. Repeat this operations until chloroform layer gives
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
no colouring. Next add 5 ml of chloroform into water layer, shake violently,
let it stand, and discard chloroform layer, Repeat this operation until water
layer shows no yellow colouring. Filtrate water layer through dried filter
paper to remove very small particles of chloroform in the solution.
Potassium cyanide-ammonium chloride solution Dissolve 1.0 g of po-
tassium cyanide specified in JIS K 8443 in water to make total 500ml.
Dissolve bit by bit ammonium chloride specified in JIS K 8116 into this,
and control its pH to 9.0 t o 9.5. Wash this solution with 8-quinolinol chlo-
roform solution and chloroform similarly to the ammonium acetate solu-
tion mentioned in (e), and purify it.
1,lO-phenanthroline solution (1 g l l ) Dissolve 1.3 g of 1,lO-phenan-
throlinium chloride monohydrate specified in JIS K 8202 in water t o make
total 11. Otherwise, dissolve 1.1g of 1,lO-phenanthroline monohydrate
specified in JIS K 8789 in 100 ml of ethanol (95) specified in JIS K 8102
and add water to make 11.
8-quinolinol solution (10 gll) Add 5 ml of acetic acid specified in JIS K
8355 in 2 g of 8-quinolinol specified in JIS K 8775, and after slightly heating
them to dissolve add water to make total 200ml.
383
K O 1 0 1 : 1998
Aluminum standard solution (0.1 mgAl/ml) Dissolve 1.76 g of alumi-

num potassium sulfate 12 hydrate [bis (sulfuric acid) potassium aluminum-
water (1/12)] specified in JIS K 8255 in 20 ml of hydrochloric acid (l+l),
transfer it into a 1O00 ml volumetric flask, and add water up t o the marked
line. Otherwise, take 0.100 g of aluminum (99.9 % or more) specified in
JIS K 8069, dissolve it in 20 ml of hydrochloric acid (l+l) by heating, let
it cool, transfer it into a 1O00 ml volumetric flask, and add water up to
the marked line.
Aluminum standard solution (1 pgAlíml) Pipet 10 ml aluminum stan-
dard solution (0.1 mgAl/ml) in a 1O00 ml volumetric flask, add 20 ml of hy-
drochloric acid (l+l),and add water up t o the marked line.
Take a suitable amount(2) (containing 5 t o 50 pg as Al) of the sample which
has been treated in 4(1), add 1 ml of hydroxylammonium chloride solution
(100 g/Z) and 5 ml of 1,lO-phenanthroline solution (1g/Z), followed by shak-
ing and control its pH to about 3 . 5 W by dripping aqueous ammonia (1+2).
Add water t o make total about 80 ml, and let it stand for about 15 min.
Add 3 ml of 8-quinolinol solution (10 g/Z) and 10 ml of ammonium acetate
solution (150 g/Z), and control its pH t o 5.2 t o 5.5(4) by dripping of aqueous
ammonia (1+2).
Transfer this solution into a separatory funnel, add water t o make total
about 100 ml, add 10 ml (or 20 ml) of chloroform, shake violently for about
1min, and let i t stand.
Separate chloroform layer into another separatory funnel, add 25 ml of po-
tassium cyanide-ammonium chloride solution, shake them, and let it stand.
Place chloroform layer into a 30 ml test tube with ground stopper, add about
1g of sodium sulfate, and shake slightly to remove moisture.
Place a part of chloroform layer into an absorption cell, and measure ab-
sorbance in the vicinity of 390nm wavelength with making chloroform a
reference solution.
Take about 70 ml of water for a blank test, carry out the operations in (a)to
(g),measure absorbance, and correct the absorbance obtained on the sample.
Find the quantity of aluminum on the working curve, and calculate the
concentration (pgAl/Z) of aluminum in the sample.
Working curve Pipet step by step 5 to 50 ml of aluminum standard so-
lution (1pgAl/ml) into as many separatory funnels, respectively add water
t o make total about 70 ml, hereafter carry out the operations in (a) to (h),
and draw the relation curve between the quantities of aluminum (Al) and
absorbances.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
384
K O101 : 1998
Notes (1) Do not use the method 4.3 mentioned in 4. I n case of the sample
containing little of organic substance, the following is allowable:
add 5 ml of hydrochloric acid per 100 ml of a sample, and heat
gently t o concentrate the liquid t o about one-fifth of its original.
(2) Generally, amount of a sample is 50 t o 100 ml, but it may be at
most 500 ml.
(3> Use bromophenol blue test paper.
(4> Use bromocresol green test paper. When pH does not fall in from
5.2 t o 5.5, adjust it by using hydrochloric acid (1+2) o r aqueous
ammonia (1+2).
Remarks When fluoride ion is contained in a sample, add 36 mg of be-
ryllium sulfate per 0.5 mg of fluoride ion for eliminating the
disturbance.
In case of existing chromium, the extraction at pH 5.2 to 5.5
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
shall be carried out at as low temperature as possible. Cool-

ing with icy water is preferable.
When a lot of manganese is contained, it shall be removed by
washing chloroform solution, which extracts complex, with acetic
acid-ammonium acetate solution of p H 7 or less in which
hydroxylammonium chloride is dissolved.
When titanium, molybdenum, or others is contained, the fol-
lowing should be done: after removing copper, nickel, cobalt
by the operation in (3)(e), wash chloroform layer with solu-
tion prepared by mixing 25 ml of ammonium chloride (50 glZ)
with pH 10, which is alkalized by aqueous ammonia, and 2 ml
of hydrogen peroxide specified in JIS K 8230.
This method allows iron to exist by 0.45 mg.
The simultaneous determination of aluminum and iron shall
be carried out as follows:
Omitting the operation t o add 1ml of hydroxylammonium
chloride solution (100 glZ) and 5 ml of 1,lO-phenanthroline so-
lution (1glZ) mentioned in (3) (a),carry out the operations in
(b) t o ( g ) , and measure the absorbance A in the vicinity of
390 nm and absorbance B in the vicinity of 470 nm wavelength.
Separately, take about 80 ml of water for a blank test, carry
out the operations in (e) t o ( g ) , correct the absorbance A and
absorbance B obtained on the sample, and make respectively
absorbance A’ and absorbance B’.
On the working curve of iron (III) near 470 nm wavelength,
find the quantity of iron (III) corresponding to absorbance B‘,
and calculate the concentration (mgFelZ) of iron.
Apply the quantity of iron (III) corresponding to absorbance
B’ to the working curve of iron (III) near 390 nm wavelength,
and find the absorbance C by iron (III) near 390 nm wavelength.
Subtract absorbance C from absorbance A’, and obtain the ab-
sorbance D by aluminum in the vicinity of 390 nm wavelength.
385
K O101 : 1998
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Using absorbance D ,find the quantity of aluminum on the

working curve of aluminum in the vicinity of 390nm wave-
length, and calculate the concentration (mgAl/Z) of aluminum
in the sample.
Working curve Pipet step by step 5 t o 50ml of aluminum
standard solution (1pgAl/ml) into as many separatory funnels,
respectively add water t o make each about 80m1, carry out
the operations in (c) t o (f), and measure absorbances in the
vicinity on 390 nm wavelength. Separately, pipet step by step
0.5 to 10 ml of iron (III) standard solution (10 pgFe/ml)(*) into
as many separatory funnels, respectively add water t o make
each about 80m1, carry out the operations in (e) to (0, and
measure absorbances in the vicinity of 470nm and 390nm
wavelength. Take about 80 ml of water for a blank test, carry
out the operations in ( c ) to (0, measure absorbances in the
vicinity of 470 nm and 390 nm wavelength, and correct the ab-
sorbances obtained on aluminum standard solution and iron
(III) standard solution. Draw relation curve between the quan-
tities of aluminum (Al) and absorbances near 390nm wave-
length, and relation curve between the quantities of iron (Fe)
and absorbances near 470 nm and near 390 nm wavelength.
Note (*) Iron (III) standard solution (10pgFe/ml)
Dissolve 8.63g of ammonium iron (III) sulfate 12
hydrate [bis (sulfuric acid) iron (III) ammonium-
water (1/12)] specified in JIS K 8982 in 20 ml of
sulfuric acid (l+l)and water, transfer it in a 1O00 ml
volumetric flask, and add water up to the marked
line. Make this solution iron (III) standard solu-
tion (1mgFe/ml), pipet 10 ml of this solution into a
1 O00 ml volumetric flask, add 10 ml of sulfuric acid
(l+l), and add water up t o the marked line.
59.2 Flame atomic absorption method After pretreatment of a sample, spray

it in an acetylene-dinitrogen monoxide flame, and measure atomic absorption by alu-
minum at 309.3 nm wavelength t o determine aluminum.
Determination range: Al 5 t o 100mglZ
(a) Potassium chloride solution (100g/Z) Dissolve 10 g of potassium chlo-
ride specified in JIS K 8121 in water t o make total 100 ml.
(b) Aluminum standard solution (0.5 mgAyml) Dissolve 8.794 g of aluminum
potassium sulfate 12 hydrate [bis (sulfuric acid) potassium aluminum water
(1/12)] specified in JIS K 8255 in 20 ml of hydrochloric acid (l+l), transfer
it into a 1O00 ml volumetric flask, and add water up t o the marked line. Oth-
erwise, dissolve 0.500 g of aluminum (99.9 % or more) specified in JIS K 8069
in 30 ml of hydrochloric acid (l+l) with heating, after cooling, transfer it into
a 1O00 ml volumetric flask, and add water up to the marked line.
386
K O101 : 1998

(b) Aluminum hollow cathode lamp
Take a suitable amount (containing 0.5 t o 10 mg as Al) of the sample which
has been pretreated in (3)into a 100 ml volumetric flask, add 1ml of hy-
drochloric acid, and add water up t o the marked line.
Place 50 ml of this solution in a dried beaker, and add 2 ml of potassium
chloride solution (100 g/O.
Spray the sample solution in (b)into acetylene-dinitrogen monoxide flame ( 5 )
according t o 6 of JIS K 0121,and read the indicated value(6) a t 309.3 nm
wavelength.
Take water the same amount as that of the sample pretreated in (3)for a
blank test, carry out the operations in (3) and (4) (a) t o ( c ) similarly to the
sample, read the indicated value, and correct the indicated value obtained
on the sample.
Find the quantity of aluminum on the working curve, and calculate the
concentration (mgAl/Z) of aluminum in the sample.
Working curve Pipet step by step 1t o 20 ml of aluminum standard solu-
tion (0.5 mgAVm1) into as many 100 ml volumetric flasks, respectively add
acid and hydrochloric acid t o make the same acid concentration as that of
the sample pretreated in (3)(a),and add water up to the marked line. Carry
out the operations in (b) and (c) on these solutions. Separately take water
for a blank test, add acid and hydrochloric acid to make the acid concentra-
tion the same as that of the sample treated in (3)(a),carry out the opera-
tions in (b) and (c), correct the indicated value obtained on the standard
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
solution, and draw the relation curve between the quantities of aluminum
(Al) and indicated values. Prepare the working curve when the sample is
measured.
Notes (5) The flame with much of fuel gives a high sensitivity.

atomize it in an electric furnace, and measure atomic absorption by aluminum at
309.3 nm wavelength t o determine aluminum.
Determination range: Al 20 to 20OpglZ
is less affected.
387
K O101 : 1998

element to be determined and verify that there is no interference t o use.
(b) Nitric acid (l+l) Highly purified nitric acid specified in JIS K 9901.
(c) Aluminum standard solution (1 pgAl/ml) Take 5 ml of aluminum stan-
dard solution ( O . 1 mgAl/ml) mentioned in 59.1 (1)ci) into a 500 ml volumetric
flask, add 10 ml of nitric acid (l+l), and add water up to the marked line.
Prepare when it is used.

(c) Aluminum hollow cathode lamp
K 0970, 10 t o 50 p1. Or automatic injection device.
(a) Inject a definite amount (for instance, 10 to 50 p1) of the sample which has
been pretreated in (3)into an exothermic body using a micropipet, hereaf-
ter according t o 6 of JIS K 0121, dry it (100 t o 120 "C for 30 t o 40 s), ash
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
it (600 to 1O00 "C for 30 to 40 s), atomize i t ( 7 ) (2 200 t o 3 O00 "C for 3 t o
6 s ) , and read the indicated value(6) at 309.2 nm wavelength(*).
(b) Take water the same volume as that of sample a t preparatory treatment
to sample side, and correct the indicated value obtained on the sample.
(c) Find the quantity of aluminum on the working curve, and calculate the
concentration (pgAl/Z) of aluminum in the sample,
Working curve Take step by step 2 t o 20 ml of aluminum standard so-
lution (1 pgAl/ml) into as many 100 ml volumetric flasks, respectively add
acid to make the same acid concentration as that of the sample pretreated
in (3)(a),and add water up to the marked line. Carry out the operation in
(a) on this solution. Separately, take water for a blank test, add acid t o
carry out the operation in (a),correct the indicated value obtained on the
standard solution, and draw the relation curve between the quantities of
aluminum (Al) and indicated values. Prepare the working curve when the
sample is measured.
Notes (7) The conditions for drying, ashing, or atomizing vary depending
upon apparatus. They may be also affected by such as injected
volume of sample and concentration of coexisting salts.
388
K O 1 0 1 : 1998
(8) Repeat successively a t least 3 times the operation in (a), and

59.4 ICP atomic emission spectrometry After the pretreatment of a sample,

spray it into an inductively coupled plasma through the sample introducing part,
and measure the emission by aluminum a t 309.271 nm wavelength t o determine alu-
minum.
Determination range: Al 80 to 4 O00 pg/Z
Repeatability: 2 to 10 % by coefficient of variation (depending o n apparatus and
K 8180.
(b) Aluminum standard solution (20 pgAl./ml) Pipet 10 ml of aluminum stan-
dard solution (0.5mgAVml) mentioned in 59.2(1)(b) into a 250ml volu-
metric flask, add 5 ml of hydrochloric acid (l+l),
and add water up t o the
marked line.
(c) Mixed standard solution [(20 pgCa, 10 pgMg, 20 pgAl)/mll Follow
49.3 (1)(cl.
Remarks 8 When the sample, which has been preparatorily treated, has
a rich concentration of sodium, potassium, calcium and mag-
nesium, but poor concentration in aluminum, the procedure
shall be as follows: Take 100 ml of the sample, carry out the
operation in which chloroform in 59.1 (3)(a) to (e) is replaced
by 3-methyl-1-butanol(isoamyl alcohol) specified in JIS K 8051,
and put the 3-methyl-1-butanol layer in a test tube with a
ground stopper. In this case, the operation to remove mois-
ture by adding sodium sulfate mentioned in 59.1 (3)(f) may
be omitted.
(a) Spray the sample which has been pretreated in (3)into a plasma accord-
ing to 5.8 of JIS K 0116, and measure emission strength at 309.271nm
wavelength(9) (10) (11).
(b) Take the same amount of water as that of the sample a t pretreatment in
(3)for a blank test, carry out the operations in (3) and (4)(a) similarly t o
sample side, and correct the emission strength on the sample.
(c) Find the quantity of aluminum on the working curve, and calculate the
concentration (pgAl/Z) of aluminum in the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
389
K O101 : 1998
Working curve Pipet step by step 0.4 t o 20 ml(12) (13) of aluminum stan-
dard solution (20 pgAl/ml) into as many 100 ml volumetric flasks, respec-
tively add acid t o make the same acid concentration as that of the sample
which has been pretreated in (3) (a), and add water up t o the marked line.
Carry out the operation in (a)on this solution. Separately, take water for
a blank test, add acid t o make the same acid concentration as that of the
sample pretreated in (3)(a),carry out the operations in (a), correct the
emission strength obtained on the standard solution, and draw the rela-
tion curve between the quantities of aluminum (Al) and emission strength.
When the apparatus capable of simultaneously measuring two
spectrums with different wavelength is used, an internal stan-
method is applied the procedures are as follows: Take a suitable
amount of the sample, which has been treated in (3)(a), into a
ml) [Follow Note (8) of 451, add acid t o make the same acid con-
centration as the sample in (4) (a),and add water up to the marked
line. Carry out the spraying operations in (4)(a) on this solu-
tion, measure emission strength a t both 309.271nm and
371.029 nm (yttrium) wavelength, and obtain the emission-strength
ratio of aluminum and yttrium.
Separately, pipet step by step 0.4 to 20 ml of aluminum stan-
dard solution (20 pgAl/ml) into as many 100 ml volumetric flasks,
respectively add 10 ml of yttrium solution (50 pgY/ml), add acid
t o make the same acid concentration as the sample of (4) (a),and
tion (4)(a)on these solutions, measure emission strengths at both
309.271 nm and 371.029 nm wavelength, draw the relation curve
between emission-strength ratio of aluminum to yttrium and the
concentration of aluminum, and make it the working curve. On
this working curve, find the quantity of aluminum corresponding
to the emission-strength ratio obtained on the sample, and cal-
culate the concentration (pgAl/l) of aluminum in the sample.
whatever sample may be used.
When, after making preparatory operations according t o Remarks
8, 3-methyl-1-butanollayer is directly sprayed, the working curve
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
shall be prepared as follows: dilute aluminum standard solution

(20 pgAl/ml) t o suitable concentration (1 t o 4 pgAl/ml), take step
by step 1 to 5 ml of the solution, make them 100 ml with water,
carry out the operations in Remarks 8 and (4) (a)and (b)similarly
390
K 0101 : 1998
to the sample, and draw the relation curve between the quanti-
ties of aluminum (Al) and emission strengths.
(13) When calcium and magnesium are simultaneously tested, use
mixed standard solution [(20ygCa, 10 ygMg, 20 pgAl)/ml], and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
391
K O101 : 1998
60 Iron (Fe) For the determination of iron, phenanthroline absorptiometry, flame

atomic absorption method, electric heating atomic absorption method, or ICP atomic
emission spectrometry shall be applied.
60.1 Phenanthroline absorptiometry After adding hydroxylammonium chloride

and 1,lO-phenanthrolineinto a faintly acidic solution, adjust its pH to be 4 t o 5 using
ammonium acetate, produce reddish orange iron (II) complex, and measure its ab-
sorbance t o determine iron.
Determination range: Fe 20 to 500pg
Hydrochloric acid (l+l) Prepare using hydrochloric acid specified in JIS
K 8180.
Nitric acid (l+l) Prepare using nitric acid specified in JIS K 8541.
Aqueous ammonia ( l + l ) Prepare using aqueous ammonia specified in
JIS K 8085.
Hydroxylammonium chloride solution (100 glZ) Follow 40.2 (i)( c ) .
1,lO-phenanthroline solution ( i g/Z) Follow 59.1 (i)(g).
Ammonium acetate solution (500 g/Z) Dissolve 500 g of ammonium ac-
etate specified in JIS K 8359 in water t o make total 1 2 .
Iron standard solution (i mgFe/ml) Put 1.000 g of iron (99.5 % or more)
heat it t o dissolve, let it cool, transfer it
in 30 ml of hydrochloric acid (l+l),
into a 1O00 ml volumetric flask, and add water up t o the marked line.
Otherwise, dissolve 7.02 g of ammonium iron (II) sulfate hexahydrate [bis
(sulfuric acid) iron (II) ammonium hexahydrate] specified in JIS K 8979 in
20 ml of hydrochloric acid (l+i) and a suitable volume of water, transfer it
into a 1O00 ml volumetric flask, and add water up to the marked line. Or,
use iron standard solution, Fe 1000, specified in JIS K 0016.
Iron standard solution (10 ygFe/ml) Pipet 10 ml of iron standard so-
lution (imgFe/ml) in a l O00 ml volumetric flask, add 20 ml of hydrochlo-
ric acid (l+l),
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Take a suitable amount (containing 20 to 500 pg as Fe) of the sample which
has been treated in 4(1>(2) in a beaker, add 1 t o 2 ml of nitric acid (l+l),
and boil it.
(b) After water is added t o make total 50 to 100m1, add aqueous ammonia
(1+1)t o make faint alkalinity. Boil this solution for several minutes t o
produce precipitate(3) (4), and then let it stand for a while.
392
K O101 : 1998
After settling down of precipitate, filtrate through filter paper 5 grade A,

and wash with warm water several times. Put the precipitate into the original
beaker with washing, add 4 m l of hydrochloric acid (l+l), heat them t o
dissolve, filtrate through the original filter paper, dissolve simultaneously
the iron (III) hydroxide clinging to the paper. Wash the filter paper with
warm water several times.
Put together filtrate and washings, add water t o make total about 70m1,
add 1 ml(5) of hydroxyl ammonium chloride solution (100 glZ) and shake them.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Add 5 ml of 1,lO-phenanthroline solution (igll), shake them, add succes-

sively 10 ml(6) of ammonium acetate solution (500 g/Z), mix by shaking again,
and let it cool.
Transfer it into a 100 ml volumetric flask, add water up t o the marked line,
and let it stand for about 20 min.
Place a part of this solution into an absorption cell, and measure its absor-
bance in the vicinity of 510nm wavelength.
Take about 50 ml of water for a blank test, add 1to 2 ml of nitric acid (l+l),
carry out the operations in (b)to (g), measure its absorbance, and correct
Find the quantity of iron on the working curve, and calculate the concen-
tration of iron (mgFelZ) in the sample.
Working curve Pipet step by step 2 t o 50ml of iron standard solution
(10 pgFe/ml) into as many 100 ml volumetric flasks, respectively add 4 ml
carry out the operations in (d)t o (h),and draw
of hydrochloric acid (l+l),
the relation curve between the quantities of iron (Fe) and absorbances.
Notes (1) When the sample has little of organic substances o r suspensoid
and no disturbing materials, the following is allowable: add hy-
drochloric acid (l+l)by 4 ml per 100 ml of sample, boil it to concen-
2
trate up t o about 3 of its original volume, and hereafter carry
out the operations in and after (d) for determination.
(2) While carrying out 4, in case of applying the decomposition by
hydrochloric acid or nitric acid at 4.2, if undissolved matter is
1
left after the solution is concentrated to about 7 of its original
volume by heating, the following procedures should be carried out.
When amount of solution is concentrated about by heating,
6
add 10ml of hydrochloric acid, and heat to almost drying up.
Dissolve this with 4 ml of hydrochloric acid (l+l) and a little wa-
ter. If there remains a lot of residue, filtrate through filter paper
5 grade B, and wash it with warmed water. Put together filtrate
and washings, and store it. Place the residue together with the
filter paper in a platinum crucible, and heat t o carbonize and up
to ash it.
If the ashed substance gives no colouring, no iron in the ashed
residue is confirmed. When it colours into brown, moisten the
ashed substance with several drops of sulfuric acid (2+1) (pre-
pare using sulfuric acid specified in JIS K 8961),add 2 to 3 ml
393
K O101 : 1998
of hydrofluoric acid specified in JIS K 8819,heat gently to vola-

tilize silica, successively heat t o dry, and then heat for about
10 min. After cooled, add 2 g of potassium pyrosulfate specified
in JIS K 8783 or 2 g of potassium hydrogen sulfate specified in
JIS K 8972 and make fusion. After cooled, dissolve the fused
substance in hydrochloric acid (l+l),put it together with the
filtrate and washings above-mentioned, and carry out the operation
in (b) and after to determine iron.
If the sample has little content of silicates (clays) and silica,
omit the elimination of silica by hydrofluoric acid, and the fu-
sion by potassium pyrosulfate or potassium hydrogen sulfate should
preferably be carried out.
(9 When the quantity of iron is extremely small (Fe 20 pg or less),
add 0.1 g of aluminum potassium sulfate 12 hydrate, as a collec-
tor, specified in JIS K 8255,add again aqueous ammonia (l+l)
to make it slightly alkaline to produce aluminum hydroxide, and
collect iron followed by filtration, In this case, because the pre-
cipitate is difficult to dissolve in hydrochloric acid, add a little
more hydrochloric acid, and heat t o concentrate up t o about 5 ml
of solution. Then, add water to make total about 70 ml, and add
0.1 g of sodium potassium tartrate tetrahydrate specified in JIS
K 8536. Hereafter, carry out the operations in and after (d).
(4) When it contains no substance t o prevent colouring, carry out
preferably the operations in and after (d)for determination. (Refer
t o Remarks 4.)
(5> Alternatively, add 0.1 g of L(+)-ascorbic acid specified in JIS K
9502.
When it colours, pH becomes about 4.8. When concentration of
hydrochloric acid is high, neutralize it with aqueous ammonia
(l+l), and control the pH t o 4 to 5 for colouring.
Carry out the pH control procedure in accordance with the
order of operations in (3)after addition of 1,lO-phenanthroline
solution (ig/Z).
Remarks 1 When determining dissolved iron, take a suitable amount (con-
taining 20 to 500 pg as Fe) of the sample filtrated in 3.2 (use
filter paper 5 grade C), add 1 to 2 ml of nitric acid (l+l) and
boil it. Hereafter, carry out the operations in (3)(b)to (i) to
determine iron. If interfering substance does not coexist, the
determination may be carried out by the operations in (3)(d)
and after with omitting the operations of (3)(b) and ( c ) .
2 For obtaining suspensoid iron, subtract dissolved iron from iron
(total).
3 To determine iron (II), carry out as follows.
Place a suitable amount (containing 20 to 50pg as Fe) of
the solution filtrated in 3.2 into a 100 ml volumetric flask, add .
5 ml of 1,lO-phenanthroline solution (1 g/Z), control pH to about
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
394
K O101 : 1998
5 by adding ammonium acetate solution (500 gL), add oxygen-

free water mentioned in 2 (12)(a) up to the marked line, and
let it stand for about 20 min. Hereafter, carry out the opera-
tions in (g) to (i) for determination of iron, and calculate the
concentration [mgFe (II)/Zl of iron (II) in the sample.
Because iron (II) is easily oxidized by oxygen in air, carry
out this test immediately after sampling, but when immedi-
ate treatment is impossible, make the operation until colouring
a t sampling site, and measure its absorbance after bringing
it back t o a laboratory.
4 When the iron is tested which is not previously separated by
being made hydroxide, such as mercury, copper, cadmium,
nickel, cobalt, zinc makes disturbance. Provided that 50 mg/Z o r
less of cadmium, 10 mg/Z or less of zinc, 1 mg/Z or less of mercury
doesn’t disturb. If colouring is made at pH 3.5, 10 mg/Z or less
of copper and 10 mg/Z or less of cobalt gives no disturbance. The
disturbance owing to about 10 mg/Z of nickel can be prevented
by adding 5 ml of EDTA solution (Dissolve 3.7 g of disodium
dihydrogen ethylenediaminetetraacetate dihydrate specified in
JIS K 8107 in 100 ml of water.) and by boiling for about 10 min.
The disturbance by coexisting a lot of zinc can be removed
by colouring after adding a lot of 1,lO-phenanthrolinesolution
(1g/Z) a t pH 9. A lot of phosphate ion also disturbs determina-
tion, but it can be lessened by controlling pH t o be 5 t o 7 when
colouring, and by measuring absorbance after 2 h standing.
60.2 Flame atomic absorption method After pretreatment of a sample, spray

it into a flame such as acetylene-air, and measure atomic absorption by iron at 248.3 nm
wavelength to determine iron.
Determination range: Fe 0.3 t o 6mglZ

(a) Iron standard solution (10 pgFe/ml) Follow 60.1 (1)(h).

(b) Iron hollow cathode lamp
Remarks 5 When determining dissolved iron, take a suitable amount of
sample filtrated in 3.2 (use filter paper 5 grade Cl, and treat
according to 4.5.
6 For suspensoid iron, follow Remarks 2.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
395
K O101 : 1998
7 When the sample, with low concentration of iron, contains

almost no interfering substances, take 100 ml of a sample, add
2 ml of nitric acid specified in JIS K 8541, boil it, carry out
the operations according to 60.1 (3)(b)and (e),and then sepa-
rate and concentrate it.
(a) Spray the sample which has been pretreated as in (3)into a flame accord-
ing to 6 of JIS K 0121,and read the indicated value(') at 248.3 nm wave-
length.
(b) Take water the same amount as that of sample pretreated in (3)for a blank
test, carry out the operations in (3)and (4)(a)similarly t o sample side,
read indicated value, and correct the indicated value obtained on the sample.
(c) Find the quantity of iron on the working curve, and calculate the concen-
tration (mgFel2) of iron in the sample.
Working curve Pipet step by step 3 t o 60ml of iron standard solution
(10 pgFelm1) into as many 100 ml volumetric flasks, add acid respectively
t o make the same acid concentration as that of the sample pretreated in
(3)(a), add water up to the marked line. Carry out the operation in (a) on
this solution. Separately take water for a blank test, add acid t o make the
same acid concentration as that of the sample pretreated in (3) (a), carry
out the operation in (a), correct the indicated value obtained on the stan-
dard solution, and draw the relation curve between the quantities of iron
(Fe) and indicated values. Prepare the working curve when the sample is
measured.
Note (7) Absorbance or its proportional value shall be valid.
Remarks 8 In case of the sample containing a lot of silica, add calcium (or
magnesium) as an inhibitor of interfering by about 200 mgll.

atomize it in an electric furnace, and measure atomic absorption by iron at 248.3 nm
wavelength t o determine iron.
Determination range: Fe 5 t o lOOpgl2
is less affected.

element t o be determined, and verify that there is no interference t o use.
JIS K 9901.
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K O101 : 1998
(c) Iron standard solution (1 pgFe/ml) Take 10 ml of iron standard solu-

tion (10 pgFe/ml) mentioned in 60.1 (1)(h) into a 100 ml volumetric flask,
add 2 ml of nitric acid (l+l),
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(c) Iron hollow cathode lamp
Remarks 10 When determining dissolved iron, follow Remarks 5.
Inject a definite amount (for instance, 10 to 50 pl) of the sample which has
been pretreated in (3)into an exothermic body using a micropipet, hereaf-
ter according to 6 of JIS K 0121, dry it (100 t o 120 "C for 30 t o 40 s), ash
it (600 t o 1O00 "C for 30 t o 40 s), atomize i t ( 8 ) (2 200 t o 2 800 "C for 3 to
6 s), and read the indicated value(') a t 248.3nm wavelength(?.
Take water the same amount as that of sample at preparatory treatment
tration (pgFelZ) of iron in the sample.
Working curve Pipet step by step 0.5 to 10 ml of iron standard solution
(1 mgFe/ml) into as many 100 ml volumetric flasks, respectively add acid
to make the same acid concentration as that of the sample pretreated in
(3)(a),and add water up t o the marked line. Carry out the operation in
(a) on these solutions. Separately, take water for a blank test, add acid to
make the same acid concentration as that of the sample pretreated in (3)(a)
used for working curve preparation, carry out the operations in (a),correct
the indicated value obtained on the standard solution, and draw the rela-
tion curve between the quantities of iron (Fe) and indicated values. Pre-
Notes (8) The conditions for drying, ashing or atomizing vary depending
upon apparatus. They may also be affected by such as injected
397
K O 1 0 1 : 1998
60.4 ICP atomic emission spectrometry After sample is pretreated, spray it

into a n inductively coupled plasma through the sample introducing part, and mea-
sure emission by iron at 238,204 nm wavelength to determine iron.
Determination range: Fe 20 t o 200 pglZ, 0.2 t o 5 mgll
Repeatability: 2 to 10 % coefficient of variation (depending on apparatus and
(a) Iron standard solution (10 pgFe/ml) Follow 60.1 ( i )(h).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Mixed standard solution [(lopgCu, 10 pgZn, 8 pgCd, 10 FgNi, 10 pgPb,

10 pgMn, 10 pgFe)/mll Follow 51.4 ( i )(b).
(a) ICP emission spectrometer
Remarks 12 When determining dissolved iron, follow Remarks 5 .
14 When the sample, which has been preparatorily treated, has
rich concentration of sodium, potassium, magnesium, and cal-
cium, and poor concentration in iron, the operation in Remarks
7 in 51 may be allowable for determination of iron.
Spray the sample which has been pretreated as in (3)into a plasma through
emission strength at 238.204nm wavelength (10) (11) (12).
Take the same amount of water as that of sample pretreated in (3)for a
blank test, carry out the operations in (3) and (4)(a)similarly to sample
side, and correct the emission strength obtained on the sample.
tration (pgFeIZ) of iron in the sample.
Working curve Pipet step by step 0.2 to 2 ml (or 2 to 50 ml)(13)(14) of
iron standard solution (10 pgFe/ml) into as many 100 ml volumetric flasks
respectively, add acid to make the same acid concentration as the sample
pretreated in (3)(a), and add water up t o the marked line. Carry out the
operation in (a)on these solutions. Separately, take water for a blank test,
add acid t o make the same acid concentration as that of the sample pre-
treated in (3) (a),carry out the operation in (a),correct the emission strength
the quantities of iron (Fe) and emission strengths. Prepare the working
curve when the sample is tested.
398
K O101 : 1998
Notes (lo) When the apparatus capable of simultaneously measuring two

method is applied, the procedures are as follows: Take a suit-
able amount of the sample, which has been treated in (3)(a),
into a 100ml volumetric flask, add 10ml of yttrium solution
(50 pgY/ml) [Follow Note (8) of 451, add acid t o make the same
acid concentration as the sample in (4) (a), and add water up to
the marked line. Carry out the spraying operations in (4)(a)
on this solution, measure emission strength a t both 238.204 nm
and 371.029 nm (yttrium) wavelength, and obtain the emission-
strength ratio of iron and yttrium.
Separately, pipet step by step 0.2 to 2 ml (or 2 to 50 mi) of
iron standard solution (10 pgFe/ml) into as many 100 ml volu-
metric flasks, respectively add 10 ml of yttrium solution (50 pgY/
mi), add acid to make the same acid concentration as the sample
of (4) (a), and add water up t o the marked line. Carry out the
spraying operation in (4) (a) on these solutions, measure emis-
sion strengths a t both 238.204 nm and 371.029 nm wavelength,
draw the relation curve between emission-strength ratio of iron
to yttrium and the concentration of iron, and make it the work-
ing curve. On this working curve, find the quantity of iron cor-
responding to the emission-strength ratio obtained on the sample,
and calculate the concentration (pgFe/Z) of iron in the sample.
high concentration of salts in a sample, the standard addition
racy have been confirmed.
prepared as follows: dilute iron standard solution (10 pgFe/ml)
to suitable concentration (0.1 to 1 pgFe/ml), take step by step
0.2 to 2 ml (or 2 to 50 ml), make them 500 ml (or definite amount
of 100 to 500ml), carry out the operations in Remarks 14 and
(4)(a) and (b) similarly to sample side, and draw the relation
curve between the quantities of iron (Fe) and emission strength.
(14) When copper, zinc, cadmium, nickel, lead, and manganese are
simultaneously tested, use mixed standard solution [(lo pgCu,
10 pgZn, 8 pgCd, 10 pgNi, 10 pgPb, 10 pgMn, 10 pgFe)/ml], and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
399
K O101 : 1998
61 Chromium (Cr) Chromium shall be classified into total chromium and chro-
mium (VI).
61.1 Total chromium For the determination of total chromium, diphenylcarbazide

absorptiometry, flame atomic absorption method, electric heating atomic absorption
method, ICP atomic emission spectrometry or ICP mass spectrometry shall be applied.
61.1.1 Diphenylcarbazide absorptiometry Oxidize chromium (III) to chromium

(VI) by potassium permanganate, add 1,5-diphenylcarbonohydrazide (diphenyl-
carbazide), and measure absorbance owing to produced reddish violet complex for
determination.
Determination range: Cr 2 to 50pg
Sulfuric acid (1+9) Prepare using sulfuric acid specified in JIS K 8951.
Potassium permanganate solution (3 g/Z) Follow 46.1 (i)(f).
Sodium nitrite solution (20 gll) Dissolve 2 g of sodium nitrite specified
in JIS K 8019 in water to make total 100 ml. Prepare this solution when
it is needed.
Urea solution (200 gll) Dissolve 20 g of urea specified in JIS K 8731 in
water to make total 100ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Diphenylcarbazide solution (10 gil) Dissolve 0.5 g of 1,5-diphenylcarbono-

hydrazide (diphenylcarbazide) specified in JIS K 8488 in 25 ml of acetone
specified in JIS K 8034, and add water t o make total 50 ml. Store it in a
cool and dark place. This is effective for 1 week.
Chromium standard solution (0.1 mgCr/ml) Dry potassium dichromate,
reference material for volumetric analysis, specified in JIS K 8006 at 150 "C
for about 1 h, and let it cool in a desiccator. Take its 0.283 g on the base
of KzCr20, 100 %, dissolve in water, transfer it into a 1 O00 ml volumetric
flask, and add water up to the marked line. Otherwise use reference ma-
terial-standard solution-chromium standard solution, Cr 100, specified in
JIS K 0024.
Chromium standard solution (2 pgCr/ml) Pipet 20 ml of chromium stan-
dard solution (0.1 mgCr/ml) into a 1O00 ml volumetric flask, and add wa-
ter up to the marked line.
(a) Photometer Spectrophotometer or photometric photometer
(a) Take a suitable amount(2) (containing 2 to 50 pg as Cr) of the sample which

has been treated in 4(1) into a beaker, add 3 ml of sulfuric acid (1+9)(3),
and heat until white fume of sulfuric acid slightly generates(*)( 5 ) . After it
is cooled, add about 30 ml of water, and heat it t o dissolve residue.
400
K O101 : 1998
Heat the solution gently, and drip potassium permanganate solution (3 g/Z)
until the solution colours. Boil it while keeping its colour by dripping po-
tassium permanganate solution ( 3 g/Z) when it is about to be decoloured, for
several minutes.
Cool it with running water, add 10 ml of urea solution (200 g/Z), while agi-
tating violently add drop by drop sodium sulfite solution (20 g/Z)(6)to decolour
its red colour, and decompose both excessively added potassium perman-
ganate and manganese (IV) oxide.
Transfer it into a 50ml volumetric flask, keep its temperature a t about
15 OC(7), add 1ml of diphenylcarbazide solution (10 g / O , and immediately
shake. Add water up to the marked line, and shake, followed by standing
for 5 min(8).
Place a part of the solution in an absorption cell, and measure its absor-
bance in the vicinity of 540 nm wavelength.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Take about 30 ml of water for a blank test, add 3 ml of sulfuric acid (1+9),
carry out the operation in (d) and (e),measure its absorbance, and correct
Find the quantity of chromium on the working curve, and calculate the con-
centration of total chromium (pgcrll) in the sample.
Working curve Pipet step by step 1 to 25 ml of chromium standard so-
lution (2 pgCr/ml) into as many beakers, respectively add 3 ml of sulfuric
acid (1+9), add water t o make about 30 ml total, carry out the operations
in (b) to (0,and draw the relation curve between the quantities of chro-
mium (Cr) and absorbances.
4.3 out of 4 shall not be used.
When sample, with low concentration of chromium has almost no
organic substances and suspensoid, take a suitable amount of
500 ml or less, add 2 ml of sulfuric acid specified in JIS K 8951
per 100 ml of a sample, heat to boil, and let it cool. Add 1 ml of
ammonium iron (II) sulfate solution (5 mgFe/ml) [Dissolve 3.5 g of
ammonium iron (II) sulfate hexahydrate specified in JIS K 8979
in 100 ml of water containing a few drops of sulfuric acid.], stir
well, add 2 ml of nitric acid specified in JIS K 8541, and boil for
a few minutes to oxidize iron (II). After standing this solution for
a while, neutralize with aqueous ammonia (1+4), boil it until no
smell of ammonia, and allow it t o stand at about 80 OC for about
20 min t o complete precipitation. Filtrate it through filter paper
5 grade A, wash it several times with warm ammonium nitrate
solution (10 g/Z), dissolve it with 5 ml of sulfuric acid (1+15),wash
the filter paper with warm water, and carry out the operation in
(b) to (f). Provided that, when preparing working curve, 5 mg of
iron [Fe (III)]should be respectively added into standard solution.
The concentration of sulfuric acid should be preferably kept near
0.1 mol/Z for the colouring of chromium (VI) by diphenylcarbazide.
When a lot of sulfuric acid is added a t pretreatment, 3 ml of sul-
furic acid (1+9) shall be added after removing sulfuric acid by
heating to generate white fume. When generating this white fume,
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
401
K O101 : 1998
never heat strongly. Because it makes the residue unsoluble due

to producing anhydride of chromium (III) sulfate. Adding 20 mg
of sodium sulfate can prevent this.
When the pretreatment makes it generate white fume, this op-
eration can be omitted.
Instead of sodium sulfite solution (20 g/Z), sodium azide solution
(50g/Z) can be used. In this case, drip carefully sodium azide
solution (50 g/Z), stir enough to decompose permanganate, and
boil it for 2 to 3min to decompose excess sodium azide.
Because the temperature of solution influences colouring, it is
important to keep it about 15 OC.
Colouring gives its highest in 2 o r 3 min, and then gradually fades
away, but practically no change occurs during 5 to 15 min.
1 When sample contains iron, absorbance shows smaller value
according t o increasing iron, but it becomes nearly constant
at about 1mg per 50 ml of colouring solution. (It is about 20 %
lower value.) Before adding diphenylcarbazide solution, how-
ever, if 2 ml of sodium diphosphate solution (Dissolve 5 g of
sodium diphosphate 10 hydrate specified in JIS K 8785 in water
t o make total 100 mi.) is added, the influence by 2.5 mg or less
of iron will be neglected.
In case of less iron than chromium, it will be neglected.
2 Applying this method, molybdenum, mercury, or vanadium gives
influence. Molybdenum gives no influence until 0.1 mg. The
disturbance by mercury is eliminated by adding chloride ion.
The influence by vanadium will be neglected if absorbance
is measured after 10 to 15 min from colouring.
3 When there are a lot of disturbances by iron or others, carry out
as follows: take a suitable amount (containing 2 to 50 pg as Cr)
in a separatory funnel, add 5 ml of sulfuric acid (l+l) per 20 ml
of sample to make the concentration of sulfuric acid about
1.8 mol/Z, and drip potassium permanganate solution (3 g/Z) to
colour faintly. Add 5 ml of cupferron solution (50 g/Z) [Dissolve
5 g of cupferron (N-nitroso-N-phenylhydroxylamine ammonium
salt) (N-hydroxy-N-nitrosobenzeneamine ammonium salt, no-
menclature by IUPAC) specified in JIS K 8289 in water to make
total 100 mi.] and 10 ml of chloroform specified in JIS K 8322,
shake them strongly for about 30 s to extract iron and others,
and settle it.
Separate chloroform layer, add again 1ml of cupferron so-
lution (50 g/Z) and 10 ml of chloroform into water layer t o re-
peat extraction, and separate chloroform layer. Remove water
layer into a 100 ml beaker, and heat to evaporate water to get
slight drying. Add a little of sulfuric acid and nitric acid, and
again evaporate to dryness for decomposing of organic sub-
stances. Dissolve it in 0.3 ml of sulfuric acid (l+l) and about
30 ml of water. Oxidize chromium with potassium perman-
ganate solution (3 g/Z), and follow the description in (3).
402
K O101 : 1998
61.1.2 Flame atomic absorption method After pretreatment of sample, spray

it in a flame such as acetylene-air, and measure the atomic absorption by chromium
a t 357.9 nm wavelength t o determine total chromium.
Determination range: Cr 0.2 to 5 mg/Z
Repeatability: 2 t o 10 % coefficient of variation (depending on apparatus and
(a) Chromium standard solution (10 pgCr/ml) Pipet. 50 ml of chromium
standard solution (0.1 mgCr/ml) mentioned in 61.1.1 (i) (f) into a 500 ml
volumetric flask, add 10 ml of nitric acid (l+l>, and add water up to the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
marked line.

(b) Chromium hollow cathode lamp
(a) Treat a sample according t o 4.5 or Remarks 4(1)(4).
Remarks 4 When the sample, with low concentration of chromium, has
almost no organic substance and suspensoid, the preparatory
operations shall be as follows:
Take a suitable amount of sample, carry out operation ac-
cording to Note ('9,and coprecipitate chromium with iron (III)
hydroxide. Filtrate through filter paper 5 grade A and dis-
solve the precipitate in a little nitric acid (1+2), and wash fil-
ter paper with warm water. Put together washings and filtrate,
and make it a definite amount having 0.1 to 1 mol/Z acidified
by hydrochloric acid or nitric acid.
(a) Spray the sample which has been pretreated as in (3)into a flame(9) accord-
ing to 6 of JIS K 0121,and read indicated value('*) at 357.9 nm wavelength.
(b) Take the same amount of water as that of the sample in (3)for a blank
test, carry out the operations in (3)and (4)(a) similarly to sample side,
measure the indicated value, and correct the indicated value obtained on
the sample.
(c) Find the quantity of chromium on the working curve, and calculate the con-
centration of total chromium (mgCrlZ) in the sample.
Working curve Pipet step by step 2 to 50 ml of chromium standard solu-
tion (10 pgCr/ml) into as many 100 ml volumetric flasks, respectively add acid
to make the same acid concentration as that of the sample pretreated in (3) (a)
and add water up t o the marked line. Carry out the operation in (a)on these
solutions. Separately take water for a blank test, add acid to make the same
acid concentration as that of the sample pretreated in (3) (a),carry out the
operation in (a)t o correct the indicated value on the standard solution, and
403
K O101 : 1998
draw the relation curve between the quantities of chromium (Cr) and indi-
cated values. Prepare the working curve when the sample is measured.
Notes (9) Use an acetylene-air flame or acetylene-dinitrogen monoxide flame
which has less fuel.
Remarks 5 When the sample, with low concentration of chromium, has
no disturbing material for extraction, the following shall be
allowable for determination.
Take a suitable amount (containing 5 t o 100 pg as Cr) in a
100 ml beaker, add 2 ml of sulfuric acid (1+2) and a few drops
of potassium permanganate (3 g/Z), and heat it. Boil for a few
minutes while keeping slight red colour in solution by drip-
ping potassium permanganate (3 glZ) when colour of perman-
ganate is about to decolour. Cool it with running water, transfer
it into a separatory funnel, and add water t o make total about
100 ml. Add 20 ml of butyl acetate solution of N,N-dioctyl-
octaneamine (trioctylamine) (30 g/Z), shake violently for about
10 min, and let it stand. Spray the butyl acetate layer as it
is into the flame, and determine chromium.
When making working curve, use suitably diluted solution
of chromium standard solution (10pgCr/ml). Instead of butyl
acetate specified in JIS K 8377, 4-methyl-2-pentanone speci-
fied in JIS K 8903 can be available.
6 In case of an acetylene-air flame, fuel-rich flame gives a high
sensitivity, but simultaneously increases the disturbance by
iron, nickel, or others. In this case, make sodium sulfate,
potassium disulfate, or ammonium hydrogen difluoride remain
in the solution by around 1 %.
In case of an acetylene-dinitrogen monoxide flame, almost
all disturbances disappear.
61.1.3 Electric heating atomic absorption method After pretreatment of a

sample, atomize it in an electric furnace, measure atomic absorption by chromium
a t 357.9 nm wavelength to determine total chromium.
Determination range: Cr 5 t o lOOpg/Z
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 7 Because this method is easily affected by coexisting acids, salts
and their concentrations, the sample which is less affected shall
be adopted.
(a) Water Water A3 specified in JIS K 0557. Carry out a blank test o n the
element t o be determined, and verify that there is no interference.
JIS K 9901.
404
K O101 : 1998
(c) Chromium standard solution (1 pgCr/ml) Pipet 10 ml of chromium stan-

dard solution (10 ygCr/ml) mentioned in 61.1.2 (i)(a)into a 100 ml volumetric
flask, add 2 ml of nitric acid (l+l),

(b) Exothermic body Made of graphite or heat-resisting metal
( c ) Chromium hollow cathode lamp
(e) Micropipet Piston operated micro-volume apparatus specified in JIS K
0970, 10 to 5 0 ~ 1 . Or an automatic injection device.
(a) Treat a sample according t o 4.5 or Remarks 4(1). The last solution, how-
ever, should be made 0.1 t o l mol/Z acidified by nitric acid.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Inject a certain amount (for instance, 10 to 50 pl) of the sample which has
been pretreated as in (3) into an exothermic body using a micropipet, here-
after according t o 6 of JIS K 0121,dry it (100 t o 120 "C for 30 to 40 s), ash
it (500 to 600 "C for 30 to 40 s), atomize it(11) (2 400 t o 2 900 "C for 5 to
10 s ) , and read the indicated value(10) at 357.9 nm wavelength(l2).
in (3) for a blank test, carry out the operations in (3) and (4) (a) similarly
Find the quantity of chromium on the working curve, and calculate the con-
centration (pgCrl2) of total chromium in the sample.
Working curve Pipet step by step 0.5 to 10ml of chromium standard
solution (ipgCr/ml) into as many 100 ml volumetric flasks, respectively add
nitric acid t o make the same acid concentration as that of the sample pre-
treated in (3)(a) and add water up t o the marked line. Carry out the op-
eration in (a) on these solutions. Separately, take water for a blank test,
add nitric acid t o make the same acid concentration as that of the sample
pretreated in (3)(a), carry out the operation in (a),correct the indicated
value obtained on the standard solution, and draw the relation curve be-
tween the quantities of chromium (Cr) and indicated values. Prepare the
working curve when the sample is tested.
Notes (11) The conditions for drying, ashing, o r atomizing vary depending
upon apparatus, and they may be also affected by such as in-
jected volume of sample and concentration of coexisting salts.
(12) Repeat successively at least 3 times the operations in (a), and
405
K O101 : 1998
61.1.4 ICP atomic emission spectrometry After sample is pretreated, spray it

into an inductively coupled plasma through the sample introducing part, and measure
the emission by chromium at 206.149 nm wavelength t o determine total chromium.
Determination range: Cr 20 t o 4000pglZ

(a) Chromium standard solution (10pgCr/ml) Follow 61.1.2 (i) (a).
(b) Mixed standard solution [(lopgCr, 10 pgV)/ml] Pipet 10 ml of chro-
mium standard solution (0.1 mgCr/ml) in 61.1.1 (i)(f) and 10 ml of vana-
dium standard solution (0.1 mgV/ml) in 62.1 (i)( g )into a 100 ml volumetric
flask respectively, add 2 ml of nitric acid (l+l),and add water up to the
marked line. Prepare this solution when it is used.
(a) Treat a sample according t o 4.5(1).
(a) Spray the sample which has been pretreated as in (3) into a plasma through
the sample introducing part according to 5.8 of JIS K 0116,and measure
emission strength at 206.149 nm wavelength(l3) (14) (15).
(b) Take the same amount of water as that of sample pretreated in (3) for a
blank test, carry out the operations in (3)and (4)(a)similarly to the sample,
and correct the emission strength obtained on the sample.
(c) Find the quantity of chromium on the working curve, and calculate the con-
centration (pgCr/Z) of total chromium in the sample.
Working curve Pipet step by step 0.2 to 40 m l ( 9 of chromium standard
solution (10 pgCr/ml) into as many 100 ml volumetric flasks, respectively
add acid to make the same acid concentration as that of the sample pre-
treated in (3)(a),and add water up t o the marked line. Carry out the
operation in (a)on these solutions. Separately, take water for a blank test,
add acid to make the same acid concentration as that of the sample pre-
treated in (3)(a),carry out the operation in (a),correct the emission strength
the quantities of chromium (Cr) and emission strengths. Prepare the working
curve when the sample is measured.
method is applied the procedures are as follows: Take a suit-
able amount of the sample, which has been treated in (3)(a),
into a 100ml volumetric flask, add 10ml of yttrium solution
(50 pgY/ml) [Follow Note (8) of 451, add acid t o make the same
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
406
K O101 : 1998
acid concentration as the sample in (4) (a), and add water up to

the marked line. Carry out the spraying operations in (4)(a)
on this solution, measure emission strength at both 206.149 nm
and 371.029 nm (yttrium) wavelength, and obtain the emission-
strength ratio of chromium and yttrium.
Separately, pipet step by step 0.2 to 40 ml of chromium stan-
dard solution (10 pgCr/ml) into as many 100 ml volumetric flasks,
respectively add 10 ml of yttrium solution (50 pgY/ml), add acid
t o make the same acid concentration as the sample of (4) (a),and
add water up t o the marked line. Carry out the spraying opera-
tions in (4) (a)on these solutions, measure emission strengths at
both 206.149 nm and 371.029 nm wavelength, draw the relation
curve between emission-strength ratio of chromium to yttrium and
the concentration of chromium, and make it the working curve.
On this working curve, find the quantity of chromium correspond-
ing t o the emission-strength ratio obtained on the sample, and
calculate the concentration (pgCr/Z) of chromium in the sample.
high concentration of salts in the sample, the standard addition
racy have been confirmed.
(16) When vanadium is simultaneously tested, use mixed standard
solution [(lo pgCr, 10 pgV)/mll, and prepare preferably the work-
ing curve for vanadium under the test condition of vanadium.
Remarks 8 When sample has rich concentration of sodium, potassium, mag-
nesium, and calcium and has poor concentration in chromium,
the following shall be allowable: take a suitable amount of
sample (containing 0.4to 80 pg as Cr) in a 100 ml beaker, and
carry out operations according t o Remarks 5 . In this case,
use a torch for organic solvent as a plasma torch.
61.1.5 ICP mass spectrometry Pretreat a sample, add an internal standard sub-
part, measure the ionic current in each number of massedelectric charges of chro-
chromium and that of internal standard substance t o determine total chromium.
Determination range: Cr 0.5 to 25 pg/Z, 10 to 500 pg/Z
(a) Water Follow 61.1.3 (1)(a).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
407
K O 1 0 1 : 1998
Nitric acid ( l + l ) Follow 61.1.3 (1) (b).

Yttrium solution (1 ygY/ml)(l7) Follow 51.5 (1)( e ) .
Chromium standard solution (1 ygCr/ml) Follow 61.1.3 (1)( c ) .
Chromium standard solution (50 ngCr/ml) Take 50 ml of chromium
standard solution (1 ygCr/ml) in a 1000 ml volumetric flask, add 3 ml of
nitric acid (1+1) and add water up to the marked line. Prepare when it is
used.
Mixed standard solution [(ipgCu, 1 ygzn, 1 ygCd, 1 ygPb, 1 ygMn,
1 ygCr)/mll Follow 51.5 (1) (f).
50 ngMn, 50 ngCr)/mll Follow 51.1 (1)(g).
(a) Treat a sample according to 4.5(l).
(b) Take a suitable amount (containing 0.05 t o 50 yg as Cr) of sample treated
in (3)(a) in a 100 ml volumetric flask, add 1ml of yttrium solution (1pgY/
mi), add nitric acid (l+l)t o make final concentration of nitric acid 0.1 to
0.5 mollE and add water up to the marked line.
(4) Operation Operations shall be carried out as follows(19).
Make the ICP mass spectrograph ready to run, spray the solution in (3) (b)
read the indicated value(21) in the number of masseslelectric charges (20) of
chromium and yttrium, and obtain the ratio between the indicated value
of chromium and that of yttrium.
in (3) (a),carry out the operations in (3) and (4)(a)similarly t o the sample,
obtain the ratio between the indicated value of chromium and that of yt-
trium, and correct the ratio of the indicated values between chromium and
yttrium obtained on the sample.
Find the amount of chromium on a working curve, and calculate the con-
centration (ygCrlZ) of chromium in the sample.
Working curve Pipet step by step I t o 50 ml of the chromium standard
solution (50 ngCr/ml or 1ygCr/ml)(22)in as many 100 ml volumetric flasks,
add 1ml of yttrium solution (1ygY/ml), add nitric acid (l+l)t o make the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
408
K O 1 0 1 : 1998
same acid concentration as the sample carried out the operation in (3)(b)
and add water up to the marked line. Carry out the operation in (a) on
this solution. Separately put 1ml of yttrium solution ( 1 pgY/ml) as a blank
test in a 100ml volumetric flask, add nitric acid (l+l) to make the same
acid concentration as the sample of (3)(b),and add water up to the marked
line. Carry out the operation in (a),correct the indicated values obtained
the indicated value to the amount of chromium (pgCr) and the indicated
value of yttrium. Prepare the working curve when the sample is measured.
61.2 Chromium (VI) [Cr (VI)] For the determination of chromium (VI), diphenyl-
carbazide absorptiometry, flame atomic absorption method, electric heating atomic
absorption method, ICP atomic emission spectrometry or ICP mass spectrometry shall
be applied.
61.2.1 Diphenylcarbazide absorptiometry Add 1,5-diphenylcarbonohydrazide

(diphenylcarbazide)into a sample, and measure the absorbance of produced reddish
violet complex to determine chromium (VI).
Determination range: Cr (VI) 2 to 50pg
(a) Sulfuric acid (1+9) Prepare using sulfuric acid specified in JIS K 8951.
(b) Ethanol (95) Specified in JIS K 8102.
(c) Diphenylcarbazide solution (10 g/Z) Follow 61.1.1 (1)(e).
(d) Chromium (VI) standard solution [2 pgCr (VI)/ml] Follow 61.1.1 (1)(g).
(a) Take two suitable amount of samples [containing 2 to 50 pg as Cr (VI)] into
two beakers (A) and (BI, and neutralize the sample with sodium hydroxide
solution (40glZ) for acidic sample or with sulfuric acid (1+35) for alkaline
sample.
(b) Transfer the solution in beaker (A) into a 50 ml volumetric flask (A), and
add 2.5 ml of sulfuric acid (1+9).
(c) Add 2.5 ml of sulfuric acid (1+9) in the solution in the beaker (BI, add a
little ethanol (95) and boil it to reduce chromium (VI) into chromium (III)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
409
K O101 : 1998
and expel excess ethanol. After it is cooled, transfer it into a 50 ml volu-

metric flask (BI.
(d) Keep both volumetric flasks (A) and (B) a t about 15 OC, respectively add
1ml of diphenylcarbazide solution (10 g/O, immediately mix them, add water
to the marked line, and let them stand for about 5 min.
(e) Place a part of the solution in volumetric flask (A) in an absorption cell,
and measure its absorbance in the vicinity of 540 nm wavelength with making
the solution in volumetric flask (B) contrast solution.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(0 Find the quantity of chromium (VI) on the working curve, and calculate
the concentration of chromium (VI) [mg C r (VI)/Zl in the sample.
Working curve Pipet step by step 1 to 25 ml of chromium (VI) standard
solution [2pgCr (VI)/mll, and respectively carry out the operations (b) t o
(d) which correspond t o those for the volumetric flask (A). Place portions
of these solutions into an absorption cell, and measure absorbance in the
vicinity of 540 nm wavelength. In this case, reference solution shall be as
follows: take about 30 ml of water, and carry out the operations ( c )and (d)
corresponding to those for the volumetric flask (B). Draw the relation curve
between the quantities of chromium [Cr (VI)] and absorbances.
Remarks 13 If sample has coloured o r coexisting matters which reduce
Cr (VI) when being acidified, determination is difficult. The
sample containing no chromium (III), however, is determined
according to 61.1.
61.2.2 Flame atomic absorption method After sample is pretreated, spray it

into a flame such as acetylene-air, and measure atomic absorption by chromium (VI)
a t 357.9 nm wavelength t o determine chromium (VI).
Determination range: Cr (VI) 0.2 t o 5 mgll

(a) Chromium standard solution (10 pgCr/ml) Follow 61.1.2 (1)(a).
(b) Chromium hollow cathode lamp
(a) Take a suitable amount of sample (In case of containing suspensoid, fil-
trate through filter paper 5 grade C or filter media with 0.45 pm pore di-
ameter, and use filtrate after discarding about the first 50ml filtrate.),
and add hydrochloric acid specified in JIS K 8180 or nitric acid specified
in JIS K 8541 t o make the solution 0.1 t o 1 mol/Z acidity. In case of sample
which contains chromium (III), however, follow the operations in Remarks
15of (2).
410
K O101 : 1998
Remarks 15 When the sample, with low concentration of chromium (VI),

has no disturbing material, operations shall be as follows.
(1) When the sample does not contain chromium (III), fol-
low the operations in Remarks 4 or Remarks 5.
(2) When the sample contains chromium (III), treat it as fol-
lows.
Take 500 ml or less of the sample, add 1ml of ammo-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
nium iron (III) sulfate solution [Dissolve 5 g of ammo-

nium iron (III) sulfate 12 hydrate specified in JIS K 8982
in 1ml of sulfuric acid (l+l), and add water to make total
100 mi.], and stir them. After making the solution slightly
alkaline by adding aqueous ammonia (1+4), and boil it
gently until almost no ammonia odor is apparent. After
maturing precipitate by keeping at warm condition near
boiling, filtrate it through filter paper 5 grade A, and wash
with warm ammonium nitrate solution (10 g/Z). Put to-
gether filtrate and washings, and add hydrochloric acid
or nitric acid to make 0.1 to 1mol/Z acidity.
(4) Operation Operations shall be as’ follows.
(a) Carry out the operations according t o 61.1.2 (4).
61.2.3 Electric heating atomic absorption method After a sample is pretreated,

atomize it in an electric furnace, and measure its atomic absorption by chromium at
357.9 nm wavelength to determine chromium (VI).
Determination range: Cr (VI) 5 to 100 pg/Z
Remarks 16 Follow Remarks 7.
(a) Water Follow 61.1.3 (1)(a).
(b) Nitric acid ( l + l ) Follow 61.1.3 (1)(b).
(cl Chromium standard solution (1 pgCr/ml) Follow 61.1.3 (1)(c).
(a) Electric heating atomic absorption apparatus Electrically heating type
(c) Chromium hollow cathode lamp
411
K 0101 : 1998

(a) Take a suitable amount of the sample, and treat it according t o the opera-
tions in 61.2.2 (3)(a) or Remarks 15. The last solution, however, should
be made 0.1 t o 1 mol/Z acidified by nitric acid.
(a) Carry out the operations according t o 61.1.3 (4).
61.2.4 ICP atomic emission spectrometry After sample is pretreated, spray it

into an inductively coupled plasma through the sample introducing part, and measure
the emission by chromium at 206.149 nm wavelength to determine chromium (VI).
Determination range: C r (VI) 20 to 4 O00 pglZ
(a) Chromium standard solution (10pgCr/ml) Follow 61.1.2 (i)(a).
(b) Mixed standard solution [(lopgCr, 10 pgV)/ml] Follow 61.1.4 (i) (b).
(a) Take a suitable amount of sample and treat according t o 61.2.2 (3)(a) o r
Remarks 15. Provided that, use nitric acid, and make the last solution 0.1
to 0.5 mol/Z acidified by nitric acid.
(a) Follow 61.1.4 (4).
61.2.5 ICP mass spectrometry Pretreat a sample, add an internal standard sub-
part, measure the ionic current in each number of masses/electric charges of chro-
chromium and that of internal standard substance t o determine chromium (VI).
Determination range: Cr(V1) 0.5 to 25pg/Z, 10 to 5OOpglZ
Repeatability: 2 t o 10 % by coefficient of variation (depending o n apparatus and
(a) Water Follow 61.1.3 (i)(a).
(b) Nitric acid (1+1) Follow 61.1.3 (i)(b).
(c) Yttrium solution (i pgY/ml)(l7) Follow 51.5 (i)(c).
(d) Chromium standard solution (i pgcrlml) Follow 61.1.3 (i)(c).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
412
K O101 : 1998
Chromium standard solution (50 ngCr/ml) Take 50 ml of chromium

standard solution (1 pgCr/ml) in a 1 O00 ml volumetric flask, add 3 ml of
nitric acid (l+l)and add water up to the marked line. Prepare when it is
used.
Mixed standard solution [(iygCu, 1 pgZn, 1 ygCd, 1 pgPb, 1 pgMn,
Mixed standard solution [(50ngCu, 50 ngZn, 50 ngCd, 50 ngPb,
50 ngMn, 50 ngCr)/ml] Follow 51.5 (1) (g).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

(a) Treat according to 61.2.3 (3)(a) o r Remarks 15. Provided that, use nitric
acid, and make the solution 0.1 to 0.5 mol/Z acidified by nitric acid.
(b) Take a suitable amount (containing 0.05 t o 50 pg as Cr) of sample treated
in (3)(a)in a 100 ml volumetric flask, add 1ml of yttrium solution ( 1 pgY/
mi), add nitric acid (l+l) t o make final concentration of nitric acid 0.1 to
0.5 mol/Z and add water up to the marked line.
(4) Operation Operation shall be carried out as follows.

(a) Carry out the operation according t o 61.1.5 (4).
413
K 0101 : 1998
62 Vanadium (V) For the determination of vanadium, N-benzoyl-N-phenyl-

hydroxylamine absorptiometry, flame atomic absorption method, electric heating atomic
absorption method, or ICP atomic emission spectrometry shall be applied.
62.1 N-benzoyl-N-phenylhydroxylamine absorptiometry Oxidize sample by

potassium permanganate to make vanadium (V), add IV-benzoyl-IV-phenylhydroxylamine
(N-hydroxy-N-phenylbenzamid,nomenclature by IUPAC) t o produce reddish violet
vanadium complex, extract it from hydrochloric acid solution using chloroform, and
measure its absorbance to determine vanadium.
Determination range: V 2 to 50pg
(1) Reagent Reagents shall be as follows.
K 8180.
Perchloric acid Specified in JIS K 8223.
Copper (II) solution (10 g/Z) Add 1 g of copper (99.9 % or more) speci-
fied in JIS K 8660 in 10 ml of nitric acid ( l + l )heat
, to dissolve, add 20 ml
of perchloric acid, and evaporate it by heating to generate white fume. After
cooling it, dilute it with water to make total 100 ml.
BPHA chloroform solution (2 glZ) Dissolve 0.2 g of N-benzoyl-N-phenyl-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
hydroxylamine specified in JIS K 9569 in 100 ml of chloroform specified

in JIS K 8322.
Potassium permanganate solution (3 g/Z) Follow 46.1 (1) (a.
Vanadium standard solution (0.1 mgV/ml) Dissolve 0.230 g of ammo-
nium vanadate (V) specified in JIS K 8747 in the mixture of 10 ml sulfu-
ric acid (l+l)and 200 ml of hot water. After cooling, transfer it into a 1 O00 ml
Vanadium standard solution (2 pgV/ml) Pipet 20 ml of vanadium stan-
dard solution (0.1 mgV/ml) into a 1 O00 volumetric flask, add 10 ml of sul-
furic acid (l+l),and add water up to the marked line.

(a) Take a suitable amount (containing 2 t o 50 pg as V) of a sample, add 5 ml
of nitric acid and 3 ml of perchloric acid, evaporate by heating to generate
white fume of perchloric acid, and concentrate it until nearly drying up.
(b) After cooling it, add 10 ml of water, add 1 ml of copper (II) solution (10 g/O,
drip potassium permanganate solution (3 g/Z) until the solution gets faint red
colour, add one drop excessively, and let it stand for about 5 min for oxida-
tion of vanadium.
414
K O 1 0 1 : 1998
Transfer it into a 100ml separatory funnel and add water to make total
about 30 ml (1).
Add 10 ml of BPHA chloroform solution (2 g/Z). Then, add 40 ml of hydro-
chloric acid (2+1)to reduce excess permanganate, immediately stir it for
about 30 s, and extract vanadium complex.
After letting it stand, filtrate chloroform layer through dried filter paper.
Place a part of chloroform layer in an absorption cell, and measure absor-
bance in the vicinity of 530 nm wavelength with making chloroform refer-
ence solution.
Take about 10 ml of water for a blank test, carry out the operations in (b)
to (fl, and correct the absorbance obtained on the sample.
Find the quantity of vanadium on the working curve, and calculate the con-
centration of vanadium (pgV/Z) in the sample.
Working curve Pipet step by step 1 to 25 ml of vanadium standard so-
lution (2 pgV/ml) into as many separatory funnels, respectively carry out
the operations in (b)t o (g), and draw the relation curve between the quan-
tities of vanadium (V) and absorbances.
Note (1) Previously put a mark on a separatory funnel.
62.2 Flame atomic absorption method After a sample is pretreated, spray it

into the flame of acetylene-dinitrogen monoxide, and measure atomic absorption by
vanadium at 318.4 nm wavelength t o determine vanadium.
Determination range: V 1 t o 20mglZ

(a) Nitric acid Specified in JIS K 8541.
(b) Aluminum nitrate solution (400gll) Dissolve 70 g of aluminum nitrate

enneahydrate specified in JIS K 8544 in a little water with heating, and
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
after cooling it, add water to make total 100 ml.

(c) Vanadium standard solution (0.1mgV/ml) Follow 62.1 (1) (g).

(b) Vanadium hollow cathode lamp

415
K O101 : 1998

Take a suitable amount (containing 0.1 t o 2 mg as V) of the sample which
has been pretreated in (3) into a 100 ml volumetric flask, add 1ml of nitric
acid, and add water up t o the marked line.
Take 50 ml of this solution into a dried beaker, and add 1ml of aluminum
nitrate solution (400 g/Z).
Spray the solution in (b) into the flame of acetylene-dinitrogen monoxide
according t o 6 of JIS K O121 (21, and read the indicated value(3) at 318.4 nm
wavelength.
Take water the same amount as that of sample for a blank test, carry out
the operations in (3) and (4) (a) to (c), and correct the indicated value ob-
Find the quantity of vanadium on the working curve and calculate the con-
centration of vanadium (mgVIZ) in the sample.
lution (0.1 mgV/ml) into as many 100 ml volumetric flasks, respectively add
acid t o make the same acid concentration as that of sample pretreated in
(3)(a), and add water up t o the marked line. Carry out the operations in
(b) and (c)on these solutions. Separately take water for a blank test, add
acid and nitric acid to make the same acid concentration as that of the sample
pretreated in (3)(a),carry out the operations in (b) and ( c ) , correct the
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
curve between the quantities of vanadium (VI and indicated values. Pre-
Notes (2) The flame with rich fuel gives a high sensitivity. Because high
sensitivity range is very limited in the flame, so in advance find
this position.
62.3 Electric heating atomic absorption method After a sample is pretreated,

atomize it in an electric furnace, measure atomic absorption by vanadium at 318.4 nm
wavelength to determine vanadium.
Determination range: V 10 to 200pglZ
Remarks 1 This method is easily affected by the kind and concentration
of coexisting acid and salt, therefore applicable t o the sample
which is less affected.
element to be determined, and verify that there is no interference.
JIS K 9901.
416
K O101 : 1998
(c) Vanadium standard solution (1pgV/ml) Pipet 2 ml of vanadium stan-

dard solution (0.1 mgV/ml) in 62.1 (1)( g ) into a 200 ml volumetric flask,

(b) Exothermic body Made of graphite or heat-resisting metal
(cl Vanadium hollow cathode lamp
K 0970,10 t o 50 pl. Or an automatic injection device.
Inject a definite amount (for instance, 10 to 50 p1) of the sample which has
been pretreated as in (3)into a n exothermic body using a micropipet, here-
after according t o 6 of JIS K 0121,dry it (100 t o 120 "C for 30 to 40 s), ash
it (500 t o 600°C for 30 to OS), atomize i t ( 4 ) (2400 t o 3 000°C for 5 to
10 s), and read the indicated value(3) at 318.4nm wavelength(5).
t o sample side, and correct the indicated value obtained on the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
centration (pgV/E) of vanadium in the sample.
lution (1pgV/ml) into as many 100 ml volumetric flasks, respectively add
acid t o make the same acid concentration as that of the sample pretreated
in (3) (a),and add water up to the marked line. Carry out the operation in
(a) on these solutions. Separately, take water for a blank test, add acid to
make the same acid concentration as that of the sample pretreated in (3)(a),
carry out the operation in (a), correct the indicated value obtained on the
standard solution, and draw the relation curve between the quantities of
vanadium (V) and indicated values. Prepare the working curve when the
sample is measured.
Notes (4) The conditions for drying, ashing, or atomizing vary depending
upon apparatus, and they may be also affected by such as in-
jected volume of a sample and concentration of coexisting salts.
417
K O101 : 1998
62.4 ICP atomic emission spectrometry After sample is pretreated, spray it

into an inductively coupled plasma through the sample introducing part, and mea-
sure the emission by vanadium at 309.311 nm wavelength to determine vanadium.
Determination range: V 20 t o 2 O00 pg/Z
(a) Vanadium standard solution (10 pgV/ml) Pipet 10 ml of vanadium stan-
dard solution (0.1 mgV/ml) in 62.1 ( i )(g) into a 100 ml volumetric flask,
(b) Mixed standard solution [(lopgCr, 10 pgV)/mll Follow 61.1.4 (1) (b).


Remarks 2 When the sample, which has been preparatorily treated, has
rich concentration of sodium, potassium, magnesium, and cal-
cium, and poor concentration in vanadium, the operation in
Remarks 7 in 51 may preferably be carried out.

Spray the sample which has been pretreated in (3)into a plasma through
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
emission strength at 309.311nm wavelength(6) (7) (a).
Take water the same amount as that of the sample which has been pre-
treated in (3)for a blank test, carry out the operations in (3)and (4)(a)
similarly t o the sample, and correct the emission strength obtained on the
sample.
centration of vanadium (pgV/Z) in the sample.
Working curve Pipet step by step 0.2 to 20 mi(9) (10) of vanadium stan-
dard solution (lOpgV/ml) into as many 100ml volumetric flasks, respec-
tively add acid t o make the same acid concentration as the sample pretreated
in (3)(a),and add water up t o the marked line. Carry out the operation in
(a)on these solutions. Separately, take water for a blank test, add acid to
carry out the operation in (a), correct the emission strength obtained on
of vanadium (V) and emission strengths. Prepare the working curve when
the sample is tested.
418
K O101 : 1998

method is applied the procedure is as follows: Take a suitable
amount of the sample, which has been treated in (3) (a), into a
ml) [Follow Note ( 8 ) of 451, add acid t o make the same acid con-
centration as the sample in (4) (a),and add water up to the marked
line. Carry out the spraying operations in (4)(a) on this solu-
tion, measure emission strength a t both 309.311nm and
371.029 nm (yttrium) wavelength, and obtain the emission-strength
ratio of vanadium and yttrium.
Separately, pipet step by step 0.2 t o 20 ml of vanadium stan-
dard solution (10 pgV/ml) into as many 100 ml volumetric flasks,
respectively add 10 ml of yttrium solution (50 ygY/ml), add acid
to make the same acid concentration as the sample in (4)(a),
and add water up t o the marked line. Carry out the spraying
operation in (4) (a)on these solutions, measure emission strength
at both 309.311 nm and 371.029 nm wavelength, draw the rela-
tion curve between emission-strength ratio of vanadium t o yt-
trium and the concentration of vanadium, and make it the working
curve. On this working curve, find the quantity of vanadium
corresponding t o the emission-strength ratio obtained on the
sample, and calculate the concentration (pgV/Z) of vanadium in
the sample.
high concentration of salts in the sample, the standard addition
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
In case of the apparatus capable of using high-order spectrum
When, after making preparatory operation according to Remarks
prepared as follows: dilute vanadium standard solution (10 pgV/
mi) t o suitable concentration (0.1 t o 1 pgV/ml), take step by step
0.1 t o 20m1, make them 500ml (or definite amount of 100 t o
500 mi), carry out the operation in Remarks 2 and (4) (a) and
(b) similarly to sample side, and draw the relation curve between
the quantities of vanadium (V) and emission strengths.
When chromium is simultaneously tested, use mixed standard
solution [( 10 pgCr, 10 pgV>/ml],and prepare preferably the working
curve under the test condition of vanadium.
419
K O101 : 1998
63 Bacterial test Bacterial tests shall be classified into the test for general bac-
teria, heterotrophic bacteria, and Escherichia coli group. This test shall be princi-
pally carried out immediately after sampling. If immediate test is impracticable,
store it a t O t o 5 "C (Do not freeze it.) in a dark place, and test it as soon as possible.
63.1 Sampling and collection of bacteria When a lot of bacteria are antici-
pated in sample water, sample using a sampling bottle or water sampler.
When the number of bacteria is anticipated t o be small, filtrate a definite amount
of sample water through a filter membrane of 0.45pm pore diameter, and collect
bacteria on the filter membrane.
(i) Instrument Instruments shall be as follows.
(a) Sampling bottle A 100 ml glass bottle with a stopper. A sampling bottle(1)
shall be sterilized by dry heat as in 63.2 (3)(a) or by autoclaving as in
63.2 (3)(b) after its neck and stoppered place have been wrapped with such
as metal foil or parchment paper. Otherwise, a 100 ml polyethylene bottle
for bacterial test which has been sterilized can be used. Be careful not to
contaminate it until being used for sampling.
(b) Water sampler Heyroth type water sampler. A sampler is set in a por-
table container and then sterilized by dry heat according t o 63.2 (3)(a). A
water sampler is exemplified in Fig. 63.1.
-7-
A: Sampling bottle
(100mi)
B: Stopper of bottle
C: Chain
D: Chain for stopper
opening
E: Chain for fastening
the holder of bottle
F: Holder of bottle
G: Portable container
H: Lid of portable con-
tainer
I: Sinker
Fig. 63.1 Example of water sampler

Note (1) In case of sample containing oxidizing material such as residual
chlorine, put 20 t o 50 mg of sodium thiosulfate pentahydrate (pow-
dered) specified in JIS K 8637 into a sampling bottle, and steril-
ize either by autoclaving mentioned in 63.2 (3)(b) o r by Oxirane
(ethylene oxide).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
420
K 0101 : 1998
(2) Operation Sampling and collection of bacteria shall be as follows.

(2.1) When a lot of bacteria are anticipated in sample water, a sample is taken in a
sampling bottle.
Sampling of surface water Surface water from such as a river o r wa-
terway shall be directly sampled in a sampling bottle.
Water in each depth Water in a definite depth shall be sampled using
a Heyroth sampler.
Sampling using a pump Sampling is carried out by pump. Sterilize an
outlet of a pump in advance, attach there a soft vinyl chloride tube previ-
ously sterilized, and after discharging sufficiently take a sample into a
sampling bottle.
Sampling from water faucet Sterilize a water faucet in accordance with
flaming in 63.2 (3) (d),open it, discharge the water sufficiently kept in piping,
and take sample into a sampling bottle.
Sampling from piping Sample in a sampling bottle similarly t o (d).
(2.2) When the number of bacteria is anticipated to be small, filtrate a definite amount
of sample water through a filter membrane of 0.45 pm pore diameter, and col-
lect bacteria on the filter membrane.
(a) When bacteria are collected from piping for such as process water where
bacteria are thought to be a few, collect directly bacteria on a filter mem-
brane according to 4.4 of JIS K 0550.
(b) When bacteria cannot be collected by the operation in (a),collect bacteria
on a filter membrane according t o Remarks 1 in 4.4 of JIS K 0550.
(c) When a sample is confined in a container, collect bacteria on a filter mem-
brane according t o Remarks 2 in 4.4 of JIS K 0550.
63.2 General bacteria General bacteria mean the living bacteria making a colony
on a culture medium when being cultivated on a standard agar culture medium at
(36+1) "C for (24+2)h, and they are counted by the number in 1 ml of a sample.
(i) Reagent and culture medium Reagents and culture media shall be as follows.
Water Water A2 or A3 specified in JIS K 0557. Use this water for prepa-
ration of reagents and culture media.
Dilution water Use physiological saline solution (Dissolve 8.5 g of sodium
chloride specified in JIS K 8150 in water t o make total 11.) o r phosphate
buffer solution (pH 7.2) [Dissolve 34 g of potassium dihydrogenphosphate
specified in JIS K 9007 in about 500 ml of water, drip sodium hydroxide
solution (i mol/Z) to adjust its pH to 7.2, and add carbon dioxide-free water
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
stated in 2 (12) (b). Pipet 1.25 ml of this solution, and add water t o make
total 11.1. Prior to use, sterilize it by autoclaving for 15 min.
Standard agar culture media(2) Put 5 g of peptone (Use hydrolysate of
casein by pancreatin.), 2.5 g of enzyme extract (powdered), 1 g of D(+)-glu-
cose specified in JIS K 8824, and 15 g of agar (powdered) specified in JIS
42 1
K O 1 0 1 : 1998
K 8263 into 1 I of water, and dissolve them by heating. Control its pH so

as t o make the pH after sterilization 7.0k0.1, transfer it into a test tube o r
flask, sterilize for about 15 min using autoclaving sterilization in (3)(b),
and store in a cold and dark place.
Note (2) Culture medium on the market is available (refer t o Remarks 1).
(2) Instrument and apparatus Instruments and apparatuses shall be as follows.

Measuring pipet 1 ml. Either wrap it in parchment paper or put its tip
innermost in a pipet sterilizer and sterilize by dry heat according to (3) (a).
Bottle for dilution This is a ground-stoppered glass bottle or a test tube
or Erlenmeyer flask with a cotton stopper whose capacity should be 2 times
o r more than that of water used. They should be sterilized by dry heat
according t o (3)(a). Previously marking a t the level of 9 ml o r 99 ml is
convenient when dilution water is put in.
Petri dish Glass made with about 90 mm in diameter and about 15 mm
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
in height. Wrap in parchment paper and sterilize in a Petri-dish sterilizer

by dry heat according to (3)(a). Otherwise use plastic sterilized Schale,
90B, specified in JIS K 0950.
Erlenmeyer flask Capacity of 300 to 500 ml and 1O00 t o 2 O00 ml. This
can be used for culture media and the preparation of dilution water, and
after being stoppered with a cotton plug, sterilize by dry heat in (3) (a) or
autoclaving in (3)(b).
Cotton plug Cotton plugs, which consist of good quality long fiber and
are not degreased, can be used for stoppers of test tubes and flasks, plugs
made of synthetic resin, metal or silicone are used instead.
Colony counter Equipped with a magnifying glass of 1.5 magnification.
Incubator Incubator specified in JIS T 1702. Capable of controlling a t
(36+1) OC.
Dry-heat sterilizer Capable of controlling from 160 t o 200 "C.
Autoclave Autoclave specified in JIS T 7322 or JIS T 7324. Capable of
heating up t o 121 OC o r higher, and of applying inside pressure as 196 kPa.
Ordinary pressure steam sterilizer Capable of heating a t 100 "C and
using under atmospheric pressure (101.325 kPa). The autoclave in (i) is
available when it is used under 101.325 kPa inside pressure and a t 100 O C .
(3) Sterilization of instruments Sterilizing of instruments shall be carried out

as follows.
(a) Dry heat sterilization This can be used to sterilize glass and metal in-
struments. The condition of sterilization is 170 "C and 1h.
(b) Autoclaving This can be used t o sterilize such as culture media, dilu-
tion water, a sampling bottle containing sodium thiosulfate pentahydrate
specified in JIS K 8637, culture media t o be disposed of. The condition is
121 O C and 30 min.
422
K O101 : 1998
(c) Steam (intermittent steam) sterilization This can be used for such as
culture medium added with molasses where autoclaving is unpractical. It
needs 101.325kPa inside pressure and successive 3 trials of 30 min at 100 "C
each (3 days). An autoclave can be used.
(d) Flame sterilization This can be used for test tubes and neck of flasks.
A test tube and flask are sterilized before and after culture operation with
being held aslant and kept in a flame and rotated for a while.
(4) Disinfection Disinfection shall be carried out as follows.
Prior to and after tests, hands, fingers and laboratory benches shall be dis-
infected. For disinfection of hands and fingers, use creosol soap water
(10 g/Z), cationic soap solution (1to 10 g/Z), or alcohol for disinfection [etha-
nol (80 vol%)].
Laboratory bench shall be disinfected by spraying cationic soap solution
(10 g/Z) o r phenol solution (30 t o 50 g/Z), or by wiping it with a cloth moist-
ened with these solutions.
The instruments such as used pipets, sampling bottles, diluting bottles should
be immersed in disinfectant solution such as creosol soap solution (30 t o
50 g/Z) for 24 h, and then wash them with water enough to remove the dis-
infectant completely.
Test tubes and Petri dishes which have been used for culture test shall be
sterilized, for every culture medium, by autoclaving sterilization in (3)(b),
and washed well with water after discarding culture media.
(5) Dilution of sample water The dilution of sample water shall be carried out
as follows.
(a) When sample water is anticipated t o contain 300 or more of general bacte-
ria in l m l , stir it sufficiently to become uniform, pipet its 1 ml with a
measuring pipet, and add it into a diluting bottle containing 9 ml o r 99 ml
of dilution water, followed by complete stirring.
(b) Next, take 1 ml, repeat the operation in (a), successively repeat these op-
erations several times, and prepare the diluted sample by which 30 to 300
colonies will be obtained after culturing.
A measuring pipet should be sterilized each time it is used.
(a) Place respectively 1 ml of a sample taken in 63.1 (2) o r diluted sample pre-
pared in ( 5 ) in two o r more Petri dishes.
(b) Melt standard agar culture medium by heating in a water bath, keep it a t
about 50 O C , add aseptically about 15 ml into each Petri dish, and before
solidifying mix them well by stirring.
(c) Spread the mixture of culture and the sample all over the dish, leave it
horizontally, after the culture is solidified cover the dish, and put them in
an incubator upside down.
(d) Culture them at ( 3 6 f l ) "C and for (24I2)h.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
423
K 0101 : 1998
(e) Count the number of colonies on the culture medium using a counter of colony,
, calculate an average t o express it with the number in 1ml of sample (each/
mi). In case of diluted sample, pick up the culture giving 30 t o 300 colo-
nies, and find the number in 1ml by multiplying it with dilution factor.
When the number of bacteria is 100 or more, round off t o get two Sig-
nificant figures.
Remarks 1 Treatment of culture media
(1) Storing of prepared culture media and aseptic test
Sterilized culture media shall be stored in a cold and dark
place with no evaporation of water. The media kept for a
long time should not be used. Prior to use, it should be
confirmed that there are no various bacteria mingled by
keeping them in an incubator for 1 night.
(2) Disposal of used culture media The culture media used
for bacteria culture shall be discarded after they were
sterilized as it was contained in a Petri dish using an
autoclave.
63.3 Heterotrophic bacteria Heterotrophic bacteria mean living bacteria which

shape colonies on culture media after culturing a t (25fl)"C and for 5 days using
standard agar culture media, and are expressed by their number in 1ml o r 1 2 of
sample water.
(1) Reagent and culture medium Reagents and culture media shall be as fol-
lows.
(b) Dilution water Follow 63.2 (1)(b).
(c) Standard agar culture media(2) Follow 63.2 (1)(c).
(2) Instrument and apparatus Instruments and apparatuses shall be as follows,

Pipet 1t o 10 ml. Either wrap it in parchment paper or put its tip inner-
most in a pipet sterilizer, and sterilize by dry heat according to (3)(a).
Measuring cylinder Capacity of 100 t o 1O00 ml. Wrap its mouth with
parchment paper, and treat it by autoclaving in (3)(b)or steam steriliza-
tion in (3)(cl.
Pincette One with smooth tips. Immediately before using, treat it with
flame sterilization as in (3)(d).
Stereomicroscope With 6 t o 80 total magnification.
Colony counter Follow 63.2 (2) (f).
Filter (Separate type) Follow 16.1 (1) (a). Wrap each part of a filter with
such as parchment paper, and sterilize them answering t o their material.
Filter membrane The organic membrane, measuring 0.45 pm pore di-
ameter and about 50 mm diameter, has printed section lines like a section
paper.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
424
K O101 : 1998
Wrap it in such as parchment paper, put in a glass Petri dish, and ster-
ilize by an autoclave of (3)(b). Otherwise use a sterilized membrane on
the market. When handling this membrane, employ a pincette.
(h) Petri dish Follow 63.2 (2)(e).
(i) Incubator Incubator specified in JIS T 1702. Capable of controlling at
(25fl) OC.
(j) Dry heat sterilizer Follow 63.2 (2)(h).
(k) Autoclave Follow 63.2 (2) (i).
(3) Sterilization of instrument Sterilizing of instruments shall be carried out

as follows.
(a) Dry heat sterilization Follow 63.2 (3)(a).
(b) Autoclaving Follow 63.2 (3)(b).
( c ) Steam (intermittent steam) sterilization Follow 63.2 (3)(e).
(d) Flame sterilization Follow 63.2 (3)(d).
(4) Disinfection Follow 63.2 (4).

(5.1) In case the number of bacteria in the sample is anticipated to be many
(a) Carry o u t the operation in 63.2 (6)(a) t o (cl.
(b) Culture it at (25+1)"C for 5 days.
(c) Count the number of colonies on the culture medium using a counter of
colony, calculate an average to express it with the number in 1ml of sample
(eacWm1). I n case of diluted sample, pick up the culture giving 30 to 300
colonies.
(d) Calculate the number of heterotrophic bacteria in 1ml of the sample ac-
cording to the following formula.
1
a=nx-
V
where, a : number of heterotrophic bacteria (eacWm1)
n : number of colonies on culture medium (each)
V : sample (ml)
When the number of bacteria is 100 or more, round off t o get two sig-
nificant figures.
(5.2) In case the number of bacteria in the sample is anticipated to be few
(a) Collect(3)bacteria on filter membrane owing t o the operation in 63.1 (2.2).
(b) Melt standard agar culture medium by heating in a water bath, and while
keeping it a t about 50 O C , take aseptically about 15 ml on a Petri dish. Let
it stand horizontally t o make it solid.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
425
K O101 : 1998
(c) Take out the filter membrane from the filter with a pincette, and make the
membrane contact closely collecting-surface upward on the Petri dish in
(b). At this time, be careful not to leave bubbles between the filter mem-
brane and culture medium.
(d) Cover the Petri dish, put it in an incubator upside down, and culture it a t
(25kl)"C for 5 days.
(e) After culturing, count the number of colonies on the membrane using a
stereomicroscope (or a counter of colony).
(f) Calculate the number of heterotrophic bacteria in 1 1 of sample according
t o the following formula.
1 O00
a=nx-
V
where, a : number of heterotrophic bacteria (each/Z)
n : number of colonies on organic filter membrane
(each)
V : sample (ml)
Note (3) Control amount of sample so as t o make the number of colonies
30 t o 300 after culturing, and therefore control preferably the
amount of filtrate step by step (for instance, 10 rnl, 100 ml and
1 O00 mi).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 2 Treat culture medium as shown in Remarks 1.
3 Instead of standard agar culture medium, M-TGE agar cul-
ture medium (M-TGE liquid culture medium) or standard liq-
uid culture medium is available,
M-TGE agar culture medium is prepared as follows.
Add 5.0 g of tryptone, 6.0 g of meat extract, 2.0 g of D(+)-
glucose specified in JIS K 8824, and 15 g of agar (powdered)
specified in JIS K 8263 in 1 I of water, heat t o dissolve them,
and control t o make the pH 7.0k0.1 after sterilization. Next,
put it into an Erlenmeyer flask, carry out autoclaving in (3)(b)
for about 15 min, and store it in a cold and dark place. When
liquid culture medium is prepared, addition of agar should be
eliminated.
The operation for test where liquid culture medium is used
shall be as follows.
Place an absorbing pad (containable 1.8 to 2.2 ml of liquid
culture medium) in every Petri dish using a pincette, and add
standard liquid culture medium o r M-TGE liquid culture
medium t o sufficiently wet the absorbing pad with standard
liquid culture medium o r M-TGE liquid culture medium (gen-
erally, 2 ml will be enough). Remove the filter membrane on
which bacteria have been collected with a pincette, and make
the membrane contact closely collecting-surface upward on the
absorbing pad in the Petri dish. At this time, be careful not
t o leave bubbles between a filter membrane and absorbing pad.
Hereafter carry out the operation in ( c ) t o (a.
426
K O101 : 1998
63.4 Escherichia coli group Escherichia coli group means aerobic (or opportu-
nistic anaerobic) bacteria which can decompose lactose to produce gas and acid, and
be judged by the presumptive test by lactose bouillon culture medium and the deter-
mination test by brilliant green lactose bile bouillon culture medium.
(i) Reagent and culture medium Reagents and culture media shall be as fol-
lows.
Water Follow 63.2 ( i )(a).
Doubly condensed lactose bouillon culture medium (doubly con-
densed LB culture medium)(2) Add 3 g of meat extract, 5 g of peptone,
5 g of lactose monohydrate specified in JIS K 8728 into 500 ml of water,
dissolve by heating, control it t o make pH 7.0IT0.2 after sterilization, and
add 12 ml of bromothymol blue solution (2 g/Z)(4). Transfer each about 10 ml
of this into Durham’s fermentation tubes (middle-sized test tube) shown in
(2) (b),sterilize by an autoclave for about 15 min as in (3)(b),immediately
dip them into cold water, and store in a dark and cold place. Do not use
Durham’s tube holding bubbles inside.
Brilliant green lactose bile bouillon culture medium (BGLB culture
medium)(2) Dissolve 10 g of peptone and 10 g of lactose monohydrate speci-
fied in JIS K 8728 in about 500 ml of water, separately dissolve 20 g of
dried cattle bile (powdered) in about 200 ml of water (pH 7.0 to 7.51, mix
them together, and add water t o make total about 970 ml, and control its
pH to 7.4. Next, add 13.3 ml of brilliant green solution ( i g/Z)(5), and add
water to make total 1 E . After filtrating through such as absorbent cotton,
pour it into Durham’s fermentation tubes (small test tube) in (2) (b)by each
3 t o 4 ml, sterilize them by an autoclave in (3)(b)for about 15 min, quickly
cool them in a cold water, and store them in a dark and cold place. Do not
use Durham’s tube holding bubbles inside.
Notes (4) Add 0.2 g of bromothymol blue specified in JIS K 8842 in about
50 ml of water, add 5 ml of sodium hydroxide solution (0.1 moll),
heat it up at about 50 “C to dissolve it, followed by cooling, and
add water t o make total 100 ml.
(5) Dissolve 0.1 g of brilliant green in water t o make total 100 ml.
Instrument and apparatus Instruments and apparatuses shall be as follows.
(a) Measuring pipet 10 ml. After being wrapped with parchment paper, put
its tip innermost into a pipet sterilizer, and carry out the dry heat sterîl-
ization according to (3)(a).
(b) Durham’s fermentation tube Put a Durham’s tube (glass tube with one
end closed), measuring about 10mm outside diameter and about 20mm
high, into a small test tube (about 11 mm diameter and about 150 mm high)
or middle-sized test tube (about 15 mm diameter and about 150 mm high)
with open end down, and stopper with a cotton plug or a cap.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
427
K O 1 0 1 : 1998
--
Cotton plug or cap
Durham's tube
Culture medium
- __ - I_ .
Fig. 63.2 Example of Durham's fermentation tube

(middle-sized tube)
(c) Platinum loop Prepare platinum or nichrome wire measuring 0.7 to 0.8 mm
diameter and about 80 mm long, make it a loop with a 2 to 5 mm inside
diameter at one end, and fix the other end to a holder. Sterilize it by a
flame as in (3)(d) before and after its use.
(d) Incubator Follow 63.2 (2)(g).
(e) Dry heat sterilizer Follow 63.2 (2)(h).
(0 Autoclave Follow 63.2 (2)(i).
(3) Sterilization of instruments Sterilization of instruments shall be carried out
as follows.
(c) Steam (intermittent steam) sterilization Follow 63.2 (3)( c ) .
(d) Flame sterilization Follow 63.2 (3)(d). The sterilization of a platinum
loop, which is for the plantation of bacteria, shall be done by red-heating
in a flame.
(5) Presumption test Presumption test shall be carried out as follows.

(a) Add respectively 10 ml of samples taken according to 63.1 with measuring
pipet into five Durham's fermentation tubes (middle-sized tube) in which
doubly condensed lactose bouillon culture medium has been put.
(b) Place them in an incubator, and culture at (36fl)"C for (24f2)h.
(c) If there is a recognized generation of gas and the colour of the culture me-
dium turns yellow, it is judged to be positive by presumption test, and then
determination test in (6)shall be carried out.
(d) If there is no recognized gas, continue the culturing on t o (48+3)h.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
428
K O101 : 1998
(e) If there is a recognized generation of gas and the colour of culture medium
turns yellow, it is judged t o be positive by presumption test, and then the
determination test in (6)shall be carried out. If there is no recognized
gas, they are judged t o be negative o n Escherichia coli group.
(6) Determination test Determination test shall be as follows.
(a) In case of presumption-test positive, plant bacterial liquid of 1 platinum
loop into a Durham's fermentation tube (small test tube) in which brilliant
green lactose bile bouillon culture medium (BGLB culture medium) has been
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
previously put.
(b) Place this in an incubator, and culture a t (36I1) "C for (2432) h or (48114) h.
(c) If there is no generation of gas, it is judged to be negative on Escherichia
coli group, and if generation of gas, it is judged t o be determination-test
positive (6).
Note (6) If the number of Durham's fermentation tubes having showed
positive is applied to the most-probable number table, the num-
ber of Escherichia coli groups can be estimated.
63.5 Fecal Escherichia coli group Fecal Escherichia coli group means the bac-
teria which, one of Escherichia coli group, generate gas or gather colony when being
cultured on EC culture medium or M-FC culture medium a t (44.510.2)"C for (24I2) h.
( i ) Reagent and culture medium Reagents and culture media shall be as fol-
lows.
(a) Water Follow 63.2 (i)(a).
(b) EC culture medium(2) Dissolve 20 g of tryptose, 5 g lactose monohydrate
specified in JIS K 8728, 1.5 g of bile acid salt (No. 31, 1.5 g of potassium
dihydrogenphosphate specified in JIS K 9007,4 g of dipotassium hydrogen-
phosphate specified in JIS K 9017,and 5 g of sodium chloride specified in
JIS K 8150 in 1 E of water, control t o make its pH 6.9 after sterilization, pour
respectively about 10 ml in Durham's fermentation tubes (middle-sized tube)
in 63.4 (2)(b), carry out autoclaving in (3)(b) for 15 to 20 min, quickly let
them cool by dipping in cold water, and store in a dark and cold place. Do
not use Durham's tube holding bubbles inside.
(c) M-FC culture medium(2) Add 10 g of tryptose, 5 g of proteose peptone
(or polypeptone), 3 g of yeast extract (powdered),5 g of lactose monohydrate
specified in JIS K 8728, 1.5 g of bile acid salt (No. 31, 5 g of sodium chlo-
ride specified in JIS K 8150, 0.1 g of aniline blue, and 15 g of agar (pow-
dered) specified in JIS K 8263 in 1 Z of water, heat it t o dissolve them,
cool it at about 45 OC, and control its pH t o 7.4. Immediately pour its 15
to 20ml in a Petri dish, and let it solidify. This culture medium is not
treated by autoclaving in (3)(b), and should be used within 96 h.
(a) Pipet Follow 63.3 (2)(a).
(b) Measuring cylinder Follow 63.3 (2)(b).
429
K O101 : 1998
(c) Pincette Follow 63.3 (2)( c ) .

(d) Stereomicroscope Follow 63.3 (2)(d).
(e) Counter of colony Follow 63.2 (2) (0.
(f) Filter (separate type) Follow 63.3 (2) (0.
(g) Filter membrane Follow 63.3 (2)(g).
(h) Petri dish Follow 63.2 (2)( c ) .
(i) Durham's fermentation tube Follow 63.4 (2) (b).
(j) Platinum loop Follow 63.4 (2)(c).
(k) Incubator Incubator specified in JIS T 1702, Capable of controlling a t
(44.51r0.2)"C.
(1) Dry heat sterilizer Follow 63.2 (2)(h).
(m) Autoclave Follow 63.2 (2)(i).
(3) Sterilization of instruments Sterilization of instruments shall be carried o u t

as follows.
(b) Autoclaving Follow 63.2 (3) (b).
(c) Steam (intermittent steam) sterilization Follow 63.2 (3)(c).
(d) Flame sterilization Follow 63.4 (3)(d).

(5.1) Case of using EC culture medium
(a) Plant an amount of a platinum loop onto EC culture medium from the
Durham's fermentation tube which has been judged positive by the pre-
sumption test mentioned in 63.4 (5)(a) t o (e).
(b) Place this in an incubator, and culture at (44.5k0.2)"C for (24k2)h.
(c) If there is recognized generation of gas, it shall be judged to be positive on
fecal Escherichia coli group, if no generation of gas, judged t o be negative
on fecal Escherichia coli group.
(5.2) Case of using M-FC culture medium
(a) Carry out the operation in 4.4 of JIS K 0550, and collect(3) bacteria on a
filter membrane. When sample water is kept in a container, use a filter
(separate type), and collect (3) on a filter membrane.
(b) Melt M-FC agar culture medium by heating in a water bath, and while
keeping it a t about 50°C, take aseptically about 15ml into a Petri dish.
Leave it horizontally to solidity it.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
430
K 0101 : 1998
( c ) Take out a filter membrane from a filter with a pincette, and make the
membrane contact closely collecting-surface upward on the Petri dish in
(b). At this time, be careful not to leave bubbles between the filter mem-
brane and culture medium.
(d) Cover the Petri dish, put it in an incubator upside down, and culture i t at
(44.5k0.2)"C for (24I2)h.
(e) When green colony is found on the filter membrane using a stereomicro-
scope (or a counter of colony), it shall be judged t o be positive on fecal
Escherichia coli group. The number of these colonies shall be the number
of fecal Escherichia coli groups.
Remarks 4 As to handling of culture media, follow Remarks 1.
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431
K O101 : 1998
64 Biological test Biological test shall be classified into special bacteria, algae
and protozoa, etc. and they shall be tested by using an optical microscope.
Moreover, if necessary, special bacteria can be tested by culture test in parallel.
Principally, carry out this test immediately after sampling, and when immediate
test is impossible, store it in a dark place, after adding formaldehyde, a t O to 5 "C
(Do not freeze it.), and carry out test as soon as possible.
64.1 Biological test For biological test, observe mainly with an optical micro-
scope, and identify its genus and species making use of materials such as an illus-
trated book on biological classification and so on.
The test about macroscopic organisms shall be contained in this test.
(a) Fixing formaldehyde solution Formaldehyde solution (formalin) specified
in JIS K 8872.
( 2 ) Instrument Instruments shall be as follows.

Sample container Wide-mouthed bottle with a stopper measuring 100
t o 500ml capacity, and made of glass or polyethylene.
Komagome pipet 1, 5, and 10ml
Measuring pipet 1, 5, and 10 ml
Measuring cylinder 10 t o 50 ml, and 100 t o 500 ml
Erlenmeyer flask Follow 63.2 (2) (d).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Dilution bottle Follow 63.2 (2) (b).
Pincette With crooked tip.
Plankton net Covered with bolting cloth of silk NXX No. 13 o r nylon NXX
No. 13. Fig. 64.1 shows its example.
Unit: cm
Tow line
approx. 500 .~
I
,
,-- Bolting Cloth
,'
Fig. 64.1 Example of plankton net
432
K O 1 0 1 : 1998
(i) Centrifugal separator With revolution 3 O00 t o 4 O00 min-1.

íj) Precipitation tube With capacity of 10 t o 30 ml, graduated by every 0.1
to 0.2m1, and tapered toward its bottom.
(k) Tube for centrifugal separator With capacity of 10 to 50m1, gradu-
ated every 1ml or 10 ml, and made of glass.
(1) Slide glass Standard type (76.0 x 26.0 mm) of quality 2 grade specified
in JIS R 3703.
(m) Slide glass with section lines Dimension 75 x 36 mm and graduated with
section lines of 1mm interval. Fig. 64.2 shows its example.
Unit: mm
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Fig. 64.2 Example of slide glass with section lines
(n) Cover glass Quality 2 grade ( 1 8 x 18mm, 2 4 x 2 4 m m , or 3 2 x 2 4 m m )
specified in JIS R 3702.
(o) Petri dish Follow 63.2 (2) ( c ) .
(p) Stereomicroscope Follow 63.3 (2) (d).
(9) Optical microscope and its accessory mirror Optical microscope, with
100 t o 1 O00 total magnification, shall consist of the following.
(i) Optical microscope Biological microscope with liquid-immersion lens
specified in JIS B 7132. To observe such as minute algae, iron bac-
teria, o r other bacteria, a phase-contrast device shall be conveniently
used.
(ii) Objective lens Liquid-immersion objective specified in JIS B 7147,
and having 90 or more nominal magnification.
(iii) Ocular Specified in JIS B 7148, and having 5 to 15 nominal mag-
nification.
(iv) Cross sample mover (mechanical stage) Attached to an optical
microscope and makes a slide glass with section lines move crosswise.
(3) Sampling Sampling shall be as follows.
(a) Sampling at water tap or faucet When number of organisms is large,
100 to 500 ml of water shall be sampled. When it is small, a definite amount
of water shall be filtered through a plankton net, and moved into a sample
container with water.
433
K O101 : 1998
Sampling from water tank, reservoir, cooling tower, etc. Sample simi-
larly to (a). Such as flock suspending in water, the attached on a wall or
construction, o r precipitate on water bottom shall be collected into a sample
container together with water making use of a Komagome pipet, pincette,
hand net or the like.
Such as swelling by rust shall be collected using a spatula into a sample
container together with water with care not t o crush the swellings.
Sampling from water catch or water channel Sample similarly t o (b).
When immediate test is impossible, add fixing formaldehyde solution t o the
sample by about 90 ml per 1I of sample water.

(a) Place a little amount of the sample taken a t (3)on a slide glass, put a cover
glass, observe it with an optical microscope, and record the species (genus)
of organisms successively from the main one. When sample gives few or-
ganisms, take a definite amount of sample in a tube for centrifuge, and
treat the sample which has been concentrated by centrifugalizing.
(b) Settle down the sample, which has been quantitatively sampled, in a pre-
cipitation tube o r measuring cylinder for a sufficient time, and read the
quantity of the precipitate.
( c ) Next, stir the sample in (b) sufficiently, take its definite amount on a slide
glass with section lines, observe it with an optical microscope similarly t o
(a), count the number of each organism, and calculate the number of each
organism in 1 ml or 1 2 of sample.
(d) In case of large size organism, transfer the sample into a Petri dish and
observe its features as t o shape or structure with a stereomicroscope.
64.2 Bacteria Bacteria shall be tested with optical microscope, and classified roughly
into zoogloea, sulfate reducing bacteria, sulfur bacteria, Sphaerotilus, iron bacteria,
fungus, and so on according t o the shape and structure of a cell, size, existence of a
sheath, precipitate of iron. Then, if necessary, carry out culture test a t the same
time.
The organisms other than iron bacteria can be generally observed in a water area
where organic contamination is getting worse, therefore they are used as an index
bacteria for contamination.
(i) Reagent and culture medium Reagents and culture media shall be as fol-
lows.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Water Follow 63.2 ( i )(a).

(b) Standard agar culture medium(1) Follow 63.2 ( i )(c).
(c) Improved ISA culture medium(1) Dissolve 1.0 g of tryptone, 5 g of so-

dium lactate (70 %), 0.5 g of sodium sulfite specified in JIS K 8061, 0.5 g
of ammonium iron (III) citrate, 2 g of magnesium sulfate heptahydrate
specified in JIS K 8995, and 0.5 g of iron (II) sulfate heptahydrate speci-
fied in JIS K 8978 in 1 2 of water, control its pH so as t o make the pH
434
K O101 : 1998
after sterilization 7.3,carry out autoclaving in (3)(b)for 15min, cool it

quickly in cold water, and submit this as soon as possible t o test.
Stork's culture medium(1) Add 1 g of D(+)-glucose specified in JIS K
8824, 0.2g of magnesium sulfate heptahydrate specified in JIS K 8995,
10 mg of iron (III) chloride hexahydrate specified in JIS K 8142, 1 g of
peptone, 50 mg of calcium chloride dihydrate specified in JIS K 8122,and
12.5 g of agar (powdered) specified in JIS K 8263 in 1 I of water, heat t o
dissolve them, control its pH so as to make the pH after sterilization 7.2,
and carry out autoclaving as in (3)(b) for 15 min.
Waksman culture medium(1) Add 5 g of peptone, 10 g of D(+)-glucose
specified in JIS K 8824, 1 g of potassium dihydrogenphosphate specified
in JIS K 9007, 0.5 g of magnesium sulfate heptahydrate specified in JIS
K 8995,and 15 g of agar (powdered) specified in JIS K 8263 in 11 of wa-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ter, heat to dissolve them, control its pH to be 3.8 to 4.0, and carry out
autoclaving in (3)(b) a t 110 "C for 15 min.
In order t o suppress the growth of bacteria, add 35 mg of rose bengal
(acid red 94)per 1I of culture medium, and when it is cultured again add
35 mg of Aureomycin per 1I of culture medium.
Czapek Dox culture medium(1) Add 30 g of sucrose specified in JIS K
8383, 3 g of sodium nitrate specified in JIS K 8562, 1 g of dipotassium
hydrogenphosphate specified in JIS K 9017, 0.5g of potassium chloride
specified in JIS K 8121, 0.5g of magnesium sulfate heptahydrate speci-
fied in JIS K 8995,0.01 g of iron (II) sulfate heptahydrate specified in JIS
K 8978,and 15 g of agar (powdered) specified in JIS K 8263 in 1 I of wa-
ter, heat it t o dissolve them, control t o make the pH 7.3 after sterilization
and carry out autoclaving in (3)(b) for 15 min.
Potato glucose agar culture medium(1) Add 4 g of potato extract, 20 g
of D(+)-glucoseSpecified in JIS R 8824,and 15 g of agar (powdered) speci-
fied in JIS K 8263 in 11 of water, heat i t to dissolve them, control its pH
so as to make the pH after sterilization 6.0 to 7.0, and carry out autoclav-
ing in (3)(b) for 15 min.
Note (1) Follow Note (2) in 63 (refer t o Remarks i).
Remarks 1 Treatment of culture medium shall conform to Remarks 1 of 63.
(2) Instrument Instruments shall be as follows.
(a) Measuring pipet Follow 64.1 (2)( e ) .
(b) Komagome pipet Follow 64.1 (2)(b).
(c) Measuring cylinder 10 to 25 ml and 50 to 500 ml
(d) Erlenmeyer flask Follow 63.2 (2)(d).
(e) Dilution bottle Follow 63.2 (2)(b).
(f) Petri dish Follow 63.2 (2)( e ) .
(g) Pincette Follow 64.1 (2)(g).
(h) Cover glass Follow 64.1 (2)(n).
435
K O101 : 1998
(i) Slide glass Follow 64.1 (2) (i).

6) Slide glass with section lines Follow 64.1 (2) m).
(k) Stereomicroscope Follow 64.1 (2) (p).
(1) Optical microscope and its accessory mirror Follow 64.1 (2) (a).
(m) Incubator Follow 63.2 (2) (g). Capable of freely controlling at 25 t o 37 "C.
(3) Sterilization of instruments Sterilization of instruments shall be carried out

as follows.
(a) Heat dry sterilization Follow 63.2 (3) (a).
(c) Steam (intermittent steam) sterilization Follow 63.2 (3) (c).
(d) Flame sterilization Follow 63.2 (3) (d).

(5.1) Zoogloea Zoogloea means a gathering of bacilli, measuring 1x 2.0 t o 4.0 pm,
which is covered with gelatinous material and forms dendritic, digitate, or in-
determinate shaped colony (size, 500 t o 1 O00 pm). In many cases, the main
constituent is of slime and flock.
Zoogloea ramigera and Zoogloea filipenduia are well known species.
(a) Take a definite amount of sample water on a slide glass with section lines,
carry out the operations similarly to 64.1 (4) (a), and observe it with an
optical microscope.
(b) Count the number of zoogloeas appearing in a field of view, and calculate
the number existing in 1 ml of a sample.
(5.2) Sulfate reducing bacteria Sulfate reducing bacteria, measuring 0.5 to 1.0 x 2.5
to 5.0 pm, are curved bacillary o r spiroid individuals giving active movement.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(a) Take a definite amount of sample water o n a slide glass with section lines,
carry out the operation similarly t o 64.1 (4) (a),and observe it with an optical
microscope.
(b) Making reference to such as an illustrated book o n biology, record the spe-
cies and number of sulfate reducing bacteria appeared in a field of view as
follows(2).
+ appear very rarely (appearance rate, 10 % or less)
++ appear rarely (appearance rate, 20 to 30 %)
+++ appear commonly (appearance rate, 40 t o 60 %)
++++ appear frequently (appearance rate, 70 t o 80 %)
+++++ appear very frequently (appearance rate, 90 % o r more)
436
K O101 : 1998
When culturing is needed, put the improved ISA culture medium(3) into a
test tube with a stopper (by about half the volume of the test tube), add a
suitable amount of sample water, moreover add improved ISA culture medium
t o fill it up, and stopper it with care not to leave bubbles. Then, culture
anaerobically it at 30 "C for 5 t o 7 days(*).
When the bottom or whole(3) of the culture medium changes black, it shall
be judged to be positive on sulfate reducing bacteria.
Notes (2) Classify sulfate reducing bacteria found, and in every section line
on the slide glass, and count their number. Repeat this trial
three times, and calculate the number and species of sulfate
reducing bacteria in 1ml of sample according t o the following
formula.
10
A =(al+ +u~)x-
3n
where, A : number of zoogloeas (number of units/ml)
al, az, u3 : number of zoogloeas counted at each trial
n : concentrating factor of sample
(3) When preparing improved ISA culture medium in ( l ) ( c ) ,it is
allowable t o test with improved ISA agar culture medium which
is prepared by adding 1 5 g of agar (powdered) specified in JIS
K 8263. In this case, when the culturing, which has been done
according to 63.2 (6) (a)t o ( c ) , gives black colony, the number of
bacteria can be counted.
(4) An anaerobic jar is advisable.
Remarks 2 Sulfate reducing bacteria live in water or slime which has
become anaerobic owing to organic contamination, and is ob-
ligate anaerobic bacteria producing hydrogen sulfide by reducing
sulfate. They are often found in industry water, and some-
times cause the corrosion of metals. DesuZfouibrio genus is
often observed.
(5.3) Sphaerotilus Sphaerotilus makes a filiform body composed of linearly aligned
several cylindrical bacilli, one of which measures 1x 2.0 t o 6.0 pm, in a trans-
parent sheath. This filiform body is seriously featured by its pseudo-ramifica-
tion.
carry out the operation similarly t o 64.1 (4) (a),and observe it with an optical
microscope.
(b) Making reference to such as an illustrated book on biology, record the spe-
cies and number of Sphaerotilus appeared in a field of view similarly t o
(5.2) (bN5).
(c) When culturing is needed, making use of Stork's culture medium or stan-
dard agar culture medium, culture it a t 25 "C for 5 days according t o the
operations in 63.2 (6) (a) to ( c ) .
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
437
K O101 : 1998
(d) The colony generated is observed with an optical microscope, and if trans-
parent sheath is confirmed, Sphaerotilus is judged t o be positive.
Note (5) Count the number of classified Sphaerotilus found in every area
between two section lines of slide glass, repeat three times this
counting, and count the species and number of Sphaerotilus in
1ml of sample according t o the formula in Note ( 2 ) .
Remarks 3 Sphaerotilus is commonly called sewage bacteria or water cotton,
and in many cases it makes slime o r flock in industrial water
o r water containing a lot of organic substances. Sphaerotilus
natans is typical one.
(5.4) Iron bacteria Iron bacteria have brown colored bodies showing characteris-
tic shape, and the species having ribbon shape, measuring 1 t o 1.5 pm in width,
is called Gallionella ferruginea. The species forming a line composed of sev-
eral long bacilli in a long straight sheath, measuring about 2 pm in width, is
called Leptothrix ochracea. These two are typical iron bacteria.
carry out the operation similarly to 64.1 (4) (a),and observe it with an optical
microscope.
cies and number of iron bacteria appeared in a field of view similarly t o
(5.2) (b)(9.
Note (6) Count the number of classified iron bacteria found in every area
between two section lines of slide glass, repeat three times this
counting, and calculate the species and number of iron bacteria
in 1ml of sample according to the formula in Note ( 2 ) .
Remarks 4 Iron bacteria live in such as underground water, subsoil wa-
ter, and spring water, and oxide iron (II) to iron (III), which
makes water red and forms slime.
(5.5) Sulfur bacteria Sulfur bacteria, whose cell measures 3 x 2.5 t o 5 pm, forms
long filiform body without ramification. This filiform body makes whitish grey
thin film (cobweb like). The shape of this body resembles that of Sphaerotilus
or Oscillatoria (blue-green algae), it can be discriminated by sulfur particles
stored in its filiform body.
carry out the operation similarly to 64.1 (4) (a),and observe it with an optical
microscope.
cies and number of sulfur bacteria appeared in a field of view similarly to
(5.2) (b)(7).
Note (7) Count the number of classified sulfur bacteria found in every area
between two section lines of the slide glass, repeat three times
this counting, and calculate the species and number of sulfur
bacteria in 1 ml of sample according t o the formula in Note ( 2 ) .
Remarks 5 The typical species giving some obstacles in industrial water
is Beggiatoa alba.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
438
K O101 : 1998
(5.6) Fungus When testing fungus, culture a sample in potato glucose agar cul-
ture medium, Waksman agar culture medium, or Czapek Dox agar culture me-
dium, produce spores, observe this together with mycelia, conidiophore, and
their coadunation condition with an optical microscope, and determine its ge-
nus and species making reference of an illustrated book on classification of
organisms .
(a) Take a suitable amount of sample water prepared in 64.1 (3),and carry
out the operations in 63.2 (6) (a) t o (c). In this case, instead of standard
agar culture medium, use such as potato glucose agar culture medium,
Waksman agar culture medium, or Czapek Dox agar culture medium.
(b) Culture it a t 20 to 25 "C for 5 t o 7 days.
(c) Cotton-like colony which is colored peculiarly to fungus grow on culture
medium. Observe with an optical microscope the shape of spores, mycelia,
conidiophores, and so on, and record its genus, species, and number, by
making use of an illustrated book on classification of organisms similarly
t o (5.2) (b)(8).
Note (8) Count the number of classified funguses found in every area be-
tween two section lines on the slide glass, repeat this counting
three times, and calculate the species and number of funguses in
1 ml of sample according to the formula in Note (2).
Remarks 6 Because there are so many kinds of funguses and they live in
every circumstances not only in water, when making culture,
it should be done with the greatest possible care to prevent
the culture from the contamination by circumstances.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Funguses are roughly classified into trichobacteria, Phyco-

mycetes, Ascomycetes, Zygomycetes and Deuteromycetes, and
many of them do not generate spores in water, therefore the
observation of their mycelia in the view of an optical micro-
scope cannot give the determination of species. Accordingly,
the culture test in agar culture medium is required.
64.3 Algae In order to test algae, observe them using an optical microscope, clas-
sify them into Chlorophyta, phycochrome, diatom, Rhodophyta, and so on making
reference to an illustrated book on algae, and determine their genus and species.
(i) Instrument and apparatus Instruments and apparatuses shall be as follows.

(a) Komagome pipet Follow 64.1 (2)(b).
(b) Centrifugal separator Follow 64.1 (2) (i).
(c) Tube for centrifugal separator Follow 64.1 (2) (k).
(d) Cover glass Follow 64.1 (2) (n).
(e) Slide glass Follow 64.1 (2) (i).
(f) Slide glass with section lines Follow 64.1 (2) (m).
(g) Optical microscope and its accessory mirror Follow 64.1 (2) (9).
439
K O101 : 1998

(a) Carry out the operations in 64.1 (4) (a), and observe the sample using an
optical microscope.
(b) Observe the size, shape and arrangement of cells and the colour tone and
shape of chromatophore, and determine the genus and species of algae
appeared in the view of the microscope making use of such as an illustrated
book on algae, and record their number as in (5.2) (b).
(c) When sample water is quantitatively taken, calculate the number of algae
found in 1ml or 1 1 of a sample(9).
Note (9) Count the number of classified algae found in every area between
two section lines on the slide glass, repeat this counting three times,
and calculate the number of algae in 1ml or 1 1 of the sample
according t o the formula in Note (2).
64.4 Animal The size of animals in water has a very wide range from microscopic
to macroscopic, therefore observe carefully their features using an optical microscope,
and determine their genus and species making use of an illustrated book on organ-
isms and so on.
(a) Komagome pipet Follow 64.1 (2) (b).
(b) Centrifugal separator Follow 64.1 (2) (i).
(c) Tube for centrifugal separator Follow 64.1 (2) (k).
(d) Cover glass Follow 64.1 (2) (n).
(e) Slide glass with section lines Follow 64.1 (2) (m).
(f) Optical microscope and its accessory mirror Follow 64.1 (2)(9).
(a) Stir enough the sample which has been treated as specified in 64.1 (4) (a)
and (b),take a definite amount (for instance, 0.1 mi) of it on a slide glass
with section lines, put on a cover glass (24x 32 mm), and observe it using
an optical microscope with 200 t o 400 total magnification.
(b) Record the species (genus) of animals and its number appearing in the view
according t o (5.2) (b),and calculate the species (genus) and number of each
animal in 1ml o r 1 1 of a sample(10).
Note (10) Count the number of classified animals found in every area be-
tween two section lines on the slide glass, repeat this counting
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
three times, and calculate the number of each animal in 1ml of

sample according t o the formula in Note (2).
440
K O101 : 1998
Annex (informative) Supplement
This Annex is only a supplement of the matter concerning the specifications in

this Standard, written in the corresponding form to the standard, therefore this is
not a part of the standard itself.
I Transparentness This is the degree indicating the transparence of sample water

and shall be tested as follows: place sample water in a transparence meter, see it
through from its top, measure the depth by which double cross on a sign board put
on the bottom can be clearly seen, and express it by the degree where one degree
means 10mm.
Measurable range: 1 to 30 degrees
(a) Transparence meter It is shown in Informative reference Fig. 1. This
is made of glass, with an outlet at the bottom, equipped with graduations
at every 5 mm from the level of a sign board to 50 mm height, and every
10 mm from 50 mm to 300 mm. Put a sign board at the bottom as shown
in Informative reference Fig. 2.
Fill the transparence meter with the sample water already agitated, see
the meter through from its top, and flow out the sample water from the
bottom outlet until double cross on the sign board can be seen(1), followed
by reading graduation of liquid level.
Repeat the operation two o r three times, obtain average of readings, and
make it transparentness with degree.
Note (1) When the sample has a lot of suspensoid, it often precipitates on
the bottom, which may give an error, therefore attention will be
needed.
Remarks : Even when using the same illuminance, the light with differ-
ent saturation may gives different transparentness. Light source
shall be principally daylight and direct sunlight will not be used.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
441
K O 1 0 1 : 1998
-.A -._
c,/
A: Cylinder with outlet at its
,- Make a notch bottom
here. B: Blackboard for screening
CI t o CB: Holding frame for cylinder
D: Sign board
E: Base
F: Rubber tube with pinchcock
Informative reference Fig. 1 Transparence meter
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
442
K O101 : 1998
Unît: mm
Cylinder with outlet
at its bottom
Sign board
A Width of black line 0.5
White plastic or
porcelain board
Detail of side view

--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Informative reference Fig. 2 Detail of transparence meter
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
443
K O 1 0 1 : 1998
II Oxygen demand by alkaline potassium permanganate ( CODOH) Alkalize

a sample, add potassium permanganate as oxidizing agent, react them in boiling water
for 20 min, obtain the quantity of potassium permanganate consumed for the reac-
tion, and express it by the amount (mgO/Z) of oxygen corresponding to it.
Carry out this test immediately after sampling, if immediate test is impossible,
store it according to 3.3 in the text, and carry out the test as soon as possible.
(1) Reagent Reagents shall be as follows.
Water Follow 17 (i)(a)in the text.

Sodium hydroxide (100 gll) Follow 22.2.1 ( i )(b) in the text.
Sulfuric acid (2+1) Take 1 volume of water, and add gradually, while
cooling and stirring, 2 volume of sulfuric acid specified in JIS K 8951.
Sodium azide solution (40 gil) Dissolve 4 g of sodium azide specified in
JIS K 9501 in water to make total 100ml.
Starch solution (10 gll) Follow 22.1.2 ( i )(i) in the text.
Potassium iodide solution (100 gll) Dissolve 10 g of potassium iodide
specified in JIS K 8913 in water t o make total 100 ml. Prepare this solu-
tion when it is used.
Potassium permanganate solution (2 mmolll) Take 0.32 g of potassium
permanganate specified in JIS K 8247 into a flat bottomed flask, and add
1050 t o 1100 ml water t o dissolve it. Boil this gently for 1 to 2 h, and let
it stand for about 16 h.
Filtrate the supernatant through a glass filter G4. (Do not wash i t with
water before and after filtration.) Place the solution in the coloured bottle
which was steam-washed for about 30 min for storing.
10 mmoYZ Sodium thiosulfate solution Follow 28.3 (i)(e) in the text.
(a) Erlenmeyer flask with a stopper 200 ml
(b) Water bath Follow 17(2)(a)in the text.
(a) Take a suitable amount(2) of sample water(1) in a 200 ml Erlenmeyer flask
with a stopper, add water t o make total 50 ml, and add 1ml of sodium
hydroxide solution (100 g/Z).
(b) Add 10 ml of potassium permanganate solution (2mmol/Z), stir them, im-
mediately put it into a boiling water bath, and heat it for 20 min, While
heating, keep the flask so as to make the level of sample surface in the
flask lower than water level in the bath and not t o let the flask touch the
bottom of the bath.
(c) Take it out from the bath, after cooling it t o room temperature with cold
water add 1ml of sodium azide solution (40 g/Z), and stir to mix them.
444
K O 1 0 1 : 1998
Add 1 ml of pottasium iodide solution (100 g/Z) and 0.5 ml of sulfuric acid
(2+1)(3),stopper it, stir it, and let it be left in a dark place for about 5 min.
Titrate isolated iodine with 10 mmoVZ sodium thiosulfate solution, when
the solution becomes pale yellow add 1ml of starch solution (10 g/Z) as
indicator, and titrate on until the blue coloured by iodide disappears.
Separately, take 50 ml of water into a 200 ml Erlenmeyer flask with a stopper,
add 1ml of sodium hydroxide solution (100 gíZ), and carry out the opera-
tions in (b)t o (e).
Calculate CODOH(mgOlZ) in accordance with the following formula.
CODOH = ( b - a ) x f X- 'Ooo x 0.08

V
where, CODOH: oxygen demand by alkaline potassium per-
manganate solution (mgOlt)
a : 10 mmolíl sodium thiosulfate solution needed
for titration (ml)
b : 10 mmol/Z sodium thiosulfate solution needed
a t the test using water (mi)
f: factor(4) of 10 mmoVZ sodium thiosulfate so-
lution
0.08 : oxygen equivalent t o 1ml of 10 mmol/Z so-
dium thiosulfate solution (mg)
V : sample (mi)
When suspensoid is contained, after making it uniform by suffi-
cient stirring, sample it.
This should be the amount of potassium permanganate solution
(2 mmol/Z) of which nearly half will be left after heating for 20 min.
However, in case where oxygen demand by alkaline potassium
permanganate is 8 mgOlZ o r less, it should be 50 ml.
The suitable amount of sample water shall be decided by the
preparatory test carried out in (3).
When rough estimation of CODOHis known, the following for-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
mula gives the suitable amount (Vml) of the sample.

1 O00 x 0.08
v=5x
CODoHestimation of the sample (mgO/Z)
where, V : sampling amount of sample (ml)
5 : reaction estimation amount of potassium perman-
ganate solution (2 mmol/Z) (mi)
0.08 : oxygen equivalent to 1 ml of potassium perman-
ganate solution (2mmol/Z) (mg)
If iron is contained, add 1 m l of potassium fluoride solution
(300 g/Z) before adding sulfuric acid (2+1).
Use the factor of 0.1 mol/Z sodium thiosulfate solution given in
22.1.2 (i)(d) of the text.
445
K O101 : 1998
III Cation surface-active agent Cation surface-active agents are classified into
such as aliphatic amine salts, quaternary ammonium compounds, and alkyl pyridium
salts. For the determination of cation surface-active agent, orange II absorptiometry
is applied, but in ordinary water a direct determination cannot be carried out be-
cause cation surface-active agent makes a stable ion pair with anion surface-active
agent. Therefore, a t first pretreat a sample t o remove anion surface-active agent
(separation by ion exchange), and then apply orange II absorptiometry.
Orange II absorptiometry Extract the ion pair with chloroform which was pro-
duced by the reaction of cation surface-active agent and anionic orange II [sodium
4-(2-hydroxy-l-naphthalenyl)azobenzenesulfonatel, measure its absorbance, and express
it as tetradecyldimethylbenzylammonium chloride.
Determination range: cation surface-active agent,
[ C H ~ ( C H ~ ) I ~ C H ~ N ( C H ~ ) 20
~ CtH
o~ C ~pg
350 H~C~]
Water Water A3 Specified in JIS K 0557.
Sodium chloride Specified in JIS K 8150.
Acetic acid-sodium acetate buffer solution (pH 3.5) Dissolve 6 ml of
acetic acid specified in JIS K 8355 and 0.9 g of sodium acetate trihydrate
specified in JIS K 8371 in water to make total 11.
Orange II solution Dissolve 0.1 g of orange II [sodium 442-hydroxy-l-
naphthalenyl) azobenzenesulfonate pentahydrate] in water to make total
100 ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Methanol solution (50 vol%) Prepare using methanol specified in JIS
K 8891.
Strongly alkaline anion-exchange resin (I type) Follow 23.2.1 (1) (k)
in the text. When using, wash it 3 t o 4 times with methanol specified in
JIS K 8891 and then 3 t o 4 times with water.
Cation surface-activeagent standard solution [0.1 mgCH3(CHd&H2N
(CH3)2CH2CsH&l/ndI Weigh O. 100 g of tetradecyldimethylbenzylammonium
chloride on the base of 100 % pure, dissolve it in water, transfer into a 1 O00 ml
volumetric flask, and add water up to the marked line, Prepare this when
it is used.
Cation surface-active agent standard solution [lo ~ ~ C H ~ ( C H ~ ) I ~ C H ~ N
(CH&CH2C6H~Cl/mll Pipet 20 ml of cation surface-active agent standard
solution [O. 1 ~ ~ C H ~ ( C H ~ ) I ~ C H ~ N ( C H ~ ) ~ Ca H200
into ~ Cml~ H ~C~/~~]
volu-
metric flask, and add water up t o the marked line. Prepare this when it is
used.
(2) Instrument and apparatus Instrument and apparatuses shall be as follows.

(a) Cylindrical drip funnel 100ml
446
K O101 : 1998
(b) Separatory funnel 200 ml

(c) Anion-exchange-resincolumn Put glass wool specified in JIS K 8251
at lower place of a glass pipe with a cock, measuring 12 mm in inside di-
ameter and about 300 mm in length, pack about 25 ml of strongly basic anion-
exchange resin (I type) while mixing with water so as not t o leave bubbles
inside, then put glass wool a t upper end of the pipe, and control water level
to be about 10 mm above from the glass wool, Make this an anion-exchange-
resin column. Set a 100 ml cylindrical drip funnel at its upper end for dripping
solution.
This anion-exchange-resin column can be used during about 20 cycles.
However, it cannot be reused after regeneration.
(3) Pretreatment Pretreatment shall be as follows.
(a) Place 100 ml of sample water (1) [containing 20 t o 350 pg as C H ~ ( C H ~ ) I ~ C H ~ N
(CH&CH2C6H&l] in the cylindrical drip funnel set at upper end of the an-
ion-exchange-resin column, flow it down into the anion-exchange-resin col-
umn a t the flow rate of about 1ml/min(2), and receive the effluent in a
300 ml Erlenmeyer flask.
(b) Stop this flowing just before the sample in the cylindrical drip funnel set at
upper end of the anion-exchange-resincolumn runs out, add 50 ml of methanol
) the cylindrical drip funnel, again flow down at the flow rate
( 5 0 ~ 0 1 %into
of about 1ml/min(2), put this effluent together with the above 300 ml Er-
lenmeyer flask.
Notes (1) When the sample is acidic, neutralize it with sodium hydroxide
solution (40gl), and when alkaline, with hydrochloric acid (l+ll).
(2) Keep the level of the solution about 10 mm higher than the glass
wool a t the upper end of the anion-exchange-resin column.
Transfer the effluent obtained a t (3)(b) into a separatory funnel and add
30 ml of acetic acid-sodium acetate buffer solution (pH 3.5), 1.5 g of sodium
chloride, and 10 ml of orange II solution into the effluent, and stir them.
Add 10 ml of chloroform, stir them violently for 3 min, and let it stand.
Transfer chloroform layer into a 20 ml measuring cylinder (with a stopper).
Add 10 ml of chloroform into water layer, again stir them violently for 3 min,
and let it stand.
Put this chloroform layer in the 20 ml measuring cylinder (with a stopper)
at (c), and add chloroform up t o a 20 ml level mark.
Add about 3 g of sodium sulfate, and stir them for dehydration.
Place a part of chloroform layer into a n absorption cell and measure absor-
bance in the vicinity of 485 nm wavelength with making chloroform a ref-
erence solution.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
447
K O101 : 1998
(h) Take 100 ml of water and 50 ml of methanol (50 vol%) in a separatory fun-
nel for a blank test, carry out the operations in (a)t o ( g ) , measure its ab-
sorbance, and correct the absorbance obtained on the sample.
(i) Find the quantity of cation surface-active agent on the working curve, and
calculate the concentration of cat-
ion surface-active agent.
Working curve Pipet step by step from 2 to 35 ml of cation surface-ac-
tive-agent standard solution [ 10 ~ ~ C H ~ ( C H ~ ) I ~ C H ~ N ( C H ~ ) ~into
CH~C~H~C
as many separatory funnels, and respectively add water to make them to-
tal 100 ml, add 50 ml of methanol (50 vol%), carry out the operations in (a)
to (h),and draw the relation curve between the quantities of cation sur-
face-active agent [CH3(CH2)i2CH2N(CH3)2CH2CsH5CI] and absorbances.
Remarks: Even when anion surface-active agent exists 100 times more
than cation surface-active agent, separation can be done. The
coexistence of sulfate ion, nitrate ion, carbonate ion, o r phos-
phate ion does not disturb the determination.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
448
K O101 : 1998
IV Ion-selective electrode method for iodide ion Control pH to be about 5 by

adding acetic acid buffer solution, and measure an electrode potential using an iodide
ion-selective electrode making as an indicator electrode t o determine iodide ion.
Determination range: I- 0.1 t o 1O00 mg/Z
Acetic acid buffer solution (pH 5) Follow 32.4 (i)(a) in the text.
Iodide ion standard solution (1 O00 mgI-ll) Follow 33.1 ( i )(f) in the
text.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Iodide ion standard solution (100 mgI-/Z) Pipet 20 ml of iodide ion stan-
dard solution (i O00 mgI-/Z) into a 200 ml volumetric flask, and add water
up to the marked line. Prepare this when it is used.
Iodide ion standard solution (10 mgI-/Z) Pipet 20 ml of iodide ion stan-
dard solution (100 mgI-/Z) into a 200 ml volumetric flask, and add water up
t o the marked line. Prepare this when it is used.
Iodide ion standard solution ( i mgI-/Z) Pipet 20 ml of iodide ion stan-
dard solution (10 mgI-/Z) into a 200 ml volumetric flask, and add water up
t o the marked line. Prepare this when it is used.
Iodide ion standard solution (0.1 mgI-/C) Pipet 20 ml of iodide ion stan-
dard solution (i mgI-/Z) into a 200 ml volumetric flask, and add water up
to the marked line. Prepare this when it is used.
(a) Potentiometer Follow 31.2 (2) (a) in the text.
(b) Indicator electrode Iodide ion-selective electrode.
(c) Reference electrode Follow 32.4(2)(c) in the text,
(d) Magnetic stirrer Follow 31.2 (2) (d) in the text.
(3) Preparation of working curve Working curve shall be prepared as follows.

(a) Take 100 ml of iodide ion standard solution (0.1 mgI-lZ) in a 200 ml beaker,
add 10 mi(') of acetic acid buffer solution (pH 5).
(b) Immerse an indicator electrode(2) ( 3 ) and reference electrode(4) (5) in this
solution, and stir it with a magnetic stirrer(6) strongly enough not t o make
bubbles touch the electrodes(7).
( c ) Measure temperature of solution, and measure electric potential by a po-
tentiometer ( 8 ) .
(d) Take 100 ml of iodide ion standard solution ( i mgI-/Z), 100 ml of iodide
ion standard solution (10 mgI-/Z), 100 ml of iodide ion standard solution
(100 mgI-/E), and 100 ml of iodide ion standard solution (iO00 mgI-lZ) into
200 ml beakers respectively, and add 10 ml(1) of acetic acid buffer solution
(pH 5). Adjust temperature of each iodide ion standard solution to become
the temperature at ( c ) + l OC, carry out the operations in (b)and (c), and
449
K O101 : 1998
measure the electric potentials given by iodide ion standard solutions (ito
1O00 mgI-ll) (8).
(e) Graduate concentrations of iodide ion on logarithmic axis of semilogarithm
graph paper and potentials on uniform axis, and draw the relation curve
between the concentrations (mgI-/Z) of iodide ion (I-)and electric potentials (9).
Notes (1) Adding acetic acid buffer solution is t o control pH to 5 when mea-
suring, and t o make ionic strength uniform.
(2) An iodide ion-selective electrode should be used for measurement
of electric potential after a pointer gets stability when immers-
ing it in iodide ion standard solution (0.1 mgI-lZ).
(3) Follow Note (12) in 31 in the text.
(8) The response time of an iodide ion-selective electrode is 2 to 3 min
a t 10 t o 30 "C solution temperature under 0.1 to 1mgI-lZ con-
centration of iodide ion, and 1min o r longer under 10 mgI-ll or
more.
(9) The potential difference between iodide ion standard solution
(0.1 mgI-lZ) and iodide ion standard solution (10 mgI-ll) falls in
110 to 120 mV (25 O C ) , and the working curve given from 0.1 t o
1O00 mgll concentration of iodide ion makes straight line.
(a) Take 100 ml of sample(l0) in a beaker, add 10 ml acetic acid buffer solu-
tion (pH 51, and adjust the temperature of solution t o the temperature at
(3)(cl* 1OC.
(b) After the operations in (3)(b)and ( c ) , find the concentration of iodide ion
on the working curve, and calculate the concentration (mgI-lZ) of iodide ion
in the sample.
Note (10) When adding acetic acid buffer solution (pH 5) is not effective t o
make the solution pH 5 because of acidity o r alkalinity of solu-
tion, add previously acetic acid (1+2) or sodium hydroxide solu-
tion (100 glZ) t o make the solution pH 5, and then add water t o
get definite volume.
Remarks 1 In case of an ionic-concentrationmeter, carry out the operations
in (3)(a)to ( c ) using iodide ion standard solution (i mgI-lZ and
100 mgI-lZ), and adjust an ionic-concentration meter t o point
1mgI-/Z and 100 mgI-ll. Furthermore, confirm the indicated
value shown on the ionic concentration meter by using iodide
ion standard solution (0.1 mgI-/l and 10 mgI-ll) and iodide ion
standard solution (1O00 mgI-/Z).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
450
K O 1 0 1 : 1998
2 Sulfide ion shall be tested as follows: in advance add zinc acetate

solution (100 g/Z), filtrate the precipitate through filter paper
5 grade C, and use the filtrate for test.
The allowance limits of main coexisting materials are ex-
pressed as follows with the maximum ratio.
F-, Cl-, Br-, Nos-, Sod2-, Cos2- : lo4
po43-:103
s 2 0 3 2 - : 10
CN-: 10-1
3 Potentiometric titration by an ion-selective electrode
Take 100 ml of sample water in a beaker, adjust its pH to be
7, use an indicator electrode (iodide ion-selective electrode or
silver ion-selective electrode), titrate it with 10 to 100 mmol/Z
silver nitrate solution while measuring potential according to
the operations in (3)(b), draw a titration curve, and find the
end point of titration. Inflection point of titration curve shows
the order of iodide ion, bromide ion, and chloride ion. Making
use of the inflection point, find the end point, and calculate
the amount of each ion.
One milliliter of 10 mmol/Z silver nitrate solution is equivalent
to 1.269 mg of I-, 0.799 mg of Br-, and 0.355 mg of Cl-.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
451
K O 1 0 1 : 1998
V Ion-selective electrode method for bromide ion Control pH to be about 5

by adding acetic acid buffer solution, and measure an electrode potential using a
bromide ion-selective electrode making as an indicator electrode to determine bro-
mide ion.
Determine range: Br- 0.5 t o 1O00 mgll
(i) Reagent Reagents shall be as follows.
Acetic acid buffer solution (pH 5) Follow 32.4 (i)(a) in the text.
Bromide ion standard solution (iO 0 0 mgBr-ll) Heat potassium bro-
mide specified in JIS K 8506 at 110 "C for 4 h, and let it cool in a desicca-
tor. Dissolve its 1.49 g in water, transfer it in a 1O00 ml volumetric flask,
Bromide ion standard solution (100 mgBr-ll) Pipet 20 ml of bromide
ion standard solution (i O00 mgBr-ll) in a 200 ml volumetric flask, and add
Bromide ion standard solution (10 mgBr-ll) Pipet 20 ml of bromide
ion standard solution (100 mgBr-ll) in a 200 ml volumetric flask, and add
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Bromide ion standard solution ( imgBr-ll) Pipet 20 ml of bromide ion
standard solution (10 mgBr-ll) in a 200 ml volumetric flask, and add water
up t o the marked line.
Bromide ion standard solution (0.5 mgBr-/@ Pipet 10 ml of bromide
ion standard solution (10 mgBr-ll) in a 200 ml volumetric flask, and add
water up t o the marked line.

(a) Potentiometer Follow 31.2 (2) (a)in the text.
(b) Indicator electrode Bromide ion-selective electrode
(c) Reference electrode Follow 32.4(2)(c)in the text.

as follows.
(a) Take 100 ml of bromide ion standard solution (0.5 mgBr-ll) in a 200 ml beaker,
add 10 ml(1) of acetic acid buffer solution (pH 5).
(b) Immerse an indicator electrode (bromide ion-selective electrode)(2) (3) and
reference electrode(4) (5) in this solution, and stir it with a magnetic stir-
rer(6) strongly enough not t o make bubbles touch the electrodes(').
(c) Measure temperature of solution, and measure electric potential by a po-
tentiometer ( 8 ) .
(d) Take 100 ml of bromide ion standard solution (i mgBr-ll), 100 ml of bro-
mide ion standard solution (10 mgBr-A), 100 ml of bromide ion standard
452
K O101 : 1998
solution (100 mgBr-/E), and 100 ml of bromide ion standard solution

(1O00 mgBr-/E) into respectively 200 ml beakers, and add 10 ml(1) of acetic
acid buffer solution (pH 5). Adjust temperature of each bromide ion stan-
dard solution t o the temperature a t ( c ) I l O C , carry out the operations in
(b) and ( c ) , and measure the electric potentials given by bromide ion stan-
dard solutions (i t o 1 O00 mgBr-/Z)(S).
(e) Graduate concentrations of bromide ion on logarithmic axis of semilogarithm
between the concentrations (mgBr-/Z) of bromide ion and electric potentials (9).
Adding acetic acid buffer solution is t o control pH to 5 when mea-
suring, and t o make ionic strength uniform.
A bromide ion-selective electrode should be used for measure-
ment of electric potential after a pointer gets stability when
immersing it in bromide ion standard solution (0.5 mgBr-4).
Follow Note (12) in 31 in the text.
The response time of a bromide ion-selective electrode is about
1min a t 10 t o 30 "C solution temperature under 0.5 mg/Z con-
centration of bromide ion.
The potential difference between bromide ion standard solution
1mgBr-/Z and 100 mgBr-lZ falls in 110 to 120 mV (25 O C ) , and the
working curve given from 0.5 t o 1O00 mgll concentration of bro-
mide ion makes straight line.

(a) Take 100 ml of sample water(l0) in a beaker, add 10 ml acetic acid buffer
solution (pH 5 ) , and adjust the temperature of solution t o the temperature
a t (3)( c ) + i"C.
(b) After the operations in (3)(b)and ( c ) , find the concentration of bromide
ion on the working curve, and calculate the concentration (mgBrlE) of bro-
mide ion,
Note (10) When adding acetic acid buffer solution (pH 5 ) is not effective t o
make the solution pH 5 because of acidity or alkalinity of solu-
tion, add previously acetic acid (1+2) or sodium hydroxide solu-
tion (100 g/E) t o make the solution pH 5 , and then add water to
get 100 ml.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Remarks 1 In case of an ionic-concentrationmeter, carry out the operations

in (3)(a)t o ( c ) using bromide ion standard solution (imgBr-/Z
and 100 mgBr-/E), and adjust an ionic-concentration meter to
point 1.0 mgBr-/Z and 100 mgBr-/Z. Furthermore, confirm the
indicated value shown on the ionic-concentrationmeter by using
bromide ion standard solution (0.5 mgBr-/Z and 10 mgBr-/E) and
bromide ion standard solution (1O00 mgBr-/E).
433
K 0101 : 1998
Sulfide ion o r cyanide ion gives disturbance, therefore these

should be eliminated beforehand. Sulfide ion shoula be fixed
by adding zinc acetate buffer solution (100 g / l ) , be filtrated
through filter paper 5 grade C, and filtrate should be used.
The allowance limits of main coexisting material- are ex-
pressed as follows with the maximum ratio.
F-, NO^-, sop: 104
c1- : l o 2
I- :
Potentiometric t i t r a t i o n by an ion-selective electrode
Follow Remarks 3 of Informative reference VI.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
454
K O101 : 1998
VI Ion-selective electrode method for nitrate ion Add phosphate buffer so-
lution into a sample to control its pH t o 6.8, and measure electric potential using a
nitrate ion-selective electrode making as a n indicator electrode to determine nitrate
ion.
Determination range: Nos- 0.5 to 1 O00 mgll
Phosphate buffer solution (pH 6.8) Dissolve 17 g of potassium dihydrogen
phosphate specified in JIS K 9007 and 17.8 g of disodium hydrogen-phos-
phate specified in JIS K 9020 in water to make total 11.
Nitrate ion standard solution (1 O00 mgNOs-lZ) Follow 37.2.3 ( i )(i) in
the text.
Nitrate ion standard solution (100 mgNO3-lZ) Take 20 ml of nitrate ion
standard solution (1 O00 mgNOs-ll) into a 200 ml volumetric flask, and add
water up to the marked line. Prepare this when it is needed.
Nitrate ion standard solution (10 mgNOs-lZ) Take 20 ml of nitrate ion
standard solution (100 mgNO3-ll) into a 200 ml volumetric flask, and add
water up t o the marked line. Prepare this when it is needed.
Nitrate ion standard solution (imgNOs-lZ) Take 20 ml of nitrate ion
standard solution (10 mgNOs-lZ) into a 200 ml volumetric flask, and add water
up the marked line. Prepare this when it is needed.
Nitrate ion standard solution (0.6 mgNOs-lZ) Take 10 ml of nitrate i o n
standard solution (10 mgNO3-lZ) into a 200 ml volumetric flask, and add water
up to the marked line. Prepare this when it is needed.

(b) Indicator electrode Nitrate ion-selective electrode
(c) Reference electrode Follow 32.4(2)(c)in the text. The liquid in a n
outer cylinder should be potassium sulfate solution (0.25 molli).

as follows.
(a) Take 100 ml of nitrate ion standard solution (0.5 mgNO3-lZ) in a 200 ml beaker,
add 5 ml(1) of phosphate buffer solution (pH 6.8).
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
(b) Immerse an indicator electrode ( 2 ) (3) and reference electrode (4) (6) in this
solution, and stir it with a magnetic stirrer(6) strongly enough not to make
bubbles touch the electrodes (7).
(c) Measure temperature of the solution, and measure electric potential by a
potentiometer (8).
455
K 0101 : 1998
(d) Take 100 ml of nitrate ion standard solution (i mgNO3-1'0, 100 ml of nitrate
ion standard solution (10 mgNOs-lZ), 100 ml of nitrate ion standard solution
(100 rngNO~-lZ),and 100 ml of nitrate ion standard solution (iO00 mgN03-/Z)
into 200 ml beakers respectively, and add 5 ml of phosphate buffer solution
(pH 6.8).
Adjust temperature of each nitrate standard solution t o the temperature
at ( c ) + iOC, and carry out the operations in (b) and (c), and measure the
electric potentials of nitrate ion standard solutions (1to 1O00 mgNOs-lZ).
(e) Graduate concentrations of nitrate ion on logarithmic axis of semilogarithm
between the concentrations (mgNO3-lZ) of nitrate ion and electric potentials (9).
Notes (1) Adding phosphate buffer solution (pH 6.8) is t o control pH when
measuring, and t o make ionic strength uniform.
(2) A nitrate ion-selective electrode should be used for measurement
of electric potential after a pointer gets stability when immers-
ing it in nitrate ion standard solution (0.5 mgNOa-lZ).
(5) Use potassium chloride solution (3 mol/Z t o saturated solution)
for the liquid of an inside cylinder of a reference electrode and
potassium sulfate solution (0.25mollZ) for the liquid of an out-
side cylinder. When saturated potassium chloride solution is used
for the liquid of inside cylinder, crystal of potassium chloride will
deposite and cling on the electrode because of lowering of solu-
tion temperature, which results in increase of resistance.
(8) Response time of a nitrate ion-selective electrode is about 3 min
at 10 to 30 O C solution temperature under 0.5 to 10 mgNO3-lZ of
nitrate ion concentration, and about 30 s under 10 mgNOg-ll or
higher.
(9) The potential difference between nitrate ion standard solution
1mgNO3-lZ and 100 mgNOs-/Z falls in 110 to 120 mV (25 "C), and
the working curve given from 0.5 mgNO3-ll t o 1O00 mgNO3-lZ
concentration of nitrate ion makes straight line.
(a) Take lOOml(10) of sample water into a 200ml beaker, add 5 m l of phos-
phate buffer solution (pH 6.8), and adjust the solution temperature to the
temperature a t (3)(c)+ 1 "C.
(b) Carry out the operations in (3)(b)and ( c ) , find the concentration of ni-
trate ion on the working curve, and calculate the concentration (mgNOs-lZ)
of nitrate ion in the sample.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
456
K O101 : 1998
Note (10) When adding phosphate buffer solution is not effective to make
the solution pH 6.8 because of acidity or alkalinity of the solu-
tion, add previously sulfuric acid (1+5) or sodium hydroxide so-
lution (40 glZ) into a suitable amount of sample water to neutralize
it, and make it a definite volume.
Remarks 1 In case of an ionic-concentration meter, carry out the opera-
tions in (3)(a)to (c) using nitrate ion standard solution (ito
100 mgNOs-lZ), adjust an ionic-concentration meter to point
1mgNO3-lZ and 100 mgNO3-lZ. Furthermore, confirm the in-
dicated value shown on the ionic-concentration meter by us-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
ing nitrate ion standard solution (0.5 mgNO3-lZ and 10 mgNOa-/

I) and nitrate ion standard solution ( i O00 mgNOs-lZ).
2 The allowance limits of main coexisting materials are expressed
as follows with the maximum ratio.
so42-, 3.3 x 104 HC03-, CN- : 1x lo2
HzP04- : 2 x 1 0 4 Noz-: 25
F-: 1 . 6 104~ Br-: 7.7
po43-: i x 104 c103- : 4 x 10-l
cos2-: 5~ 103 I-: 5 x
CH3COO- : 2.5 x lo3 cio4-: i x 10-3
Cl-: 2 x lo2
457
K 0101 : 1998
VI1 Ion-selective electrode method for sulfide ion Add sodium hydroxide buffer
solution into a sample to make it strong alkaline about 13 of pH, and measure elec-
tric potential using a sulfide ion-selective electrode making as an indicator electrode
t o determine sulfide ion.
Determination range: S2- 0.1 t o 100 mg/Z
Sodium hydroxide buffer solution (pH 13) Dissolve 40 g of sodium hy-
droxide specified in JIS K 8576 in about 400 ml of water, after cooling it
add 10 g of L(+)-ascorbicacid specified in JIS K 9502 and 9.3 g of dihydrogen
disodium ethylenediaminetetraacetate dihydrate specified in JIS K 8107,
dissolve them, further add 500 ml of glycerol specified in JIS K 8295, and
add water to make total 11.
Sulfide ion standard solution (100 mgS2-lZ) Pipet 20 ml of sulfide ion
standard solution ( iO00 mgS2-/Z)stated in 40.1 (1)(e)in the text into a 200 ml
volumetric flask, add 20 ml(1) (2) of sodium hydroxide buffer solution (pH 131,
and add water up to the marked line. Prepare this when it is used. Cal-
culate the concentration of this solution from the concentration of the sul-
fide ion standard solution (i O00 mgS2-/Z)in 40.1 ( i )(e) in the text.
Sulfide ion standard solution (10 mgS2-lZ) Pipet 20 ml of sulfide ion
standard solution (100 mgS2-/Z)into a 200 ml volumetric flask, add 20 ml(1) (2)
of sodium hydroxide buffer solution (pH 13), and add water up to the marked
line. Prepare this when it is used. Calculate the concentration of this solution
from the concentration of the sulfide ion standard solution (1O00 mgS2-lZ)
in 40.1 (i)(e) in the text.
Sulfide ion standard solution (i mgS2-/Z) Pipet 20 ml of sulfide ion stan-
dard solution (10 mgS2-/Z)into a 200 ml volumetric flask, add 20 ml(1) (2) of
sodium hydroxide buffer solution (pH 13), and add water up t o the marked
from the concentration of the sulfide ion standard solution (iO00 mgS2-/Z)
in 40.1 ( i )(e) in the text.
Sulfide ion standard solution (0.1mgS2-/Z) Pipet 20 ml of sulfide ion
standard solution (1mgS2-/Z)into a 200 ml volumetric flask, add 20 ml(1) (2)
of sodium hydroxide buffer solution (pH 131, and add water up to the marked
from the concentration of the sulfide ion standard solution ( i O00 mgS2-/Z)
in 40.1 (i)(e) in the text.
Notes (1) Sodium hydroxide buffer solution (pH 13) should be added enough
t o let 100ml of sulfide ion standard solution contain 10ml of
sodium hydroxide buffer solution (pH 13).
(2) During measurement, alkalinity must be kept a definite value,
12 of pH or more.

--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
458
K O101 : 1998
(b) Indicator electrode Sulfide ion-selective electrode (3).

(c) Reference electrode Follow 32.4(2)(c) in the text.
Note (3) Sulfide ion-selective electrode shall be the fixed membrane elec-
trode made of silver sulfide.

as follows.
Take 100 ml of sulfide ion standard solution (0.1 mgS2-/Z)in a 200 ml bea-
ker, and add 10 ml of water(4).
Immerse an indicator electrode (5) (6) and reference electrode (7) (ß) in this
solution, and stir it with a magnetic stirrer(9) strongly enough not t o make
bubbles touch the electrodes(10).
Measure the temperature of the liquid, and measure its potential using a
potentiometer (11).
Take 100 ml of sulfide ion standard solution ( i mgS2-/Z), 100 ml of sulfide
ion standard solution (10 mgS2-/Z),and 100 ml of sulfide ion standard solu-
tion (100 mgS2-/Z)respectively into 200 ml beakers, and add 10 mi(4) of water.
Adjust the temperature of each sulfide ion standard solution to the tem-
perature a t ( c ) I l O C , carry out the operations in (b)and ( c ) , and measure
the potential of each sulfide ion standard solution (i to 100 mgS2-/Z).
Take a semilogarithm paper, graduate the concentration of sulfide ions o n
a logarithm axis and potentials on a uniform axis, and complete the rela-
tion curve between concentrations of sulfide ion (mgS2-/E)and potentials (12).
When measuring sample, 10 ml of sodium hydroxide buffer solu-
tion (pH 13) is added into the sample per 100 ml of the sample,
therefore 10 ml of water is added to meet this quantity of solution.
Immerse a sulfide ion-selective electrode in sulfide ion standard
solution (0.1 mgS2-/Z)when it is used, and measure its potentials
after its indicated value is stabilized.
If the sensitive membrane of an indicator electrode (ion-selec-
tive electrode of sulfide ion) is dirtied, the potential gradient of
working curve becomes small, and response speed also becomes
slow. The dirt of a sensitive membrane should be removed by
circular polishing, holding the sensitive membrane vertically, with
fine sand paper (about ## 1000) and a few drops of water, and
then wipe it with soft paper, then wash with water.
Follow Note (14) in 3 1 in the text.
Response time is about 3 min for sulfide ion standard solution
(0.1 mgS2-/Z), 1 t o 2 min for that of 1 mgS2-/Z, and about 1min
for that of 100 mgS2-/E.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
459
K O101 : 1998
(12) Sulfide ion standard solution (0.1 mgS2-/Z) gives about -680 mV
and sulfide ion standard solution (100 mgS2-lZ) gives about
-770 mV, and the working curve in this interval becomes linear.

(a) Take 100 ml of a sample in a 200 ml beaker, add 10 ml of sodium hydrox-
ide buffer solution (pH 131, and control the temperature of the solution to
the temperature a t (3)( c l I l O C .
(b) After the operations in (3)(b)and ( e ) , find the quantity of sulfide ion from
the working curve, and calculate the concentration of sulfide ion (mgS2-/Z)
in the sample.
Remarks 1 Ion-selective electrode method cannot determine the sulfide
staying in suspensoid.
2 The allowance limits of main coexisting materials are expressed
as follows with the maximum ratio.
Cl-, Br-, I-, Nos- : lo4
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
460
K O101 : 1998
VI11 Barium-sulfate turbidimetry for sulfate ion Add stabilizer and barium
chloride in a sample, and let it react to sulfate ion under a definite condition t o make
barium sulfate, and determine it making use of turbidimetry.
Determination range: so42- 1 to 5 mg
Repeatability: 10 % by coefficient of variation
(a) Glycerol solution (l+l) Prepare using glycerol specified in JIS K 8295.
(b) Sodium chloride solution Dissolve 240 g of sodium chloride specified
in JIS K 8150 into a mixture of 20 ml of hydrochloric acid specified in JIS
K 8180 and water t o make total 11.
(c) Barium chloride Shift barium chloride dihydrate specified in JIS K 8155
t o catch the part that goes through 710 pm opening and is stopped on 500 pm
opening.
(d) Sulfate ion standard solution ( i mgS0d2-/ml) Follow 42.1 (1) (f) in the
text.
(2) Apparatus Apparatuses shall be as follows.
Take two suitable amounts (containing each 1 to 5 mg as S042-)of filtered
sample(1) into two 100 ml conical beakers, and add water t o make each 50 ml
respectively.
Add 10 ml of glycerol solution (1+1)respectively and 5 ml of sodium chlo-
ride solution(2), and stir them with a magnetic stirrer.
Add 0.3 g of barium chloride into one of them(?, stir them for 1 min, leave
them for 4 min, and stir them again for 15 s.
Immediately remove these solutions into absorption cells, and measure ab-
sorbance(3) in the vicinity of 450 nm wavelength within 1min making the
solution without barium chloride a reference solution.
Take two 50 ml waters into 2 conical beakers as a blank test, carry out the
operations in (b) to (d),and correct the absorbance obtained on the sample.
concentration of sulfate ion (mgS042-/Z)in the sample.
Working curve Pipet step by step from 1 t o 5 ml of sulfate ion standard
solution (i mgS042-/Z),add water t o make them up t o 50 ml respectively,
carry out the operations in (b) t o (e),and draw the relation curve between
the quantities of sulfate ion (sod2-)and absorbance.
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
Notes (1) When sample shows acidity or alkalinity, adjust its pH to about 7.
(2) I n this time, pH becomes 1.4 t o 1.6.
(3) This is an apparent absorbance.
461
K O101 : 1998
Attached Table 1 Normative references
JIS B 7132 Biological microscopes for immersion objectives

JIS B 7147 Biological microscope objectives
JIS B 7148 Microscope eyepieces
JIS B 7411 Solid-stem general purpose liquid-in-glass thermometers
JIS C Direct acting indicating analogue electrical measuring
1102-1
instruments and their accessories Part 1 :Definitions
and general requirements common to all parts
JIS C 1102-2 Direct acting indicating analogue electrical measuring
instruments and their accessories Part 2 : Special re-
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
quirements for ammeters and voltmeters
JIS K O010 Reference materials-Standard solution-Copper
JIS K O011 Zinc standard solution
JIS K 0012 Cadmium standard solution
JIS K 0013 Nickel standard solution
JIS K 0015 Lead standard solution
JIS K 0016 Iron standard solution
JIS K 0018 Reference material-pH Standard solution-Oxalate
JIS K O019 Reference material-pH Standard solution-Phthalate
JIS K 0020 Reference material-pH Standard solution-Equimolal
phosphate
JIS K O021 Reference material-pH Standard solution-Tetraborate
JIS K O022 Reference material-pH Standard solution-Carbonate
JIS K 0024 Reference material-Standard solut ion-Chromium
JIS K 0026 Reference material-Standard solution-Arsenic
JIS K 0027 Reference material-Standard solution-Manganese
JIS K 0028 Sulfate ion standard solution
JIS K O029 Chloride ion standard solution
JIS K 0030 Fluoride ion standard solution
JIS K 0031 Nitrate ion standard solution
JIS K 0032 Nitrite ion standard solution
JIS K 0033 Reference materials-Standard solution-Phosphate ion
JIS K 0034 Reference materials-Standard solution-Ammonium
ion
JIS K 0050 General rules for chemical analysis
JIS K 0094 Sampling methods for industrial water and industria1
wastewater
JIS K O102 Testing methods for industrial wastewater
JIS K 0114 General rules for gas chromatographic analysis
JIS K 0115 General rules for molecular absorptiometric analysis
462
K O101 : 1998
Attached Table 1 (continued)

~~
JIS K 0116 General rules for atomic emission spectrometry

JIS K 0117 General rules for infrared spectrophotometric analysis
JIS K 0121 General rules for atomic absorption spectrochemical
analysis
JIS K O122 General rules for ion selective electrode method
JIS K 0127 General rules for ion chromatographic analysis
JIS K O211 Technical terms for analytical chemistry (general part)
JIS K 0215 Technical terms for analytical chemistry (analytical
instrument part)
JIS K 0550 Testing methods for detection and estimation of micro-
biological contaminants in highly purified water
JIS K 0552 Testing methods for electric conductivity in highly puri-
fied water
JIS K 0557 Water used for industrial water and wastewater analy-
sis
JIS K 0805 Continuous total organic carbon analyzer
JIS K 0950 Sterilized plastic petri dishes
JIS K 0970 Piston operated micro-volumetric apparatus
JIS K 1101 Oxygen
JIS K 1105 Argon
JIS K 1107 High purity nitrogen
JIS K 2251 Crude petroleum and petroleum products-Sampling
JIS K 8005 Reference materials for volumetric analysis
JIS K 8012 Zinc
JIS K 8013 Zinc powder
JIS K 8019 Sodium nitrite
JIS K 8032 Acetonitrile
JIS K 8034 Acetone
JIS K 8044 Diarsenic trioxide
JIS K 8046 Sodium metaarsenite
JIS K 8048 4-Amino antipyrine
JIS K 8051 3-Methyl-1-butanol
JIS K 8059 Sodium hydrogensulfite
JIS K 8061 Sodium sulfite
JIS K 8069 A l uminum
JIS K 8085 Ammonia solution
JIS K 8101 Ethanol (99.5)
JIS K 8102 Ethanol (95)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
463
K O101 : 1998
JIS K 8103 3iethyl ether

JIS K 8107 3isodium dihydrogen ethylenediamine tetraacetate
iihydrate
JIS K 8116 Qmmonium chloride
JIS K 8121 Potassium chloride
JIS K 8122 Jalcium chloride dihydrate
JIS K 8123 Talcium chloride
JIS K 8124 Jalcium chloride (for drying)
JIS K 8129 Zobalt (II) chloride hexahydrate
JIS K 8132 Strontium chloride
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
JIS K 8136 Tin (II) chloride dihydrate
JIS K 8139 Mercury (II) chloride
JIS K 8142 ìron (III) chloride hexahydrate
JIS K 8150 Sodium chloride
JIS K 8155 Barium chloride dihydrate
JIS K 8159 Magnesium chloride hexahydrate
JIS K 8163 Potassium hexachloroplatinate
JIS K 8180 Hydrochloric acid
JIS K 8197 N-1-Naphthylethylenediaminedihydrochloride
JIS K 8201 Hydroxylammonium chloride
JIS K 8202 I , 1 O-Phenanthrol inium chloride monohydrate
JIS K 8223 Perchloric acid
JIS K 8228 Magnesium perchlorate
JIS K 8230 Hydrogen peroxide
JIS K 8247 Potassium permanganate
JIS K 8249 Potassium periodate
JIS K 8252 Ammonium peroxodisulfate
JIS K 8253 Potassium peroxodisulfate
JIS K 8255 Aluminium potassium sulfate 12-water
JIS K 8263 Agar
JIS K 8267 Sodium formate
JIS K 8271 Xylene
JIS K 8272 Xylene cyano1 FF
JIS K 8283 Citric acid monohydrate
JIS K 8284 Diammonium hydrogen citrate
JIS K 8288 Trisodium citrate dihydrate
JIS K 8289 Cupferron
464
K O101 : 1998
JIS K 8291 Xycine

JIS K 8295 Xycerol
JIS K 8306 ,-Cresol
JIS K 8312 Dotassium chromate
JIS K 8318 $odium p-toluenesulfonchloramide trihydrate
JIS K 8322 7hloroform
JIS K 8355 4cetic acid
JIS K 8356 Zinc acetate dihydrate
JIS K 8359 4mmonium acetate
JIS K 8361 Ethyl acetate
JIS K 8371 Sodium acetate trihydrate
JIS K 8374 Lead (II) acetate trihydrate
JIS K 8377 Butyl acetate
JIS K 8383 Sucrose
JIS K 8410 Calcium oxide
JIS K 8432 Magnesium oxide
JIS K 8443 Potassium cyanide
JIS K 8454 Sodium N,N-diethyldithiocarbamate trihydrate
JIS K 8465 1,2-Dichloroethane
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
JIS K 8474 Potassium trihydrogen dioxalate dihydrate

JIS K 8488 1,5-diphenyEcarbonohydrazide
JIS K 8489 s-Diphenylcarbazone
JIS K 8490 Dithizone
JIS K 8491 2,6-Dibromo-N-chloro-p-benzoquinone monoimine
JIS K 8495 p-Dimethylaminobenzylidene rhodanine
JIS K 8498 Dimethylglyoxime
JIS K 8500 N,N-Dimethylformamide
JIS K 8501 Sodium disu lfite
JIS K 8506 Potassium bromide
JIS K 8514 Sodium bromide
JIS K 8517 Potassium dichromate
JIS K 8519 Oxalic acid dihydrate
JIS K 8522 Potassium oxalate monohydrate
JIS K 8528 Sodium oxalate
JIS K 8529 Bromine
JIS K 8530 Potassium bromate
JIS K 8532 L(+)-Tartaric acid
465
K O 1 0 1 : 1998
Attached Table ,1 (continued)
JIS K 8533 Sis[(+)-tartratoldiantimonate (III) dipotassium

.rihydrate
JIS K 8536 Potassium sodium (+)-tartrate tetrahydrate
JIS K 8541 Vitric acid
JIS K 8544 4luminium nitrate enneahydrate
JIS K 8545 4mmonium nitrate
JIS K 8548 Potassium nitrate
JIS K 8550 Silver nitrate
JIS K 8552 2obalt (II) nitrate hexahydrate
JIS K 8558 ‘Mercury (II) nitrate n-hydrate
JIS K 8562 Sodium nitrate
JIS K 8563 Lead (II) nitrate
JIS K 8568 Pia nganese (II) nit rat e hexahydrate
JIS K 8574 Potassium hydroxide
JIS K 8576 Sodium hydroxide
JIS K 8580 T’in
JIS K 8586 Sulfanilic acid
JIS K 8588 Ammonium amidosulfate
JIS K 8603 Soda lime
JIS K 8617 Calcium carbonate
JIS K 8622 Sodium hydrogen carbonate
JIS K 8625 Sodium carbonate
JIS K 8635 Thiourea
JIS K 8637 Sodium thiosulfate pentahydrate
JIS K 8646 Dextrin hydrate
JIS K 8653 Devarda’s alloy
JIS K 8659 Starch, soluble
JIS K 8660 Copper
JIS K 8669 3,3’-Dimethylbenzidinium dichloride
JIS K 8680 To1uene
JIS K 8701 Lead
JIS K 8721 p-Nitrophenol
JIS K 8722 Sodium pentacyanonitrosylferrate (III) L ‘rate
JIS K 8728 Lactose monohydrate
JIS K 8731 Urea
JIS K 8736 Eriochrome black T
JIS K 8747 A m m o n i u m vanadate W)
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
466
K O101 :1998
JIS K 8775 9-Quinolinol

JIS K 8776 2-Hydroxy-1-(2-hydroxy-4-sulfo-1-naphthylazo)-3-
uaphthoic acid
JIS K 8780 Pyrogall o1
JIS K 8783 Potassium pyrosulfate
JIS K 8785 Sodium diphosphate decahydrate
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
JIS K 8789 1 , l O-Phenanthroline monohydrate
JIS K 8798 Phenol
JIS K 8799 Phenolp ht ha1ein
JIS K 8801 Potassium hexacyanoferrate (III)
JIS K 8809 Potassium hydrogen phthalate
JIS K 8810 1 -Butanol
JIS K 8819 Hydrofluoric acid
JIS K 8824 D(+)-glucose
JIS K 8826 Sodium hydroxide for nitrogen compounds analysis
JIS K 8830 Uranine
JIS K 8832 Brucine n-hydrate
JIS K 8839 2-Propanol
JIS K 8840 Bromocresol green
JIS K 8842 Bromothymol blue
JIS K 8844 Bromophenol blue
JIS K 8847 Hexamethylenetetramine
JIS K 8848 Hexane
JIS K 8858 Benze ne
JIS K 8863 Boric acid
JIS K 8866 Sodium tetraborate decahydrate
JIS K 8872 Formaldehyde solution
JIS K 8885 Silicon dioxide
JIS K 8889 Metacresol purple
JIS K 8891 Methanol
JIS K 8893 Methyl orange
JIS K 8896 Methyl red
JIS K 8897 Methylene blue
JIS K 8900 2-butanone
JIS K 8903 4-Methyl-2-pentanone
JIS K 8905 Hexaammonium heptamolybdate tetrahydrate
JIS K 8913 Potassium iodide
467
K O101 : 1998
JIS K 8920 lodine

J I S K 8949 Sodium sulfide nonahydrate
JIS K 8951 Sulfuric acid
J I S K 8953 Zinc sulfate heptahydrate
JIS K 8960 Ammonium sulfate
JIS K 8962 Potassium sulfate
JIS K 8965 Silver sulfate
JIS K 8972 Potassium hydrogen sulfate
JIS K 8978 Iron (II) sulfate heptahydrate
JIS K 8979 Ammonium iron (II) sulfate hexahydrate
JIS K 8980 Mercury (II) sulfate
JIS K 8982 Ammonium iron (III) sulfate 12-water
JIS K 8983 Copper (II) sulfate pentahydrate
JIS K 8987 Sodium sulfate
JIS K 8990 Ammonium nickel (II) sulfate hexahydrate
JIS K 8992 Hydrazi ni um s u1fate
JIS K 8995 Magnesi u m sulfate heptahydrate
JIS K 8997 Manganese (II) sulfate pentahydrate
JIS K 9000 Ammonium thiocyanate
JIS K 9003 Liquid paraffin
JIS K 9005 Phosphoric acid
JIS K 9007 Potassium dihydrogenphosphate
JIS K 9009 Sodium dihydrogenphosphate dihydrate
JIS K 9016 Diammonium hydrogenphosphate
JIS K 9017 Dipotassium hydrogenphosphate
JIS K 9019 Disodium hydrogenphosphate 12-water
JIS K 9020 Disodium hydrogenphosphate
JIS K 9047 L-Glutamic acid
JIS K 9062 Nickel
JIS K 9066 Sulfanilamide
JIS K 9501 Sodium azide
JIS K 9502 L(+)-Ascorbic acid
JIS K 9512 Silver N,N-diethyldithiocarbamate
JIS K 9519 Mercury (II) thiocyanate
JIS K 9520 1,1,2,2-Tetrachloroethane
JIS K 9525 p-Hydrazinobenzenesulfonicacid hemihydrate
JIS K 9547 Phenylfluorone
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---
468.
K O101 : 1998
Attached Table 1 (concluded)
JIS K 9548 3-Methyl-I -phenyl-5-pyrazolone

JIS K 9550 Polyvinyl alcohol
JIS K 9569 N-Benzoyl-N-phenylhydroxylamine
JIS K 9703 2,2,4-Trimethylpentane
JIS K 9704 2-Amino-2-hydroxymethyl-1,3-propanediol
JIS K 9901 Highly purified nitric acid
JIS K 9902 Highly purified hydrochloric acid
JIS K 9904 Highly purified perchloric acid
JIS K 9905 Highly purified sulfuric acid
JIS P 3801 Filter paper (for chemical analysis)
JIS R 1301 Porcelain crucibles for chemical analysis
JIS R 1302 Porcelain basins for chemical analysis
JIS R 3503 Glass apparatus for chemical analysis
JIS R 3505 Volumetric glassware
JIS R 3702 Cover glasses for microscopes
JIS R 3703 Slide glasses for microscope
JIS T 1702 Incubator
JIS T 7322 High-pressure steam sterilizers for medical use
JIS T 7324 High-pressure steam sterilizers for medical use (small
size)
JIS T 9107 Surgical rubber gloves
JIS Z 0701 Silicagel desiccants for packaging
JIS Z 8719 Metamerism index-Evaluation method of degree of
metamerism for change i n illuminant
JIS Z 8801 Test sieves
JIS Z 8802 Methods for determination of p H of aqueous solutions
--`,``````,`,`,,,`,,`,,````,`-`-`,,`,,`,`,,`---

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ICs: 13.060.

Testing methods for industrial water

Japanese Standards Association

1-24, Akasaka 4, Minato-ku Tokyo 107-8440 Japan

Page 261 line 1 1

Error: 20 ml of sulfuric acid(1+1)

JAPANESE INDUSTRIAL STANDARD

Page 11, line 17

In Notes( 7), replace “--.andsuspended matters’’ by “*--or

Error V :sample used for dilution (mi)

Page 32, line 4

Page 59, line 9

Page 68, line 19

Page 94, line 6

Page 254, line 3

Error Put 10 ml of sample (containing 2 to 50 pg as SO:-) in a precipitation tube for

Page 255, line 10

42.2( 3 )( a ) is changed as follows:

Error Put 10 ml of sample (containing 50 t,o 500 pg as SO,2-) in a precipitation tube

Page 268, line 1

Error chloride ion

Page 268, line 4

2 Common items ................................................................................................... 1

4 Pretreatment of sample .................................................................................... 8

4.4 Decomposition with nitric acid and sulfuric acid ..................................... 10

5 Marking of results ............................................................................................. 11

8 Odour and threshold odour number (TON) .................................................. 15

12 Electric conductivity ......................................................................................... 42

13 Acid consumption .............................................................................................. 47

14 Alkali consumption ........................................................................................... 50

18 Oxygen demand by potassium dichromate (CODcr) .................................... 67

19 Biochemical oxygen demand (BOD) ............................................................... 70

20 Organic carbon (TOC) ....................................................................................... 79

21 Total oxygen demand (TOD) ........................................................................... 85

22 Phenols and p-cresols ....................................................................................... 88

23 Surface active agents ........................................................................................ 97

24 Dissolved oxygen ............................................................................................... 112

25 Total carbonate .................................................................................................. 125

26 Hexane extracts ................................................................................................. 132

27 Missing number ................................................................................................. 137

28 Residual chlorine ............................................................................................... 138

28.2 Diethyl-p-phenylenediamine (DPD) colorimetric method ....................... 141

29 Required amount of chlorine ........................................................................... 152

30 Hydroxide ion (OH-) .......................................................................................... 154

31 Fluorine compounds .......................................................................................... 155

Chloride ion (Cl-) ...............................................................................................

33 Iodide ion (I-) ..................................................................................................... 174

34 Bromide ion (Br-) ............................................................................................... 179

35 Cyanide compounds ........................................................................................... 183

36 Ammonium ion (NH4+) ...................................................................................... 196

36.2 Indophenol Blue absorptiometry .................................................................. 199

37 Nitrite ion (Noz-) and nitrate ion (NOS-) ...................................................... 210

37.2.1 Reducing distillation-Indophenol Blue absorptiometry ........................ 213

38 Organic nitrogen ................................................................................................ 225

39 Total nitrogen .................................................................................................... 229

41 Sulfite ion so^^-) .............................................................................................. 249

43 Phosphorus compound and total phosphorus ............................................... 259

44 Silica (SiOs) ........................................................................................................ 275

44.1 Ionic silica ........................................................................................................ 275

45 Boron (B) ............................................................................................................. 284

46 Arsenic (As) ........................................................................................................ 289

47 Sodium (Na) ........................................................................................................ 300

48 Potassium (K) ..................................................................................................... 306