Wat. Res. Vol. 30, No. 11, pp.

2771-2775, 1996
~ Pergamon Copyright © 1996ElsevierScienceLtd
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SPECTROPHOTOMETRIC DETERMINATION OF WATER

COLOUR IN HAZEN UNITS

D A G HONGVE* and G U N V O R AKESSON

National Institute of Public Health, P.O. Box 4404 Torshov, N-0403 Oslo, Norway

(First received May 1995; accepted in revisedform May 1996)

Abstract--Colour is a useful index of dissolved humic substances in water. The reference method for
colour determination is a visual comparison of samples with reference solutions of hexachloroplatinate,
the Hazen method. The precision of this method is poor in relation to common drinking water criteria.
A spectrophotometric method with better precision that gives results on the same scale as the Hazen
method is described in this paper. Filtration is crucial before spectrophotometric measurement. We found
no effect of filtration on true colour. The method has been validated through interlaboratory comparisons.
Forty laboratories using this method obtained the same mean values as 20 laboratories using visual
judgement. The standard deviations of spectrophotometric measurements were half the values of visual
determinations. Copyright © 1996 Elsevier Science Ltd

Key words--colour, drinking water, spectrophotometer, interlaboratory comparison

INTRODUCTION (Thienemann, 1925; Ohle, 1934) are not much used

Colour is a property of natural and conducted water
that is perceivable to most people without any kind
of instrumentation. Colour per se represents an
aesthetic problem in water supplies and it shows the
presence of organic components that may be
potentially harmful in connection with water
chlorination. Colour is a useful index of dissolved
humic matter (Thurman, 1983) and is often used as
such in scientific literature. It is therefore essential to
have methods for objective description of the visual
impression of colour. A simple method for assess-
ment of water colour is visual comparison with a
fixed scale. Reference solutions made of hexachloro-
platinate and cobalt ions in hydrochloric acid (Pt-Co
solutions) mimic the hues of most natural waters
(Hazen, 1892). Such comparison is one of the oldest
methods in common use within water science. The
colour value is given as the concentration of Pt in the
matching reference solution. 1 rag/1 Pt = 1 TCU
(true colour unit) (World Health Organization, 1984).
Modern methods for measurement of optical
properties and chemical determination of the
coloured components have replaced the traditional
colour determination in many connections. However,
the visual judgement still finds its use both because of
its simplicity and because some proposed instrumen-
tal methods have given ambiguous results. Most
countries have established criteria for colour in
drinking water based on the Hazen method. Similar
methods that are based on other reference solutions
*Author to whom all correspondence should be addressed.
today. The Hazen method is adopted as reference
method by organisations that set standards for water
quality examination, like International Organization
for Standardization and the European Committee for
Standardization (1994), World Health Organization
(1984) and American Public Health Association
(1992). In daily practice most laboratories omit use
of reference solutions and compare instead with
calibrated coloured glass discs. This modification is
also included in the standard method.
The visual assessment is not a very precise method.
Most operators find it difficult to distinguish between
colours that differ with < 5 TCU. The standard
deviation in interlaboratory comparisons is usually
near 5 TCU. According to usual drinking water
standards, for example in the European Union (EU)
and the USA, this uncertainty represents 25-33% of
maximum permitted colour. The method therefore
does not fulfil the EU requirement for precision
better than 10%. In order to make more precise
measurements, various photometric methods have
been used. A major obstacle for these has been to
match apparatus readings with visual judgements.
This is because photometers are confined to measure
at defined spectral lines or bands. However, Crowther
and Evans (1981) have shown that photometer
readings can be calibrated to agree with visual
judgements. Cuthbert and Giorgio (1992) reviewed
spectrophotometric methods for colour determi-
nation and found that wavelengths from 240
to 450nm had been used. They showed that
absorbances at different wavelengths can usually be
fitted with a multiplicative linear model, but the slope

WR30/]I--I 2771
2772 D. Hongve and G./~kesson
Table 1. Results from a Nordic interlaboratory comparison of 11 laboratories (Stark, 1985). Mean values _+ standard deviation. All values
are as TCU (=mg/I Pt)
Spectrophotometer
Visual
method 410 nm 420 nm 436 nm 450 nm 465 nm
Settled/filtered S F S F S F S F S F S F
Synthetic
40mg/IPt 37_+3 38_+3 40+1 40+1 40+2 40_+1 40-+2 40_+2 4 0 + 2 40_+2 40-+3 40_+2
Tapwater <5 <5 4_+2.5 3.5_+1.5 6+3.5 4.5_+1.5 6_+4.5 4.5_+1.5 4.0_+3 3_+1.5 4.5+3.5 2_+1.5
Lakewater 1 9 _ + 4 15_+4 26_+2 1 8 _ + 1 41_+4 27_+2 44+4 26-+3 30_+2 18-+2 28_+4 15-+3
Lakewater 52_+8 44_+8 59_+7 44_+4 91_+11 67_+5 95_+10 67_+6 67_+9 46_+4 61_+9 40_+4
Industrial
wastewater 80_+7 77_+8 85_+8 75_+5 130_+10 114_+11 140_+10 123_+6 99+6 87_+8 87_+7 73_+4

of the regression lines varies so much between
samples that calculations of colour from absorbance
measurements at arbitrary wavelengths give large
errors. They state that estimation of colour by
relating absorption coefficients of water samples to
those of standard solutions is a more precise method
that is considerably easier to use than the traditional
visual comparative method. It is therefore peculiar
that spectrophotometric Pt-colour determinations
performed in this way sometimes give significant
overestimates of concentrations of humic substances.
SPECTROPHOTOMETRIC COLOUR MEASUREMENT
Development method
Extensive testing of various possibilities for
spectrophotometric colour measurement took place
in the Nordic committee for standardization of water
analysis (INSTA) during the early eighties. Some
leading water laboratories in Sweden, Finland and
Norway participated in method development and
laboratory intercalibration. During this work
Hongve (1984, and unpublished) found that spectro-
photometer calibration at 410 nm gave very good
correlation between visual and spectrophotometric
determinations for filtered samples. The influence of
turbidity increased as the wavelength increased.
Bj/irnborg et al. (personal communication)
made spectrophotometric colour determinations of
unfiltered and filtered natural samples at wavelengths
400, 410, 420, 432 and 450 nm. Best agreements with
comparator values were achieved with readings at 410
and 450 nm, although the correlations were relatively
poor. The correlation coefficients improved with
filtration of the samples. Filtration reduced both
comparator and spectrophotometric colours. Starck
(1985) conducted an interlaboratory comparison with
11 participating laboratories. The colours of synthetic
and natural samples were determined as comparator
readings and using spectrophotometer calibrated
against Pt-Co solutions. The wavelengths were 410,
420, 436, 450 and 465 nm. Good agreements were
found between comparator and spectrophotometer
readings of filtered samples at 410nm and 450-
465 nm. The relative contribution of particles in
samples cleared only by 30 rain settlement increased
with increasing wavelength. Relatively poor agree-
ment between absorbance readings indicated that
some spectrophotometers should have been recali-
brated. The results of the colour determinations in
this intercomparison are given in Table 1.
A standardised method
In 1988 The Norwegian Standards Association
approved a method for spectrophotometric colour
determination as an alternative to the visual method.
This is based on the following principles: The water
is filtered through a membrane filter with pore size
0.45 #m and therefore only true colour is determined.
The absorbance is measured at 410nm in a
spectrophotometer with band width not more than
5 nm. Length of cuvettes shall be 40 or 50 mm. The
absorbance of a reference solution with 100 mg/l Pt is
used for conversion of the results to TCU. It is crucial
that the spectrophotometer employs correct wave-
length and control of the monochromator adjustment
function must be performed at regular intervals.
DISCUSSION
Wavelength
When comparing the spectral absorption in the UV
and visible range for Pt-Co solutions and dissolved
humic substances, the reason seems obvious for the
inconsistencies in spectrophotometric determinations
at various wavelengths. Figure 1 shows the absorp-
tion spectrum for visible light in a reference solution
containing 100 mg/1Pt and a matched solution of
fulvic acid. Visually, both solutions have the same
colour intensity and hue although their absorption
10
8 I~ Pt-Cosolution
-- fulvic acid
0
400 500 600 700
wavelength, nm
Fig. 1. Absorbance of visible light in a reference solution
(Pt-Co) with 100 mg/l Pt compared with a visually matched
solution of fulvic acid.
 Spectrophotometric colour determination 2773
.~ 1,2
spectra are different. Specific for the platinum
solution is a strong absorption of short waved light
up to 410nm. The two solutions have equal ~ 0,8,
absorption coefficients at circa 410 nm. In the visible
range, the reference solution has a minimum in
t
"0

absorbance around 430 nm. We have tested a wide ~-0,4.

m
range of natural coloured samples that after filtration
have shown equal absorbance with matched reference ! mica

solutions around 410, 450-460 and 490-500nm. ~0,0

Thus, spectrophotometer readings have to be
calibrated against reference solutions in one of these
ranges if direct comparison of absorbance readings
shall be used to assess colour. If the spectropho-
tometer readings are calibrated at other wavelengths,
the colour determinations may be erroneous in the
sense that they do not agree with comparator
readings. In Fig. 2 the ratio of the two curves
from Fig. 1 is given for the range 400-500 nm. This
shows how the choice of wavelength may alter the
spectrophotometer colour readings relative to the
visual values with a factor between 0.7 and 1.8.
Readings in the range 430-440 nm occur quite often
in the literature and these give significant overesti-
mates. Filter photometers with broad band optical
filters may also give variable results, depending on the
specifications of the filter and the photosensor.
Filtration
In some standards for visual determination of
colour "true colour" is defined as the visible colour
in samples without visual turbidity or in samples
where the turbidity has been removed by filtration or
centrifugation (e.g., American Public Health Organ-
ization, 1992). In case of turbid samples, the results
are reported as "apparent colour". Particles in
natural waters cause usually a relatively constant
attenuation of light through the visible part of the
spectrum in spectrophotometric measurements
(James and Birge, 1938). The operators are most
often able to compensate for the cloudy impression
produced by a slight turbidity of this type, and
filtration may not change the judgement. However, if
particles are present that cause most light attenuation
in the short waved range, they may contribute
significantly to the apparent colour. Suspended flocks
of humus colloids and hydrous ferric oxides act in
this way. Simultaneously, such particles contribute
0 ......
4OO 450 5OO
wavelength, nrn
Fig. 2. Variation with wavelength in the absorbance ratio
between fulvic acid and a visually matched Pt-Co reference
solution.
400 500 600 700
wavelength, nm
Fig. 3. Differences in spectrophotometer absorbance
readings before and after filtration of water with added
turbidity of ground minerals (white feldspar, black mica)
and brown humus flocks.
little to nephelometric turbidity because they cause
little light scattering. In Fig. 3 examples of differences
between absorption spectra of filtered and unfiltered
samples containing mineral turbidity and humus
flocks are given. Filtration of strongly coloured
samples often reduce the observed colour. Many
researchers have interpreted reduction in colour by
filtration as adsorption of dissolved or colloidal
substances to the filter, may be because little
reduction in turbidity is observed together with a
significant reduction in colour. Our opinion is that
the reduction in visible colour observed after rapid
filtration of appropriate volumes of water (e.g., 25 ml
with suction) is normally caused by separation of
particles. For demonstration purposes we have
filtered various types of coloured water through
different filters. Figure 4 shows results as differences
in absorbance of visible light due to filtration. The
"stored water" was a sample of stream water that has
been stored for eight years in a tank at about 4°C and
is virtually free of particles; colour was 75 TCU and
turbidity 0.04 NTU (nephelometric turbidity unit).
The "lake water" was a fresh sample with colour
0.8
0.6
.~ 0.4
0.2
~ . ~ stored water
0.0 n0 ~ , ,,~
m
-0.2
400 500 500 700
wavelength, nm
0.22 pm membrane ~ 0,45 pm membrane
, . . ~ w 0 . 7 IJrn m e m b r a n e - - - glass fibre
paper
Fig. 4. Differences in spectrophotometer absorbance
readings before and after filtration of two water samples
through various filters. 0.7~m: Millipore HC; 0.45/Jm:
Gelman supor; 0.22/~m: Millipore GS; Glass fibre:
Whatman GF/C; Paper: Schleier & Schuell 597.
2774 D. H o n g v e a n d G . / ~ k e s s o n

prescribes simple filtration through glass fibre filter

before colour determination (EEC Council Directive
79/869). Since no specifications are given for the
"D filters, employment of membrane filters with pore size
0.45 #m will be a better alternative.

<
0 . . . . . . i . . . . . . , . . . . . . .. . . . . . . , . . . . . ".......
400 500
wavelength, nm
0.2 pm membrane ,,
---x-- 0.8 pm membrane - -
...... paper . . . .
Fig. 5. Differences in s p e c t r o p h o t o m e t e r a b s o r b a n c e
readings o f s t r e a m w a t e r before a n d after c e n t r i f u g a t i o n
(3000 rpm, 10 min) and filtration through various filters.
0.2/am: Gelman metricel; 0.45 #m: Gelman supor; 0.8 #m:
Millipore AA; Glass fibre: Whatman GF/C; Paper: Schleier
& Schuell 597.
75 TCU and turbidity 0.65 NTU. The stored water
showed no significant spectral change with any of the
chosen filters. Negative differences with paper and
glass fibre filters were probably caused by released
fibres. All types of filters reduced the attenuation of
short waved light in the lake water and there was no
obvious relationship between pore size and retention
of light absorbing substances. Figure 5 shows similar
results for a turbid stream sample (colour 60 TCU,
turbidity 8.5NTU) which was centrifugated and
filtered. The particles in this sample were less
efficiently withheld by the glass fibre filter and almost
not at all by the paper. The effects of membrane
filtration were the same as for centrifugation but
depended to some extent on pore size.
Although the coloured particles in the two turbid
samples contribute to both visual and spectrophoto-
metric measurements in much the same way as
dissolved humic substances, the calibration of optical
properties of suspended particles against Pt-Co
solutions is not feasible. This is because these
properties depend on physical variables as particle
size and surface structure, and probably also on the
chemical composition. Meaningful spectrophotomet-
ric colour determinations can therefore be made only
on optically clear samples. Filtration through
membrane filter with pore size 0.45 #m has been
prescribed as mandatory for determination of
spectral absorption (EN-ISO 7887) and will usually
be adequate. The EU drinking water standard
INTERLABORATORY COMPARISONS
Interlaboratory comparisons of the two standard-
600 700
ised methods in Norway have been performed in 1993
and 1994 (/~kesson, 1994, 1995). The participants
0.45 prn membrane were around 75 municipal and regional laboratories
- glass fibre engaged in the control of public water supply. One
centrifugation
third of these have used comparators and two thirds
have used the spectrophotometric method, Large
variances in colour determinations have been noted
during earlier trials. Therefore all participants were
furnished in advance with a calibrated coloured
solution made from commercial humic acid (Fluka)
in order to calibrate their readings. The provided
solution had to be diluted 1:100 to make a working
solution with colour 30 TCU. The result of the
internal calibration was reported with the results for
the unknown samples. The test samples were natural
lake waters with turbidity around 1 NTU. Results
from 17 laboratories who did not report to have
established internal quality assurance or were not
able to adjust their reading of the known reference
solution to 30 _+ 3 TCU have been rejected from the
statistical treatment. The results are summarised in
Table 2.
The average results for the two methods were
statistically different only for one sample (B-1993).
For the other samples the differences were not
statistically significant (p < 0.05). The distributions
of the results from 1994 for the two methods are
shown in Fig. 6. The reduction in standard deviation
for the spectrophotometric measurements from 1993
to 1994, seen in Table 2, indicates that operator
training and quality assurance has improved the
precision for this method.
CONCLUSION
Spectrophotometric determination of colour gives
results that are comparable with the Hazen method
if spectrophotometers are calibrated against Pt-Co
reference solutions, the samples are filtered, and
wavelengths are around either 410 or 450--465 nm.
410 nm is recommended because of better sensitivity.
Other wavelengths may cause large discrepancies

Table 2. Results of national interlaboratory comparisons of colour determination. Each laboratory has
given results for one method. S = spectrophotometric method, V = visual method; values as TCU
Sample no. A-1993 B-1993 A-1994 B-1994
Method S V S V S V S V
Number of laboratories 38 20 38 20 40 20 40 20
Average 41.7 38.8 33.8 31.9 26.0 25.2 18.2 17.1
Median 43.0 40 34.0 30 25.5 25 18.0 15
Standard dev. 3.5 4.7 3.4 3.8 2.4 4.7 1.6 3.9
Coeff. of var. (%) 8.5 12.2 10.1 11.8 9.4 18.5 8.8 22.6
 Spectrophotometric colour determination 2775

Sample A
is a critical parameter for many water works that
30 depend on surface water, and the spectrophotometric
Spmctl-opho~ommtmP n-40 method is recommended for control of drinking
water quality.
15

REFERENCES

0 /~kesson G. (1984) Laboratory intercomparison report.

Chemical analysis of drinking water. Norwegian Food
Control Authority. Report 7/I994. 84 pp. (in Norwegian).
Comparator n-20 ,~kesson G. (1985) Laboratory intercomparison report.
15 i i , 1 ~ 1 1 1 i , i . : ;ll , Chemical analysis of drinking water. Norwegian Food
14 18 22 28 30 34 38 Control Authority. Report. In press (in Norwegian).
Co I o u r
American Public Health Organization (1992) Standard
Methods for the Examination of Water and Wastewater.
18th edn. Washington DC.
Sample B Bjiirnborg B., Frrberg E. and Hietala M. (Personal
Communication) National Enviromental Board of
24 " : " " ' r . . . . . . . .' " " " ,' I
Sweden. Communication to a Nordic working group for
SPlCtrOphotoml+ir n-40
I standardization of colour measurement.
Crowther J. and Evans J. (1981) Estimating color in Hazen
units by spectrophotometry. J. A WWA 73, 265-270.
• 12 Cuthbert I. D. and Giorgio P. D. (1992) Toward a standard
L
0 method of measuring color in freshwater. Limnol.
Oceanogr. 37, 1319-1326.
Hazen A. (1892) A new color-standard for natural waters.
~ o Am. Chem. J. 14, 300-310.
_1
Hongve D. (1984) Water colour should be measured by
Complrator nl20 410 nm after filtration. Refbla' 2/84. Norwegian Institute
12
for Water Research, Oslo. pp 6-8 (in Norwegian).
8 12 18 20 24 28
International Organization for Standardization and Euro-
ColouP pean Committee for Standardization (1994) EN-ISO
Fig. 6. Distribution of results of colour measurement for 7887. Water Quality--Examination and Determination of
two lake water samples in a laboratory intercomparison in Colour. 10 pp.
Norway, 1994. James H. R. and Birge E. A. (1938) A laboratory study of
the absorption of light by lake waters. Trans. Wis. Acad.
Sci. Arts Lett. 31, 1-154.
Norwegian Standards Association (1988) Norwegian
between spectrophotometric and visual colour deter- Standard 4787. Water examination. Determination of
mination. Particle separation is essential for the colour. Spectrophotometer method. 3 pp. (in Norwegian).
spectrophotometric method because light attenuation Ohle W. (1934) Chemische und physikalische Untersuchun-
caused by particulates cannot be calibrated against gen norddeutscher Seen. Arch. Hydrobiol. 26, 386-464.
visual colour. One interlaboratory comparison Starck B. (1985) Determination of water colour. Centrallab-
oratorium AB, Helsinki. 28 pp. (in Swedish).
between Nordic countries and two interlaboratory Thienemann A. (1925) Die Binnegew~isser Mitteleuropas.
comparisons in Norway have proved that spectre- Die BinnengewLisser 1, 255 pp.
photometer readings at 410nm and comparator Thurman E. M. (1983) Determination of aquatic humic
readings give identical results and that the former substances in natural waters. U.S. Geol. Surv. Pap.
Hydrol. Sci. W2262. pp. 47-52.
method has best precision. The maximum colour World Health Organization (1984) Guidelines for drinking-
allowed in drinking water is low compared with the water quality. Vol. 2. Health Criteria and Other
sensitivity and precision of the visual method. Colour Supporting information. 335 pp.