4 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [ Vol. VI, No.

REPORT ON EGGS AND EGG PRODUCTS.

By H. L. LOURIB (Food and Drug Inspection Station, New York, N. Y.),

Referee.
At the last meeting your referee reported that there seemed no pos-

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sibility of determining whether or not dried egg products were decomposed
unless a study of their composition was made at the point of production
in order to obtain definite data as to the composition and character of
the raw eggs used and the changes that take place during manufacture.
It is well known, generally, that dried eggs are produced almost
exclusively at the present time in China. The methods of production
have been radically changed owing to the action of the Department of
Agriculture in refusing admission to eggs which were contaminated with
large amounts of zinc.
It is to be regretted that no methods which will conclusively show
the use of decomposed material are available at the present time.
The analyst must depend largely on the physical characteristics of the
dried-egg product to determine whether or not it has been made from
decomposed material. The methods for the determination of heavy
metals are satisfactory as are those for the presence of preservatives.
As it was considered that it would be of great value, a tentative
chapter on the methods of analysis generally used in government
laboratories for the determination of the sanitary quality of liquid eggs
has been prepared, including methods of analysis for liquid eggs, dried
eggs and egg products. It was impossible for your referee to obtain
collaborative action which would decide conclusively whether the
methods proposed are acceptable or not. However, it should be pointed
out that the Bureau of Chemistry has conducted a valuable piece of
research work with respect to the examination of frozen egg products
and interpretation of results under the direction of H. W. Redfield1.
With respect to the analysis of egg products, the work of your referee
has been confined to the analysis of materials such as egg noodles, in
which the important consideration is to determine whether or not the
quantity of eggs present is in compliance with the standards formulated
by the Standards Committee for this commodity. There has been a
great deal of discussion as to the accuracy of the Juckenack method for
the determination of the amount of eggs and work done at the Bureau
of Chemistry and at the New York and San Francisco Food and Drug
Inspection Laboratories shows that this method does not give results
which indicate the true amount of egg present. The difficulty that the
investigator who attempts to devise new methods for the estimation of
eggs in products of this type meets is that it is necessary to work out
' U. S. Dept. Agr. Bull. 846: 1920.
1922] LOURIE: REPORT ON EGGS AND EGG PRODUCTS 5

new ratios with respect to the lecithin P205 content of the various
flours. There is a growing practice among manufacturers of egg noodles
and similar products to use dried yolk or so-called dried whole egg mix-
tures which contain excessive amounts of yolk instead of whole egg
which the standard set forth by the Department of Agriculture requires.
The problem here is to prove that there is a deficiency in albumen

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derived from egg. It can readily be seen that a manufacturer using
5 per cent of dried yolk would obtain a product which, on analysis,
would indicate from the lecithin P205 content that it contained much
more than 5 per cent of whole eggs. Yet this product would be in
violation of the standard which calls for 5 per cent of whole eggs.
The New York and San Francisco Stations and the Bureau of Chem-
istry have been working on methods to determine whether egg noodles
are made from whole egg, yolky mixtures or yolk. This work has not
been completed. With respect to the determination of lecithin P205,
Jacobs and Rask have devised a method, based on the saponification of
the fat, which recovers practically all of the lecithin P205 content in
egg noodles. It has been noted that the lecithin P205 of flour and of
egg solids, when determined by this method, is considerably higher
than when determined by the Juckenack method; hence, different
values for these constants must be inserted in the formula which is used
for calculating the equivalent of egg solids from the lecithin P205 con-
tent of noodles. I t should be noted that the values for this method,
which are given in the following compilation of methods, are not based
upon a sufficiently large number of determinations to be accepted as
final.
FROZEN AND LIQUID EGG PRODUCTS.
TAKING OF SAMPLES.

Frozen egg samples shall consist, when possible, of original unopened packages.
They shall consist of representative containers of the product in any individual ship-
ment. Enough containers shall be taken to represent a whole shipment fairly. All
samples shall be sent to laboratories in the quickest possible way. When transported
by common carriers, samples shall be so packed as to prevent thawing and every pre-
caution shall be taken to prevent delay in transit. They shall be delivered to the
analyst immediately upon arrival at the laboratory and no sample shall be examined
which does not arrive in a frozen condition. If the material is slightly melted around
the circumference, the subsamples for bacteriological and chemical examination must
be withdrawn from the portion which is still frozen.
When samples are opened, the bacteriologist, chemist, and microscopist shall all be
present and, in case of official samples, shall initial and date seals and cans for identi-
fication in the regular manner.
In order to preclude all possibility of a claim of contamination during sampling, the
bacteriologist shall always withdraw subsamples first when a container is opened.
The chemist shall then withdraw subsamples. The remainder shall be turned over to
the microscopist. Each analyst shall give a receipt to the one from whom the con-
tainer was received, in the case of official samples.
6 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [ Vol. VI, No i

For withdrawing subsamples, it will be found convenient to use a sterile butter

sampler of sufficient length to enable one to remove a core of frozen material from the
top to the bottom of the container, after first removing with a sterilized instrument
(chisel) the surface layer of the frozen material.
If the sample is frozen very solidly, it may be found necessary to use a brace and
long-shanked bit or ship-builder's auger and to collect the shavings for the sample.
Cores shall be taken midway between the center and circumference, from at least

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three widely separated parts of the container to form a composite sample.
Liquid egg samples shall be thoroughly mixed with a sterilized utensil such'as a long-
handled dipper which has been immersed (including the handle) in alcohol and flamed,
and then subsamples withdrawn for examination. The subsamples for bacteriological
examination must be placed in sterilized containers.
The chemical examination shall be started immediately after the taking of sub-
samples and all determinations shall be made in duplicate.
The subsamples are thawed by partially immersing the containers in warm water,
the temperature of which should not be above 50CC. They must then be thoroughly
mixed, preferably with an electric stirrer. In the absence of such an instrument in
the chemical laboratory, the mixing may be quite satisfactorily accomplished by suck-
ing the melted subsample several times through a Gooch crucible containing no asbes-
tos, using very moderate suction. This mixed composite sample is examined by the
methods which follow:
TOTAL SOLIDS.
Weigh approximately 5 grams of the sample into a tared lead dish of 2\ inches to
3 inches diameter and dry in a vacuum of not less than 25 inches at 55°C. until there is
no further loss in weight. This drying usually requires about 4 hours. It is recom-
mended that weighings be made at the end of 3 \ hours' drying, and thereafter at inter-
vals of about 30 minutes. Weigh to 3 decimal places. There is an appreciable gain
in weight after the minimum has been reached. Express the results as per cent on
the wet basis.
ETHER EXTRACT.
Extract the dry residue from the determination of solids with absolute ether, pref-
erably in a Knorr apparatus, but if this is not available, in a Johnson extractor. Cut
through the sides of the lead dish containing the solids at 4 equidistant points. Place
the dish upon a fat-free filter paper. Flatten down the sides of the dish. Place another
fat-free filter paper on top of the flattened dish and roll the papers and dish into a
cylinder which will fit the extraction tube fairly snugly. In making the cylinder,
turn in the ends in such a way as to prevent solid particles from dropping into the
extraction flask. Place the cylinder in the extraction tube without any asbestos plug
below it. If the extractor is working rapidly, 3 hours is sufficient to insure a proper
extraction.
Distil off the ether from the extraction flask and dry the extract for one hour at 55 °C.
in a vacuum of not less than 25 inches. Weigh to 3 decimal places. Express the results
as per cent on the wet basis.
The ether used should not contain any alcohol or water, as a higher result is
obtained when either is present. It is, therefore, understood that ether freshly
distilled from sodium will be used.
ACIDITY OF THE FAT.
Dissolve the fat obtained in the determination of ether extract in 50 cc. of neutral
benzine to which has been added 2 drops of phenolphthalein indicator. Titrate with
0.05N sodium ethylate. Express the results as the number of cc. of 0.05N sodium
ethylate required to neutralize 1 gram of fat.
1922] LOURIE: REPORT ON EGGS AND EGG PRODUCTS 7

AMMONIA NITROGEN.
Titration Method.
This method is an adaptation of Folin's method for the determination of ammonia
in urine1. It consists essentially in making the sample alkaline, removing the liberated
ammonia by aeration and catching it in a measured quantity of standardized acid.
The excess acid is then titrated.

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The apparatus consists of the following:
1. A wash bottle containing dilute sulfuric acid (about 35 per cent) to remove any
ammonia that may be present in the air entering the system.
2. Some form of trap to prevent sulfuric acid being carried over mechanically.
3. An aerating cylinder about 50 mm. in diameter and 350 mm. high, fitted with a
2-hole rubber stopper carrying a right-angle air-inlet tube, open at the bottom and
extending to within J inch of the bottom of the cylinder and a trap containing either
a cotton or glass wool plug to prevent any liquid from being carried over mechanically.
4. An 8-ounce, wide-mouthed bottle fitted with a delivery tube coming from the
trap on the aerating cylinder. It is not essential that the special ammonia absorption
tubes be used. An ordinary glass tube with a small bulb blown on the end, through
which a few holes are punctured, answers very well. The method of making these is
given by Folin and Farmer2.
5. A means of passing air through the system. This is best done by a pump which
will furnish a blast with a pressure of 10 pounds per square inch and which discharges
into a tank of sufficient size to compensate for the pulsations of the pump and to deliver
a steady blast.
Suction may be used but it is not recommended.
Each of the first four parts enumerated is fitted with a 2-hole rubber stopper and
all are connected by glass tubes of suitable shape and length to permit the proper pas-
sage of air through the apparatus. The tube leading into the acid wash bottle should
contain a stop-cock for regulating the air supply.
Weigh approximately 25 grams of sample in a convenient container. Pour as much
as possible of this material into the aeration cylinder and transfer the remainder by
means of four 25 cc. portions of ammonia free water, stirring each time with a rubber-
tipped glass rod to remove any egg adhering to the sides of the weighing vessel. Add
75 cc. of alcohol, mix well, let stand for 15 minutes. Add approximately 1 gram of
sodium fluoride, 2 cc. of 50% potassium carbonate solution and 1 cc. of kerosene. Con-
nect the apparatus and aerate into the receiving bottle which should contain 10 cc. of
0.02N sulfuric acid, 2 drops of methyl red indicator (saturated solution in 95% alcohol)
and about 75 cc. of ammonia free water.
The aerations should be carried on for 4 hours or as long as necessary to remove all
of the ammonia, using as rapid a current of air as possible.
Titrate the excess of acid with 0.02N sodium hydroxide (free from carbon dioxide).
Express the results obtained as milligrams of ammonia nitrogen per 100 grams of
sample on the wet basis.
If there is insufficient time to complete the determination, the sample may be left
overnight in the cylinder with the alcohol and sodium fluoride added. The potassium
carbonate should, of course, not be added until ready to proceed.
If the sample has a bad odor, it may be necessary to use more than 10 cc. of 0.02N
sulfuric acid.
It is essential that a blank experiment be run to determine the percentage recovery
of ammonia using a known amount of pure ammonium sulfate (containing about 3 mgs.
1
Z . physiol. Chem., 1902, 37: 161.
' J. Biol. Chem., 1912, 11: 499.
8 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [ Vol. VI, No. 1

of nitrogen) and 25 cc. of water, instead of the egg. For method of preparing pure
ammonium sulfate see Folin and Farmer1. The recovery should be over 95 per cent.
It is also essential to run blank experiments with the reagents and water used.

REDUCING SUGARS.
Wash 25 grams of sample into a 200 cc. graduated flask with 75 cc. of water. Make
slightly acid by adding 2 cc. of 5% acetic acid for white or whole egg and 1 cc. for yolk.

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Mix and immerse the flask in boiling water until the egg material is thoroughly coagu-
lated. This requires about 15 minutes. Cool to room temperature and make up to
the mark with washed alumina cream. Shake vigorously for 1 minute, allow to stand
5 minutes and then shake 1 minute. Filter through a dry folded filter and determine
the reducing sugar in 50 cc. of the filtrate by the Munson and Walker method2. Calcu-
late as dextrose and express the results as per cent on the wet basis.
The washed alumina cream is prepared by washing ordinary alumina cream 5 times
by decantation, using enough water so that at least half of the total volume may be
syphoned off each time.
On account of the volume occupied by the precipitate, the results are a trifle high.
This error amounts to about 1.4 per cent for white, 3.3 per cent for whole egg and
7.0 per cent for yolk. It is not customary to correct for this.
INDOL AND SKATOL.
A 200 cc. portion of sample is diluted with 500 cc. of water, acidified with 40 cc. of
5% acetic acid for white or whole egg and 20 cc, for yolk, coagulated in boiling water
or live steam and filtered. The filtrate is steam distilled as rapidly as possible. Ex-
tract the distillate (approximately 300 cc.) with ether in a separatory funnel. To the
ether extract add about 3 cc. of water and evaporate before an electric fan until the
smell of ether has almost but not entirely disappeared. A trace of ether does not
affect the result. Add 10 cc. of water, filter and apply the following test to the filtrate:

Vanillin Test for Indol and Skalol.

To the solution to be tested add a few drops of a 5% solution of vanillin in 95%
alcohol, and concentrated sulfuric acid. The acid should be added in the proportion
of 2 cc. for each 5 cc. of solution being tested. If indol is present, an orange color will
be formed which is soluble in chloroform, amyl acetate or amyl valerianate. If skatol
is present, a deep red to violet color will be formed which is readily soluble in chloro-
form, amyl acetate or amyl valerianate3.
As confirmatory tests, if needed, the following are suggested:

p-Dimethylaminobenialdehyde Test.
To the solution to be tested, add 1 cc. of a solution consisting of 4 parts of paradi-
methylaminobenzaldehyde, 380 parts of absolute alcohol and 80 parts of concentrated
hydrochloric acid, in such a way as to form two liquid layers. If indol is present, a
purplish red color will be formed. If skatol is present, a blue violet color will be formed4.

Herler's B-Naphtha Quinone Test.

Make test solution slightly alkaline with potassium hydroxide. Add 1 drop of a
2% solution of B-naphtha quinone sodium monosulfonate. If indol is present, a blue or
green blue color will be formed6.
• X Biol. Chem., 1912, 11: 496.
2 Assoc. Official Agr. Chemists, Methods. 1920, 78.
! J. Biol. Chem., 1916, 24: S28.
< Z. physial. Chem., 1906, 47: 25.
r
'J. Biol. Chem., 1906, 1: 257.
1922] LOURIE : REPORT ON EGGS AND EGG PRODUCTS 9

Pyruvic Aldehyde Test.

To the solution to be tested add a small crystal of ferric sulfate and a few crystals of
pyruvic aldehyde. Then carefully run sulfuric acid down the inside of the container
in such a way as to form a layer at the bottom. If indol is present, a red violet ring
will form1.
Dimethylaniline Test.

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To the test solution add a few drops of dimethylaniline and sulfuric acid. If skatol
is present, a deep red violet coloration is formed1.

Glycolic Acid Test.

To the solution to be tested add a few crystals of glycolic acid and an equal volume
of sulfuric acid. If skatol is present, a red violet color is formed2.

PRESERVATIVES*.

DRIED EGGS.
PHYSICAL CHARACTERISTICS.

Perform organoleptic tests. Note particularly the taste and odor of the product as
it is. Place about 10 grams in a 100 cc. beaker, add 50 cc. of water, stir, cover with
watch glass and let stand one-half hour. Note odor.

2THC.

Place 25 grams of a well-mixed sample in an 800 cc. Kjeldahl flask; add 5 grams of
zinc-free potassium sulfate, 3 to 4 glass beads to prevent bumping, 30 cc. of concen-
trated sulfuric acid, in the case of yolks or whole eggs (25 cc. of the acid in the case of
albumens) and 30 cc. of concentrated nitric acid. Do not heat. When spontaneous
action subsides, add 10 cc. of concentrated nitric acid. After 2 or 3 additions of con-
centrated nitric acid the action becomes less violent. Heat gently, at first, continuing
the addition of concentrated nitric acid and increasing the temperature as the digestion
proceeds until the contents of the flask are straw colored or colorless, after the nitric
acid fumes have' been boiled off. This digestion may be accomplished in the case of
albumen in 40 minutes and in the case of yolks or whole eggs in 1 hour. To warm
digestion add 100 cc. of water; pour into a 400 cc. beaker and rinse flask with two suc-
cessive 50 cc. portions of water. To the combined water solution add concentrated
ammonia to faintly alkaline. Pass hydrogen sulfide gas through solution for 15 min-
utes which should be sufficient to saturate. (At this point the majority of albumens
indicate the presence or absence of zinc. In the case of albumen, if zinc is present,
add 1 cc. of a diluted solution of ferric chloride containingt0.5 gram of solid ferric chlo-
ride per 100 cc. This will assist in retaining zinc sulfide on the paper when filtering.
Pass hydrogen sulfide gas through the solution for 15 minutes.) Heat beaker on steam
bath for one-half hour. Remove. Allow to settle from 5 to 10 minutes. Decant
through 9 cm. filter paper allowing as much of the precipitate as possible to drain
thoroughly. Dissolve the zinc sulfide from this precipitate with 10% hydrochloric
acid, the solution after passing through the filter paper being returned to the original
beaker. Copper and lead sulfide are insoluble at this point, and may be determined
by the A. O. A. C. methods4. To the hydrochloric acid solution add 5 grams of ammon-
ium chloride and excess of bromine water and a slight excess of concentrated ammonia.

i J. Biol. Chan., 1916, 24: 527.

'Biochcm. Z., 1919,19: 523.
» Assoc. Official Agr. Chemists, Methods, 1920,117.
• Ibid., 161, 285.
10 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [ Vol. VI, No. 1

Neutralize carefully with 10% hydrochloric acid adding 2 cc. in excess; add 10 cc. of
50% by weight of ammonium acetate and 8 to 10 drops of 10% ferric chloride solution,
or enough to give a good reddish tinge. Dilute to about 300 cc. with water and boil
for 1 minute. Settle; filter hot and wash with hot 5% ammonium acetate. Pass
hydrogen sulfide gas through filtrate for 15 minutes. Heat one-half hour on steam
bath; filter through weighed heavily padded Gooch crucible using gentle suction. Wash
with hot 5% ammonium acetate solution. Dry in oven; then ignite, roasting first.

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Increased weight of Gooch is due to oxide of zinc. This multiplied by 0.8034 gives
zinc present in 25-gram sample.

PRESERVATIVES'.

EGG PRODUCTS.
EGG NOODLES.
MOISTURE.
Dry a convenient quantity of the sample, 2 to 3 grams, at the temperature of boiling
water, in a current of dry hydrogen or in vacuo, until it ceases to lose weight (approxi-
mately 5 hours). The loss in weight is the moisture content.
ASH.

Char a convenient quantity of the sample, 5 to 10 grams, in a previously ignited and

weighed platinum or silica dish and burn at the lowest possible heat until free from
carbon. If a carbon-free ash can not be obtained in this manner, exhaust the charred
mass with water, collect the insoluble residue on an ashless filter, burn until the ash
is white or nearly so, and then add the filtrate to the ash and evaporate to dryness.
Heat the residue to redness, cautiously at first to avoid decrepitation or spattering,
cool in a desiccator and weigh.
SODIUM CHLORIDE IN ASH.

Determine chlorides by the silver nitrate gravimetric or the Volhard volumetric

method. These methods are described in detail in any of the standard texts on quanti-
tative analyses. Calculate to the equivalent of sodium chloride. Total ash, minus
sodium chloride, is the salt-free ash.
NITROGEN'.
FAT,

Treat 5 grams of the sample in a loosely stoppered 200 cc. Erlenmeyer flask with a
mixture of 10 cc. of alcohol (95%), 2 cc. of concentrated ammonium hydroxide, and
3 cc. of water, keeping the contents of the flask at the boiling point for 2 minutes, pref-
erably on the steam bath. . After cooling, extract the contents of the flask with 3 suc-
cessive 25 cc. portions of ethyl ether, mixing and tamping the material thoroughly
each time with a glass rod flattened at the end and pouring the extracts off by decan-
tation into a 250 cc. beaker. The last 25 cc. portion of ether should be drained out as
completely as possible, after which another 15 cc. portion of the same ammoniacal
alcohol solution is added to the flask and the matted material disintegrated as thor-
oughly as possible by means of the flattened glass rod which may be left in the flask for
this purpose. The flask is then returned to the steam bath and the entire procedure
repeated, the second set of ether extracts being poured into the beaker containing the
first set. The second treatment with the ammoniacal alcohol mixture should be more
gradual and somewhat longer than the first, so that the ether remaining in the flask

' Assoc. Official Agr. Chemists, Methods, 1920, 117.

' Ibid., 7.
1922] LOOBIE: BEPOBT ON EGGS AND EGG PBODUCTS 11

may be evaporated off and the ammoniacal alcohol brought to the required boiling
point without results disastrous to the determination.
Evaporate the combined extracts to dryness on the steam bath and extract the fat
from the residue left in the beaker with successive portions (5 or 6 treatments, using
about 15 cc. each time) of a mixture of equal volumes of ethyl ether and petroleum
ether. Collect the extracts in a tared platinum dish (do not try to filter), and evap-
orate to dryness on the steam bath. Dry the residue in a water-jacketed oven at

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the temperature of boiling water for 30 to 45 minutes, cool in a desiccator and weigh.
LECITHIN P20S. METHOD I.

Add 3 cc. of a concentrated alcoholic solution of potassium hydroxide to the fat in

the dish, as obtained in the preceding method. Evaporate to dryness, char, and
determine total P205 by the volumetric method'. Owing to the small quantity of
P205 in the fat from a 5 gram sample of noodles, it may be advisable in some cases to
use the nephelometric method2.
The following formula is irsed for calculating the percentage of egg solids in noodles
on the moisture-free basis:
(A-0.0548) X 1 0 0 = X ,
(1.38 = 0.0548)
in which A =percentage of lecithin P205 in the sample,
0.0548=percentage of lecithin P205 in flour (dry basis),
and 1.38 = percentage of lecithin P205 in whole egg solids.
GASOLINE COLOR VALUE.
Place 20 grams of the sample in a wide-mouthed, glass-stoppered bottle of about
120 cc. (4 oz.) capacity, and add 100 cc. of colorless gasoline. Stopper tightly and
shake vigorously for 5 minutes. Let stand for 16 hours, shake again for a few seconds
until the flour has been loosened from the bottom of the bottle and thoroughly mixed
with the gasoline, and then filter immediately through a dry 11 cm. paper into an
Erlenmeyer flask, keeping the funnel covered with a watch glass to prevent evaporation.
In order to secure a clear filtrate a certain quantity of the flour should be allowed to
pass over onto the paper and the first portion of the filtrate passed through the filter a
second time. It will be found convenient to fit the filter paper to the funnel by means
of water, after which it should be dried thoroughly, either by letting stand overnight
in a well-ventilated place or by heating.
Determine the color value of the clear gasoline solution in a Schreiner or Campbell-
Hurley colorimeter, using for comparison a solution of potassium chromate containing
0.05 gram of potassium chromate per liter, conveniently prepared by making 10 cc. of
an aqueous solution containing 0.5 gram of potassium chromate per 100 cc. up to 1
liter. The colorimeter tube containing the gasoline solution should be adjusted to
read 50 mm., then the tube containing the standard chromate solution raised or lowered
until the shades of yellow in both tubes match. The reading of the chromate solution
divided by the reading of the gasoline solution gives the gasoline color value, the color
of the standard chromate solution being considered as unity.

NOTES.
Standing for a longer time than that prescribed does not appear to affect the results.
In fact, the filtration may be dispensed with entirely if the solution is allowed to settle
after the second shaking until perfectly clear, which usually requires at least 24 hours.
The color value may be determined also in Nessler tubes, using for comparison
1
Assoc. Official Agr. Chemists, Methods, 1920, 3.
'J. Biol Chem., 1918, 36: 335.
12 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [Vol. VI, No. i

potassium chromate solutions of various dilutions prepared from t h e stock solution

(containing 0.5 gram per 100 c c ) , and filling the tubes in all cases to t h e height of
50 mm., or (less accurately) in a Lovibond tintometer. In the latter case the color
value is obtained directly, the solution of potassium chromate containing 0.05 gram per
liter corresponding in color to the 1.0 of t h e yellow slide. In order to avoid confusion
the color value should be referred to the potassium chromate solution containing 0.05
gram per liter in all cases.

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If an approximate result is sufficient for the purpose, it m a y be obtained by carefully
pipeting off the clear, supernatant gasoline solution before t h e second shaking. T h e
results thus obtained are about 10% lower than those obtained by t h e standard method
as just given.
ARTIFICIAL COLOR.
Herlwig's Method1.
Macerate with a pestle in a 6-inch mortar 75 grams of the finely ground sample with
40 cc. of hydrochloric acid (1 to 1). A fairly stiff mass results but no more acid should
be added. Continue the maceration until all particles are moistened and the entire
mass is homogeneous. Then add 60 to 70 cc. of amyl alcohol and again macerate well,
in the cold, until most of the color is apparently extracted. T h e time necessary for
this depends upon the kind of color and its amount, but 5 to 10 minutes is usually
sufficient. Filter the mixture on a folded filter paper or on a Biichner funnel using
suction. Much of the excess of alcohol may be forced out by pressing down on t h e
material on t h e filter with t h e pestle. The filter paper and residue may then be enclosed
in a cloth and the greater part of the remaining alcohol pressed out. In order t o remove
all color the exhausted sample may be returned to the mortar and re-extracted with
30 to 40 cc. of fresh amyl alcohol and filtered as before. Two such extractions will
remove most of the color and three will leave the material being extracted almost
perfectly white. The amyl alcohol will contain the color in a fairly pure condition.
Wash the amyl alcohol with 20 cc. of hydrochloric acid (1 to 1), then remove t h e
color fractionally by successive washings with hydrochloric acid of decreasing strengths
such as 4 N , N , 0.25 N , etc. Warming the acid is recommended. T h e color or colors
so removed m a y be identified by Mathewson's method 2 . Attention is particularly
called t o t h e directions on pages 18 to 20 of this bulletin under the caption of
"Abridged Procedure for Permitted Dyes Only".
Many colors will be colorless in acid amyl alcohol and not apparent to the eye until
washed out and the solution neutralized.

NOTES.
This method seems capable of extracting small quantities of color very thoroughly
from pastes. T h e procedure is very simple and the extraction takes little time. T h e
extracted color is fairly free from foreign interfering substances and in a condition
suitable for specific tests for its identification.

Jablonski Method3.
Macerate 250 grams of the sample (depending upon t h e quantity of color present),
with alcohol of a strength of about 8 0 % by volume and m a d e slightly alkaline with
ammonia. Continue t h e maceration until t h e color is extracted, if necessary warming
on t h e steam b a t h . Filter and evaporate the filtrate until t h e alcohol is expelled a n d
then add about one-fourth its volume of 2 5 % salt solution, cool if necessary, and extract
with gasoline or ether. T h e solvent will extract saffron, annatto, turmeric a n d all
1
Devised by Raymond Hertwig.
23 U. S. Dent. Agr. Bull. 448: (1917).
Devised by C. F. Jablonski.
1922] LOURIE: REPORT ON EGGS AND EGG PRODUCTS 13

oil-soluble colors, also fats. Draw off the aqueous layer and wash t h e solvent with
5 % salt solution adding t h e washings to the water solution. If t h e aqueous layer and
washings are colorless no other colors are present.
Saffron, a n n a t t o and turmeric may be removed from t h e gasoline (or ether) with
70% alcohol and the usual tests applied. Any oil-soluble colors, if suspected, may be
fractionally separated and identified b y Mathewson's method.
If colored, extract t h e remaining aqueous solution and washings with amyl alcohol.

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This will remove the common orange colors, Orange I, Orange I I , and Crocein Orange
(S. & J. Nos. 85, 86 and 13, respectively), also Martius Yellow (S. & J. No. 3). Sepa-
rate the aqueous layer and wash t h e amyl alcohol 2 or 3 times with a 5 % salt solution
to remove small amounts of Naphthol Yellow S and add t h e washings t o t h e aqueous
solution. If the aqueous solution is colored Naphthol Yellow S or Tartrazine may be
suspected. Naphthol Yellow S may be removed b y extraction with amyl alcohol
after acidifying to approximately 1/64N with hydrochloric acid. If, after this treat-
ment, any color remains in t h e aqueous solution strongly acidify it with hydrochloric
acid and again extract with amyl alcohol. Tartrazine will be extracted.
The advantage of the procedure as outlined above is the easy separation of the various
color groups as well as a partial elimination of the protein material which usually
causes trouble with other alcoholic extraction methods.

Modification of the Jablonski Method'.

Macerate 250 to 500 grams of the sample, depending upon the quantity of color
present, with alcohol of a strength of about 8 0 % by volume made slightly alkaline
with ammonia. Warm on the steam bath in order to insure sufficient extraction of
color. Filter with suction upon a Buchner funnel and evaporate the filtrate until most
of the alcohol has been expelled and the consistency is t h a t of cool molasses. Take
from the steam b a t h and add about an equal volume of low boiling petrolic ether. Also
add a large pinch of coarse sand. Stir the mixture until homogeneous and again place
upon t h e steam bath, taking care t h a t the petrolic ether does not evaporate too quickly.
From time to time when it is noticed t h a t practically all of the petrolic ether has evap-
orated add additional quantities, performing this operation until such time as the mass
assumes a spongy, jelly-like consistency and the odor of alcohol is faint. Take from
the steam bath, spread t h e product as well as possible upon a large watch glass and
place in t h e h o t water oven. Continue t h e heating until such time as t h e product is
dry and tough. I t may then be scraped from t h e glass and ground in a mortar. Pour
the ground mass into a separatory funnel and add about half of its volume of 2 5 % salt
solution. At this point it is advisable to pour into the separatory a few cc. of small
lead shot. This helps t o disintegrate the mass upon extraction with gasoline. The
method may then proceed in t h e same manner as stated in the Jablonski method.
The advantage of this modification depends upon the fact t h a t emulsions due to
gluten are avoided. No special care need be observed in t h e amount of added am-
monia and, provided the alcohol is finally removed, separations proceed easily.

RECOMMENDATIONS.
It is recommended—
(1) That the following methods given under the examination of
frozen and liquid egg products be adopted as tentative methods:
Total Solids Titration Method for Ammonia Nitrogen
Ether Extract Reducing Sugars
Acidity of F a t Indol and Skatol.

>ByM. G. Wolf.
14 ASSOCIATION OF OFFICIAL AGRICULTURAL CHEMISTS [ Vol. VI, No. 1

(2) That the following methods of analysis under "Egg Products",

sub-heading "Egg Noodles", be adopted as tentative methods:
Fat Lecithin P205, method by Jacobs and Rask.
(3) That during the coming year, the referee conduct collaborative
work with respect to the determination of heavy metals, especially zinc,

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in dried eggs, and that collaborative work be conducted with respect to
the determination of fat and lecithin P205 in egg noodles using the
methods of Jacobs and Rask.
(4) That no further work be done on the Juckenack method for
lecithin P205 since investigative work at three widely separated labora-
tories of the Bureau of Chemistry has shown this method to give low
results.
NOTE.—Your referee can not see the value of conducting, collaborative work with
respect to the methods of analysis employed in the examination of frozen and liquid
eggs since the research work conducted and already published by the Bureau of Chem-
istry1 amply covers the field and represents a type of collaborative work that would be
practically impossible for any referee to conduct for the association.

R E P O R T ON PRESERVATIVES (SACCHARIN).
By MILTON G. W O L F (U. S. Food and Drug Inspection Station, New
York, N. Y.), Referee.
Your referee regrets that he was unable to conduct any collaborative
work on methods for the determination of saccharin in foods. How-
ever, the details of a new method for the determination of saccharin in
bread, cake and similar products have been worked out. This method
deals largely with improvements with respect to the preparation of the
sample before the final extraction with ether.
An improved method for the detection of small amounts of coal-tar
dye in alimentary products led to the belief that the same ideas might
be used to advantage in the quantitative estimation of saccharin in
baked-food products. The directions are as follows:
A water-alcohol mixture appears to be the only efficacious solvent of the saccharin
which has been incorporated in the bread; the water moistens the bread while the
alcohol dissolves the saccharin. When this mixture, containing the saccharin filtered
from the bread, is evaporated, the gluten, which at first was soluble, forms into a charac-
teristic gelatinous mass from which ether, the final solvent, does not extract the sac-
charin quantitatively. To obviate this, heat the hydroalcoholic solution on the
steam bath in a beaker until most of the alcohol is evaporated and the product has the
consistency of molasses.. Take from the steam bath and add about an equal volume
of low boiling petrolic ether and a few pinches of coarse sand. Stir the mixture until
homogeneous and again place upon the steam bath, taking care that the petrolic ether

' U. S. Dept. Agr. Bull. 846: (1920).