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-I---. 1 2 CueO, mg . . . . . . . . . . 299.6 298.0 Dextrose, mg . . . . . . . 137.0 136.2 Starch, mg . . . . . . . . . 123.3 122.6 Percentage of starch 41.09
7111 2 297.5 298.4 135.95 136.40 122.4 122.7 40.91
7-111-----. 1 2 300.1 299.4 137.25 136.90 123.5 123.2 40.98
Average percentage of s t a r c h . . . . . . . . . . . . . 4 0 . 9 6
METHOD)-Threegram samples were used and t h e diastase conversion carried out as outlined in Leach’s “Food Inspection a n d Analysis,” p. 284. The Munson-Walker method was used for determining t h e amount of dextrose. The reductions were determined on an aliquot p a r t of a definite volume equivalent t o 0 . 2 4 0 g. of material. For t h e conversion 0 . 6 0 g. of commercial diastase with a reduction equivalent of 33.1 mg. of cuprous oxide was used. STARCH D E T E R M I N A T I O N (DIASTASE
CuzO, mg . . . . . . . . Dextrose, mg .....
..
starch
.........
7 1 7 1 2 233.9 234.1 89.40 89.50 80.46 80.55 33.53
33.56
Average percentage of starch
----II----. 1 2 232.8 233.3 88.85 89.10 79.96 80.19 33.32
...
7-1117 1 2 232.8 232.0 88.85 88.46 79.96 79.61
33.41
33.32
33.17
33.39
D E T E R M I N A T I O N O F S U G A R ( R E D U C I N G SUGARS)-The determination of sugar was difficult on account of the colloidal condition of t h e sugar extract. This difficulty was finally overcome b y keeping t h e solution just slightly alkaline, which seemed t o settle t h e colloids. Filtering was avoided as far as possible b y increasing t h e volume of t h e solution and pipetting an aliquot volume. Five-gram samples were boiled in 1 5 0 cc. of 5 0 per cent neutral alcohol for a n hour on a steam bath, with reflux condenser. The solution was cooled t o room temperature and t h e volume made u p t o 500 cc. with 9 5 per cent alcohol made just alkaline. After thoroughly mixing and settling over night, 400 cc. were pipetted off with continuous suction and evaporated on a water bath t o 20 cc. This volume was made u p t o 2 5 0 cc., using 2 cc. of lead acetate t o clarify. After clarifying, 2 0 0 cc. were pipetted with continuous suction into a beaker and t h e excess of lead precipit a t e d with anhydrous sodium carbonate. T h e solution was filtered a n d jo cc. of t h e filtrate used for determining t h e sugar b y t h e Munson-Walker method. From a n aliquot p a r t of t h e solution equivalent t o 0.80 g. of material only a mere trace of CuzO was formed. DETERMINATION
OF
SUGAR
(AFTER
INVERSION)-
Fifty cubic centimeters of t h e solution in t h e preceding determination from which t h e excess lead was precipitated were pipetted into a 100-cc. graduated flask, 5 cc. of concentrated hydrochloric acid added, t h e volume made up t o I O O cc. with distilled water, and allowed t o remain over night a t about z o o C. --I---. 1
CuzO, mg . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 21.00 9.40 Dextrose, mg.. . . . . . . . . . . . . . . . . . . . . . . . . . Percentage of dextrose,. , , . , , , , , . , , , . , , . 2 . 3 5 Average percentage of dextrose.. . . . . . . . . . Percentage calculated as cane sugar.. . . . . .
2 21.40 9.56 2.39
c---II--. 1 2 21.90 21.10 9.44 9.76 2.44 2.36 2.39 2.15
The acid was nearly neutralized and t h e sugar determined in an aliquot part of t h e solution, equivalent t o 0.40 g. of material, b y t h e Munson-Walker method.
Vol.
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D E T E R M I N A T I O N OF C R U D E FIBER-This determination was made on 2-g. samples containing 8 . 5 7 per cent of moisture. Kennedy’s modification’ of Sweeney’s method was used and modified b y filtering a n d igniting in alundum crucibles. Crude fiber and ash, grams..
I I1 I11 IV 0.2585 0.2650 0.2590 0.2580 0.0358 0.0380 0.0398 0.0382 0.2127 0.2220 0.2192 0.2198 10.63 11.1100 10.96 10.99 10.92
....
,
TANNIr;-Qualitative
tests showed tannin.
SUMMARY -PERCENTAGES ON20-Mesh 72-Mesh OvenCONSTITUENTS as Received Air-Dried Dried Moisture. 11.28 8.60 0.00 . . . . . . . . . . 4.33 4.46 4.88 Oil (ether extract). . . . . . . . . . 7.03 7.24 7.92 20.93 Protein. 19.13 Starch (diastase). 33.39 36.52 40.98 44.83
.................
........
F
DesiccatorDried
....
.... 8.46
.... ....
---PERCENTAGES ON-20-h.Iesh 72-Mesh OvenCONSTITU~NTS as Received Air-Dried Dried Hemicellulose (starch by acid conversion . . . . . . 7.37 7.59 8.31. . . . . . . trace trace trace Sugar (after inversion). . . . . . . . . . . . . . . . . . 2.32 2.39 2.31 Sugar (after inversion computed t o cane sugar) ....................... 2.08 2.15 2.35 Crudefiber ............................ 10.59 10.92 11.92 Tannin and other undetermined substance by difference ......................... 6.35 6.52 7.17
The proximate analysis shows t h a t the seeds would make a good component part of a stock food a n d a s seeds of related species have been found t o contain considerable amounts of potassium nitrate, t h e rather high protein content would suggest t h a t t h e y might. be valuable as a chicken or bird food. CHEMICALLABORATORY UNIVBRSITY OF MINNESOTA MINNEAPOLIS
THE DETECTION OF VEGETABLE GUMS IN FOOD
PRODUCTS By A. A. COOK AND A. G. WOODMAN Received M a y 2, 1918
The use of gums in food products is dependent mainly on their physical properties, t h e most noteworthy of which is their colloidal nature. This property enables t h e gum substance t o hold within itself relatively large quantities of water and still impart a decided “body” t o t h e mixture. Their use is specifically, then, as thickeners and binders in such food products as marshmallow preparations, ice cream, custards, pie fillings, egg substitutes, a n d flavoring emulsions. The gums ordinarily employed are gum arabic, gum tragacanth, Indian gum, agar-agar, a n d commercial dextrin. Gelatin, egg albumin, a n d . commercial glucose, as well as starch, are also used for t h e same purpose. EXISTING METHODS
The methods which have been proposed for t h e detection of this class of materials are based for t h e most p a r t on isolated reactions for a particular gum, depending on some color or solubility test of t h e crude gum itself , a n d having no -reference t o t h e detection of small amounts of t h e gum in a complex food mixture. Of t h e few t h a t are more general perhaps t h e best . 1
THIS JOURNAL, 4 (1912), 600.
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T H E JOURNAL OF INDUSTRIAL A N D ENGINEERING CHEMISTRY
known are those proposed b y Patrick,’ and Howard,z and t h e group scheme devised b y C ~ n g d o n . ~The first two of these were suggested only for ice cream and, while quite simple a n d easily performed, are unsatisfactory for complicated food mixtures and entirely useless so far as identification of t h e gum is concerned. The more pretentious scheme of Congdon appeared quite promising and was given a thorough trial on food products and on known mijYtures with gums. The results obtained were, however, very disappointing. Congdon’s procedure is evidently based on qualitative tests made on t h e crude gums, no provision being made for separating t h e gum from t h e other components of t h e complex food mixture, no matter how seriously these interfere with precipitation or color tests. Further, some of t h e tests included in t h e scheme were found t o be neither specific nor conclusive even with t h e gum itself. SEPARATION O F T H E
GUM
As t h e basis for a workable scheme i t was decided a t t h e outset t h a t on account of t h e complex food mixtures t o which gums are added, all tests for t h e identification of t h e thickener should be limited t o tests made on t h e relatively pure gum substance, previously separated from t h e food product. This, of necessity, eliminates many of t h e tests described in t h e literature for t h e individual gums, most of which are dependent on impurities naturally occurring in t h e raw gums, a n d limits t h e available reactions for a n orderly scheme largely t o t h e precipitation tests. All such tests t h a t could be found in t h e literature, a n d t h e action of all available solvents were carefully studied on solutions of gum arabic, agar, gum tragacanth, Indian gum, dextrin, gelatin, a n d egg albumin. Since Indian gum is not so specific a term as “arabic” or “tragacanth” and includes a t least two different species, two samples of this g u m , of entirely different appearance a n d obtained from different sources, were used. After much experimentation, which need not be detailed here, t h e following systematic procedure was finally adopted for t h e separation of t h e gum in a comparatively pure condition from t h e food product. This procedure coiisists, in brief, in precipitating t h e protein of t h e food mixture b y heating with acetic acid and tannin, and t h e n precipitating t h e gums from the filtrate b y acetone. I n this way t h e sugars and other- acetone-soluble materials are left in t h e filtrate. Since milk is a common ingredient of t h e class of foods in question, soluble phosphates have also t o be removed b y an extra precipitation with ammonia. Finally, t h e redissolved gums are precipitated relatively pure b y alcohol. The procedure is summarized in Table I. TABLEI-THE SEPARATION OF GUMS A-ELIMINATION
OF PROTEINS
I-Dilute
sample to suitable concentration with water, add 5 cc. dilute acetic acid and 25 cc. of IO per cent tannin solution, and heat mixture for 20 to 30 min. Centrifuge and filter. Discard precipitate.
1
U. S. Dept. of Agr., Bur. of Chem., Bull. 116, 24.
2
J. A m . Chem. Soc., 29 (1907). 1622.
8
THISJOURNAL, 7 (1915), 606.
5.3 1
Note-Casein, coagulable proteins, and some of the gelatin precipitated. Fats and other insoluble substances included in precipitate. 2-Add 40 t o j o cc. more tannin solution t o filtrate from X I and heat for short time. Centrifuge and filter. Discard precipitate. Note-Remainder of gelatin and soluble proteins precipitated. B-SEPARATIONOF
GUMS AND DEXTRIN FROM SUGARS
I-Treat clear filtrate from Az with twice its volume of acetone. Centrifuge and filter. Discard filtrate. Wash precipitate twice with acetone. Note-Precipitate includes gums and dextrin. h-0 precipitate shows absence of gums, dextrin, and milk solids. z-Dissolve precipitate from B I in 50 cc. of warm water slightly acidified with acetic acid and add I O cc. of ammonia (sp. gr. 0.90). Centrifuge and filter. Discard precipitate. Note-Calcium phosphate from milk solids precipitated. C-ISOLATIONOF
PURE GUM SUBSTANCE
Add acetic acid to filtrate from Bz until slightly acid. Add alcohol, one volume at a time, until a well defined precipitate appears. Note-Gums and dextrin precipitated in fairly pure condition. No precipitate with five volumes of alcohol indicates absence of gums and dextrin. Within certain limitations, which will be discussed later, this procedure is capable of separating gums and dextrin from complex food mixtures. I n t h e numerous experiments on which i t was based t h e amount of gum present varied from 0.1 t o over 1.0 g. and t h e weight of sample from j o t o 2 0 0 g. It is certain t h a t amounts of gum as small as 0.1 g. can be separated b y t h e procedure from ordinary food mixtures. It should be remembered in this connection, however, t h a t some gums are more readily detected t h a n others when present in equivalent amounts. Tragacanth, for example, is much easier t o detect in small quantities t h a n either arabic or agar. The relation of t h e amounts of other precipitable matter, especially protein, is also of some importance since t h e gums tend t o be carried down mechanically in t h e precipitation of protein, hence t h e ratio of protein t o gum may be so great t h a t t h e procedure will fail t o detect t h e gum through mechanical loss. T.4BLE 11
Approximate Volumes of Alcohol Necessary for Precipitation Characteristic Vols. Al- Vol. Gum Appearance of coho1 Solution Gum Precipitate Agar.. . . . , 3-4 1 Finely divided white precipitate; settles very slowly 1 White flocculent preArabic.. . . , 2 cipitate. settles quickly: neither sticky nor coherent Indian, . . , 2-3 1 Stringy precipitate; becomes very coherent after settling Tragacanth 2 1 Coherent, jelly-like mass; floats in clots in upper part of solution Dextrin.. . 3 1 White, fine precipit a t e ; settles slowly; very sticky IDENTIFICATION
Characteristics of Gum Precipitate After Standing for Some Time in Air Usually remains soEt and non-coherent Becomes dry powdery
and
Becomes dark colored: toueh coherent layer Flatteps down, becoming a semitransparent coherent layer Tends t o become hard on long standing
O F T H E GUM
Certain of t h e precipitation tests for t h e gums, which have been used as t h e basis for t h e foregoing method of separation, serve also fairly well for t h e
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identification of the gums. It suffices in general t o note t h e approximate volume of alcohol required for the final precipitation and the nature and appearance of t h e alcohol precipitate. I t is advantageous in this connection t o pour off most of t h e alcohol after t h e precipitate has settled and allow the moist gum precipitate t o stand exposed t o the air for a short time, noting its behavior when drying. Table I1 presents in concise form the characteristic differences which ’ are shown b y the common gums. C 0 N F I R M AT 0 R Y T E S T S
While the characteristic differences described in the preceding table have been the chief reliance in identifying the gums, a number of the tests described in the literature have been examined t o determine their value as confirmatory tests. Most of these, as previously stated, depend on impurities present in the crude gum and hence, as would be expected, proved of little value when applied t o t h e separated gum precipitate. Several procedures, however, were found even under these conditions t o be distinctly helpful. Chief of these was t h e presence of characteristic diatoms in t h e agar. The test is a well-known one and consists in identifying under the microscope the peculiar diatoms which are associated with agar. chiefly Arachnoidiscus Ehrenbergii and various species of Cocconeis, after destroying the organic matter b y digestion with acid. The characteristic appearance of these diatoms will be found figured in most standard texts on food analysis. The procedure consisted in destroying t h e organic matter of the sample by heating with nitric and sulfuric acids until t h e solution became colorless, diluting the concentrated acid solution with water, centrifuging t o collect the siliceous residue, and examining this under the microscope. The test may be applied t o the original material, b u t much time will be saved by using the tannin precipitate obtained in A I of Table I. Since this precipitate is separated b y the centrifuge i t will obviously contain all t h e relatively heavy particles, including the diatoms, and its use will eliminate t h e interference due t o soluble carbohydrates, as cane sugar, commercial glucose, etc., which use up time and acid in the digestion. This procedure was tried repeatedly on many samples of agar including t h e purest bacteriological material, and on food mixtures containing agar, and the presence of the characteristic diatoms noted in every case. Although this test actually depends on t h e presence of ‘(impurities” in t h e gum, it was found t h a t t h e diatoms are so widely distributed in commercial samples and remain so consistently in the tannin precipitate t h a t t h e test was most useful. The volatile acidity of Indian gum was also found of value as a confirmatory test. It has been noted by several authors t h a t the species of gum coming under the classification of ‘Indian gum have the characteristic property of developing an acetic odor when exposed t o t h e air, and Emery’ has made this characteristic the basis of a method for t h e detection of 1 T H I S JOURNAL,
4 (1912), 374.
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Indian gum as an adulterant of gum tragacanth. The method consists, in brief, of accelerating t h e hydrolysis of t h e gum by heating with acid, distilling, and titrating the acetic acid produced. Emery gives t h e following typical figures, expressed as cc. of N / r o acid per gram of gum: Tragacanth, . . . . , , . . 3.2- 4.2 Indian gum
. . . . 25.4-28.3
Emery’s method w o t h e dried gum precipitates obtained ’ in the systematic procedure of Table I and t h e following results were obtained: TABLE111 Volatile Acidity
.......... ...........
0.25
These figures show t h a t t h e differences found by Emery hold for the precipitated gums, althbugh t o a less marked degree. The method, although rather long and tedious, has distinct value as a confirmatory test on gum precipitates when Indian gum is suspected and t h e characteristics described in Table I1 are not conclusive. LIMITATIONS O F T H E M E T H O D
The delicacy of t h e method under favorable conditions has been pointed out in a previous paragraph, There are, however, certain limitations t o its usefulness which should be definitely noted. FiFst, t h e successful use of the procedure for t h e identification of t h e gums depends primarily upon t h e ability t o recognize t h e different visible characteristics of t h e gum precipitates. It is, therefore, highly desirable t h a t before using t h e procedure for analytical purposes the analyst should know the various gum precipitates (‘by sight.” This can be readily accomplished with t h e aid of a few prepared solutions of t h e gums concerned. The identification of t h e gums where more t h a n one is present in the food mixture is a more difficult matter. With some combinations of gums a partial separation can be accomplished by the fractional precipitation of t h e procedure, b u t with such a combination as dextrin and agar-agar, or Indian gum and tragacanth, this would be practically impossible, although there might be indications as t o t h e presence of both gums. A more serious matter is t h e possible presence of two substances which interfere distinctly with t h e separation and identification of the true gums. These are pectin and commercial glucose. The first of these is peThaps less important since, although it is somewhat similar t o t h e gums and would t o a certain extent be precipitated with them in t h e procedure, few of t h e commercial food products in which gums are used ordinarily would be likely t o contain fruit pectins. The most likely combinations would be fruit pie fillings and jellies and jams made from apple stock, food products in which t h e presence of gums h a s been noted.
July, 1918
T H E J O C R N A L O F I N D U S T R I A L A *VD E N GI N E E R I N G C H E M I S T R Y
I n t h e few cases where from t h e source of t h e sample t h e presence of pectin might be expected, its removal prior t o t h e alcoholic separation of t h e gums may be aided b y observing t h e following precautions: ( I ) I n adding t h e acetic acid t o t h e ammoniacal solution (C of Table I ) , i t should be added slowly, t h e mixture stirred, and allowed t o stand for some time with occasional shaking. T h e removal of pectin a t this point may be accelerated b y adding a few drops of a tannin-iron solution before t h e ammonia and acetic acid treatment. ( 2 ) Small amounts of alcohol, one-quarter t o onehalf of a volume, are added with stirring t o t h e ammoniacal solution. (3) When t h e amount of iron present is slight, judging from t h e color of t h e solution, as well as the precipitates, a few drops of a ferric chloride solution are added t o t h e aqueous solution of t h e acetone precipitate ( B ~ - o fTable I). Of greater practical importance is t h e presence of commercial glucose in products which also contain gum, a combination which is common in commercial products like marshmallow creams. The interfering factor here is of course dextrin a n d this is precipitated with t h e final gum precipitate b y alcohol, although the scheme of fractional precipitation outlined in Table I should give some indication of its presence. Numerous tests have shown t h a t in t h e case of Indian gum and gum tragacanth commercial glucose interferes b u t little with their detection a n d identification, even when t h e ratio between t h e quantities of glucose and gum is as high as 40 t o I for Indian gum and 1 2 0 t o I for gum tragacanth. I n t h e case of gum arabic t h e interference of commercial glucose is distinctly noticeable when t h e ratio of glucose t o gum is 2 0 t o I, a portion of t h e dextrin precipitating with 2 volumes of alcohol along with t h e gum arabic, and b y its sticky character masking t h e flocculent, noncoherent characteristic of t h e gum arabic. With agar, although no experiments were carried out, t h e interference would be still greater. Since t h e amount of commercial glucose present in a marshmallow paste, for example, is likely t o exceed t h e ratio given for gum arabic, in such cases an additional step in t h e procedure may be needed. This additional step is based on t h e fact t h a t dextrin is more readily hydrolyzed b y boiling with dilute acid t h a n are t h e gums, I t is carried out on t h e precipitate obtained with two volumes of alcohol, which will contain t h e greater part of t h e gum arabic and a portion of t h e dextrin. 0.5 g. of t h e dried gum precipitate is heated for j min. with 50 cc. of water and 2 . j cc. of concentrated hydrochloric acid (sp. gr. 1.20). The hydrolysis is conveniently carried out in a large test t u b e immersed in boiling water. Experiments on known mixtures have shown t h a t in this way one p a r t of gum arabic may be detected in a mixture with 4 parts of dextrin, a delicacy which allows t h e detection of t h e gum in t h e presence of a considerable proportion of commercial glucose. Further, i t must be remembered t h a t t h e precipitate ob-
533
tained with 2 volumes of alcohol would not contain t h e whole of t h e dextrin involved, for t h e larger p a r t of this, as has been shown, would come down only with t h e third volume of alcohol. The precipitate t o be hydrolyzed would, therefore, contain t h e gum with a small amount only of t h e dextrin present in t h e original mixture. Without question. then, t h e procedure is quite delicate and is capable of detecting relatively small quantities of gum arabic in t h e presence of commercial glucose. SU M M A R Y
A method is described for t h e separation of t h e more common gums from food products based upon elimination of proteins b y acetic acid and tannin and precipitation of the gums b y acetone a n d finally alcohol. The separated gums are identified mainly b y their fractional precipitation with alcohol and t h e characteristic appearance of t h e precipitated pure gum. The necessary modification of t h e method in t h e presence of such interfering substances as milk solids, pectin, and commercial glucose is described. T h e method described is capable of detecting with ordinary commercial products 0.1 g. of gum in IOO g . of a complex food mixture. MASSACHUSETTS INSTITUTE O F CAMBRIDGE,
TECHNOLOGY
MASS
UNIFORM NITROGEN DETERMINATION IN COTTONSEED MEAL By J. S. MCHARGUE Received April 13, 1918
Chemists often have trouble in obtaining uniform results in duplicating nitrogen determinations on Cottonseed meal. Since cottonseed meal is so extensively used as a source of protein i n feeds, i t is a matter of considerable importance whether or noc all serious errors have been eliminated in a nitrogen determination on this material. The object of this paper is t o call attention t o procedures common among chemists which are often t h e cause of considerable error in t h e determination of nitrogen in cottonseed meal. During t h e past year t h e writer has been called upon t o check a number of cottonseed meal samples in which t h e amount of nitrogen was in question. On a few of these samples duplicate determinations of nitrogen showed variations ranging from as much as 0.50 t o 1.50 per cent of protein, while other samples of cottonseed meal gave almost identical results upon duplication. The samples upon which varying results were obtained naturally suggested further investigation in regard t o t h e cause of t h e variations. Self1 has shown t h a t nitrogen can be lost b y volatilization during t h e digestion if a large excess of potassium sulfate has been added. He attributes this loss t o t h e formation of K H S 0 4 when a n excess of HzS04 has been boiled off. The following experiments were made t o determine whether or not nitrogen was lost in a cottonseed meal 1
Pharm. J . , 88, 384-5; Chem. A b s . ( A m e r . ) ,6 (1912), 2048.
