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955
probably close enough for most practical purposes, and also t o t h e thousandth of a milligram (the figures in t h e parentheses) where t h e amount given or found was not in even hundredths. The average error (neglecting t h e algebraic signs) was practically t h e same (0.003 mg.) whether we accept the figures expressing t h e amounts of aniline t o t h e nearest hundredth of a milligram or t o t h e thousandth of a milligram. These results further confirm t h e general principle t h a t from the point of view of percentage accuracy i t is well t o work with as large amounts as possible within t h e given range, since a n error of only 0 . 0 0 3 mg. in t h e case of No. 6 shows up as a much greater percentage error t h a n even over three times this amount ( 0 . O I O mg.) in t h e case of No. 3. Finally, when we recall t h a t by t h e ordinary application of t h e hypochlorite test for aniline even t h e solution with t h e highest of t h e above amounts of aniline (No. 3) would not show even qualitatively t h e presence of any aniline while t h e proposed method measures such small amounts quantitatively, and the further fact t h a t this method does not require elaborate apparatus but is very simple and quickly and easily carried out, its advantages when one has occasion t o detect and estimate such small amounts of aniline, become very apparent.
used; 2 0 cc. of this solution was then mixed with I cc. of calcium hypochlorite solution,* t h e available chlorine of which was 0 . I per cent. After standing 2 minutes, i t was mixed with I cc. of N S a O H and allowed t o s t a n d I O minute^.^ The color of this solution was then compared with those of standards, which were obtained by treating similarly known amounts of a standard aniline solution. The mixing with t h e reagents a n d t h e final reading of t h e color was carried out in t h e same tube, t h e narrow form j o cc. Nessler tubes having been found convenient for this purpose. The mixing of t h e unknown solution and t h e standards with t h e reagents was effected nearly simultaneously with t h e aid of bulbed glass rods, b y means of which i t was found comparatively very easy t o mix thoroughly t h e contents of four or five tubes practically simultaneously. The standards were prepared from a stock solution of pure aniline in distilled water, which contained I gram aniline in 1,000cc. of solution. The regular4 standards contained 0, 0.01,0 . 0 2 , 0 . 0 3 , 0.04,0.05, 0 . 0 6 and 0.07 mg. aniline, respectively, a n d t h e volume of each, exclusive of t h e reagents, was 2 0 cc. RESULTS OBTAINED
The following results were obtained by t h e above procedure when working with aqueous solutions of aniline, t h e amount of aniline in which was unknown5 t o t h e writer a t t h e time t h e work was done.
HYGIENICLABORATORY U. S. PUBLICHEALTHSERVICE WASHINGTON, D. C.
MILLIGRAMS ANILINEIN 20 Cc Given Found Error 1 .......... 0.04 0.04 0.00 (0.000) 2. 0.02 (O:Oi4) 0.02 ( 0 : 0 2 3 ) 0.00 (-0.001) 3 0.18 ... 0.17 -0.01 (-0.010) .......... 0 . 1 1 ... 0.11 (o:ios) 0.00 (-0.00s) 3 .......... 0.03 (0.032) 0 . 0 3 (0.032) 0.00 (0.000) 6 0.01 (0.014) 0.02 (0.017) 4-0.01 (C0.003) AVERAGE. .................................... 0.003 0.003
KO.
A MODIFICATION OF THE PRICE METHOD FOR THE SEPARATION OF THE PERMITTED COAL-TAR
......... ..........
4 ..........
As will be noted, t h e amounts of aniline are expressed t o t h e nearest one-hundredth of a milligram, which is Since i t is important that the unknown solution and the standards be mixed with the reagents as nearly simultaneously as possible, it is well to use pipettes or burettes with comparatively large outflow openings, so as to consume as little time as possible for the adding of the reagents. 2 This solution was prepared by suitably diluting with water a stock solution, the available chlorine of which was about 3 per cent, which was obtained by shaking thoroughly for 10 minutes 20 g. of a sample of commercial chlorinated lime with 100 cc. water and then filtering. This stock solution shouid not be prepared from a chlorinated lime which has lost much of the available chlorine that it originally had, since it might leave too much calcium in the final reagent and thus cause a precipitate or turbidity when the excess alkali is added. 8 In the case of the standard containing the smallest amount of aniline (0.01 mg.), reducing this time of standing to 5 minutes appears advantageous. 4 For most practical purposes these standards are probably sufficient, since in case the color of the unknown solution happens to fall between two of the above standards one can, especially after some experience, judge which of the two standards it approaches nearer and assign i t a proper intermediate value which will probabiy be close enough for most practical purposes. When dealing with the middle and especially the lower end of the above series of standards, however, since there would be greater possibility for a larger percentage error, i t is advisable to confirm any such assumed value by actually preparing t h e corresponding standard and one a little above and another a little below t h a t value and repeating the simultaneous treatment with the reagents. In order to enable one t o prepare quickly and easily such intermediate standards, i t is advisable to have on hand a more dilute solution of aniline than the above mentioned stock solution Such a solution was prepared by diluting 10 cc. of the stock solution to 1000 cc. with distilled water,which has also the advantage t h a t the volume used in the preparation of any standard, expressed in cc., gives also t h e value of t h a t standard in hundredths of a milligram of aniline. By keeping some of this dilute solution in an appropriate burette, any desired intermediate standard was quickly and easily prepared. These solutions were submitted t o the writer as “unknowns” by Dr. A. Seidell. of this laboratory. 1
COLORS TO INCLUDE TARTRAZINE By E. H. INCERSOLL
,
Received July 2, 1917
Food Inspection Decision 76, issued by t h e U. S. Department of Agriculture in 1907, enumerated seven coal-tar colors t h a t could be used in foods under certain restrictions. Price’ in 1911 described a method for t h e separation of these colors when they occurred in mixtures. Recently, under Food Inspection Decision 164, there has been another coal-tar dye added t o t h e seven previously permitted, namely, Tartrazine. The including of this additional color among t h e permitted ones makes i t necessary for t h e analyst t o have available a method whereby these eight coal-tar dyes may be separated and identified in cases where they occur in mixtures. Estes2 has stated t h e necessity for such a method and gives a modification of t h e method described by Price t o include this additional color. By this method Amaranth is separated from Tartrazine by saturating the aqueous solution of these two dyes with sodium chloride; however, i t has been observed t h a t when b u t small amounts of dye have been taken out of t h e original mixture on extraction with t h e ammonium sulfate reagent, t h e separation is very difficult if a t all attainable. Furthermore, while some Tartrazine, like Naphthol Yellow S, is soluble in saturated ammonium sulfate solution, t h e larger part is not extracted b y this reagent, if t h e Price direc1
U. S. Department of Agriculture, B. A. I. Circular 180.
2
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T H E J O U R N A L OF I N D U S T R I A L A N D ENGINEERING C H E M I S T R Y
Vol. 9 , No.
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tions t o wash until t h e washings are no longer red are adhered t o , b u t remains insoluble on the filter. As t h e method of separation described by Estes proved unsatisfactory in t h e hands of t h e author, work was undertaken t o devise a method which would be entirely satisfactory. I n undertaking this work, it was thought best t o take t h e Price method as a basis, and, if possible, so t o modify it as t o include all of t h e eight permitted coal-tar colors. The method described below is a result of this work and is given only after it has been tried by several analysts working independently and with satisfactory results on samples of unknown mixtures of coal-tar colors. R u b from 0.I t o 0 . 2 g. of t h e dye sample, depending upon t h e amount of foreign salts in t h e mixture, with 2 5 cc. of saturated ammonium sulfate solution in a mortar and filter through a dry filter. If t h e filtrate comes through red, wash t h e color residue in t h e mortar a n d on t h e filter with successive I O t o 15 cc. portions of t h e ammonium sulfate solution until t h e washings are no longer colored red. The filtrate and washings contain t h e greater part of the Amaranth together with some Naphthol Yellow S and also some Tartrazine. Combine t h e filtrate and washings and shake with successive portions of acetic ether until t h e acetic ether is no longer colored yellow. The acetic ether removes t h a t portion of t h e Naphthol Yellow S which was dissolved b y t h e ammonium sulfate solution and may be discarded, since t h e greater part of this dye is recovered later in t h e scheme. Shake t h e ammonium sulfate solution containing Amaranth and some Tartrazine with acetone t o remove these colors; discard t h e ammonium sulfate solution, dilute t h e acetone portion with an equal volume of water and drive t h e acetone off on a steam bath. Saturate with
in excess; cool and filter through a dry filter. Wash with saturated sodium chloride solution until the washings are colorless. When a bulky precipitate is obtained here, which is difficult t o wash, it may be timesaving t o redissolve t h e precipitate and excess salt in water and repeat t h e salting and washing process, adding t h e filtrate and washings t o those of t h e first saturation. T h e combined filtrate and washings contain Light Green S F Yellowish, Naphthol Yellow S, Tartrazine, trac'es of Orange I , and possibly Amaranth, since t h e latter dye may not be entirely removed by the first extraction of t h e d r x sample with t h e ammonium sulfate reagent. To separate the Naphthol Yellow S,extract with successive portions of acetone until t h e acetone fails to remove any more color. Combine t h e acetone extracts and wash with several portions of saturated sodium chloride solution t o remove traces of Tartrazine and Light Green S F Yellowish from t h e acetone. Add t o t h e acetone solution an equal volume of water and drive off acetone on t h e steam bath. Acidify t h e water solution and shake with amyl alcohol t o remove traces of Orange I t h a t may be present; discard this amyl alcohol solution. Drive off all t h e amyl alcohol mechanically held in the aqueous solution by warming on t h e steam bath and test this solution for Naphthol Yellow S. T o separate the Light Green S F Yellowish from t h e Tartrazine, remove the acetone from t h e aqueous salt solution by heating on t h e steam bath, and add fuller's earth in t h e proportion of 0 .5 g. t o each I O cc. of warm dye solution. After mixing well and heating, allow t o settle; filter and wash with water. The Light Green S F Yellowish remains on t h e filter and can be dissolved in strong, hot acetic acid and further
warm, settle, filter a n d wash with warm saturated sodium chloride solution until t h e washings are no longer colored yellow. T o recover t h e Amaranth, suspend the' alumina cream precipitate in saturated ammonium sulfate solution a n d shake with acetone. The filtrate contains Tartrazine and is t o be discarded, or, when dealing with small amounts, if desired, can be saved and t h e Tartrazine identified with t h e greater portion of this color separated further in t h e scheme. Dissolve t h e portion of t h e original sample not dissolved b y ammonium sulfate, in water, acidify with acetic acid and shake with successive portions of ethyl ether until t h e ether is no longer colored. T h e ether contains erythrosine, which it is very essential t o remove completely from t h e other dyes before proceeding further. Wash t h e ether solution several times with water and finally extract t h e erythrosine from t h e ether with dilute ammonia solution. Remove t h e ammonia by evaporation on t h e steam bath a n d observe if this solution, when very dilute, has a n y fluorescence which might indicate t h e presence of prohibited colors having similar reaction. Remove t h e ether from t h e acetic acid aqueous solution by warming on a steam bath, saturate with sodium chloride a t steam b a t h temperature and add sodium chloride
mixture, t h e filtrate from t h e precipitation of t h e Light Green S F Yellowish will be yellow or golden yellow, not decolorized by hydrochloric acid. Imperfect removal of Naphthol Yellow S, previously, would result in a yellow filtrate here which could be decolorized by hydrochloric acid. The Tartrazine can be further isolated from a possible trace of Amaranth by adding I O cc. of alumina cream t o each IOO cc. of solution, mixing, warming and filtering, when t h e Tartrazine will be found in t h e sodium chloride filtrate. T o isolate from t h e salt, evaporate and redissolve in alcohol. Dissolve t h e precipitant containing Orange I , Ponceau 3R and Indigo Disulfo Acid, together with excess sodium chloride o n t h e filter paper, in water and extract with three successive portions of acetic ether. Orange I is taken up by acetic ether. Combine t h e acetic ether extracts and wash with saturated sodium chloride solution, until no more color is removed. Extract t h e acetic ether solution with water t o obtain the Orange I in a n aqueous solution and free from acetic ether by warming on the steam bath. Warm t h e water solution .containing Ponceau 3R and Indigo Disulfo Acid from which t h e greater part of the Orange I has been removed, on t h e steam bath until free from acetic ether, cool, add I O g. gran-
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T H E J O U R N A L OF I N D U S T R I A L A N D E N G I N E E R I N G C H E M I S T R Y
ulated calcium chloride, allow t o s t a n d 1 5 minutes a n d then a d d 1 5 cc. of a freshly prepared stannous chloride solution containing t h e equivalent of 3 per cent metallic t i n a n d 1 2 per cent of hydrochloric acid (sp. gr. I . 19). Mix well a n d allow t o s t a n d until t h e solution shows no blue color. If Ponceau 3R is present, i t will be precipitated. Filter immediately, wash t h e precipitate twice with 2 5 per cent calcium chloride solution t o remove all t h e reduced Indigo Disulfo Acid, dissolve t h e remaining residue in dilute ammonia solution a n d test for Ponceau 3 R . T o t h e filtrate, which should be practically colorless, a d d 3 per cent hydrogen peroxide solution. A deep blue coloration indicates t h e presence of Indigo Disulfo Acid. MEATINSPECTION LABORATORY D.C. WASAINGTON.
DETECTION OF ADDED WATER IN MILK BY MEANS OF A SIMPLIFIED MOLECULAR CONCENTRATION CONSTANT' By LESLIEW. FERRIS Received May 31, 19 17
It is a known fact t h a t t h e osmotic pressure of t h e blood serum of a healthy cow is practically constant, a n d naturally t h e osmotic pressure of t h e milk must be regulated b y t h a t of t h e blood. The bulk of t h e osmotically active substances in t h e milk are lactose a n d soluble salts, chiefly t h e chlorides of t h e alkali metals. Since t h e quantity of lactose in milk varies considerably t h e chlorides would be expected t o vary inversely t o t h e lactose. Therefore, a figure t h a t would represent both lactose a n d chlorides in isotonic equivalence ought t o be fairly constant. Mathieu a n d Ferre2 have embodied this principle in t h e calculation of a simplified molecular concentration constant which t h e y use as t h e basis of a method for t h e detection of added water in milk. Giving I g. of sodium chloride t h e isotonic equivalence of 1 1 . 9 g. of crystallized lactose they obtain a n apparent cons t a n t by adding t h e grams of lactose per liter of milk t o t h e grams of sodium chloride per liter X 11.9. They t h e n correct this apparent constant for t h e volu m e of t h e f a t a n d casein, using 0 . 9 4 as t h e specific gravity of fat and I . 35 as t h e specific gravity of casein. Mathieu a n d Ferre conclude t h a t this simplified molecular constant varies between 74 a n d 79 for genuine milk and falls below 73 if t h e milk contains 5 t o 8 per cent added water. Since t h e completion of this work i t is noted t h a t Mathieu3 has found values between 79 a n d 82 for 13 out of a new series of 93 samples taken in summer. Monier-Williams4 has calculated t h e Mathieu a n d Ferre constant for a number of samples of milk from individual cows. After eliminating certain samples t h a t were known t o be of abnormal character h e found values for t h e constant varying from 7 0 . 0 t o 7 8 . 1 . 1 Published by permission of the Secretary of Agriculture; read at the meeting of the American Chemical Society, New York City, September 25 to 30. 1916. Ann. Pals., 7 (1914). 12-21. 8 I b i d . . 9 (1916). 45-8. 4 Report of the Local Government Board, Great Britain. No. 22 (1914), f
27-9.
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The object of this work was t o test t h e procedure upon different grades of American milk. T h e molecular concentration constant was determined upon 3 I samples of milk, representing complete milkings from several breeds of cows. The different samples of milk varied considerably in quality, as is shown b y t h e fact t h a t t h e percentage of solids-not-fat ranged from 7 . 6 3 t o 9 . 5 2 . On all of t h e samples analyzed fat was determined b y t h e Babcock Method, casein b y t h e Official Method of t h e Association of Official Agricultural Chemists, and t h e specific gravity taken with t h e lactometer. Sodium chloride was determined b y t h e Volhard Method on a copper sulfate serum, as used b y Poetschke,' with t h e exception t h a t i t was found best t o weigh instead of measure t h e sample of milk. Lactose was determined in t h e copper sulfate serum b y t h e Munson and Walker Method. No correction was made for t h e volume of t h e precipitate produced by t h e copper sulfate, since this error was found t o be well within t h e error of t h e manipulation. Duplicate determinations of lactose and sodium chloride were made on 30 samples and t h e two corresponding figures for t h e molecular concentration constant calculated, as shown ip Table I. The greatest difference was 0 . 6 (Sample 16) and t h e average experimental error on t h e 30 samples was 0 . 2 2 . F/G. I-Re/ation of Lactose and Sodium 6h/on& B
A
14 IS 16 I7 /8 I9 20 2/ 22 23 24 25 26 27 28 29 30 3/ 32 33 54 3SfC
Fig. I shows t h a t within certain limits t h e per cent of sodium chloride gradually rises as t h e lactose decreases. T h e grams per liter of lactose are represented as ordinates a n d t h e grams per liter of sodium chloride multiplied by I I , 9 as abscissae. When t h e experimental results are plotted it is seen t h a t they approach a constant represented b y t h e line A - B . Table I gives t h e analyses of t h e samples of milk with t h e three figures t h a t are used as a basis for t h e detection of added water: the percentage of solidsnot-fat, t h e refraction and ash of t h e sour serum a n d t h e molecular concentration constant. The values for t h e molecular concentration constant vary between 7 1 . 1 and 8 2 . 6 . T h e two high figures for t h e constant as shown in Samples 28 and 30, C O W NO. 3 , were obtained upon milk t h a t was abnormally high in salt. Cow No. 3 was near t h e end of t h e lacta1
THISJOURNAL, 3 (1910), 210-2.
