November, 1924
INDUSTRIAL AND EATGINEERING CHEMISTRY
1181
Vitamin Potency of Cod-Liver Oils’ X-Medicinal Cod-river Oils By Arthur D.Holmes THE E I,. PATCHCo., BOSTON,MASS.
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INCE it has been genA number of bottles of oil were purchased on the open market, and erally accepted that the chemical and physical characteristics of these oils were detercod-liver oilis the richmined by the usual analytical methods. A study of their vitamin est known source of the fatpotency showed that the vitamin content of medicinal cod-lioer oils soluble vitamins, those who may vary as much as tenfold. Apparently, there is little, if any, reuse cod-liver as a source lationship between the chemical and physical characteristics of codof the fat-soluble vitamins liver oils and their oitamin potency. These results show the need of for either prophylactic or information concerning the amount of the fat-soluble vitamins prescurative purposes are much ent in cod-liver oil to be used in vitamin therapy. interested inhaving as exact i n f o r m a t i o n as possible concerning the vitamin content of edible cod-liver oils. Unfortunately, as yet there is relatively little information of this character available. Recently Drummond, Zilva, and Golding reported2 the results of tests of the vitamin potency of a group of cod-liver oils that ranged in quality from medicinal to cattle or industrial oils. Their unit for comparison was the weight in milligrams of oil which just maintained steady growth in a test rat of about 100 grams body weight. These authors found their most potent oil to have a “growth dosage” of 5 mg., and their poorest oil a “growth dosage” of I O 50 mg. Obviously, the first oil had ten times the vitamin potency of the second, and consequently could be administered in doses of one-tenth the volume required of the second oil. Information of this character naturally raises a question as to the vitamin potency of the medicinal cod-liver oils on the market in this country. It was to secure data in this connection that the present investigation was undertaken.
PHYSICAL AND CHEMICAL AND VITAMIN PROPERTIES POTENCY
In studying the vitamin A content of this group of Norwegian cod-liver oils, it appeared of interest to determine whether any rela-
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PHYSICAI, AND CHEMICAL CHARACTERISTICS OR NORWEGIAN OILS COMPARED WITH THEIRVITAMIN POTENCY Specific Refrac- SaponifiAcid Cod-liver gravity tive index cation Iodine value oil feda Sample a t 25’ C. a t 20O C. value number Per cent Mg. 1.4786 8 184.0 1.2760 164.4 0.788 185.7 9 0.6725 1.4790 165.6 0.105 1.4779 10 186.1 2.3110 151.7 0.336 1.4784 11 181.4 1,4300 157.3 1.815 26 1.4780 188.3 0.5520 154.7 0.176 27 186.8 1.4798 169.0 0.072 0.6278 37 1.4778 187.4 0.7205 152.3 0.296 43 185.4 1.4790 164.3 0.5481 0.144 44 186.5 1.4790 0.5147 164.0 0.340 1.4794 80 186.1 0.7391 171.4 0.278 a This figure represents merely the smallest amount of oil given t o an animal whirh completed the 45-day experimental period. I t does not represent the amount of oil required t o produce good growth for, as shown in the charts, some of the rats receiving these amounts of oil lost weight during the entire experimental period.
To provide samples that would be fairly representative of the present-day product, ten lots of cod-liver oil were obtained on the open market. Samples 10 and 26 were from the same concern; the remainder were distributed by different companies. The label on each package guaranteed the oil to be pure h’orwegian oil of highest quality. Apparently, the majority of the oils were offered to the customer merely as a definite volume of cod-liver oil, for the labels carried no statement concerning the vitamin potency of the oil contained in the package. I n all instances in which a statement was made concerning the vitamin potency of the oil, tests in this laboratory showed that the oil had a higher vitamin A potency than was guaranteed. 1 Presented before the Division of Biological Chemistry a t the 67th Meeting of the American Chemical Society, Washington, D. C . , April 21 to 26, 1924. 2 J. A g r . Sci., IS, 157 (1923).
index is quite uniform, and the saponification value and iodine number are both within the U. S. P. specifications (saponification between 180 and 190, and iodine number between 140 and 180). The greatest variation is in the acid value of the oils, which is from 0.514 to 2.311 per cent. In the absence of data concerning the condition of the livers from which the oils were obtained, the manufacturing and storage conditions, and especially whether any of the oils have been
Vol. 16, No. 11
INDUSTRIAL A N D ENGINEERING CHEMISTRY
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VITAMIN POTENICY OF LIVER O I L
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Chart 27
N O R W E G I A N OIL * I 1
STARTED L l v E R O I L
STARTEO LIVER O I L
subjected to the alkali-heat renovating process, it is impossible t o draw definite conclusions from the acid values. Assuming that there has been no acid neutralization, the acid values reported indicate that all the oils, except possibly Nos. 8, 10,and 11, represented good grade, medicinal cod-liver oils.
The amount of oil necessary to supply sufficient vitamin A for laboratory animals to live for the usual 45-day experimental period varied from 0.715 to 18.15 mg. On inspection, however, it will be found that the amount of oils essential for minimum vitamin requirement bears no relation to
November, 1924
I N D U S T R I A L A N D ENGINEERING CHEMISTRY
1183 I ChDIt32
VITAMIN POTENCY OF LIVER OIL NORWEGIAH O I L 1 4 3
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VITAMIN POTEN C Y I o F LIVER 01 N O R W E G I A N OIL * 4 4
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Chart 33
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I I I I VITAMIN POTENCY STARTE0,LIVER OIL
any of the physical or chemical values obtained for this series of oils.
TESTSFOR VITAMIN A POTENCY To provide data concerning the vitamin A potency of the oils under consideration, each oil was fed in varying amounts
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to a group of laboratory animals. Ten groups of animals from the laboratory colony were fed the same basal ration, inadequate as regards vitamin A. This diet consisted of casein 18 per cent, peanut oil 22 per cent, cornstarch 28 per cent, lactose 28 per cent, and salt mixture 4 per cent, supplemented by a 0.2-gram tablet of dried brewer's yeast. When the
I N D U S T R I A L A N D ENGINEERING CHEMISTRY
1184
animals commenced to lose weight and gave other evidence that their body reserve of vitamin A was exhausted, their diet was supplemented by uniform daily additions of cod-liver oil. As in other studies of this character, the experimental period was 45 days, a t the end of which the test was concluded regardless of whether the animal was gaining or failing. The amount of oil administered, the amount of food ingested during 5-day periods, and the change in body weight are reported in the charts. Nine animals were used to test the vitamin potency of Oil 8. Of these, four which received less than 5 mg. a day were unable to recover from their malnutrition. The five animals that received 7.88 mg. or more of the oil daily recovered from the effects of their previous vitamin-deficient diet. From these data it appears that about 8 mg. of this oil are required to supply sufficient vitamin A to meet the needs of an albino rat for growth. None of the eight animals that were fed 3.32 mg. or less of Oil 9 were able to recover from the vitamin A privation to which they had been subjected. Considering the results of the series as a whole, one would conclude that between 7 and 14 mg. of this oil would be required daily to produce good growth in this type of laboratory animal. According to the limited results obtained concerning the
Vol. 16, No. 11
vitamin A potency of Oil 10, 3.36 mg. of this oil daily supplied sufficient vitamin A to produce fairly good growth in albino rats. The tests of Oil 26, which was distributed by the same concern, show it to be a more potent oil. Inasmuch as nine animals received amounts of Oil 11 varying from 0.9 to 14.5 mg. daily, and all failed to recover from the malnutrition occasioned by an inadequate supply of vitamin A, it is concluded that more than 14.5 mg. of this oil are needed to meet the vitamin A requirements of the laboratory animals. The charts which report the results of the tests of Oils 27, 37,43,44, and 80, show a quite distinct line of demarcation between those animals that recovered from malnutrition induced by an inadequate supply of vitamin A and those that did not recover. It will also be noted that the amount of the different oils required to effect recovery of the laboratory animals is not identical for the different oils. Accordingly, it is evident that the vitamin content of these oils varies appreciably. I n brief, the tests reported here show that the medicinal cod-liver oils on the market are not uniform in vitamin content, and that if one wishes to be assured of a cod-liver oil of high vitamin potency, it is essential to insist on an oil whose vitamin potency has been determined.
Quantitative Estimation of Anthracene in Anthraquinone' By Harry F. Lewis CORNELL COLLEGE,MT.VERNON.IA.
HE estimation of small amounts of anthracene in an-
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thraquinone is of interest in connection with the use of the anthraquinone in sulfonation. This value can be obtained by analyzing the sample for anthraquinone, then oxidizing a fresh sample, using the Luck method, and estimating the total anthraquinone after oxidation. By subtracting the first value from the second, the amount of anthraquinone formed from the anthracene present may be determined. This procedure is long and tedious, and gives unsatisfactory results. Among the properties of anthraquinone described in the literature may be found the fact that pure anthraquinone dissolves in oleum, producing a yellow color which does not change upon heating the oleum. This is in contrast with the behavior of anthracene, which chars rapidly in hot oleum with the evolution of sulfur dioxide. These facts form the basis of a qualitative test for the purity of anthraquinone as devised by Nelson and Scott. They found that the color of the filtrate obtained after diluting the hot oleum mixture could be used in determining whether a sample of anthraquinone was of satisfactory quality, comparing the color with that obtained in a similar manner with a satisfactory anthraquinone. This qualitative test has been developed into an analytical method for the quantitative estimation of small amounts of anthracene in anthraquinone. PROCEDURE
A 2-gram sample of anthraquinone is taken from a wellground, dry mixture of the unknown and placed in a 15.2 X 2.5-cm. (6 x 1 in.) hard-glass test tube, 25 cc. of 10 per cent oleum added, the stopper containing a thermometer and capillary vent tube inserted, and the tube heated in a bath a t 150' C. for 5 minutes. The contents of the tube are then poured into 500 CC. of distilled water, stirring meanwhile. 1
Received May 15, 1924.
Sufficient of the suspension is filtered to give a color comparison against standards made u p with solutions of potassium dichromate and cobalt chloride. These can be easily made by running known mixtures of anthracene and anthraquinone under the conditions described above and matching their colors in the preparation of standards. The char solution darkens on standing and the color comparison should therefore be made immediately after filtering. ACCURACY The method as described will easily detect as small an amount of anthracene as 0.1 per cent and differentiates easily u p to 7 * 0.2 per cent. This is sufficiently high, for few commercial anthraquinones have an anthracene content greater than 5 per cent, whereas purified anthraquinones have even smaller amounts.
RESULTS The best color results were obtained with 10 per cent oleum. Oleum of 15 per cent strength causes too great a charring to differentiate easily, whereas with hot concentrated sulfuric acid an how's heating is not sufficient to produce much color. Such possible impurities as carbazol, phenanthrene, phenanthraquinone, acenaphthaquinone, and fluorene in amounts up to 2 per cent have no influence on the depth of color. Acenaphthene and naphthalene both char rapidly in 10 per cent oleum, but their presence in commercial anthraquinone is extremely improbable. Naphthalene would sublime in the driers and acenaphthene would be oxidized to the quinone, which does not char. Needless to say, they would be as undesirable in the anthraquinone to be sulfonated as would anthracene itself. ACKNOWLEDGMENT The author is indebted to the National Aniline & Chemical Company for aid in carrying out this work.
