April, 1924
INDUSTRIAL A N D ENGINEERING CHEMISTRY
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Studies of the Vitamin Potency of Cod-Liver Oil’ VII-The
Vitamin A Potency of Hake-Liver Oil By Arthur D. Holmes E. L. PATCH CO., BOSTON, MASS.
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To secure data concerning the relative vitamin potency of cod and A colorimetric analysis3 N THE c o m m e r c i a l hake-her oils, tests were made of hake-lioer oil known to be true to of this Oil by means of manufacture of codLovibond glasses showed its liver oil it is a t times name. Nine young albino rats were given hake-liver oils in amounts color to be yellow 15 and oarying from 0,00018 to 0.004 gram daily. almost ilnpossible to &miFour animals received 0.8 mg. of oil daily and failed to recouer red 1.5. nate the tendency of fishermen to include occasional from uitamin A FEEDING TESTS stray pollock, haddock or Five animals receioed from 0.8 to 4.0 mg. of hake-her oil daily and recooered, indicating that 0.8 mg. of this oil contained suficient citaNine young, growing alhake livers with cod livers bin0 rats were fed the usual destined for the manufacmin A to promote growth of young albino rats. ture of cod-liver oil. Such a synthetic diet, consisting of situation naturally raises casein, peanut oil, cornthe question as to the effect on the vitamin potency of cod-liver starch, milk sugar, and a salt mixture, supplemented by dried oil of the addition of livers of these other fish, Accordingly, brewers’ yeast, which served as a source of vitamin B. When it has been necessary to secure information concerning the the animals showed positive evidence of vitamin A starvavitamin A potency of fish-liver oils obtained from fish other tion, they were given graduated doses of hake-liver oil as a than cod. The tests reported in this paper were undertaken supplement to the diet. to determine the vitamin potency of hake-liver oils. As will be noted from the accompanying chart, four animals I n order to provide a hake-liver oil known to be true to name, (Nos. 224, 230, 240, and 245) received less than 0.8 mg. of oil a supply of hake livers were obtained by the author from daily, and died after they had been receiving oil for periods hake that had been out of the water only a few hours. The varying from 20 to 45 days. These results indicate very hake livers were secured in the early fall, a t a time when the definitely, for these animals at least, that less than 0.8 mg. fish from which they were taken were in good physical con- per day of hake-liver oil does not supply sufficient vitamin-A dition. .As a result of the excellent condition of the fish, to meet the needs of a young albino rat for growth when it is the livers were plump, of good color, and contained a rela- maintained on a diet adequate except for vitamin A. tively large amount Of Oil’ It is the ‘pinion that 3 The author is indebted to Herbert s, Bailey of the Southern Cotton Oil the oil rendered from these “prime” hake livers is typical of company for this colorimetric analysis. the highest grade of hake-liver oil. The hake livers were rendered under laboratory conditions by a process which Chart 16 VITAMIN PO~ENCY’OFHAKE-LIVER OIL has been described elsewhere.2 The oil received no purifiEARLY FALL OIL I STARTED HAKC.LIVfROIL 1 cation other than filtering and decanting processes to remove the water and liver residue. It was noted in an earlier paper that in the preparation of medicinal cod-liver oil stearin is removed from the crude oil so that the so-called refined oil will remain clear during the winter season. It is obvious, however, that it would be difficult, even under laboratory conditions, to remove identical amounts of stearin from different lots of oil, and inasmuch as the purpose of this investigation was to test the relative vitamin A potency of hake and cod-liver oils, it was judged best to test the potency of the crude oils, since these would be more iiearly comparable. Accordingly, the tests which follow were made on crude hake-liver oil. This oil was of a straw-yellow color and possessed much the same odor and flavor as that of oil which exudes from mackerel or other fat fish when they are broiled. To provide information concerning the general nature of the hake-liver oil under consideration, it was analyzed in the usual manner. The results obtained concerning the chemical and physical characteristics are given in the accompanying table: CHARACTERISTICS OF CRUDE HAKE-LIVER OIL Specific gravity at 25O C. 0 9191 Refractive index at 20° C. I 4757 Saponification value 185 9000 Iodine number 134 1000 Acid value 0 5458 Clouding point 25O C. 1 Presented before the Division of dhemistry of Medicinal Products a t the 66th Meeting of the American Chemical Society, Milwaukee, WIS , September 10 to 14, 1923. * J . Melabolzc Research, 2, 113 (1922).
I N D U S T R I A L A N D ENGINEERING CHEiMISTRY
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The five animals receiving amounts of hake-liver oil varying from 0.000808 to 0.00404 gram daily recovered from vitamin A starvation and grew continuously during the 45-day experimental period. Comparing the rate of growth of the animal receiving 0.000808 gram of hake-liver oil daily with that of the animals receiving two, three, four, and five times this amount, it will be noticed that this animal did not increase its body weight quite so rapidly as those receiving the larger amounts of hake-liver oil. However, since Rat 134, which received 0.000808 gram of oil daily, grew a t a very slightly less rapid rate than the other animals, it seems fair to assume that 0.000808 gram of hake-liver oil contains nearly, if not actually, enough vitamin A to meet the needs of albino rats for growth. The tests here reported were undertaken primarily to ascertain what effect the fisherman’s habit of including hake livers with his cod livers had on the potency of cod-liver oil.
Vol. 16, No. 4
The results in this series of tests show that 0.8 mg. daily of crude hake-liver oil, carefully prepared from hake livers secured from fish in good physical condition, contains sufficient vitamin A to meet the body requirements of growing albino rats maintained on a synthetic diet lacking in vitamin A. I n the investigation of which the present study forms a part, a large number of commercial, medicinal cod-liver oils procured from a variety of sources have been tested for their vitamin A potency. The results of these tests show that a medicinal cod-liver oil which is so potent that 1 mg. daily will meet the body requirements of a growing albino rat may well be considered as having a very high vitamin A content. In view of these facts, it is evident that including hake livers with cod livers which are to be used for the manufacture of medicinal cod-liver oil does not decrease the vitamin A potency of the resulting cod-liver oil.
Analysis of Binary Mixtures’ Volumetric Turbidity Method By Charles D. Bogin COMXERCIAL SOLVENTS CORP., TERREHAUTE, IND.
Binary mixfures of two liquids, one of which has a much greater solubility in water fhan the other, are easily analyzed by titrating with water until turbid, the appearance of turbidity furnishing a sharp end point. The method can be applied to fernary mixtures if an independent method for estimating the proportion of the third subsfance is aoailabk. Solufions of salts, mixfures of organic liquids and water, and pure organic liquids can be used as tifrafing media in a similar way. The method can offen be used in the analysis of mixtures of two
liquids, bofh of which are miscible in wafer, by adding a definite volume of a third insoluble liquid and then titrating with water. I t can also be applied to mixfures of solids by dissobing them in a liquid and titrafing with a second liquid. It is useful for identificafion of pure organic substances. The percenfage of water in organic liquids can be defermined by titrating with benzene. Moisture in solids can be defermined by exfracfing it with organic liquids and determining it by a benzene tifration.
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HE volume of water necessary to saturate a unit volume
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of a binary mixture of two liquids, one of which has a much greater solvent action for water than the other, varies greatly with the composition of the mixture, increasing considerably with any increase in the proportion of the more water-soluble component. The attainment of saturation in such binary mixtures takes place very rapidly and smoothly, the added water dissolving instantaneously up to the very point of saturation, while at that point the addition of a minute excess of water separates the hitherto homogeneous liquid into two phases, producing an intense turbidity. This appearance of the turbidity can thus serve as an admirable end point, fully comparable in sharpness and definiteness with some of the best end points known in volumetric analysis, being a very sharp and sudden transition from a clear, transparent liquid to a turbid emulsion and produced by the addition of as little as the one last drop of water. The volume of water that can be added to a unit volume of a binary mixture of any certain composition before turbidity appears is very definite and invariable, absolutely independent of the time of contact, amount of shaking, or speed of addition, and only slightly dependent on the temperature, while the technic involved is that of ordinary volumetric analysis. The measurement of the volume of water used, therefore, offers great possibilities for a quick and accurate determination of 1
Received November 13, 1923.
the composition of binary and, under certain conditions, ternary mixtures. The turbidity end point has been used considerably by physical chemists for determining the solubility relations of liquid mixtures, such as the volume of water that will dissolve in various mixtures of ethyl alcohol and benzene, and the simplicity and accuracy of the method have been very favorably commented upon.2 The use of the solubility data thus obtained for the identification and analysis of unknown mixtures of the same materials, however, has not received the attention that it deserves. A method employing a turbidity end point has been suggested for the analysis of aqueous solutions of ethyl alcohol by Dubaux and Dutoit3 and others. I n those methods the critical solution temperature of mixtures containing definite proportions of the sample and certain reagents is used to determine the percentage of ethyl alcohol in the sample. The writer has been using several modifications of the turbidity method for the analysis of mixtures of ethyl and butyl alcohols, as well as mixtures of the two alcohols together with acetone, for which analysis in connection with the Weizmann fermentation process the turbidity method has been developed. These methods will be described in detail.
* Seidell, “Solubilities,” 2nd ed., p. 776. 8
Ann. chim. anal., 13, 4 (1908).
