10. It is concluded that a satisfactory fat liquor can be made from menhaden oil. BIBLIOGRAPHY ANONYMOUS 1958. British Patent 776, 375, Journal of the American Leather Chemists Association, vol. 53, p. 65. Published by the Department of Leather Research, University of Cincinnati, Cincinnati, Ohio. ORTHMANN, A. C. 1945. Tanning Process. Hide and Leather Publishing Company, Chicago, Ill., pp. 277-297. WATSON, M. A. 1950. Economics of Cattle Hide Leather Tanning. The Rumpf Publishing Company, Chicago 6, Ill., p. 221. WILSON, M. A. 1941. Modern Practices in Leather Manufacture. Reinhold Publishing Corporation, New York, N. Y., pp. 482 and 490. Note: ACKNOWLEDGEMENT: We gratefully acknowledge the aid given to us by Albert Trostel and Sons Company of Milwaukee, Wisc., by NOPCO Chemical Company of Harrison, N. J., and by the Department of Applied Science in Tanning Research Foundation, Tanners' Council of America, University of Cincinnati. FISH OIL RESEARCH MAY SHED LIGHT ON HEART DISEASES A research project to study the relationship of fish oil in the diet to cholesterol deposits in the circulatory system of the body has been authorized by the U. S. Department of the Interior, Bureau of Commercial Fisheries. The research is expected to contribute to the fund of information being compiled on various phases of arteriosclerosis, particularly the coronary types. It is the unsaturated fatty acids (unstable and susceptible to chemical change) which abound in fish oil that are the center of attention. Studies already made under the Saltonstall-Kennedy Act for the improvement of domestic commercial fisheries have shown that fish oils contain a greater amount and a greater diversity of these unsaturated fatty acids than do many other food fats. The current investigations are also being made under the Saltonstall-Kennedy program. In one experiment miniature pigs are being used to determine the deposition of cholesterol in the body. Fish oil fatty acids of known degrees of unsaturation will be fed to the test animals. Ultimately the animal will be killed and the arteries examined to evaluate the effects of the several diets employed. Another test will be made on rats to determine which of the many fish oil fatty acids are essential to physiological welfare. One objective is attempting to determine the relationship of fish oils to metabolism and fat transport in the body, while still another is probing the properties of fish oil that may have pharmaceutical applications. The nutritive qualities of fish in reference to heart disease and current related dietary research is explored in some detail in the July 1958 issue of the Commercial Fisheries Review, published monthly by the Bureau of Commercial Fisheries. Reprints of this article --Separate 515--are available through the Office of Information, U. S. Fish and Wildlife Service, Department of the Interior, Washington 25, D. C. SIGNIFICANCE OF ULTRAVIOLET ABSORPTION DATA OF FISH-OIL FATTY ACIDS By Edward H. Gruger, Jr.* ABSTRACT Several ultraviolet absorption characteristics exist that are peculiar to the doublebond character of polyunsaturated fatty compounds and that make ultraviolet absorption measurements a means of evaluating the degree of unsaturation of compounds derived from fish oils. Accordingly, when pure polyunsaturated fatty acids from fish oils become available as analytical standards, a practical quantitative analysis will be possible. INTRODUCTION Fish oils are made up largely of triglycerides that contain straight-chain fatty acid groups having both saturated and unsaturated carbon-carbon chain linkages. The unsaturated fatty acids obtained from fish oils may possess from one to six carbon-carbon double bonds per molecule. Experimental results indicate that these double bonds are separated by single methylene (-CH,-) groups. That is, the unsaturated portions of the fatty acids have nonconjugated structures. An example of a nonconjugated unsaturated fatty acid is 9,12,15-octadecatrienoic acid: CH3CH2CH=CH_CH-CH-CH CH2-CH=CH(CH2)7COOH (a) The absorption of ultraviolet light at certain wavelengths is characteristic of definite chemical structures. The ultraviolet absorption caused by carbon-carbon unsaturation is brought about only by a conjugated double-bond system. The fatty acids in fish oils occur naturally with nonconjugated unsaturation. For the ultraviolet absorption method of analysis to be effective with fish-oil fatty acids, the unsaturation in these acids therefore must be converted to the conjugated form. For example, the following equation (b) depicts the isomerization process of nonconjugated 9,12-octadecadienoic acid to the acid possessing conjugated unsaturation: CH3(CH2)4CH=CH_CH2_CH=CH(CH2),COOH alkali CH3(CH2)4-CH2-CH-CH-CH-CH(CH2),COOH heat (b) ANALYTICAL METHOD: The double bonds in unsaturated fatty acids are conjugated by alkali-isomerization. The analytical procedure commonly employed is that of the American Oil Chemists' Society Tentative Method Cd 7-48: An 80-milligram1/ sample of fatty material is mixed with 11 grams of 21-percent potassium hydroxide in ethylene glycol that is preheated to 180° C. (3560 F.) in a suitable isomerization flask. An atmosphere of nitrogen is passed over the mixture during the entire isomerization process. The mixture is held at 180° C. for exactly 15 minutes, after which time the reaction is stopped by immediately cooling it to room temperature. The cooled mixture is diluted with an appropriate solvent to a known volume such that the concentration of isomerized material permits adquate measurement of the ultraviolet absorption spectra. Spectral absorption peaks at 233, 268, 315, 346, and 374 millimicrons are the result of the presence within the molecules of two, three, four, five, and six conjugated carbon-carbon double bonds, respectively. From the relative heights of the absorption peaks can be calculated specific extinction coefficients. The extinction coefficients are used to determine the quantity of material present in the analyzed mixture which contribute to the particular absorptions. (This is discussed in the next section.) * Chemist, Fishery Technological Laboratory, Division of Industrial Research and Services, U. S. Bureau of Commercial Fisheries, Seattle, Wash. 1/This figure is based on the unsaturation commonly found in commercial fish oils. The specific extinction coefficients are calculated from the equation K = A (c) where k is the specific extinction coefficient, A is the absorbance (or optical density) 1/Herb and Riemenschneider, Anal. Chem. 25, 953 (1953). Isomerization in 21 percent KOH in ethylene glycol at 1800 C. (356° F.) for 15 minutes under nitrogen. at a given wavelength, c is the concentration of isomerized substances in grams perliter, and 1 is the cell length in centimeters. 20 than would a C22 fatty acid with six double bonds. This reasoning is based on the fact that a C22 fatty acid with five double bonds has a lower extinction coefficient at 346 millimicrons than has a C, fatty acid with five double bonds. Also, this decrease in extinction coefficient can be explained by the effect of increasing chain length, which in turn increases the molecular weight. An increase in molecular weight causes a lowering of the molar concentration of double bonds for a given weight of sample. This then lowers the spectral absorption due to the double bonds and consequently lowers the specific extinction coefficient. These changes can be seen by examining equation (c). Fig. 1 Chemist using an automatic-recording spectrophotometer to measure the ultraviolet absorption characteristics of alkali-isomerized fatty acids from fish oil. With mixtures of highly unsaturated compounds from fish oils, such as fatty acids and fatty alcohols, the specific extinction coefficient calculated for a given wavelength is an additive value resulting from each compound in the mixture. Also, if six double bonds is the maximum number found in fish oils, there are no other ultraviolet absorptions at higher wavelengths contributing to the absorption at 374 millimicrons. One would expect, therefore, that a comparison of extinction coefficients at 374 millimicrons to the value of 29.3 (in table 1) will give a fair quantitative approximation of the content of heraenoic acids present. A pure C24 fatty acid of high unsaturation has not as yet been reported, thus the more complete analysis is not possible at this time. One further matter to consider is the possibility of the existence of fatty acids of C20 to C24 chain lengths having only two and three carbon-carbon double bonds. Untif the contrary has been unquestionably proven and reported, complete fatty acid analysis by ultraviolet absorption will not be possible. APPLICATIONS: The ultraviolet absorption data are valuable as a means of determining the success of methods of separating fish-oil fatty acids or their derivatives. The data also can be used to determine the effect of storage conditions on the high degrees of unsaturation; that is, whether or not storage treatment has affected the pentaene and hexaene content. Extinction coefficients, from absorption data of the type described above, are used in a set of simultaneous equations for solving quantitatively the percentage of each compound contributing to the particular absorptions. For quantitative analyses of this type to be accurate, however, pure compounds (the pure fatty acids in the case of fish oils) must be available for use as reference standards. Contract work being carried out by Dr. Orville Privett at the Hormel Institute, University of Minnesota, Austin, Minn., is designed to prepare the necessary "standard" fatty acids and to analyze the major commercial fish oils for their fatty acid composition. When this contract is completed, it will be possible to obtain a practical quantitative measurement of the relative proportions of the different fatty acids of various degrees of unsaturation present in all fish oils by the use of ultraviolet absorption measurements. BIBLIOGRAPHY HAMMOND, E. G., and LUNDBERG, W. O. HERB, S. F., and RIEMENSCHNEIDER, R. W. American Oil Chemists' Society, vol. 32, pp. 616-624. OFFICIAL AND TENTATIVE METHODS OF THE AMERI- 1956. Polyunsaturated Acids, Tentative Method Cd RIEMENSCHNEIDER, R. W. 1954. Analytical Methods and Composition of Fatty de SURVILLE, B. M. A.; RIVETT, D. E. A.; and 1957. The Synthesis of a 1:5-dienoic Acid and Its VARIATION IN PHYSICAL AND CHEMICAL CHARACTERISTICS OF HERRING, MENHADEN, SALMON, AND TUNA OILS Raymond O. Simmons* ABSTRACT Refractive index, iodine number, free fatty acid, saponification number, nonsaponifiable matter, stearine fraction, and Gardner color index were determined for herring, menhaden, salmon, and tuna oils. The data for menhaden oil are given for the various geographical areas along the East and Gulf coasts ranging from Long Island to the Mexican border. INTRODUCTION Processing industrial products from fish is an extensive industry. During 1956, in the menhaden industry alone, for example, over 2 billion menhaden were reduced to fish meal, solubles, and oil. An adequate domestic market exists for the fish meal and solubles as components of commercial mixed feeds for poultry and swine, but the domestic demand for fish oils has declined during the last several years. One of the reasons for this decline was the preconceived concept that commercially-available fish-body oils varied considerably in physical and chemical characteristics. The purpose of this research project was to determine the normal variation in physical and chemical characteristics of fish oils produced in the United States and Alaska. This study is a part of the over-all research program on fish oils initiated by the U. S. Bureau of Commercial Fisheries. The anticipated practical result is to extend the market for fish oils through a better knowledge of their physical and chemical properties. EXPERIMENTAL During the 1955 and 1956 season, samples of fish-body oils were analyzed for refractive index, iodine number, content of free fatty acid, saponification number, content of nonsaponifiable matter, and Gardner color number. During the 1956 and 1957 season these same analyses were made. In addition, the oils were separated into a stearine-oil fraction and a winterized-oil fraction, and the relative amounts were determined of these fractions. Also, the refractive index and iodine numbers were determined for these fractions. A Bausch and Lomb Precision Refractometer was used to determine refractive index, and the Gardner color number of the oils was determined with the 1953 series Gardner color standards for liquids. All iodine values were determined by the Wijs method using a reaction time of 1 hour. Commonly-accepted procedures as outlined in the Official and Tentative Methods of the American Oil Chemists' Society were used for the other determinations. Stearine was determined by a defined winterizing process that consisted of stepwise lowering the temperature of the oil to 5° C. and separating the solid phase and the liquid phase by centrifugation. A total of 126 menhaden and 14 herring body oils and 12 tuna and 12 salmon cannery byproduct oils were analyzed. The menhaden samples were received from plants located on the Atlantic and Gulf of Mexico Coasts, extending from Port Monmouth, N. J., to Port Arthur, Tex.; the tuna samples came from California, and the herring and the salmon samples came from Alaska. *The research reported in this paper was conducted at North Carolina State College, Department of Chemistry, under contract with the U. S. Fish and Wildlife Service. It was financed by funds made available under provisions of Public Law 466, 83rd Congress, approved July 1, 1954, generally termed the Saltonstall-Kennedy Act. This article was prepared by Dr. Donald G. Snyder, Biochemist, Fishery Technological Laboratory, College Park, Md., from progress reports submitted by the contractor to the Service. |