R. W. Riemenschneider

Pancreatic lipase hydrolysis of triglycerides by a semimicro technique

Procedures are described for rapid lipase hydrolysis of triglycerides, isolation of the hydrolytic products by TLC and their conversion to methyl esters and fatty acid analysis by GLC. The techniques are applicable to a few mg of triglycerides or fats. Examples of data obtained with purified triglycerides indicate that the specific action of pancreatic lipase for the 1,3 ester groups is nearly absolute and the technique may be used as a criterion of purity of di- and tri-acid triglycerides. Ca. 83% of the palmitic but only 10~12% of stearic and C18 unsaturated acids of commercial lard occur in 2-position.

Direct conversion of lipid components to their fatty acid methyl esters

SummaryMethyl esters were prepared from cholesteryl esters, phospholipids, and glycerides in substantially quantitative yields by methanolysis with large excess of sodium or potassium methoxide in absolute methanol.A silicic acid chromatographic adsorption column technique was described, which was effective in separating methyl esters from unsaponifiables such as sterols, pigments, etc., and free acids.Conditions for complete methanolysis of glyceride fats and oils requiring only 5 min. of reflux time were described.Quantitative conversion of fatty acids to methyl esters was accomplished by direct esterification with absolute methanol containing 4% HCL or H2SO4 and by methylation with diazomethane.

Standardization of spectrophotometric methods for determination of polyunsaturated fatty acids using pure natural acids

SummarySpectrophotometric methods of analysis for the polyunsaturated constituents of oils and fats have been carefully restandardized for several conditions of alkali-isomerization, using purified methyl esters of linoleic, linolenic, and arachidonic acids prepared by physical rather than by chemical means. A number of vegetable oil and animal fat samples were subjected to spectrophotometric analysis, and the results based on natural and on bromination-debromination fatty acid standards were compared. The natural fatty acid standards lead to significantly higher accuracy and their use in the spectrophotometric analysis of natural fatty materials is strongly recommended. Results obtained under different conditions of isomerization were in satisfactory agreement. An isomerization time of 45 minutes is recommended rather than 25 or 30 minutes. The glycerol-air technique is preferred for general use because of its simplicity and high precision. The ethylene-glycol-nitrogen technique is a close second choice because of the greater transparency of reagent blanks.

Fatty acids of cows' milk. A. Techniques employed in supplementing gas-liquid chromatography for identification of fatty acids

Milk fat methyl esters were subjected to distillation and silicic acid column chromatography to provide fractions of less complexity for gas-liquid chromatographic analysis. It was still necessary however to employ supplemental techniques for identification. Chromatograms were obtained with polyester columns of different polarity on all the fractions and necessary reference samples. While many of the components were identified in the usual way by plots of relative retention time versus number of carbon atoms, iodine values for total unsaturation and ultraviolet spectrophotometry for conjugated and nonconjugated polyunsaturated acids were essential for positive identification of some components. Similarly, examination by infrared spectrophotometry confirmed the presence or absence of conjugated diene ascis-trans, trans-trans or both. Isolatedtrans or terminal double bonds were also determined in this way. Gas-liquid chromatograms of some fractions showed incompletely resolved peaks attributable to the presence of methyl esters of odd-carbon atom, branched-chain, and unsaturated acids. Hydrogenation and rechromatographing provided more positive determination of the structure of these components. Further confirmation of identity of some peaks on the chromatogram was achieved by collection of the appropriate fractions and examination of the collected material. At least 60 fatty acids were identified, including several not previously reported, such as odd-numbered carbon chain length monoethenoid acids from C15 to C23.

Fractionation of lard and tallow by systematic crystallization

SummaryLard and edible tallow were subjected to a series of fractional crystallizations from acetone at temperatures from 20° to -45°C. Six recrystallized precipitate fractions and a filtrate residue were obtained from each fat. In addition to determining the more common physical and chemical characteristics, fatty acid composition of each fraction was calculated from spectrophotometric data, iodine value, and thiocyanogen value. The consistent results obtained throughout by the spectrophotometric method of fatty acid analysis lend further confirmation to the reliability of this method for composition studies of natural fats. The approximate amounts of tri-saturated, di-saturated, mono-saturated, and tri-unsaturated glycerides of the lard and tallow were estimated from the analysis of each fraction on the assumption that not more than two of these classes of glycerides were present in any one fraction. The tallow contains much higher proportions of tri-saturated and di-saturated glycerides and correspondingly lower proportions of the mono-saturated and tri-unsaturated glycerides than does lard. The amount of tri-unsaturated glycerides in lard was found to be significantly greater than meager information in the literature would indicate. The data indicate that the general pattern of glyceride formation in animals such as the pig and cow is probably of random character.

Vernonia anthelmintica (L.) Willd. trivernolin, 1,3-divernolin and vernolic (Epoxyoleic) acid from the seed oil

The epoxy fatty acid components isolated from the seed oil ofV. anthelmintica, Indian ironweed, where the seed had been allowed to undergo lipolysis after grinding, were trivernolin, 1,3-divernolin, and vernolic acid. By inactivation of the hydrolytic enzyme system present in the seed, oil containing more than 50% trivernolin may be obtained. This species has potentialities as a replacement crop for those now in surplus; its seed contains 20 to 26% of an oil rich in epoxyoleic (vernolic) acid combined as glycerides amounting to 70 to 75%.

Analysis of fats and oils by gas-liquid chromatography and by ultraviolet spectrophotometry

Summary1.Known mixtures of pure fatty acid methyl esters and a number of fats and oils as their methyl esters have been analyzed by conventional GLC with thermal conductivity detectors.2.Percentage of fatty acid distribution determined by GLC agreed well with known percentages in model mixtures and with analysis by the spectrophotometric method for fats and oils.3.Determination of very small amounts of arachidonic and pentacnoic acids in lard by GLC was not successful.

Fat composition and in vitro oxygen consumption of marrow from fed and fasted rabbits.

Abstract 1. 1. Spectrophotometric analyses of the marrow fat of six fed and six fasted rabbits have been compared with their respective in vitro oxygen consumptions. 2. 2. Unsaturated fatty acids constitute 62% by weight of the total marrow fat. The quantity of linoleic acid in marrow from fed rabbits is as great as that of all combined saturated fatty acids. 3. 3. The absolute quantities of all components of marrow fat diminish during starvation. The per cent of linoleic and linolenic acids decreases during fasting, whereas that of arachidonic, pentaenoic, and oleic acids increases. In terms of per cent, total saturated fatty acid in marrow fat is essentially unaffected by starvation. 4. 4. Rate of oxygen consumption in vitro is not related to the concentration of any fatty acid component of marrow fat. 5. 5. It is concluded from in vitro evidence that polyunsaturated fatty acids of marrow are not oxidized in situ but are transported elsewhere for utilization or discard.

Effect of deodorization and antioxidants on the stability of lard

SummaryDEODORIZATION produced no appreciable increase in the stability of steam-rendered lard but significantly increased the stability of kettlerendered lard. A substantial increase in the stability of lard was produced by tocopherol, regardless of whether it was added as a pure compound, as a concentrate, or as a tocopherol-bearing oil. Accelerated tests showed that this increase was more than doubled when small amounts of synergists also were added. Deodorization of the lard prior to addition of the synergistic antioxidant compositions produced even greater stability.Nordihydroguaiaretic acid was more effective than tocopherol as an antioxidant for lard, as determined by both the active oxygen and the oxygen-absorption methods. Deodorization of the lard prior to addition of this antioxidant and synergists did not effect further increase in stability over that obtained by the addition of these materials to undeodorized lard.

Autoxidation of fatty materials in emulsions. I. Pro-oxidant effect of histidine and trace metals on the oxidation of linoleate esters

Aqueous emulsions of methyl or ethyl linoleate (sodium dodecylsulfate as emulsifier) together with such added components as 1-histidine, metal chlorides, buffers, and acid or alkali, were oxidized in the dark with shaking in an oxygen atmosphere. Under optimum conditions (pH 6.5), the linoleate peroxide content, after 2 hr autoxidation at 20C, was increased more than 3-fold by the addition of 1 ppm of ferrous or ferric ions, approximately 20-fold by a 0.01 M concentration of histidine and more than 60-fold by the addition of both histidine and ionic iron. The pro-oxidative effect of other transition metal ions (Cu++, Co++, Cr+++, Mn++, and Ni++) also was investigated. None of these ions had a significant effect alone. Combined with 0.01 M histidine, only Mn++ increased peroxidation over that when when histidine alone was added.The pro-oxidative action of histidine was retarded approximately 60% by 0.1 N acetate buffer and completely repressed by 0.05 M phosphate, nonionic emulsifiers, and low and high pH. The threshold concentration of histidine necessary for pro-oxidative action was greater than 0.0001 M.