Ralph T. Holman

The thiobarbituric acid reaction and the autoxidations of polyunsaturated fatty acid methyl esters.

Abstract The TBA reaction of autoxidized single methylene-interrupted diene, triene, tetraene, pentaene, and hexaene fatty acid esters has been measured at various stages of autoxidation. Linoleate free of higher polyunsaturates gave no TBA color at early stages of oxidation. At early stages of oxidation TBA values of triene, tetraene, pentaene, and hexaene were found to vary linearly with diene conjugation, peroxide value, and oxygen uptake. TBA values of triene, tetraene, pentaene, and hexaene were found to vary linearly with diene conjugation throughout the course of the oxidation. A mechanism is proposed for TBA reactant formation from methylene-interrupted polyunsaturated compounds with cyclic peroxide as an intermediate in the formation of malonaldehyde. The existence of a cyclic five-membered ring peroxide in the oxidation products of polyunsaturated acids is postulated. The mechanism of oxidation of methylene-interrupted trienes, tetraenes, pentaenes, and hexaenes postulates an additional step in chain propagation involving intramolecular rearrangement of the hydroperoxy free radical to a free radical bearing a closed five-membered ring peroxide. The scope of usefulness and the limitations of the TBA test are discussed.

Effect of Linolenic Acid Upon the Metabolism of Linoleic Acid

Weanling rats were fed a fat-free diet supplemented with various ratios of corn and linseed oils to furnish a constant dietary level of linolenate at 1% of calories and levels of linoleate from 0.3 to 17.3% of calories. The fatty acid composition of the total liver lipids was analyzed by gas chromatography. Increasing amounts of dietary linoleate suppressed the levels of the 20:5, 22:5 and 22:6 metabo lites of linolenic acid in the liver lipids. The level of dietary vitamin E had no effect upon this phenomenon. When the level of dietary linoleate was increased, the level of 22:4 in the liver lipids, as well as the other metabolites of linoleate, was shown to increase.

Pyrrolidides for mass spectrometric determination of the position of the double bond in monounsaturated fatty acids.

Mass spectra of pyrrolidides of monounsaturated straight chain fatty acids are presented and discussed. The spectra of pyrrolidides contain mainly ions from the polar part of the molecule. This gives simple spectra from which double bond positions can be deduced directly. If an interval of 12 atomic mass units is observed between the most intense peaks of clusters of fragments containing n and n−1 carbon atoms of the acid moiety, the double bond occurs between carbons n and n+1 in the molecule This rule is valid for double bonds occurring at positions Δ5-Δ15 in an 18-carbon chain and has been applied to acids having 10–24 carbon atoms.

The rates of oxidation of unsaturated fatty acids and esters

SummaryThe rates of autoxidation of oleic acid, ethyl oleate, linoleic acid, 10,12-linoleic acid, ethyl linoleate, trilinolein, pentaerythritol linoleate, dipentaerythritol linoleate, elaidolinolenic acid, linolenic acid, ethyl linolenate, trilinolenin, and methyl arachidonate have been studied by oxygen uptake in a Warburg respirometer and the results are compared with the rates of enzymatic oxidation of lipoxidase substrates.The increase in the number of double bonds in a fatty acid by one increases the rate of oxidation of the fatty acid or its esters by at least a factor or two.Earlier findings that acids oxidize more rapidly than their esters have been confirmed.The initial rates of lipoxidase oxidation of ethyl linoleate, ethyl linolenate, and methyl arachidonate were found to be essentially the same.

Characterization of unsaturated fatty acids by gas-liquid chromatography

The equivalent chain length (ECL) has been determined on 79 methyl esters of unsaturated fatty acids and on 7 ethyl esters by gas chromatography. Ethylene glycol succinate (EGS), diethylene glycol succinate, β-cyclodextrin acetate and Apiezon L were chosen as the liquid phases to be used. For methyl esters of mono- and polyenoic acids, the differences between ECL on EGS and ECL on Apiezon L approximate 0.84 per double bond. For positional isomers, the ECL on both EGS and Apiezon L are usually greater for the isomer having the longer proximal end of the molecule (smallest ω value). In these terms a triple bond is approximately equal to three double bonds. Esters of nonconjugated dienoic and trienoic acids of the same chain length are not separable on Apiezon L if their proximal structures are the same. This also applies to tetraenoic and pentaenoic acids of the same chain length and the same proximal structure. Conjugation of double bonds, either with the ester carbonyl group or with themselves, yields ECL values on Apiezon L greater than the number of carbon atoms in the acid. Monounsaturated and nonconjugated polyunsaturated esters have ECL values on Apiezon L lower than the number of carbon atoms of the acid. The ECL values of ethyl esters of 18 and 20 carbon acids are greater than the corresponding methyl esters on Apiezon L.

The preferred metabolic pathway from linoleic acid to arachidonic acid in vitro

Abstract 1. 1. The two possible pathways which lead from linoleic acid to arachidonic acid were simultaneously studied in liver microsomes from fat-deficient rats and from fat-fed rats. 2. 2. The overall yields of the conversion of linoleic acid to eicosa-8,11,14-trienoic and arachidonic acids were calculated and gave a ratio of 16 to 1 for the pathway via γ-linolenic acid over the pathway via eicosa-11,14-dienoic acid in the fat-fed rat, and a ratio of 6 to 1 in the fat-deficient rat. 3. 3. The side reactions to the alternate pathways were also investigated. A relatively high rate of chain elongation was found for eicosa-8,11-dienoic acid which minimizes the importance of the pathway via this acid. 4. 4. The inhibition and enhancement of the different reactions in the two pathways were tested under the same conditions using either mixtures of fatty acids extracted from rat livers or single fatty acids of the linoleic acid family. 5. 5. The interactions of the different acids in chain elongation and desaturation reactions are of small magnitude. However, the desaturation of eicosa-8,11,14-trienoic acid is strongly enhanced by linoleic and γ-linolenic acids which may account for its non-accumulation in tissue lipids.

Mass spectra of the picolinyl esters of isomeric mono- and dienoic fatty acids

The picolinyl ester derivatives of the complete series of isomeric octadecenoates, methylene-interrupted octadecadienoates, and of octadec-9-ynoate have been subjected to gas chromatography-mass spectrometry. A fused-silica capillary column, coated with a cross-linked methyl silicone, was used for the separation. Electron impact spectra were determined at 70 eV. Earlier observations with a limited series of model compounds were confirmed, and it was shown that the picolinyl ester derivatives were of almost universal value in the location of double bonds in such isomers. Difficulties of interpretation arose mainly when the double bonds were close to the carboxyl group.

Omega 3 but not omega 6 fatty acids inhibit AP-1 activity and cell transformation in JB6 cells.

Epidemiological and animal-based investigations have indicated that the development of skin cancer is in part associated with poor dietary practices. Lipid content and subsequently the derived fatty acid composition of the diet are believed to play a major role in the development of tumorigenesis. Omega 3 (ω3) fatty acids, including docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), can effectively reduce the risk of skin cancer whereas omega 6 (ω6) fatty acids such as arachidonic acid (AA) reportedly promote risk. To investigate the effects of fatty acids on tumorigenesis, we performed experiments to examine the effects of the ω3 fatty acids EPA and DHA and of the ω6 fatty acid AA on phorbol 12-tetradecanoate 13-acetate (TPA)-induced or epidermal growth factor (EGF)-induced transcription activator protein 1 (AP-1) transactivation and on the subsequent cellular transformation in a mouse epidermal JB6 cell model. DHA treatment resulted in marked inhibition of TPA- and EGF-induced cell transformation by inhibiting AP-1 transactivation. EPA treatment also inhibited TPA-induced AP-1 transactivation and cell transformation but had no effect on EGF-induced transformation. AA treatment had no effect on either TPA- or EGF-induced AP-1 transactivation or transformation, but did abrogate the inhibitory effects of DHA on TPA- or EGF-induced AP-1 transactivation and cell transformation in a dose-dependent manner. The results of this study demonstrate that the inhibitory effects of ω3 fatty acids on tumorigenesis are more significant for DHA than for EPA and are related to an inhibition of AP-1. Similarly, because AA abrogates the beneficial effects of DHA, the dietary ratio of ω6 to ω3 fatty acids may be a significant factor in mediating tumor development.

A comparison of pyrrolidide and picolinyl ester derivatives for the identification of fatty acids in natural samples by gas chromatography-mass spectrometry.

The pyrrolidide and picolinyl ester derivatives of the fatty acids in two natural lipid samples rich in unsaturated fatty acids, pig testis lipids and cod liver oil were satisfactorily resolved on capillary columns of fused silica coated with stationary phases of varying polarity. The picolinyl esters, in particular, when subjected to gas chromatography-mass spectrometry on a column containing a cross-linked methyl silicone, gave distinctive mass spectra, which could be interpreted in terms of both the numbers and positions of the double bonds.

Mass spectrometric determination of positions of double bonds in polyunsaturated fatty acid pyrrolidides

Low resolution mass spectra of pyrrolidides of isomeric octadecadienoic acids and other polyunsaturated straight chain fatty acids are presented and discussed. The spectra of the pyrrolidides contain mainly ions from the polar part of the molecule and give spectra that are specific for each isomer. The interpretation follows, in most cases, the rule developed for monounsaturated fatty acid pyrrolidides.