Henry W.-S. Chan

Autoxidation of methyl linoleate. Separation and analysis of isomeric mixtures of methyl linoleate hydroperoxides and methyl hydroxylinoleates

The mixture of conjugated diene hydroperoxide isomers obtained from autoxidation of methyl linoleate was separated by high performance liquid chromatography (HPLC). Four major isomers were obtained from adsorption chromatography and identified as the 9 and 13 positional isomers having thetrans-trans andcis-trans configurations. The latter geometrical isomers have thetrans double bond adjacent to the hydroperoxide group. The hydroxy compounds (methyl hydroxylinoleates) obtained from the hydroperoxides by NaBH4 reduction were similarly separated but with improved resolution. This is the first instance of the complete separation of these compounds and provides a rapid method for their analysis. Unlike adsorption chromatography, reversed-phase chromatography separates the mixtures only according to the geometrical isomerism of the double bonds and not according to the position of the hydroxy or hydroperoxide function.

A simple method for the preparation of pure 9-D-hydroperoxide of linoleic acid and methyl linoleate based on the positional specificity of lipoxygenase in tomato fruit.

Incubation of linoleic acid with crude homogenate of tomato fruit gave a high yield (69%) of linoleic acid hydroperoxides with a ratio of 9- to 13-hydroperoxide isomers of 96∶4. After chromatography of the products, as free acids or methyl esters, hydroperoxides with 9- to 13-isomeric ratios of >99∶1 were obtained. The major product was characterized as 9-d-hydroperoxy-octadeca-trans-10,cis-12-dienoic acid. The results demonstrate the positional specificity of lipoxygenase from tomato fruit.

The mechanism of the rearrangement of linoleate hydroperoxides

Linoleate hydroperoxides undergo rearrangement leading to their isomerisation in which the OOH group is relocated or the stereochemistry of a double bond changed, or both. The reaction was studied mainly with pure isomers of methyl hydroperoxylinoleates since conditions could be found in which rearrangement occurred with little accompanying decomposition. The rearrangement was found to be non-stereoselective and took place by a free-radical chain mechanism. Using 18O-labelled hydroperoxide on 18O2, it was shown that the oxygen atoms of the OOH group of the hydroperoxides exchanged with surrounding molecular oxygen during the rearrangement. A mechanism for the rearrangement is proposed.

Autoxidation of methyl linolenate: Analysis of methyl hydroxylinolenate isomers by high performance liquid chromatography

The mixture of methyl hydroxylinolenates obtained by reduction of the hydroperoxide isomers formed by autoxidation of methyl linolenate was resolved by high performance liquid chromatography into eight major components. These are positional isomers with the hydroxyl group at positions 9, 12, 13, and 16. Two geometrical isomers of each positional isomer are present; these differ in the configuration of the conjugated double bonds (cis-trans andtrans-trans). Autoxidation of methyl linolenate is regioselective and favors the formation of positional isomers 9 and 16.

Formation, isolation and structure determination of methyl linolenate diperoxides

Abstract Autoxidation of methyl linolenate gives rise to isomeric mono-hydroperoxides by reaction with one mole of oxygen but further reaction with a second mole of oxygen readily occurs to produce an isomeric mixture of diperoxides. Autoxidation of individual pure methyl hydroperoxylinolenate isomers has been used as a method of obtaining less complex diperoxide mixtures which can be separated into their pure components by preparative high-pressure liquid chromatography (HPLC). The major diperoxide isomers arising from the autoxidation of pure 9R- and 13S- hydroperoxides of methyl linolenate have been isolated and characterised as isomeric epidioxyhydroperoxides of methyl linolenate. These same compounds have been identified as components of the more complex mixture of diperoxides produced during methyl linolenate autoxidation. The structures of the isolated diperoxides have been determined by physico-chemical methods and a mechanism for their formation is proposed.

Oxidative reactions of unsaturated lipids

Abstract Monohydroperoxides, which are the products of free radical autoxidation of unsaturated fatty acids, can under go further free radical chain reactions initiated by abstraction of hydrogen to reform peroxy radicals. In the autoxidation of methyl linolenate, the peroxy radicals corresponding to ‘inner’ hydroperoxides may cyclise and, after further oxygenation, yield diperoxides. The mixtures of the two peroxidic species, monohydroperoxides and diperoxides, obtained from linolenate can be identified and determined chromatographically. Mechanistic studies showed that the oxygenation of pentadienyl radicals to form peroxy radicals—an important propagation step in autoxidation—is reversible. H-donors can be used to control the relative rates of the forward and reverse reactions and so determine the relative proportions of different peroxidic species formed. α-Tocopherol was shown to be an effective H-donor and drastically altered the distribution of products formed in linolenate autoxidation. Such control of peroxide formation may be a factor determining the manifestations that are specific to the oxidative deterioration of a food product.

Thermal isomerisation of methyl linoleate hydroperoxides. Evidence of molecular oxygen as a leaving group in a radical rearrangement

Using an 18O-enriched hydroperoxide, the pentadienyl hydroperoxides (1–4) were shown to undergo rearrangement in which the oxygen atoms of the hydroperoxy group exchanged with atmospheric oxygen.

Characterization of a panel of monoclonal anti-gliadin antibodies.

A panel of monoclonal antibodies has been raised against a total gliadin fraction prepared from the hard bread wheat cultivar Avalon. The antibodies have been characterized by enzyme-linked immunosorhent assay and immunoblotting techniques for their interactions with a range of cereal prolamins, purified gliadins and short synthetic peptides. The antibodies showed broad, intermediate and narrow specificities. The immunological approach offers the protein chemist an additional means of studying the structures and interrelationships of cereal proteins.

A hydroperoxy-epidioxide from the autoxidation of a hydroperoxide of methyl linolenate

Unlike autoxidation of linolenic acid or methyl linolenate which yields mixtures of hydroperoxy-epidioxides, sequential oxidation employing an enzymic first step followed by an autoxidation step can give rise to a single hydroperoxy-epidioxide.

Thermal decomposition of individual positional isomers of methyl linolenate hydroperoxides, hydroperoxy cyclic peroxides and dihydroperoxides.

The methyl esters of 4 individual positional isomers of hydroperoxides, 2 positional isomers of hydroperoxy cyclic peroxides and a 9,16-dihydroperoxide were prepared by autoxidation of methyl linolenate and separated by preparative high pressure liquid chromatography. Isolated hydroperoxide isomers were thermally decomposed and the resulting volatile components analyzed. Each hydroperoxide or hydroperoxy cyclic peroxide isomer yielded characteristic volatile products. The major volatiles from each acyclic hydroperoxide corresponded with those predicted to arise by carboncarbon scission on either side of the corresponding alkoxy radical intermediate and little evidence was found of isomerization between the various positional isomers occurring during the process. A similar mechanism would account for the volatile products obtained from the cyclic peroxides. 2,3-Pentanedione was a significant odor contributor arising from the 13,15-epidioxy-16-hydroperoxide isomer although it was only a minor decomposition product.