Reversed-phase high-performance liquid chromatographic separation of tert.-butyl hydroperoxide oxidation products of unsaturated triacylglycerols

Triacylglycerols containing monounsaturated fatty acids are known to be relatively resistant to autoxidation and require long periods of exposure to dilute oxidants. Use of concentrated solutions of synthetic hydroperoxides, however, yields in addition to the hydroperoxides also unidentified oxidation by-products. In the present study we have employed synthetic triacylglycerols containing one (18:0/18:1/18:0 and 18:1/16:0/16:0) and two (18:0/18:0/18:2 and 18:1/18:1/18:0) double bonds per molecule to reinvestigate the formation of oxotriacylglycerols using tert.-butyl hydroperoxide as an oxidant. Reversed-phase HPLC was used to separate and tentatively identify the oxidation products based on relative retention times of standards and the estimated elution factors for functional groups and their positional distribution. Hydroperoxides, diepoxides and hydroxides were the major components of the oxidation mixtures (50–95% of total). Previously unidentified peroxide-bridged tert.-butyl adducts were present in significant amounts (5–50% of total oxidation products) in all preparations. In several instances more than one functional group was present on a single fatty chain. The tentative reversed-phase chromatographic identification of the adducts was confirmed by determination of the molecular mass of each component by on-line LC with electrospray MS. The oxidation products were quantified by HPLC with light scattering detection.