HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY THERMOSPRAY MASS-SPECTROMETRY OF EPOXY POLYUNSATURATED FATTY-ACIDS AND EPOXYHYDROXY POLYUNSATURATED FATTY-ACIDS FROM AN INCUBATION MIXTURE OF RAT-TISSUE HOMOGENATE

A method for the analysis of epoxy polyunsaturated fatty acids (EpPUFA s) and epoxyhydroxy polyunsaturated fatty acids (EpHPUFAs) in rat tiss ue homogenate, with homo-gamma-linolenic acid (20:3, n - 6), arachidon ic acid (20:4, n - 6), eicosapentaenoic acid (20:5, n - 3) or docosahe xaenoic acid (22:6, n - 3) as a substrate, has been developed. Extract ion with dichloromethane at pH 4-5 and concentration in the presence o f pyridine were performed. Spectral analysis of chromatograms obtained with high-performance liquid chromatography-thermospray mass spectrom etry showed the presence of EpPUFAs, EpHPUFAs and dihydroxy metabolite s (DiHPUFAs) of EpPUFAs corresponding to each precursor fatty acid. On a selected-ion monitoring chromatogram, many EpPUFAs, EpHPUFAs and Di HPUFAs in an extract from an incubation mixture of each precursor fatt y acid in aged rat tissue homogenate were detected simultaneously with in 70 min. EpPUFAs and DiHPUFAs derived from 20:3 (n - 6) or 20:5 (n - 3) were detected in significant amounts. From these results, a highly active cytochrome P450 system or non-enzymic oxidative reactions in a ged rat tissue homogenate were suggested.