Radical adducts of nitrosobenzene and 2-methyl-2-nitrosopropane with 12,13-epoxylinoleic acid radical, 12,13-epoxylinolenic acid radical and 14,15-epoxyarachidonic acid radical. Identification by h.p.l.c.-e.p.r. and liquid chromatography-thermospray-m.s

Abstract

Linoleic acid-derived radicals, which are formed in the reaction of linoleic acid with soybean lipoxygenase, were trapped with nitrosobenzene and the resulting radical adducts were analysed by h.p.l.c.-e.p.r. and liquid chromatography-thermospray-m.s. Three nitrosobenzene radical adducts (peaks I, II and III) were detected; these gave the following parent ion masses: 402 for peak I, 402 for peak II, and 386 for peak III. The masses of peaks I and II correspond to the linoleic acid radicals with one more oxygen atom [L(O).]. The radicals are probably carbon-centred, because the use of 17O2 did not result in an additional hyperfine splitting. Computer simulation of the peak I radical adduct e.p.r. spectrum also suggested that the radical is carbon-centred. The peak I radical was also detected in the reaction of 13-hydroperoxylinoleic acid with FeSO4. From the above results, peak I is probably the 12,13-epoxylinoleic acid radical. An h.p.l.c.-e.p.r. experiment using [9,10,12,13-2H4]linoleic acid suggested that the 12,13-epoxylinoleic acid radical is a C-9-centred radical. Peak II is possibly an isomer of peak I. Peak III, which was observed in the reaction mixture without soybean lipoxygenase, corresponds to a linoleic acid radical (L.). The 12,13-epoxylinoleic acid radical, 12,13-epoxylinolenic acid radical and 14,15-epoxyarachidonic acid radical were also detected in the reactions of linoleic acid, linolenic acid and arachidonic acid respectively, with soybean lipoxygenase using nitrosobenzene and 2-methyl-2-nitrosopropane as spin-trapping agents.