Direct ESR detection of pentadienyl radicals and peroxyl radicals in lipid peroxidation: Mechanistic insight into regioselective oxygenation in lipoxygenases

Well-resolved ESR spectra of free pentadienyl radicals have been observed under photoirradiation of di-tert-butylperoxide (Bu^tOOBu^t) and polyunsaturated fatty acids in the absence of O₂, allowing us to determine the hfc values. The hfc values of linoleyl radical indicate that the spin density is the largest at the C-11 position. The linoleyl radical is readily trapped by O₂ to produce the peroxyl radical (11-HPO ^•) in which O₂ is added mainly at the C-11 position of the pentadienyl radical as indicated by the comparison of the ESR spectra of peroxyl radicals derived from linoleic acid and [11,11-²H ₂]linoleic acid. The peroxyl radical (13-HPO^•), which is initially formed by the hydrogen abstraction from 13-(S)-hydroperoxy-9(Z), 11(E)-octadecadienoic acid (13-HPOD) by Bu^tO^•, is found to isomerize to 11-HPO^• via removal of O₂ from 13-HPO^• and addition of O₂ to linoleyl radical to produce 11-HPO^•. This finding supports an idea of O₂ entering via a specific protein channel, which determines the stereo- and regiochemistry of the biradical combination between O₂ and linoleyl radical in lipoxygenases.