Evidence against the 1:2:2:1 quartet DMPO spectrum as the radical adduct of the lipid alkoxyl radical

Abstract

It was reported that the electron paramagnetic resonance (EPR) spectrum of 5,5-dimethyl-1-pyrroline N-oxide (DMPO)/lipid alkoxyl radical exhibited a quartet with 1:2:2:1 relative intensity that is identical to that of DMPO/hydroxyl radical ( K. M. Schaich and D. C. Borg, 1990, Free Radicals Res. Commun. 9, 267–278 ). We repeated these EPR experiments using HPLC separation of radical adducts and isotope substitution. We found that the HPLC EPR chromatogram of the radical adduct with a 1:2:2:1 quartet obtained by the reduction of methyl linoleate hydroperoxide (MLOOH) with Fe 2+ exhibited identical retention time to that of the DMPO . OH radical adduct obtained from the Fenton reaction in two different solvent systems. Upon performing the same reaction in 17O-enriched water, the 17O-hyperfine coupling constants due to DMPO .17 OH were identified. Ultimately, ~80–90% of the total DMPO . OH is derived from water by an iron-dependent nucleophilic addition reaction. Initially, a water-independent mechanism also significantly contributes to DMPO/.OH formation. Regardless of its mechanism of formation, the 1:2:2:1 quartet radical adduct of DMPO formed during the reduction of MLOOH by Fe 2+ is in fact DMPO . OH .