FORMATION OF REACTIVE OXYGEN SPECIES DURING ONE-ELECTRON REDUCTION OF NORADRENOCHROME CATALYZED BY NADPH-CYTOCHROME P-450 REDUCTASE

One-electron reduction of noradrenochrome catalyzed by NADPH-cytochrome P-450 reductase resulted primarily in the formation of o-semiquinone and, probably, also o-hydroquinone. Under aerobic conditions these reduced form(s) autoxidize, accompanied by the formation of reactive oxygen species, as revealed by continuous NADPH oxidation and oxygen consumption, The presence of manganese-pyrophosphate complex contributed to autoxidation of the o-semiquinone during the reduction of noradrenochrome catalyzed by NADPH-cytochrome P-450 reductase, since the addition of the metal chelator diethylenetriamine pentaacetic acid (DETAPAC) resulted in a 34% inhibition of NADPH oxidation. Oxygen in the ground state was found to be predominantly involved in the autoxidation of o-semiquinone during the reduction of noradrenochrome catalyzed by NADPH-cytochrome P-450 reductase, since the addition of superoxide dismutase (SOD) to the incubation mixture only inhibited NADPH oxidation 13% and 6% in the absence and presence of DETAPAC, respectively, The addition of catalase to the incubation mixture resulted in a slight increase in NADPH oxidation, both in the absence and in the presence of diethylenetriamine pentaacetic acid, However, no effect of catalase and SOD together on NADPH oxidation was observed, either in the absence or presence of DETAPAC.