Effect of electron-transport inhibitors on the generation of reactive oxygen species by pea mitochondria during succinate oxidation

The effect of inhibitors of the cytochrome pathway and alternative oxidase on the rate of respiration and generation of reactive oxygen species by pea mitochondria was studied. Respiration of mitochondria from pea cotyledons was inhibited by 70-80% by salicylhydroxamate (SHAM). The rate of hydrogen peroxide production by pea cotyledon mitochondria during succinate oxidation was 0.15 nmol/min per mg protein. SHAM considerably accelerated the hydrogen peroxide production. The SHAM-dependent H2O2 production was stimulated by 2 muM antimycin A and inhibited by 5 mM KCN and 1 muM myxothiazol. The study of the rate of O-2(radical anion) generation by pea mitochondria using EPR spin traps and epinephrine oxidation showed that H2O2 accumulation can be accounted for by a significant increase in the rate of O-2(radical anion) production.