ENZYMATIC AND MOLECULAR ASPECTS OF THE ANTIOXIDANT EFFECT OF MENADIONE IN HEPATIC MICROSOMES

The enzymatic features and molecular species of the inhibitory action of menadione on lipid peroxidation in rat liver microsomes were examin ed. In an ascorbate-supported system or a NADH-supported reconstituted system containing NADH-cytochrome b(5) reductase and cytochrome b(5), menadione was not an inhibitor of lipid peroxidation at pH 7.5, while some antioxidant ability was observed at lower pH ranges. Lipid perox idation in the presence of menadione in the NADH-supported reconstitut ed system at pH 7.5 was markedly inhibited by adding lipoamide dehydro genase. NAD(P)H-supported lipid peroxidation in microsomes with increa sed DT-diaphorase activity from 3-methylcholanthrene-treated rats was highly susceptible to menadione. These inhibitions were abolished by d icoumarol, an inhibitor of DT-diaphorase. Cumene hydroperoxide-depende nt lipid peroxidation in microsomes, with desferal and NADP(+) to prev ent nonheme iron-dependent reactions and oxygen radical generation, wa s inhibited by menadione in the presence of NADPH, and the inhibition was also more effective in the microsomes with increased DT-diaphorase activity. Menadiol reacted with 1,1-diphenyl-2-picrylhydrazyl (DPPH) in ethanol at a molar ratio of DPPH/menadiol at 1.9. In an iron suppor ted reconstituted enzymatic or a nonenzymatic system at pH 7.5, menadi ol showed an antioxidant effect at an early stage, followed by a proox idant effect, which was prevented by SOD, probably by protecting menad iol autooxidation. These results show that menadione exerts an antioxi dant effect through participation of microsomal DT-diaphorase by gener ating menadiol with a radical scavenging ability, while menadiol also has a prooxidant property. (C) 1996 Academic Press, Inc.