Inactivation of microsomal NADPH-cytochrome c reductase by sulfhydryl-reactive derivatives of menadione

Five derivatives of menadione (2-methyl-1,4-naphthoquinone) having electronegative substituents on allylic carbons were prepared for study as sulfhydryl-reactive inactivators of mouse liver microsomal NADPH-cytochrome c reductase. Each of these naphthoquinones, incubated with dilute suspensions of microsomes, produced a loss of NADPH-cytochrome c reductase activity proportional to the initial naphthoquinone concentration. Each of the compounds also reacted with cysteine, as evidenced in the case of the halogenated compounds, by loss of reactive sulfhydryl groups and, in the case of 2-p-nitrophenoxymenadione, by the displacement of the leaving group, p-nitrophenol. Menadione, incubated under identical conditions, did not inactivate NADPH-cytochrome c reductase and was unreactive with cysteine. The requirement for a halogen or a nitrophenoxy substituent on at least one of the allylic carbons suggested that the mechanism of NADPH-cytochrome c reductase inactivation involves attack on critical microsomal nucleophiles, possibly sulfhydryl groups. The possible significance of these findings is discussed in relation to the antitumor activity and bioactivation of the halomethyl naphthoquinones.