Effect of superoxide dismutase on the autoxidation of substituted hydro- and semi-naphthoquinones

Abstract

The effect of superoxide dismutase on the autoxidation of hydro- and semi-1,4-naphthoquinones with different substitution pattern and covering a one-electron reduction potential range from −95 to −415 mV was examined. The naphthoquinone derivatives were reduced via one or two electrons by purified NADPH-cytochrome P-450 reductase or DT-diaphorase, respectively. Superoxide dismutase did not alter or slightly enhanced the initial rates of enzymic reduction, whereas it affected in a different manner the following autoxidation of the semi- and hydroquinones formed. Autoxidation was assessed as NADPH oxidation in excess to the amounts required to reduce the quinone present, H 2O 2 formation, and the redox state of the quinones. Superoxide dismutase enhanced 2–8-fold the autoxidation of 1,4-naphthosemiquinones, following the reduction of the oxidized counterpart by NADPH-cytochrome P-450 reductase, except for the glutathionyl-substituted naphthosemiquinones, whose autoxidation was not affected by superoxide dismutase. Superoxide dismutase exerted two distinct effects on the autoxidation of naphthohydroquinones formed during DT-diaphorase catalysis: on the one hand, it enhanced slightly the autoxidation of 1,4-naphthohydroquinones with a hydroxyl substituent in the benzene ring: 5-hydroxy-1,4-naphthoquinone and the corresponding derivatives with methyl- and/or glutathionyl substituents at C 2 and C 3, respectively. On the other hand, superoxide dismutase inhibited the autoxidation of naphthohydroquinones that were either unsubstituted or with glutathionyl-, methyl-, methoxyl-, or hydroxyl substituents (the latter in the quinoid ring). The inhibition of hydroquinone autoxidation was reflected as a decrease of NADPH oxidation suppression of H 2O 2 production, and accumulation of the reduced form of the quinone. The enhancement of autoxidation of 1,4-naphthosemiquinones by superoxide dismutase has been previously rationalized in terms of the rapid removal of O 2 • by the enzyme from the equilibrium of the autoxidation reaction ( Q 2 • + O 2 Q + O 2 • ), thus displacing it towards the right. The superoxide dismutase-dependent inhibition of H 2O 2 formation as well as NADPH oxidation during the autoxidation of naphthohydroquinones — except those with a hydroxyl substituent in the benzene ring — seems to apply to those organic substrates which can break down with simultaneous formation of a semiquinone and O 2 • . Inhibition of hydroquinone autoxidation by superoxide dismutase can be interpreted in terms of suppression by the enzyme of O 2 • -dependent chain reactions or a direct catalytic interaction with the enzyme that might involve reduction of the semiquinone at expense of O 2 • . It is suggested that the complimentary activities of DT-diaphorase and superoxide dismutase may represent a detoxication mechanism in the cellular disposal of several quinones.