The Role of NADH in Uncoupled Microsomal Monoxygenations

Abstract

The stoichiometry of the liver microsomal monoxygenase system from phenobarbital‐pretreated rabbits has been investigated with cyclohexane, n ‐hexane and perfluoro‐ n ‐hexane as substrates. With cyclohexane an almost 1:1:1 stoichiometry for cyclohexanol: NADPH: O 2 was obtained. With the uncoupler perfluoro‐ n ‐hexane no product was formed and 2 moles of NADPH were oxidised per mole of oxygen. The stoichiometry of n ‐hexane hydroxylation suggested that partial uncoupling with this substrate had occurred. NADH addition together with NADPH had no significant effect on the hydroxylation of cyclohexane but led to a more than additive hydroxylation in the case of n ‐hexane. Oxygen uptake with perfluoro‐ n ‐hexane and NADPH was almost doubled in the presence of NADH. It was shown that during uncoupling an increased oxidation of NADH via cytochrome b 5 occurs. It was concluded that active oxygen which is not used for monoxygenations is reduced to water by the NADH‐cytochrome b 5 system and that this sparing effect could be the main mechanism of the well‐known synergistic action of NADH on microsomal hydroxylations.