Stereospecificity in the oxidation of phorate and phorate sulphoxide by purified FAD-containing mono-oxygenase and cytochrome P-450 isozymes

Abstract

1. Both the cytochrome P-450-dependent mono-oxygenase system and the FAD-containing mono-oxygenase catalyse the sulphoxidation of thioether-containing organo-phosphate insecticides. Using purified FAD-containing mono-oxygenase and purified cytochrome P-450 isozymes isolated from mouse liver microsomes, the Stereospecificity of the oxidation of phorate to (+)-and (−)-phorate sulphoxide and the further oxidations of the (+-)- and (−)-phorate sulphoxides to the sulphone, the oxon sulphoxide and the oxon sulphone were examined. 2. The FAD-containing mono-oxygenase catalysed the formation of (−)-phorate sulphoxide, while two cytochrome P-450 isozymes (cytochrome P-450-B2, a constitutive form, and cytochrome P-450-PB, the principal form induced by phenobarbital) produced (+)-phorate sulphoxide. The other three constitutive cytochrome P-450 isozymes examined yielded racemic mixtures. 3. The FAD-containing mono-oxygenase had the lowest K_m for the sulphoxidation reaction, 32 μM, while the K_m values for the cytochrome P-450 isozymes ranged from 67 μM to 250 μM. No additional oxidation of phorate sulphoxide by the FAD-containing mono-oxygenase was detected using either (+)-phorate sulphoxide or (−)-phorate sulphoxide as substrates. 4. In contrast, all five cytochrome P-450 isozymes tested formed additional oxidation products; the (+)-phorate sulphoxide was the preferred substrate for all cytochrome P-450 forms. 5.The final oxidation product, phorate oxon sulphone, was derived by desulphuration of phorate sulphone, with the formation of the oxon sulphoxide being a terminal pathway