Hydroxylation of 5,5-diphenylhydantoin (phenytoin) by dog liver microsomes

Chang and Glazko in 1972 had reported failure to demonstrate any production from 5-5-diphenylhydantoin (phenytoin, DPH) by dog liver microsomes of either 5-(m-hydroxyphenyl-5-phenylhydantoin (m-HPPH) or 5-(P-hydroxyphenyl-5-phenylhydantoin (p-HPPH), metabolites of DPH produced by the dog in vivo. We have incubated DPH with 9,000 X g supermatants of dog liver homogenates and with suspensions of separated microsomes with added NADPH generating system, Mg(2+), and nicotinamide and have demonstrated the production of both m-HPPH and p-HPPH. Both metabolities were detected by thin-layer chromatography of extracts of the incubation mixtures. Detection and roughly quantitative measurement of m-HPPH were also accomplished with gas chromatography.