Oxygen-18 incorporation studies of the metabolism of phenytoin to the catechol.
Phenytoin (PHT) is metabolized primarily to 5-(4-hydroxyphenyl)-5-phenylhydantoin (4-HPPH), a 3,4-dihydrodiol metabolite (DHD), and a catechol, 5-(3,4-dihydroxyphenyl)-5-phenylhydantoin (Cat). The objective of the present studies was to determine the mechanism of Cat formation. The experiments were conducted with isolated rat hepatocytes and the 9000g supernatant fraction of mouse liver. Incubations of PHT were done under an 18O2 atmosphere and the incorporation of 18O into 4-HPPH and Cat was determined by mass spectrometry. It was found that the amount of Cat formed relative to the amount of 4-HPPH and DHD formed varied among the enzyme sources employed. However, in all cases, most of Cat formed from PHT contained two atoms of 18O. These results show that PHT is converted to Cat primarily by hydroxylation of 4-HPPH rather than by oxidation of DHD. Cat formation via DHD would add an 16O atom from hydrolysis of a 3,4-epoxide. Only 14-36% of the Cat formed contained one atom of 18O and one atom of 16O. Kinetic studies of Cat formation from 4-HPPH and DHD with rat liver subcellular fractions showed that the Vmax is higher and the Km is an order of magnitude lower with 4-HPPH than with DHD. These data suggest that DHD is only slowly converted to Cat and provide an explanation for the paucity of Cat formation via DHD when PHT is incubated with intact cells.
Billings
R E
RE
Fischer
L J
LJ
D000818:Animals; D002396:Catechols / Q000378:metabolism*; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008297:Male; D051379:Mice; D010103:Oxygen Isotopes; D010672:Phenytoin / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D002396:Catechols; D010103:Oxygen Isotopes; D010672:Phenytoin; C034221:catechol