PROTEIN-REACTIVE METABOLITES OF CARBAMAZEPINE IN MOUSE-LIVER MICROSOMES

The character of reactive metabolites formed from carbamazepine (CBZ) was sought in incubations of [C-14]CBZ in hepatic microsomes prepared from adult female mice of a strain (SWV/Fnn) susceptible to CBZ-induce d teratogenicity, The formation of radiolabeled protein adducts was us ed as an index of reactive metabolite exposure, A dependence on cytoch rome P450 was shown by a requirement for NADPH and inhibition by carbo n monoxide, 1-aminobenzotriazole, piperonyl butoxide, and stiripentol. The addition of ascorbic acid, caffeic acid, N-acetylcysteine, and gl utathione decreased the rate of binding of the radiolabel from [C-14]C BZ to microsomal protein by more than 50%. The addition of glutathione transferases diminished protein adduct formation beyond that seen wit h glutathione alone. Evidence for the formation of an arene oxide was sought through the use of inhibitors of epoxide hydrolases, including cyclohexene oxide, chalcone oxides (with the addition of cytosol as ap propriate), and by the addition of recombinant human soluble and micro somal epoxide hydrolases and recombinant rat microsomal epoxide hydrol ase. The microsomal epoxide hydrolases decreased the velocity of C-14- labeled protein adduct formation by approximately 23%, whereas inhibit ors had no effect, most likely because of the low native activity of m icrosomal epoxide hydrolase in mice. Both DT-diaphorase and catechol-O -methyltransferase diminished C-14-labeled protein adduct formation by 54% and 45%, respectively, The data suggest that the major reactive m etabolites formed from CBZ by adult female SWV/Fnn liver microsomes ar e quinones and arene oxides.