To identify cytochrome P450 (CYP) isoform(s) involved in the major metabolic pathways of mianserin (MS) enantiomers in humans, we examined the metabolites formed from S-(+)- and R-(-)-MS using human liver microsomes and eight recombinant human CYP isoforms (i.e., CYP1A2, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1 and 3A4). The mean V(max)/K(m) values of the 8-hydroxylation and N-oxidation were greater for S-(+)- than for R-(-)-MS, whereas that of the N-demethylation gave the opposite observation. When relationships were evaluated in microsomes from 10 human livers between the metabolism of substrates toward the respective CYP isoforms and stereoselective metabolism of MS at a low (5 microM) and at a high (200 microM) MS concentration, significant correlations existed in: 2-hydroxylation of desipramine vs. 8-hydroxylation of S-(+)-MS (r = 0.94, P < .01), O-deethylation of phenacetin vs. N-demethylation of S-(+)-MS (r = 0.85, P < .01) and R-(-)-MS (r = 0.69, P < .05) or N-oxidation of S-(+)-MS (r = 0.94, P < .01) at the low concentration; and 6 beta-hydroxylation of testosterone vs. three metabolic reactions of both MS enantiomers at the high concentration (r = 0.68-0.93, P < .05-.01). Quinidine inhibited the 8-hydroxylation of both enantiomers by < 40% of the respective control values. Furafylline and alpha-naphthoflavone showed a potent inhibitory effect on the N-demethylation and N-oxidation of S-(+)-MS (by up to < 50% of the respective control activities). In addition, troleandomycin inhibited the N-demethylation and N-oxidation of R-(-)-MS by < 50% of the respective control activities. Among the recombinant human CYP isoforms, CYP2D6, 2B6, 3A4 and 1A2 catalyzed the 8-hydroxylation, and CYP1A2 and 3A4 were involved exclusively in the N-oxidation, whereas CYP2B6, 2C19, 1A2, 3A4 and 2D6 showed a catalytic activity for the N-demethylation, for either or both of MS enantiomers. Taken together, the 8-hydroxylation for both MS enantiomers is mediated mainly via CYP2D6, whereas the N-demethylation for both the enantiomers and N-oxidation for S-(+)-MS are catalyzed mainly via CYP1A2. The two in vitro experiments suggested that CYP3A is involved to a certain extent in each of the stereoselective MS metabolic pathways.