STEREOSELECTIVITY AND REGIOSELECTIVITY IN THE METABOLISM OF 7,8-DIHYDROBENZO[A]PYRENE BY CYTOCHROME-P450, EPOXIDE HYDROLASE AND HEPATIC MICROSOMES FROM 3-METHYLCHOLANTHRENE-TREATED RATS

The active site of cytochrome P450 1A1 has been probed with the substr ate, 7,8-dihydrobenzo[a]pyrene using a purified, reconstituted system composed of cytochrome P450 1A1, NADPH-cytochrome c reductase and lipi d in the presence or absence of epoxide hydrolase. The turnover of the substrate was found to be 38 nmol/nmol of cytochrome P450/min. The me tabolic products that were identified are: a phenolic 7,8-dihydrobenzo [a]pyrene (20-29%); 9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (17-2 8%); benzo[a]pyrene (12-19%); 7-hydroxy-7,8-dihydrobenzo[a]pyrene (13- 16%); 8-hydroxy-7,8-dihydrobenzo[a]pyrene (7-15%); 3-hydroxybenzo[a]py rene (7-15%); 4,5-epoxy-4,5,7,8-tetrahydrobenzo[a]pyrene (0-4%); and a triol of 7,8,9,10-tetrahydrobenzo[a]pyrene (0-4%). 9,10-Epoxy-7,8,9,1 0-tetrahydrobenzo[a]pyrene undergoes rapid hydrolysis to cis- and s-9, 10-dihydroxy-7,8,9,1O-tetrahydrobenzo[a]pyrene (2:1) by benzylic attac k of water at C-10. Approximately 71% of the trans diols are derived f rom 10R)-9,10-epoxy-7,8,9,1O-tetrahydrobenzo[a]pyrene, indicating that cytochrome P450 1A1 has more than a 2:1 preference for selective epox idation of an enantiotopic face of 7,8-dihydrobenzo[a]pyrene. This ste reo-selectivity agrees with the postulated stereo-selectivity predicte d by a previously described active site model for cytochrome P450 1A1. Epoxide hydrolase in pure form or in hepatic microsomes catalyzes the hydrolysis of 9,10-epoxy-7,8,9, 10-tetrahydrobenzo[a]pyrene, which is inhibited by 1,1,1-trichloropropane 2,3-oxide. The (+)-(9S,10R)-isome r of the epoxide is slightly preferred as a substrate over its enantio mer and is cleaved by benzylic and nonbenzylic attack. Only benzylic a ttack was found with 10S)-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene .