Induction of a rat liver benzo[a]pyrene-trans-7,8-dihydrodiol glucuronidating activity by oltipraz and β-naphthoflavone

Abstract

We report here that rats possess a hitherto unrecognized xenobiotic-inducible hepatic 7,8-dihydro-7,8-diol-benzo[a]pyrene (BPD) UDP-glucuronosyltransferase (UGT) activity. BPD UGT activity is induced in female F334 rat liver by treatment with the selective Phase 2 conjugation enzyme inducer oltipraz [4-methyl-5-(2-pyrazinyl)-1,2-dithiole-3-thione at 75-450 mg/kg per day for 3 days] and also by a polycyclic aromatic hydrocarbon-type inducer, β-naphtho-flavone (80 mg/kg per day for 3 days). Incubations of oltipraz-treated rat liver microsomes with racemic trans BPD (100 μM) resulted in formation of two fluorescent glucuronides that were resolved by silica thin layer chromatography (R(f) 0.5 and 0.6). Incubations with either the (-) or (+) trans BPD isomers resulted in selective formation of the R(f) 0.5 [designated -DS, for (-) diol specific] or R(f) 0.6 [designated +DS, for (+) diol specific] glucuronide, respectively. The -DS and +DS BPD glucuronides were fluorescent under long wave ultraviolet irradiation, dependent on the presence of UDP-glucuronic acid in the incubation, and were β-glucuronidase-sensitive. The inducing effect of oltipraz on BPD UGT activity was dose-dependent. The mean BPD UGT activity of the vehicle-treated control group was 0.05 ± 0.02 nmol/mg per min compared with 0.53 ± 0.07 nmol/mg per min in the group treated with oltipraz (450 mg/kg per day for 3 days) (P < 0.001). The apparent K(m) of the induced BPD UGT for BPD was 20 μM, suggesting that the enzyme has the capacity to bind and turnover BPD under physiological conditions. Pretreatment with β-naphthoflavone, but not phenobarbital, induced BPD UGT activity to approximately the same extent as oltipraz. Neither oltipraz nor β-naphthoflavone exhibited induction of BPD UGT in livers of homozygous Gunn rats, which lack functional UGT1-encoded isozymes. We conclude that the oltipraz- and polycyclic hydrocarbon-responsive BPD UGT is a member of the UGT1 family. The role of this isoform as a modifier of susceptibility to carcinogenesis elicited by B[a]P remains to be determined.