The biochemical basis for the species difference in hepatic microsomal 4-vinylcyclohexene epoxidation between female mice and rats

Mice but not rats are susceptible to 4-vinylcyclohexene (VCH)-inducedovarian toxicity and carcinogenicity. This is due in part toa 4- to 6-fold greater rate of hepatic microsomal bioactivationof VCH to the ovotoxicant VCH-1,2-epoxide. The biochemical basisfor this difference was investigated in microsomes using enzymeinduction, enzyme inhibition with chloramphenicol or specificinhibitory antibodies, and correlation with marker steroid hydroxylaseactivities to associate VCH epoxidation with particular cytochromeP450 forms. Testosterone 6ß- and 15-hydroxylase activitiesand VCH epoxidation were decreased in microsomes from chloramphenicol-treatedmice, initially suggesting the possible involvement of P450IIIAand P450IIA forms in VCH metabolism. Although both testosterone6ß-hydroxylase and VCH epoxidase activities were increasedby dexamethasone treatment (P450IIIA inducer), anti-rat P450IIIAIgG inhibited testosterone 6ß-hydroxylase (68%) butnot VCH epoxidase activity. These latter results do not supportthe involvement of mouse P450IIIA forms in VCH epoxidation.However, results were obtained which indicated that mouse P450IIIAforms are involved in VCH epoxidation. In microsomes from untreatedfemale mice VCH epoxidase activity was inhibited 48% by antibodiesto mouse P450₁₅ (P450IIA3) at a concentration that inhibitedtestosterone 15-hydroxylase activity by 86%. No protein immunochemicallyrelated to mouse P450₁₅ was detected in female rat hepatic microsomes.VCH epoxidation by hepatic microsomes was increased in femalemice and rats by phenobarbital treatment and was inhibited byapproximately one-third by anti-rat-P450IIB1 IgG in microsomesfrom untreated animals of both species. Furthermore, microsomalVCH epoxidase and testosterone 16-hydroxylase activities werelower (34%) in female 129/J mice (deficient in constitutiveexpression of P450IIB forms) than in B6C3F1 mice. These resultssuggested partial involvement of P450IIB forms in the microsomalepoxidation of VCH. Therefore, P450 forms IIA and IIB accountfor the majority of VCH bioactivation in female mouse liver,which explains in part the susceptibility of mice to VCH-inducedovarian toxicity and carcinogenicity.     

Effects of manganese compounds on carcinogenicity of nickel sub-sulfide in rats

The effects of manganese compounds upon the carcinogenicityof Ni₃S₂ were tested in male Fischer rats. In Experiment I,rats were given i.m. injections of Ni₃S₂ (2.5 mg) and Mn dust(2.0 mg), singly or in combination. By 100 weeks, sarcomas occurredat the injection site in 0 of 24 rats in the vehicle controlgroup, in 0 of 24 rats that received Mn dust alone, and in 23of 24 rats that received Ni₃S₂ alone. Combined administrationof Ni₃S₂ plus Mn dust as a single i.m. injection resulted insarcomas in 14 of 23 rats (p <0.05 versus Ni₃S₂ alone). Inrats that received injections of Ni₃S₂ in one thigh and Mn dustin the opposite thigh, the sarcoma incidence at the site ofNi₃S₂ injection was 24 of 24 rats. In Experiment II, rats weregiven i.m. injections of Ni₃S₂ (1.2 mg) and Mn compounds (MnS,Mn₂O₃, MnO₂ or MN₂(CO)₁₀, in dosages equivalent to 1.0 mg ofMn), singly or in combination. No sarcomas occurred at the injectionsite in rats that received the vehicle or any of the manganesecompounds alone. Sarcomas occurred in 13 of 27 rats that receivedNi₃S₂ alone; this sarcoma incidence was not reduced by admixtureof any of the Mn compounds. The median tumor latent period andthe me- Wan survival period were significantly longer (p <0.05)in rats that received MnS plus Ni₃S₂, compared with rats thatreceived Ni₃S₂ alone, suggesting that MnS may have weak anticarcinogeniceffect. These experiments demonstrate that inhibition of Ni₃S₂-carcinogenesisby Mn dust is a local rather than a systemic effect, and that,with the possible exception of MnS, the other manganese compoundsthat were tested are ineffective as inhibitors of Ni₃S₂-carcinogenesis.     

Autocrine production of TGF-{alpha} and TGF-{beta} during tumour progression of rat oral keratinocytes

This study describes a new technique to separate transforminggrowth factor- (TGF-) and transforming growth factor-ß(TGF-ß) from culture supernatants using ion exchangechromatography; assays of competitive inhibition of ligand bindingwere used to quantify the amount of growth factor. The methodwas simple, inexpensive and did not require large volumes ofculture medium. The autocrine production of TGF- and TGF-ßwas examined in oral keratinocyte cell lines derived from thepalatal and lingual mucosa of rats painted with the carcinogen4-nitroquinoline N-oxide (4NQO). Escape from cellular senescence(immortality) was associated with a marked increase in TGF-production (cell line R2P) but tumour progression, as reflectedby the development of anchorage independence in agarose gelsand tumorigenicity in athymic mice, did not result in a consistentincrease or decrease of TGF- production compared to normals.Four cell lines(R8AP, R1T, R3T, R1P), with different functionalcellular phenotypes, produced two to three times more TGF- thannormals. TGF- production was inversely correlated to epidermalgrowth factor cell surface receptor expression. The autocrineproduction of TGF-ß was variable with the majorityof cell lines producing markedly little TGF-ß threecell lines (R4T, R8BP, R9T) produced more TGF-ß thannormals. The production of TGF-ß was unrelated totumour progression, the expression of TGF-ß cell surfacereceptors or TGF- production. The results indicate that theautocrine production of TGF- and TGF-ß are not accuratemarkers of tumour progression in the rat 4NQO model of oralcarcinogenesis.     

Supplemental administration of 1{alpha}-hydroxyvitamin D3 inhibits promotion by intrarectal instillation of lithocholic acid in N-methyl-N-nitrosourea-induced colonic tumorigenesis in rats

The effect of 1-hydroxyvitamin D₃ [1(OH)D₃) on promotion byintrarectal instillation of lithocholic acid (LC) in N-methyl-N-nitrosourea(MNU)-induced colonic tumorigenesis was studied in a rodentmodel. Ninety-two female F344 rats received intrarectal injectionof 2.5 mg of MNU twice in one week followed by 1 mg of LC orits vehicle alone three times weekly for 48 weeks. Those whichreceived LC were given a concomitant intragastric administrationof 0.04 µg of 1(OH)D₃ or its vehicle alone three timesweekly. In the group receiving MNU alone (n=30) five rats borecolomc tumors; in the MNU + LC group (n=32) 15 and in the MNU+ LC + 1(OH)D₃ group (n=30) six rats bore colonic tumors (MNU+ LC versus MNU + LC + 1(OH)D₃ group, P<0.05). These resultsindicated that promotion of MNU-induced colonic tumorigenesisby LC was suppressed by supplemental administration of 1(OH)D₃.     

Hormonal regulation of sex differentiated parameters in liver nodules from rats treated in the resistant hepatocyte model

Sex differentiation of liver functions has been shown to beattenuated in preneoplastic rat liver nodules. The present studywas performed to investigate whether nodules from male ratsare to some extent withdrawn from the normal growth hormone(GH) regulation of these functions. Male and female Wistar ratswere treated according to a modified resistant hepatocyte model(RH-model), with diethylnitrosamine initiation and promotionwith intragastric administration of 2-acetylaminofluorene (2-AAF)combined with partial hepatectomy (PH). Eleven months post-initiationmale rats were treated with either human (hGH) or bovine growthhormone (bGH) or ovine prolactin (oPRL) by continuous infusionfor 1 week. The mRNA expression of a number of genes known tobe sex differentiated in liver from adult control rats was comparedin nodular and surrounding tissue from nodule-bearing male,female and hormone-treated male rats. The basal mRNA expressionof the female-predominant cytochrome P4502C12 (CYP2C12) wasincreased and the male-predominant CYP2C11 was decreased inliver nodules from male rats compared with the surrounding liver.Expression of the prolactin receptor (PRL-r; female > male)and the steroid 5-reductase (female > male) genes was decreasedin male nodules, whereas no difference was observed with respectto GHreceptor (GH-r; female > male) expression in nodulesversus surrounding tissue. Early nodules obtained from malestreated according to the original RH-model (dietary 2-AAF, 0.02%)and isolated 2 weeks after completion of the 2-AAF/PH treatmentshowed significantly lower GH-r mRNA levels than the total livertissue. In hepatocellular carcinomas from hormonally unmanipulatedmales 11 months post-initiation the decrease in PRL-r expressionwas even more pronounced than in the nodules and a significantdecrease in GH-r expression was seen. In female nodules theonly significant difference with respect to the sex differentiatedparameters was a lower 5-reductase expression than in the surroundingtissue. Continuous infusion of both hGH and bGH feminized theexpression of all the sex differentiated genes in male tissuesand eliminated the previously detected differences between nodulesand surrounding tissue. oPRL also eliminated the differencesbetween nodules and surrounding tissue in males and partly feminizedthe expression of both the 5reductase and the PRL-r genes. Althoughthe expression of several sex differentiated parameters in livernodules is altered compared with surrounding tissue, the preneoplasticlesions respond adequately to the feminizing effect of continuousGH infusion, thereby indicating that nodular tissue is not withdrawnfrom the normal endocrine control of rat liver.     

Transforming growth factor-{alpha} and epidermal growth factor expression in the exocrine pancreas of azaserine-treated rats: modulation by cholecystokinin or a low fat, high fiber (caloric restricted) diet

Expression of transforming growth factor- (TGF-) and epidermalgrowth factor (EGF) was studied in normal pancreatic tissueand in (pre)neoplastic pancreatic lesions of azaserine-treatedrats. They were given either a low fat, high fiber (low caloric)diet, to inhibit carcinogenesis, or a low fat diet combinedwith injections of the cholecystokinin analog caerulein to enhancecarcinogenesis. The control groups, maintained on a low fatdiet, were injected with azaserine or were not treated at all.Autopsy was performed at 6 and 15 months after the last azaserineinjection. After both 6 and 15 months immunohistochemistry revealeda weak expression of EGF and TGF- peptides in the acinar cellsand a stronger expression in the ductular and centroacinar cells.TGF- peptide expression was reduced in both putative preneoplasticand neoplastic acinar cell lesions, but no differences in EGFpeptide expression were observed between the various stagesof exocrine pancreatic carcinogenesis. After 16 months an increasein TGF- mRNA due to treatment with azaserine was detected bysemi-quantitative PCR in total pancreatic homogenates, whereasEGF mRNA expression had decreased. TGF- mRNA levels in macroscopicallyisolated tumors were significantly lower, but EGF mRNA levelswere significantly higher, than in total pancreatic homogenatesfrom azaserine treated rats. Furthermore, EGF and TGF- mRNAlevels in isolated tumors did not differ significantly frommRNA levels in non-carcinogen-treated rats. Neither with immunohistochemistrynor with PCR were differences in EGF or TGF- expression observeddue to either inhibition or stimulation of carcinogenesis. Itis concluded that putative preneoplastic acinar cell lesionsinduced in rat pancreas by azaserine may develop into acinaradenocarcinomas independently of TGF- and EGF. The results suggestinvolvement of these growth factors at the early stage of thecarcinogenic process, during the initiation of normal acinarcells into putative preneoplastic cells. However, modulationof azaserine-induced pancreatic carcinogenesis by cholecystokininor a low fat, high fiber (caloric restricted) diet appearednot to be regulated by EGF or TGF-.     

Metabolic predisposition of a novel mutant (LEC rats) to hereditary hepatitis and hepatoma: alterations of the drug metabolizing enzymes

Previously we established that ‘LEC rats’ have displayedspontaneous fulminant hepatitis with severe jaundice, whichprogressed to liver cancer, and a single autosomal recessivegene is responsible for the cause of the diseases. The activitiesof drug metabolizing enzymes were assayed in livers from LECand control (LEA) rats at 4 weeks and 3 months before the onsetof liver cancer. At 4 weeks the cytochrome P-450 content ofthe LEC rat livers was 43% of the control (LEA) value. At 3months the level was 65% of the control. Epoxide hydrolase,-glutamyltranspeptidase and UDP-glucuronyl-transferase activitieswere 2.6-, 6.9- and 2.4-times higher than those in the LEA ratsat 4 weeks, respectively, while glutathione S-transferase activitywas not significantly different between the two strains. Theenzyme changes in the LEC rats are quite similar to those observedin hyperplastic foci and nodules in chemical carcinogenesisof hepatocytes.     

UVR induction of TGF{alpha}: a possible autocrine mechanism for the epidermal melanocytic response and for promotion of epidermal carcinogenesis

The occurrence of the epidermal growth factor homologue, transforminggrowth factor (TGF), in embryonic and neoplastic tissues suggeststhat it may he an oncofetal version of epidermal growth factor.A strong case is developing for TGF to have an autocrine modeof action in sustaining the autonomous growth of several typesof tumour. We propose that TGF normally has an autocrine rolenot only in stimulating the growth of some fetal tissues butalso with postnatal epidermal cells in response to local stimuli—inparticular ultraviolet radiation (UVR). As a first step to testthis hypothesis we have checked whether UVR will induce theproduction of TGF, measured by radioimmunoassay, using a highlyspecific monodonal antibody which recognizes native, biologicallyactive human TGF. We found that cultures of normal foreskinmelanocytes do not produce detectable amounts of TGF when grownunder routine conditions, but, within 12 h of exposure to lowdoses of short-wavelength UVR, significant quantities of TGFare produced. The UVR-induced TGF is both cell associated andreleased into the medium of these cultures. Also, UVR has apromoting action on epidermal cells which have been initiatedby carcinogenic activity. A significant part of the promotingactivity may be due to autocrine stimulation of multiplicationof partially transformed epidermal cells. In this regard wefound that UVR induced TGF in HeLa cells and all human melanomalines so far tested. Induction was complete within 24 h of asingle exposure. Dose-response curves of TGF induction in amalignant melanoma cell line showed a distinctive peak of factorinduced by low (2 J/m²) doses of UVR. Higher doses which inhibit[³H]thymidine incorporation resulted in lower levels of inducedTGF. These findings are consistent with the participation ofTGF as an autocrine mediator of UVR-induced tumour promotion,as well as cell multiplication, in sun-exposed skin.     

Existence of multiple forms of microsomal epoxide hydrolases with radically different substrate specificities

Evidence for the existence in rat and rabbit liver of two microsomalepoxide hydrolases with radically different substrate specificitieswas obtained, one with a broad specificity (EH_b), whilst theother catalyzed the hydrolysis of cholesterol 5,6-oxide (EH_ch),a reaction taken as diagnostic since it was not observed withpure fractions of EH_b. The two enzymes were physically separatedby immunoprecipitation using antibodies which had been raisedagainst EH_b purified to apparent homogeneity. The substratespecificity of the two enzymes is radically different and mutuallycomplementary. Cholesterol 5, 6-oxide has a trisubstituted oxiranering. All epoxides of this nature tested to date were not, orvery poor, substrates of EH_b. The two enzymes can also effectivelybe discriminated by inhibitors, in that 5, 6-imino-5-cholestane-3ß-olpotently inhibits EH_ch but not EH_b whilst 1, 1, 1-trichloropropeneoxide has the opposite specificity. The cytosolic EH did notsignificantly contribute to the catalysis of the hydrolysisof cholesterol 5, 6-oxide.     

Involvement of transforming growth factor-{alpha} and its receptor in a model of DES-induced renal carcinogenesis in the Syrian hamster

This study explores the role played by TGF in estrogen-inducedrenal tumors. Tumors were induced in male Syrian hamster bychronic administration of diethylstilbestrol (DES). Six experimentalgroups (n = 5–9) were chronically exposed to DES and sacrificedafter 1, 2, 4, 6, 9 and 11 months, respectively. In the courseof treatment, the nephrons were the site of an important increaseof cell turnover, which was characterized by a process of hyperplasia/dysplasiain proximal tubules preceding the neoplastic transformation.In treated animals and in controls, the analysis of renal tissueby Western blot revealed the presence of a 6 kDa polypeptidecrossreacting with anti-TGF antibody. In controls, TGF immunoreactivitywas localized in proximal and in distal tubules. Before tumordevelopment (1–4 months), TGF RIA showed an increase ofTGF concentration in renal tissue, in parallel with the increasedcell proliferation observed in proximal tubules. In addition,Western blot analysis also demonstrated in kidney tissue thepresence of a 165 kDa protein displaying the immunoreactivityof EGF/TGF receptor. The receptor immunoreactivity was localizedin proximal tubular cells suggesting an involvement of TGF intubular epithelial growth through autocrine or paracrine pathways.In large neoplasms, immunocytochemistry revealed only clustersof transformed cells intensely stained by the anti-TGF antibody.These cells displayed the appearance of stellate or polyhedriccells infiltrating adjacent neoplastic tissues. Antisera raisedagainst intra-or extracytoplasmic domains of the EGF/TGF receptorwere assayed to localize this receptor in the tumors. In contrastwith tubular structures, immunoreactivity to EGF/TGF receptorwas never detected in tumoral tissue. The apparent absence ofEGF/TGF receptor immunoreactivity in malignant cells seems toexclude an involvement of this growth factor in the tumorigenicprocess, although it could be involved in tumor neovascularization.     

The metabolism of nitrosodi-n-propylamine, nitrosodiallylamine and nitrosodiethanolamine

Nitrosodiallylamine has been reported to be non-carcinogenicin rats while nitrosodipropylamine and nitrosodiethanolamineare liver carcinogens. That nitrosodipropyiamine is metabolizedat the -position by liver microsomes from Fischer-344 rats supportsthe widely held contention that such metabolism is responsiblefor the carcinogenicity of nitrosamines. Nitrosodiallylamineis also metabolized at the a-position by the same microsomalpreparations. Thus, although -oxidation may be responsible forthe carcinogenicity of some nitrosamines, this mechanism alonecannot account for tumorigenicity. Nitrosodiethanolamine isnot metabolized by rat liver microsomes, but is metabolizedby hepatocytes for Fischer-344 rats. In this case, a mechanismother than the oxidation at the -position may be responsiblefor the carcinogenic action.     

Initiating activity in a two-stage mouse skin model of nine mutagenic pyrolysates of amino acids, soybean globulin and proteinaceous food

Trp-P-1, Trp-P-2, MeAC, AC, Glu-P-1, Glu-P-2, Lys-P-1, IQ andPhe-P-1 were tested for tumour initiating activity in a two-stageskin carcinogenesis model using 12-O-tetradecanoylphorbol-13-acetate(TPA) as the promoter. The total initiating doses were 20 mgfor Trp-P-1, Trp-P-2, Glu-P-1 and Glu-P-2, 40 mg for MeAC andAC, 5 mg for Lys-P-1, 7.5 mg for IQ and 100 mg for Phe-P-1.7,12-Dimethylbenz[a]anthracene was used as a positive controlcompound at a total dose of 100 µg. All compounds weretopically applied twice weekly for 5 weeks on the dorsal skin,and then followed by similar TPA administration for 47 weeks.Trp-P-2 induced skin tumours in 30% of the mice (0.35 tumours/mouse),and Trp-P-1, MeAC and Phe-P-1 in 10–20% (0.20–0.25tumours/mouse). The smaller amounts of Lys-P-1 and IQ appliedinduced tumours at an incidence of 10 and 5% respectively. Notumours appeared in the groups treated with test chemicals aloneor TPA alone. Statistical analysis according to either the Fisherexact test or Peto trend test revealed significant differencesfor tumour appearance in the Trp-P-1, Trp-P-2, MeAC and Phe-P-1followed by TPA groups as compared with that given TPA alone.The data were used to generate ID₅₀ (50% initiating dose) valuesfor each of the compounds.     

Inhibitory effect of {alpha}-tocopherol on the formation of nitrosomorpholine in mice treated with morpholine and exposed to nitrogen dioxide

The possibility that -tocopherol (vitamin E) inhibits the formationof nitrosomorpholine (NMOR) in vivo was investigated in miceorally pretreated with -tocopherol (2.5–100 mg/kg bodywt) once daily for 6 days. Twenty-four hours later, the animalswere injected i.p. with 2 mg of morpholine (MOR) per animalfollowed by exposure txo 47 p.p.m. of NO₂ for 2 h. Under theseconditions, low oral doses of -tocopherol (2.5–5 mg/kgbody wt) significantly decreased NMOR formation in vivo. Astotal body -tocopherol levels increased, in vivo NMOR formationdecreased, and a maximal 50–70% inhibition of NMOR formationoccurred at -tocopherol levels of 5 µg/g body wt. Additionalresults showed that NMOR was rapidly eliminated in mice, sothat studies which measure the levels of NMOR found in animalstreated with MOR and then exposed to NO₂ may underestimate theamount of NMOR that is actually formed. Finally, oral pretreatmentof up to 100 mg of -tocopherol/kg body wt had no effect on NMORelimination.     

Enhancement of BHA-induced proliferative rat forestomach lesion development by simultaneous treatment with other antioxidants

Synergistic effects of butylated hydroxyanisole (BHA) and otherantioxidants on induction of rat forestomach lesions were investigated.Groups of F344 male rats were treated with 1% BHA phis 0.7%butylated hydroxytoluene (BHT), 1% BHA phis 1% propyl gallate(PG), 1% BHA plus 1% sodium L-ascorbate (SA), 1% BHA phis 1%DL--tocopherol (-TP), 0.4% BHT phis 0.4% BHA plus 0.4% PG plus0.4% SA plus 0.4% -TP, 1% BHA or 2% BHA. Further groups of 10rats each received antioxidants without BHA as controls. Histo-togfcalexamination revealed significantly increased incidences of hyperplasiain the groups given BHA together with SA or PG at the prefundicregion or at the mid region respectively. The forestomach changesinduced by BHA together with SA were equal to those inducedby 2% BHA. On the other hand, simultaneous treatment with BHAand PG or -TP reduced the incidence of hyperplasia at the prefundkregion. It is concluded that mixed treatment with BHA and otherantioxidants exerted enhancing or inhibitory effects on theinduction of hyperplasia at different sites of the forestomachepithelium.     

Differential regulation of two microsomal epoxide hydrolases in hyperplastic nodules from rat liver

Two microsomal epoxide hydrolases of the rat liver were foundto be differentially regulated in hyperplastic nodules. Whilstthe activity for substrates of the well-known microsomal epoxidehydrolase with a broad substrate specificity (EHb), benzo[a]pyrene4,5-oxide and androstene oxide (16,17-epoxyandrosten-3-one),was greatly (5-fold) increased in the nodule microsomes andmoderately (2-fold) increased in the surrounding tissue, thatfor the substrate of the novel microsomal epoxide hydrolase,cholesterol 5, 6-oxide (EH_ch) remained unchanged. Since bothenzymes convert endogenous steroid epoxides but with distinctstructural features, this differential regulation may indicatea role of endogenous steroid epoxide(s) of a defined structureduring hepatocarcinogenesis. Alternatively, this differentialregulation may serve as a marker during hepatocarcinogenesis.     

TGF-{alpha} and EGF-receptor mRNAs in human oral cancer

Transforming growth factor alpha (TGF-) and epidermal growthfactor receptor (EGFR) have been shown to be present in mostsquamous cell carcinomas. Using the Syrian hamster oral cancermodel, we have recently demonstrated the consistent presenceof TGF- and EGFR mRNAs in chemically transformed hamster oralkeratmocytes. We now present evidence that in human oral cancer(in vivo and in vitro), TGF- and EGFR mRNAs can also be consistentlydetected. No TGF- mRNA can be detected in normal human oralepithelium by Northern blot analysis. These findings reinforcethe use of the hamster cheek pouch as an experimental modelfor the study of oral cancer development, at least in referenceto the possible participation of TGF- in the malignant transformationprocess.     

Hepatocarcinogenic heterocyclic aromatic amines that induce cytochrome P-448 isozymes, mainly cytochrome P-448H (P-450IA2), responsible for mutagenic activation of the carcinogens in rat liver

Male F344 rats were treated with hepatocarcinogenic heterocydicaromatic amines such as amino acid-and protein-pyrolysate components(Trp P-1, Trp P-2, Glu P-1, Glu P-2, AC, MeAC, IQ and MeIQx)and changes in microsomal cytochrome P-450 isozymes in the liverswere examined by means of immuno-Western blotting using anti-ratcytochrome P-450 monoclonal antibodies. The results suggestedthat all chemicals tested induce cytochrome P-448 isozymes,particularly cytochrome P-448H (P-450IA2), which efficientlymediate mutagenic activation of the carcinogens. This was substantiatedby the enzymatic analyses with the substrates showing differentcharacters to rat cytochrome P-450 isozyme-mediated mutagenesis.     

Studies by fluorescence and n.m.r. spectroscopy of the major product formed after further metabolism of ({+/-})-7{beta}, 8{alpha}-dihydroxy-9{alpha}, 10{alpha}-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene

Racemic 7ß, 8-dihydroxy-9, 10-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene(anti-BPDE) is further metabolized by liver microsomes obtainedfrom 3-methylcholanthrene pretreated rats in the presence ofNADPH to at least four products as revealed by h.p.l.c. Dataobtained from measurements by fluorescence spectroscopy underneutral and alkaline conditions and high resolution two-dimensional¹H n.m.r. spectroscopy on the major metabolite derived fromanti-BPDE are consistent with aromatic hydroxylation at the3-position either directly or indirectly via transient epoxideintermediates.