Mutagenicity of 7H-dibenzo[c,g]carbazole and metabolites in Salmonella typhimurium

7H-Dibenzo[c,g]carbazole (DBC) is a potent carcinogen of environmental import. Reverse-mutation plate-incorporation assays for mutagenicity were undertaken in Salmonella typhimurium strains TA98 and TA100. Results were negative when no exogenous activation system was used, as well as when assays incorporated liver homogenates (S9) from rats, mice and rabbits. By contrast DBC was mutagenic in a forward mutation assay in Salmonella strain TM677 using resistance to 8-azaguanine for selection. Metabolites of DBC were generated by incubation with rat-liver microsomes and separated by HPLC. Two of these metabolites were directly mutagenic for Salmonella strain TM 677 while two others were mutagenic upon addition of S9. Synthetic phenolic derivatives of DBC were also mutagenic in this assay when further metabolized. It is likely that metabolites of DBC phenols constitute the biologically active forms.     

Comparative metabolism of 7H-dibenzo[c,g]carbazole and dibenz[a,j]acridine by mouse and rat liver microsomes.

The comparative metabolism of the carcinogenic pollutants 7H-dibenzo[c,g]-carbazole (DBC) and dibenz[a,j]acridine (DBA) was investigated in vitro using 3-methylcholanthrene (3MC) induced Sprague-Dawley rat and Hsd:ICR(Br) mouse liver microsomal preparations with benzo[a]pyrene (BaP) as the positive control. Metabolites were isolated and separated by HPLC and identified by spectroscopic and co-chromatographic techniques using synthetic standards. The major metabolites of DBC were the phenols: the 5-OH-DBC, 3-OH-DBC, and 2-OH-DBC. Traces of 1-OH-DBC were also found yet no dihydrodiols were identified. The major metabolites of DBA were the 3,4-diol-DBA and 5,6-diol-DBA, 1,2-diol-DBA, DBA-5,6-oxide and 4-OH-DBA. Treatment of both mice and rats with 3MC resulted in significant (P less than or equal to 0.05) increases relative to control in the microsomal metabolism of DBA to dihydrodiol and phenol metabolites, similar to that observed for BaP. 3MC-induced rat liver microsomes significantly (P less than or equal to 0.05) increased DBC metabolism relative to control microsomes whereas DBC metabolism was not increased with 3MC-induced mouse liver microsomes. These data indicate that different enzymatic pathways are involved in the metabolic activation of DBC in the Hsd:ICR(Br) mouse and Sprague-Dawley rat.     

7H-Dibenzo[c,g]carbazole and 5,9-dimethyldibenzo[c,g]carbazole exert multiple toxic events contributing to tumor promotion in rat liver epithelial 'stem-like' cells

Immature liver progenitor cells have been suggested to be an important target of hepatotoxins and hepatocarcinogens. The goal of the present study was to assess the impact of 7H-dibenzo[c,g]carbazole (DBC) and its tissue-specific carcinogenic N-methyl (N-MeDBC) and 5,9-dimethyl (DiMeDBC) derivatives on rat liver epithelial WB-F344 cells, in vitro model of liver progenitor cells. We investigated the cellular events associated with both tumor initiation and promotion, such as activation of aryl hydrocarbon receptor (AhR), changes in expression of enzymes involved in metabolic activation of DBC and its derivatives, effects on cell cycle, cell proliferation/apoptosis and inhibition of gap junctional intercellular communication (GJIC). N-MeDBC, a tissue-specific sarcomagen, was only a weak inhibitor of GJIC or inducer of AhR-mediated activity, and it did not affect either cell proliferation or apoptosis. DBC was efficient GJIC inhibitor, while DiMeDBC manifested the strongest AhR inducing activity. Accordingly, DiMeDBC was also the most potent inducer of cytochrome P450 1A1 (CYP1A1) and CYP1A2 expression among the three compounds tested. Both DBC and DiMeDBC induced expression of CYP1B1 and aldo-keto reductase 1C9 (AKR1C9). N-MeDBC failed to significantly upregulate CYP1A1/2 and it only moderately increased CYP1B1 or AKR1C9. Only the potent liver carcinogens, DBC and DiMeDBC, caused a significant increase of p53 phosphorylation at Ser15, an increased accumulation of cells in S-phase and apoptosis at micromolar concentrations. In addition, DiMeDBC was found to stimulate cell proliferation of contact-inhibited WB-F344 cells at 1 microM concentration, which is a mode of action that might further contribute to its hepatocarcinogenicity. The present data seem to suggest that the AhR activation, induction of enzymes involved in metabolic activation, inhibition of GJIC or stimulation of cell proliferation might all contribute to the hepatocarcinogenic effects of DBC and DiMeDBC.     

Genotoxicity of 7H-dibenzo[c,g]carbazole and its methyl derivatives in human keratinocytes

Differences between tissues in the expression of drug-metabolizing enzymes may substantially contribute to tissue-specificity of chemical carcinogens. To verify this hypothesis, the spontaneously immortalized human keratinocytes HaCaT were used, in order to evaluate the genotoxic potential of 7H-dibenzo[c,g]carbazole (DBC), a known hepatocarcinogen and sarcomagen, and its synthetic tissue-specific derivatives, 5,9-dimethyl-DBC (DiMeDBC) and N-methyl-DBC (N-MeDBC), which manifest specific tropism to the liver and skin, respectively. HaCaT cells mainly express cytochrome P4501A1 (CYP1A1), which is involved in metabolism of DBC and N-MeDBC, but not DiMeDBC [10]. Both DBC and the sarcomagen N-MeDBC induced significant levels of DNA strand-breaks, micronuclei, and DNA adducts followed by the phosphorylation of the p53 protein and histone H2AX in HaCaT cells. In contrast, the specific hepatocarcinogen DiMeDBC was devoid of any significant genotoxic activity in this cell line. Our study demonstrates that the absence of drug-metabolizing enzyme(s) involved in DiMeDBC metabolism may contribute substantially to the tissue-specific genotoxicity of this hepatocarcinogen.     

Mutagenicity of some derivatives of dipyrido[1,2-a:3',2'-d]imidazoles in Salmonella typhimurium with metabolic activation by rat liver and small intestine subcellular fractions

The mutagenic effect of 2-amino-dipyrido[1,2-a:3',2'-d]imidazole (Glu-P-2) was compared with that of the 3-amino, 3-nitro, or 3-N-hydroxylated derivatives of the same base ring with methyl groups at positions 4 and 6 of the molecule. The compounds were tested in Salmonella typhimurium strain TA98 without metabolic activation and in the presence of different concentrations of subcellular fractions from livers or small intestines of rats pretreated with different P448/P450 inducers. The 4,6-dimethyl compounds are always more mutagenic than Glu-P-2. Pretreatment with Aroclor 1254 (ARO) is the most effective inducer in the activation of the 2- and 3-amino compounds by liver S9, whereas the same fraction decreases the mutagenicity of the 3-nitro derivative. S9 from small intestine increased the mutagenic effect of the 3-nitro and 3-N-hydroxylated compounds, but it was unable to activate the amino compounds.     

A comparison of DNA adduct formation in white blood cells and internal organs of mice exposed to benzo[a]pyrene, dibenzo[c,g]carbazole, safrole and cigarette smoke condensate

Measurement of tissue/cell DNA adducts represents a suitable monitor of carcinogen exposure because the majority of chemical mutagens/carcinogens react with DNA, forming covalent adducts, a key event in the initiation of chemical carcinogenesis. Investigations of DNA-adduct formation in vivo in white blood cells (WBC) versus target tissues, i.e. internal organs for most carcinogens, is expected to yield useful information about the suitability of WBC for biomonitoring and risk assessment. For this purpose, female ICR mice were given 0.4 mmole/kg benzo[a]pyrene (BP), 0.045 mmole/kg dibenzo[c,g]carbazole (DBC) or 2.47 mmole/kg safrole by oral gavage or 4 daily doses (equivalent to 3 cigarettes) of cigarette-smoke condensate (CSC) by topical application. At 24 h after dosing, DNA adducts were detected by a nuclease P1-enhanced 32P-postlabeling assay [M.V. Reddy and K. Randerath, Carcinogenesis, 7 (1986) 1543] in WBC and internal tissues treated with individual carcinogens, while CSC treatment elicited aromatic adducts in most tissues but not in WBC. Adduct patterns of WBC DNA were qualitatively similar to those of internal organs, but adduct amounts varied. BP, a systemic carcinogen, bound nearly as much to WBC DNA as to target-tissue DNA samples; whereas the liver carcinogens, DBC and safrole, bound to WBC DNA considerably less (22- and 51-fold, respectively) compared with liver DNA. The number of adducts in 10(7) nucleotides of WBC, liver, lung, kidney and spleen DNA, respectively, were: 2, 5, 3, 2 and 3 with BP; 6, 131, 6, 14 and 4 with DBC; 5, 238, 3, 5 and 0.6 with safrole. For CSC, these values were 0, 1 and 0.02 in WBC, lung and spleen, respectively. Our results show that carcinogen binding to WBC DNA does not reflect binding to target-tissue DNA in a quantitative sense for the carcinogens studied except for BP, and that WBC are not suitable surrogates for monitoring CSC exposure by DNA-adduct measurement after topical application. The CSC data in mice was consistent with the previous findings in humans that smokers' tissues but not WBC show smoking-related bulky/aromatic DNA adducts, as measured by 32P-postlabeling.     

Induction of micronuclei by 7H-dibenzo[c,g]carbazole and its tissue specific derivatives in Chinese hamster V79MZh1A1 cells

The clastogenicity/aneugenicity of N-heterocyclic polycyclic aromatic pollutant 7H-dibenzo[c,g]carbazole (DBC) and its two synthetic derivatives N-methyl DBC (MeDBC) and 5,9-dimethyl DBC (diMeDBC) was evaluated in the genetically engineered Chinese hamster V79 cell line V79MZh1A1 with stable expression of human cytochrome P4501A1 and in the parental V79MZ cell line without any cytochrome P450 activity. While none of the three carbazoles changed significantly the level of micronuclei in the parental V79MZ cells, a variable, but statistically significant rise of micronucleus frequencies was assessed in V79MZh1A1 cells. DBC induced dose-dependent increase in the number of micronuclei at harvest times of 24 and 48h and MeDBC at sampling time of 48h in V79MZh1A1 cells in comparison to untreated cells, however, no significant time-dependent increase in micronucleus frequencies was found. The use of the antikinetochore immunostaining revealed that DBC and MeDBC induced approximately equal levels of both kinetochore positive (C+) and kinetochore negative (C-) micronuclei. DiMeDBC, a strict hepatocarcinogen, did not manifest any effect on micronucleus induction in V79MZh1A1 cells.These studies suggest that genetically engineered Chinese hamster V79 cell lines expressing individual CYP cDNAs are a useful in vitro model for evaluation the role of particular cytochromes P450 in biotransformation of DBC and its tissue and organ specific derivatives.     

DNA adduct formation and persistence in liver and extrahepatic tissues of northern pike (Esox lucius) following oral exposure to benzo[a]pyrene, benzo[k]fluoranthene and 7H-dibenzo[c,g]carbazole.

The formation and persistence of DNA adducts in liver, intestinal mucosa, gills and brain of juvenile northern pike (Esox lucius) following oral exposure to benzo[a]pyrene (BaP), benzo[k]fluoranthene (BkF) and 7H-dibenzo[c,g]carbazol (DBC) were analysed by 32P-postlabelling. The dosage was 25 micromol/kg body weight of each substance, administered on 5 occasions with an interval of 12-14 days. Sampling was carried out 9 days after the second treatment, and 9, 16, 33 and 78 days after the fifth treatment. Pikes were also fed with the substances singly for comparison of adduct patterns. A complex pattern of adducts was detected in all examined tissues from fish treated with the mixture. Total adduct levels were highest in intestine (347+/-17.4 nmol adducts/mol nucleotides, mean+/-SE), followed by liver (110+/-9.3), gills (69+/-6) and brain (14+/-4.2). In pike treated with BaP alone, one major adduct was detected in all examined tissues. This BaP-adduct made up approximately 50% of the total amount of adducts in the brain. Corresponding values in liver, intestine and gills were 23, 31 and 34%, respectively. One relatively weak BkF-adduct and at least 10 different DBC-adducts were detected in all analysed tissues. Total adduct level in the intestine declined to 29.4% of the maximum value 78 days after the last exposure, while there was no significant decline in adduct levels in liver, gills or brain. The results suggest that intestine is more susceptible to adduct formation than liver after oral exposure, and that adduct levels in the intestine represent ongoing or relatively recent exposure. DNA adducts in the other investigated tissues were much more persistent and may therefore accumulate during long-term exposure.     

Mutagenicity of 3a,8a-dihydrofuro[2,3-b]benzofuran, a model of aflatoxin B1, for Salmonella typhimurium TA100.

Racemic 3a,8a-dihydrofuro[2,3-b]benzofuran has been chemically synthesized as a model of the vinyl ether structure of aflatoxin B₁ (AFB₁) and tested for mutagenicity. In the presence of 9000g rat liver supernatant fraction the compound induced his⁺ revertant colonies in $\text{S. typhimurium}$ TA 100 but with only one five-thousandth the activity of AFB₁. No mutagenicity was found when strain TA98 was used. Omission of the rat liver preparation abolished mutagenic activity. The reduced compound, tetrahydrofurobenzofuran, was inactive as a mutagen either in the presence or absence of the rat liver supernatant.     

Mutagenicity of benzo[a]pyrene 7,8-dihydrodiol and 7,12-dimethylbenz[a]anthracene 3,4-dihydrodiol in S. typhimurium mediated by microsomes from rat liver and mouse skin

The mutagenic activities of trans-7,8-dihydro-7,8-dihydroxybenzo[a]-pyrene (BP 7,8-diol) and of trans-3,4-dihydroxy-7,12-dimethylbenz[a]-anthracene (DMBA 3,4-diol) towards S. typhimurium TA100 were measured in assays that were carried out on a micro-scale in liquid medium in the presence of microsomal fractions prepared from mouse skin or rat liver. In the presence of an NADPH-generating system, microsomal enzymes converted both diols into mutagens that were probably the respective 'bay-region' diol-epoxides. The rate of the enzyme-catalysed conversion of the BP 7,8-diol into mutagens by microsomal preparations from mouse epidermis was similar to that occurring with microsomes from rat liver. Pretreatment of mice by the topical application of benz[a]anthracene (BA) or 7,12-dimethylbenz[a]-anthracene (DMBA) increased the mutagenic activity of BP 7,8-diol mediated by mouse skin microsomal preparations by 2-fold and this was paralleled by a 4-fold increase in epidermal aryl hydrocarbon (benzo[a]pyrene) hydroxylase (AHH) activity. The results are discussed in relation to the high susceptibility of mouse skin to polycyclic aromatic hydrocarbon (PAH) carcinogenesis.     

Mutagenicity of 7H-dibenzo[c,g]carbazole and its tissue specific derivatives in genetically engineered Chinese hamster V79 cell lines stably expressing cytochrome P450

The biological mechanisms responsible for aging remain poorly understood. We propose that increases in DNA damage and mutations that occur with age result from a reduced ability to repair DNA damage. To test this hypothesis, we have measured the ability to repair DNA damage in vitro by the base excision repair (BER) pathway in tissues of young (4-month-old) and old (24-month-old) C57BL/6 mice. We find in all tissues tested (brain, liver, spleen and testes), the ability to repair damage is significantly reduced (50-75%; P<0.01) with age, and that the reduction in repair capacity seen with age correlates with decreased levels of DNA polymerase beta (beta-pol) enzymatic activity, protein and mRNA. To determine the biological relevance of this age-related decline in BER, we measured spontaneous and chemically induced lacI mutation frequency in young and old animals. In line with previous findings, we observed a three-fold increase in spontaneous mutation frequency in aged animals. Interestingly, lacI mutation frequency in response to dimethyl sulfate (DMS) does not significantly increase in young animals whereas identical exposure in aged animals results in a five-fold increase in mutation frequency. Because DMS induces DNA damage processed by the BER pathway, it is suggested that the increased mutagenicity of DMS with age is related to the decline in BER capacity that occurs with age. The inability of the BER pathway to repair damages that accumulate with age may provide a mechanistic explanation for the well-established phenotype of DNA damage accumulation with age.     

Distribution of polypeptides binding guanosine 5''-[gamma-[35S]thio]triphosphate and anti-(ras protein) antibodies in liver subcellular fractions. Evidence for endosome-specific components.

Acetyl-CoA synthetase was purified 800-fold from Bradyrhizobium japonicum bacteroids. A specific activity of 16 mumol/min per mg of protein was achieved, with a 30-40% yield. The purification scheme consisted of only three consecutive chromatography steps. The enzyme has a native Mr of 150,000, estimated by gel-permeation chromatography, and a subunit Mr of 72,000, determined by SDS/polyacrylamide-gel electrophoresis. The optimum pH and temperature are 8.5 and 50 degrees C respectively. The Km values for acetate, CoA and ATP were 146, 202 and 275 microM respectively. The reaction was specific for acetate, as propionate and oleate were used very poorly. Likewise, the enzyme used only ATP, ADP or dATP; AMP, GTP, XTP and UTP could not replace ATP. Acetyl-CoA synthetase showed a broad specificity for metals; MnCl2 could replace MgCl2. In addition, CaCl2 and CoCl2 were approx. 50% as effective as MgCl2, but FeCl3, NiCl2 or ZnCl2 could not effectively substitute for MgCl2. The enzyme may be regulated by NADP+ and pyruvate; no effect was seen of amino acids, glucose catabolites, reduced nicotinamide nucleotides or acetyl-CoA. Inhibition was seen with AMP, PPi, FMN and pyridoxal phosphate, with Ki values of 720, 222, 397 and 1050 microM respectively.     

DNA adduct formation in northern pike (Esox lucius) exposed to a mixture of benzo[a]pyrene, benzo[k]fluoranthene and 7H-dibenzo[c, g]carbazole: time-course and dose-response studies

The time-course and dose dependent formation of DNA adducts in juvenile northern pike (Esox lucius) following a single exposure to a mixture of benzo[a]pyrene (BaP), benzo[k]fluoranthene (BkF) and 7H-dibenzo[c,g]carbazole (DBC) were investigated by use of the (32)P-postlabelling assay. A complex adduct pattern was detected in liver and intestine of exposed fish. For the time-course studies fish were exposed either by oral administration or by intraperitoneal (i.p.) injection. Following a single i.p. injection of the mixture (40micromole/kg body weight of each substance) significantly elevated DNA adduct levels were detected in the liver after 1 day. Adduct levels were higher in liver than in intestine, in which significant elevation were detected from day 3 to 12. Following exposure via food (80micromole/kg body weight of each substance), adduct levels were detected in both liver and intestine 1 day after exposure, and continued to increase until day 3 in liver and day 6 in intestine. Calculation of a binding index, which compensates for differences in dosage, resulted in much higher adduct formation (five times in liver and 22 times in intestine) following oral exposure. Pikes receiving single oral doses of 12.5, 50, 100 or 200micromole/kg body weight of each substance exhibited significantly higher adduct levels in both liver and intestine compared to controls. Hepatic adduct levels were also higher in fish given 100 and 200micromole/kg compared to 12.5micromole/kg. Results from this study show that DNA adducts are rapidly formed in juvenile northern pike following both i.p. injection and feeding of a mixture of BaP, BkF and DBC. A maximum level was reached within a few days, which then persisted at approximately the same level for at least 9-12 days. The results also shows that higher levels of adducts were obtained following oral administration compared to i.p. injection, particularly in the intestine.     

Heterocyclic polycyclic aromatic hydrocarbon carcinogenesis: 7H-dibenzo[c,g]carbazole metabolism by microsomal enzymes from mouse and rat liver

The metabolism of dibenzo[c,g]carbazole (DBC), was studied in vitro using microsomal fractions of mouse and rat liver from animals, which were treated with 3-methylcholanthrene (MC). The separation of extractable metabolites by high pressure liquid chromatography (HPLC) and thinlayer chromatography (TLC) as well as identification of most of them by nuclear magnetic resonance, mass spectrometry and comparison with synthetically obtained products are described. The microsomes of both species produced the same twelve compounds of which the following have been identified: five monohydroxylated derivatives (phenols), the product of further oxidation of one of them, and a dihydrodiol. The 5-OH-DBC (60% including its spontaneously-formed dimer) and the 3-OH-DBC (14%) are the main metabolites. Three minor metabolites cochromatographed with synthetically prepared 2-OH-DBC, 4-OH-DBC and 6-OH-DBC. The dihydrodiol detectable in small quantity (4–6%) was tentatively identified as 3,4-dihydroxy-3,4-dihydro-DBC by the sensitivity of its formation to very low concentrations of the inhibitor of microsomal epoxide hydrolase, 1,1,1-trichloropropene oxide, by its molecular ion and major fragment in mass spectrometry and by its dehydration product 3-OH-DBC. No other dihydrodiols were detected. The qualitative and quantitative effects of various modulators of metabolism (enzyme inhibitors, apparently homogeneous epoxide hydrolase, glutathione, supernatant fraction) were investigated. The results are discussed with respect to possible ultimate carcinogens.     

Mutagenicity of the phenacetin metabolites: N-hydroxy-p-phenetidine and nitrosophenetol in S. typhimurium TA100 and derivatives deficient in nitroreductase or O-acetylase: probes for testing intrabacterial metabolic activation

Two mutagenic metabolites of phenacetin, p-nitrosophenetol and N-hydroxy-p-phenetidine, were tested in S. typhimurium strains TA100, its nitroreductase-deficient derivative TA100NR, and O-acetylase-deficient strains TA100 Tn5-1-8-DNP1011 and -DNP1012 in the presence or absence of an exogenous metabolic activation system. The results indicate that bacterial nitroreductase(s) and O-acetylase(s), shown to be involved in the conversion of certain nitroarenes, are not required for the intrabacterial activation of the two phenacetin metabolites to bacterial mutagens. In view of the low reactivity of nitrosoarenes towards nucleophiles at neutrality, the mechanism by which they exert such a high mutagenic effect in S. typhimurium strains remains to be clarified, but is discussed.     

Suitability of L-[35S]methionine for studying the biosynthesis of the polypeptides of mouse liver endoplasmic reticulum membrane fractions in vivo.

The use of L-[35S]methionine (500-700 Ci/mmol (1 Ci = 37 GBq) for labelling the polypeptides of liver rough (R) and smooth (S)endoplasmic reticulum (ER) membrane fractions in vivo was studied. Adult mice were injected intraperitoneally with 400 muCi of the isotope and killed at various times (2'min to 24 h) thereafter. RER and SER fractions were prepared, stripped of ribosomes, and treated with Triton X-100 to remove intravesicular contents. Sufficient radioactivity was present in individual aliquots (75 microgram protein) of the ER membrane fractions to permit their analysis by fluorography after separation by electrophoresis in polyacrylamide gels containing sodium dodecyl sulphate. By 3 min, although the majority of the labelled components were of intravesicular origin, some 12 membrane polypeptides were labelled in the RER fraction (including one corresponding in migration to cytochrome P-450); some 6 of these latter polypeptides were labelled to a lesser degree in the SER membrane fraction at this time. By 5 min, the patterns of radioactive polypeptides of the RER and SER fractions (including both membrane and intravesicular components) were identical. By 7 min, some 28 labelled membrane polypeptides were detectable in the total microsomal membrane. Analysis of the 24-h samples revealed that all the membrane polypeptides seen by staining with Coomassie blue were visualised by fluorography. Other studies revealed the applicability of the approach used for producing highly labelled cell sap and serum proteins. The overall results demonstrate the suitability of L-[35S]methionine administered in vivo for producing mouse liver ER membrane polypeptides of relatively high radioactivity and are consistent with a rapid conversion of RER to SER by ribosome detachment or membrane flow.     

UKEMS trial: Bacterial mutation tests of 4-chloromethylbiphenyl, 4-hydroxymethylbiphenyl,and benzyl chloride,using E. coli WP2uvrA(pKM101) and S. typhimurium TA98 and TA100

4CMB, 4HMB and BC were assayed in plate tests, using E. coli WP2uvrA(pKM101), and S. typhimurium TA98 and TA100, in the presence or absence of microsomal activation. 4CMB was also assayed in fluctuation tests using E. coli WP2uvrA(pKM101). 4HMB was uniformly negative, and 4CMB was mutagenic to all 3 strains. BC was negative in TA98 and positive in TA100 and WP2uvrA(pKM101). The presence or absence of S9 made no substantial difference to the mutagenicity of 4CMB or BC.     

Epoxy derivatives of aromatic polycyclic hydrocarbons. The synthesis of dibenz[a,c]anthracene 10,11-oxide and its metabolism by rat liver preparations

The synthesis of dibenz[a,c]anthracene 10,11-oxide is described. The oxide was unstable and was rapidly decomposed with cold mineral acid into a mixture of 10- and 11- hydroxydibenz[a,c]anthracene. The oxide was converted by rat liver microsomal preparations and homogenates into a product that is probably 10,11-dihydro-10,11-dihydroxydibenz[a,c]anthracene and which was identical with the metabolite formed when dibenz[a,c]anthracene was metabolized by rat liver homogenates. The oxide did not react either chemically or enzymically with GSH. 10,11-Dihydrodibenz[a,c]anthracene and 10,11-dihydrodibenz[a,c]anthracene 12,13-oxide were both metabolized by rat liver preparations into trans-10,11,12,13-tetrahydro-10,11-dihydroxydibenz[a,c] anthracene and the oxide was converted chemically into this dihydroxy compound, and it reacted chemically but not enzymically with GSH. In the alkylation of 4-(p-nitrobenzyl)pyridine, the ;K-region' epoxide, dibenz[a,h]anthracene 5,6-oxide, was more active than either dibenz[a,c]anthracene 10,11-oxide or 10,11-dihydrobenz[a,c]anthracene 12,13-oxide.     

Mutagenic activation of 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline (MeIQ) by subcellular fractions and cells isolated from small intestine,kidney and liver of the rat

The mutagenic activity of the pyrolysis products 2-amino-3-methylimidazo[4,5-f]-quinoline and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline in Salmonella typhimurium TA98 using rat intestinal and renal subcellular fractions as activation systems was approximately 1 and 5 revertants per nmol, respectively. This was 1,000 times less than the activity with a subcellular fraction from rat liver. The mutagenic activity of both compounds was considerably increased using intestinal, renal and hepatic preparations isolated from PCB (Aroclor 1254)-pretreated rats, compared to preparations from control animals. In addition, both compounds displayed a moderate direct-acting mutagenic activity at concentrations above 10^-5 M. Isolated cells from small intestine, kidney and liver incubated in nucleopore chambers were able to convert both compounds into products which mutated bacteria outside the chambers. The concentrations of chemicals required to yield responses of a similar magnitude were approximately 3 orders of magnitude higher in the intestinal and renal systems compared to the hepatic system. The formation of metabolites mutagenic for Salmonella typhimurium by hepatic subcellular and cellular systems was shown to be superior to the respective intestinal and renal systems.Abbreviations AHH arylhydrocarbon hydroxylase - IQ 2-amino-3-methylimidazo[4,5-f]-quinoline - MC 3-methylcholanthrene - MeIQ 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline - PCB polychlorinated biphenyls (Aroclor 1254) - S9 the 9,000 g supernatant tissue fraction - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin