Epoxy derivatives of aromatic polycyclic hydrocarbons. The preparation and metabolism of epoxides related to 7,12-dimethylbenz[a]-anthracene

The syntheses of 7,12-dimethylbenz[a]anthracene 5,6-oxide, 7-acetoxymethyl-12-methylbenz[a]anthracene 5,6-oxide and a product that appears to be mainly 7-hydroxymethyl-12-methylbenz[a]anthracene 5,6-oxide are described. The compounds readily rearranged to phenols in the presence of mineral acid, and 7,12-dimethylbenz[a]anthracene 5,6-oxide and its 7-hydroxymethyl derivative reacted slowly with water to yield trans-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a] anthracene and trans-5,6-dihydro-5,6-dihydroxy-7-hydroxymethyl-12-methylbenz [a]anthracene respectively. Both epoxides were converted enzymically by rat liver microsomal fractions and homogenates into the related trans-dihydrodiols. The epoxides reacted chemically with GSH to form conjugates that were identical with the conjugates formed when the epoxides were incubated with rat liver homogenates. The GSH conjugates were more stable to acid than conjugates derived from other arene oxides. In the alkylation of 4-(p-nitrobenzyl)pyridine, 7,12-dimethyl-benz[a]anthracene 5,6-oxide was more active than the 5,6-oxides of 7-methylbenz[a]-anthracene and benz[a]anthracene.     

Metabolism of benzo[a]pyrene and 7,12-dimethylbenz[a]anthracene in cultured human fetal aortic smooth muscle cells.

Cultured human fetal aortic smooth muscle cells derived from the abdominal aorta converted benzo[a]pyrene (BaP) and 7,12-dimethylbenz[a]anthracene (DMBA) $\text{via}$ cytochrome P-450-dependent monooxygenation to metabolites detectable by both a highly sensitive radiometric assay and high pressure liquid chromatography (HPLC). Cells incubated with ³H-BaP transformed this substrate primarily to phenols. ¹⁴C-DMBA was converted to metabolites that cochromatographed with 12-hydroxymethyl-7-methylbenz[a]anthracene, 7-hydroxymethyl-12-methylbenz-[a]anthracene, 7,12-dihydroxymethylbenz[a]anthracene, and trans-8,9-dihydrodiol-7,12-DMBA. Exposure of cells in culture to 13 μM 1,2-benz[a]anthracene resulted in increased oxidative metabolism of both BaP and DMBA. In the case of BaP, total phenol formation was increased, while with DMBA all metabilities detected by HPLC were increased. Support for the potential role of metabolism of polycyclic aromatic hydrocarbons by aortic smooth muscle cells in the etiology of atherosclerosis was obtained.     

The metabolism of 7- and 12-methylbenz[a]-anthracene and their derivatives

1. 7- and 12-Methylbenz[a]anthracene were converted by rat-liver homogenates into the corresponding hydroxymethyl derivatives, products that are probably the 8,9-dihydro-8,9-dihydroxy and the 5,6-dihydro-5,6-dihydroxy derivatives, and a number of phenolic products. 2. Both hydrocarbons were converted into glutathione conjugates; that from 7-methylbenz[a]anthracene was also formed, together with 5,6-dihydro-5,6-dihydroxy- and 5-hydroxy-benz[a]anthracene, from 5,6-epoxy-5,6-dihydro-7-methylbenz[a]anthracene. 3. 7- and 12-Hydroxymethyl-benz[a]anthracene were converted into products that are probably 8,9-dihydro-8,9-dihydroxy derivatives, and into phenols. 4. The preparation of a number of derivatives of the hydrocarbons is described. 5. The oxidation of the hydrocarbons with lead tetra-acetate was investigated.     

Metabolism of 7,12-dimethylbenz[a]anthracene and its methyl-hydroxylated metabolites: formation of phenolic metabolites at the 2-positions.

The 1- and 2-positions of 7,12-dimethylbenz[a]anthracene (DMBA) were thought not to be involved in biotransformation to 1,2-epoxide and 1,2-dihydrodiol because of steric hindrance from the 12-methyl group (Biochem. Biophys. Res. Commun. $\text{85}$: 357–362, 1978). However, we have identified four 2-phenols as rat liver microsomal metabolites of DMBA and its methyl-hydroxylated metabolites, 7-hydroxymethyl-12-methylbenz[a]anthracene, 7-methyl-12-hydroxymethylbenz[a]-anthracene, and 7,12-dihydroxymethylbenz[a]anthracene. Our findings suggest that neither the 12-methyl group nor the 12-hydroxymethyl group blocks the microsomal oxygenations of the 1,2 positions of DMBA or its methyl-hydroxylated derivatives. The 2-phenols may be formed as nonenzymatic rearrangement products of the 1,2-epoxide intermediates, although their formations by a direct hydroxylation mechanism cannot be ruled out.     

The formation of dihydrodiols by the chemical or enzymic oxidation of 7-hydroxymethyl-12-methylbenz[alpha]anthracene and the possible role of hydroxymethyl dihydrodiols in the metabolic activation of 7,12-dimethylbenz[alpha]anthracene.

The formation of dihydrodiols from 7-hydroxymethyl-12-methylbenz[alpha]anthracene by rat-liver microsomal fractions, by mouse skin in short-term organ culture and by chemical oxidation in an ascorbic acid/ferrous sulphate/EDTA system has been studied using a combination of thin-layer chromatography and high pressure liquie chromatography. The 3,4-, 8,9- and 10,11-dihydrodiols were formed in all three systems. The 5,6-dihydrodiol was formed in rat-liver microsomal fractions and in chemical oxidation but was not detected as a metabolite of [7-3H]hydroxymethyl-12-methylbenz[alpha]anthracene when this compound was incubated with mouse skin in short-term organ culture. The possible role of hydroxymethyl dihydrodiols in the in vivo metabolic activation of 7,12-dimethylbenz[alpha]anthracene in mouse skin has been studied using Sephadex LH-20 column chromatography. The results show that the hydrocarbon-nucleic acid products formed following the treatment of mouse skin in vivo with [7,12-3H]dimethylbenz[alpha]anthracene are not the same as those that are formed following the treatment of mouse skin under the same conditions with either 7-hydroxymethyl-12-methylbenz[alpha]anthracene or 7-methyl-12-hydroxymethylbenz[alpha]anthracene.     

The effect of pretreatment with adrenal-protecting compounds on the metabolism of 7,12-dimethylbenz[a]anthracene and related compounds by rat-liver homogenates

1. 7,12-Dimethylbenz[a]anthracene is converted by rat-liver homogenates into products with the properties of the 7- and 12-hydroxymethyl derivatives, the 7,12-dihydroxymethyl derivative, the related carboxylic acids and ring-hydroxylated products such as the 8,9-dihydro-8,9-dihydroxy derivative and phenols. Ring-hydroxylated products and products arising from the further oxidation of the hydroxymethyl groups were formed when the hydroxymethyl derivatives were themselves incubated with rat-liver homogenates. 2. Pretreatment of the animal with 3-methylcholanthrene or with Sudan III, which can protect rat adrenal glands from damage by 7,12-dimethylbenz[a]anthracene or by its 7-hydroxymethyl derivative, led to an increased rate of metabolism of 7,12-dimethylbenz[a]anthracene and its hydroxymethyl derivatives. The metabolic routes mainly affected were those involving the formation of ring-hydroxylated products. 3. Pretreatment with phenobarbitone led to a small increase in the rate of metabolism of the hydrocarbon and of its hydroxymethyl derivatives, but the increase appeared mainly to involve increased metabolism of the methyl and hydroxymethyl groups. 4. Pretreatment with metyrapone increased the rate of metabolism of the hydrocarbon mainly by increasing the amounts of products resulting from hydroxylation of the methyl groups: small increases in the amounts of ring-hydroxylated products were also produced. 8. Of a number of hydrocarbons and of derivatives of 3-methylcholanthrene tested as enzyme inducers, 3-methylcholanthrene itself was the most effective.     

Stereoselective fungal metabolism of 7,12-dimethylbenz[a]anthracene: identification and enantiomeric resolution of a K-region dihydrodiol

Syncephalastrum racemosum UT-70 and Cunninghamella elegans ATCC 36112 metabolized 7,12-dimethylbenz[a]anthracene (7,12-DMBA) to hydroxymethyl metabolites as well as 7-hydroxymethyl-12-methylbenz[a]anthracene trans-3,4-, -5,6-, -8,9-, and -10,11-dihydrodiols. The 7,12-DMBA metabolites were isolated by reversed-phase high-performance liquid chromatography and identified by their UV-visible absorption, mass, and nuclear magnetic resonance spectral characteristics. A comparison of the circular dichroism spectra of the K-region (5,6-position) dihydrodiol of both fungal strains with those of the 7,12-DMBA 5S,6S-dihydrodiol formed from 7,12-DMBA by rat liver microsomes indicated that the major enantiomer of the 7-hydroxymethyl-12-methylbenz[a]anthracene trans-5,6-dihydrodiol formed by both fungal strains had a 5R,6R absolute stereochemistry. Direct resolution of the fungal trans-5,6-dihydrodiols by chiral stationary-phase high-performance liquid chromatography indicated that the ratios of the R,R and S,S enantiomers were 88:12 and 77:23 for S. racemosum and C. elegans, respectively. These results indicate that the fungal metabolism of 7,12-DMBA at the K region (5,6-position) is highly stereoselective and different from that reported for mammalian enzyme systems.     

Stereoselective fungal metabolism of 7,12-dimethylbenz[a]anthracene: identification and enantiomeric resolution of a K-region dihydrodiol.

Syncephalastrum racemosum UT-70 and Cunninghamella elegans ATCC 36112 metabolized 7,12-dimethylbenz[a]anthracene (7,12-DMBA) to hydroxymethyl metabolites as well as 7-hydroxymethyl-12-methylbenz[a]anthracene trans-3,4-, -5,6-, -8,9-, and -10,11-dihydrodiols. The 7,12-DMBA metabolites were isolated by reversed-phase high-performance liquid chromatography and identified by their UV-visible absorption, mass, and nuclear magnetic resonance spectral characteristics. A comparison of the circular dichroism spectra of the K-region (5,6-position) dihydrodiol of both fungal strains with those of the 7,12-DMBA 5S,6S-dihydrodiol formed from 7,12-DMBA by rat liver microsomes indicated that the major enantiomer of the 7-hydroxymethyl-12-methylbenz[a]anthracene trans-5,6-dihydrodiol formed by both fungal strains had a 5R,6R absolute stereochemistry. Direct resolution of the fungal trans-5,6-dihydrodiols by chiral stationary-phase high-performance liquid chromatography indicated that the ratios of the R,R and S,S enantiomers were 88:12 and 77:23 for S. racemosum and C. elegans, respectively. These results indicate that the fungal metabolism of 7,12-DMBA at the K region (5,6-position) is highly stereoselective and different from that reported for mammalian enzyme systems.     

The effect of the bay-region 12-methyl group on the stereoselective metabolism at the K-region of 7,12-dimethylbenz[a]anthracene by rat liver microsomes.

The enantiomers of a trans-5,6-dihydrodiol formed in the metabolism of 7,12-dimethylbenz[a]anthracene by rat liver microsomes (microsomal fractions) were resolved by chiral stationary-phase high-performance liquid chromatography. The major 7,12-dimethylbenz[a]anthracene trans-5,6-dihydrodiol enantiomer and its hydrogenation product 5,6,8,9,10,11-hexahydro-trans-5,6-diol were found to have 5S,6S absolute configurations by the exciton chirality c.d. method. The R,R/S,S enantiomer ratios of 7,12-dimethylbenz[a]anthracene trans-5,6-dihydrodiol formed in the metabolism of 7,12-dimethylbenz[a]anthracene by liver microsomes from untreated, 3-methylcholanthrene-treated and phenobarbital-treated male Sprague-Dawley rats were found to be 11:89, 6:94, and 5:95 respectively. These findings and those reported previously on the metabolic formations of trans-5,6-dihydrodiols from 7-methylbenz[a]anthracene and 12-methylbenz[a]anthracene suggest that the 12-methyl group in 7,12-dimethylbenz[a]anthracene plays an important role in determining the stereoselective metabolism at the K-region 5,6-double bond. Furthermore, the finding that formation of 5S,6S-dihydrodiol as the predominant enantiomer was not significantly affected by the isoenzymic composition of cytochrome P-450 present in microsomes prepared from the livers of the rats pretreated with the different inducing agents indicates that the stereoselectivity depends on the substrate metabolized rather than on the precise nature of the metabolizing-enzyme system.     

Regio- and stereoselective metabolism of 7,12-dimethylbenz[a]anthracene by Mycobacterium vanbaalenii PYR-1

The degradation of 7,12-dimethylbenz[a]anthracene (DMBA), a carcinogenic polycyclic aromatic hydrocarbon, by cultures of Mycobacterium vanbaalenii PYR-1 was studied. When M. vanbaalenii PYR-1 was grown in the presence of DMBA for 136 h, high-pressure liquid chromatography (HPLC) analysis showed the presence of four ethyl acetate-extractable compounds and unutilized substrate. Characterization of the metabolites by mass and nuclear magnetic resonance spectrometry indicated initial attack at the C-5 and C-6 positions and on the methyl group attached to C-7 of DMBA. The metabolites were identified as cis-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a]anthracene (DMBA cis-5,6-dihydrodiol), trans-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a]anthracene (DMBA trans-5,6-dihydrodiol), and 7-hydroxymethyl-12-methylbenz[a]anthracene, suggesting dioxygenation and monooxygenation reactions. Chiral stationary-phase HPLC analysis of the dihydrodiols showed that DMBA cis-5,6-dihydrodiol had 95% 5S,6R and 5% 5R,6S absolute stereochemistry. On the other hand, the DMBA trans-5,6-dihydrodiol was a 100% 5S,6S enantiomer. A minor photooxidation product, 7,12-epidioxy-7,12-dimethylbenz[a]anthracene, was also formed. The results demonstrate that M. vanbaalenii PYR-1 is highly regio- and stereoselective in the degradation of DMBA.     

The antimutagenic effect of selenium on 7,12-dimethylbenz(a)anthracene and metabolites in the amesSalmonella/microsome system

The antimutagenic effect of selenium as sodium selenite, sodium selenate, selenium dioxide, and seleno-methionine was studied in the AmesSalmonella/microsome mutagenicity test using 7,12-dimethylbenz(a)anthracene (DMBA) and some of its metabolites. Selenium (20 ppm) as sodium selenite reduced the number of histidine revertants on plates containing up to 100 μg DMBA/plate. Increasing concentrations of selenium as sodium selenite, sodium selenate, and selenium dioxide up to 40 ppm Se progressively decreased the number of revertants caused by 50 μg DMBA. DMBA and its metabolites 7-hydroxymethyl-12-methylbenz(a)anthracene, 12-hydroxymethyl-7-methylbenz(a)anthracene, and 3-hydroxy-7,12-dimethylbenz(a)anthracene were mutagenic forSalmonella typhimurium TA100 in the presence of an S-9 mixture. Selenium supplementation as Na₂SeO₃ reduced the number of revertants induced by these metabolites to background levels. The antimutagenic effect of inorganic selenium compounds cannot be explained by toxicity of selenium as determined by viability tests withSalmonella typhimurium TA100. Selenium supplementation in all forms examined, except sodium selenate, decreased the rate of spontaneous reversion. Selenium as sodium selenate was slightly mutagenic at concentrations of 4 ppm or less. Higher concentration of Na₂SeO₄ inhibited the mutagenicity of DMBA. The present studies support the anticarcinogenic potential of selenium and indicate that form and concentration are important factors in this trace element's efficacy.     

Selective oxidation of mitochondrial glutathione in cultured rat adrenal cells and its relation to polycyclic aromatic hydrocarbon-induced cytotoxicity

Primary cultures of rat adrenal cells, as well as rat adrenals in vivo, are sensitive to the potent carcinogen 7,12-dimethylbenz[a]anthracene and its liver metabolite 7-hydroxymethyl-12-methylbenz[a]anthracene, whereas unmethylated polycyclic aromatic hydrocarbons like benzo[a]pyrene or benzo[a]anthracene are ineffective. The adrenocorticolytic potencies of the hydrocarbons are affected by adrenocorticotrophic hormone and various steroids, cytochrome P450 inhibitors, and antioxidants. In the present investigation digitonin was used to fractionate cultured rat adrenal cells. It was found that the mitochondria and cytosol of the cells contained 3-5 nmol/10(6) cells (approximately 15%) and 20-30 nmol/10(6) cells (approximately 85%) of the total soluble cellular glutathione equivalents, respectively. After exposing the cells to 7-hydroxymethyl-12-methylbenz[a]anthracene in the culture medium, a time- and concentration-dependent selective oxidation of mitochondrial glutathione was observed, whereas the effect on the cytosolic glutathione was negligible. Under the same conditions, 7,12-dimethylbenz[a]anthracene and benzo[a]pyrene were unable to alter the redox levels of the subcellular pools of glutathione. Omission of adrenocorticotrophic hormone lowered the oxidation of mitochondrial glutathione induced by 7-hydroxymethyl-12-methylbenz[a]anthracene about twofold. The results suggest that rat adrenal cells contain two separate pools of glutathione, one cytosolic and one mitochondrial, of which the latter is selectively influenced by 7-hydroxymethyl-12-methylbenz[a]anthracene. Moreover, it is concluded that rat adrenal cells offer a unique model system for general studies of the effects of a selective oxidation of mitochondrial glutathione on various cell functions. These effects may constitute early changes in cytotoxicity, preceding, e.g., membrane damage and loss of cytosolic components.     

The metabolism of 7,12-dimethylbenz[a]anthracene and 7-hydroxymethyl-12-methylbenz[a]anthracene by rat liver and adrenal homogenates and by rat adrenocortical cells

The metabolism of 3H-labelled 7,12-dimethylbenz[a]anthracene (DMBA) and of 7-hydroxymethyl-12-methylbenz[a]anthracene (7-OHM-12-MBA) into solvent- and water-soluble and protein-bound derivatives has been examined in rat liver and adrenal homogenates and in rat adrenocortical cells in culture. Although the overall extents of metabolism of the substrates by the two types of homogenate were similar, there was twice as much binding to protein in incubations with the 7-hydroxymethyl derivative. Rat adrenal cells in culture metabolized DMBA more extensively than 7-OHM-12-MBA and converted much more of the parent hydrocarbon into water-soluble derivatives. Both hydrocarbons were metabolized to yield dihydrodiols that were separated and identified by high performance liquid chromatography (HPLC). The 8,9-dihydrodiol was the major dihydrodiol formed from DMBA but, with 7-OHM-12-MBA as substrate, metabolism was diverted to the 10,11- and 3,4-positions in adrenal and hepatic preparations respectively. The viability of rat adrenocortical cells in culture, as measured by trypan blue exclusion, did not appear to be affected by treatment with DMBA, 7-OHM-12-MBA, the sulphate ester of 7-OHM-12-MBA or by 3,4-dihydro-3,4-dihydroxy-7-hydroxymethyl-12-methylbenz[a]anthracene.     

Regio- and Stereoselective Metabolism of 7,12-Dimethylbenz[a]anthracene by Mycobacterium vanbaalenii PYR-1

The degradation of 7,12-dimethylbenz[a]anthracene (DMBA), a carcinogenic polycyclic aromatic hydrocarbon, by cultures of Mycobacterium vanbaalenii PYR-1 was studied. When M. vanbaalenii PYR-1 was grown in the presence of DMBA for 136 h, high-pressure liquid chromatography (HPLC) analysis showed the presence of four ethyl acetate-extractable compounds and unutilized substrate. Characterization of the metabolites by mass and nuclear magnetic resonance spectrometry indicated initial attack at the C-5 and C-6 positions and on the methyl group attached to C-7 of DMBA. The metabolites were identified as cis-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a]anthracene (DMBA cis-5,6-dihydrodiol), trans-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a]anthracene (DMBA trans-5,6-dihydrodiol), and 7-hydroxymethyl-12-methylbenz[a]anthracene, suggesting dioxygenation and monooxygenation reactions. Chiral stationary-phase HPLC analysis of the dihydrodiols showed that DMBA cis-5,6-dihydrodiol had 95% 5S,6R and 5% 5R,6S absolute stereochemistry. On the other hand, the DMBA trans-5,6-dihydrodiol was a 100% 5S,6S enantiomer. A minor photooxidation product, 7,12-epidioxy-7,12-dimethylbenz[a]anthracene, was also formed. The results demonstrate that M. vanbaalenii PYR-1 is highly regio- and stereoselective in the degradation of DMBA.     

The metabolism of a series of polycyclic hydrocarbons by mouse skin maintained in short-term organ culture

Investigations on the metabolism of ³H-labelled chrysene, benz[a]anthracene, 7-methylbenz[a]anthracene, 7,12-dimethylbenz[a]anthracene, 3-methylcholanthrene, benzo[a]pyrene, dibenz[a,c]anthracene and dibenz[a,h]anthracene by mouse skin maintained in short-term organ culture were carried out. Estimations of the distribution of the metabolites of each hydrocarbon present after 24 h showed that there were wide variations both in the rates at which the hydrocarbons were metabolised and in the amounts of metabolites covalently bound to skin macromolecules. All the hydrocarbons were metabolised to dihydrodiols, which were identified by comparison on high pressure liquid chromatography (HPLC) with the authentic compounds, and these were the same diols as those that were formed in previous experiments with rat-liver microsomal fractions. However, free dihydrodiols represented only relatively small proportions of the total amounts of metabolites formed. All the hydrocarbons yielded dihydrodiols of the type that could give rise to bay-region diol-epoxides, when further metabolised, some of which are thought to be involved in hydrocarbon carcinogenesis.     

The formation of dihydrodiols by the chemical or enzymic oxidation of benz[a] anthracene and 7,12-dimethylbenz[a] anthracene.

When benz[a] anthracene was oxidised in a reaction mixture containing ascorbic acid, ferrous sulphate and EDTA, the non-K-region dihydrodiols, trans-1,2-dihydro-1,2-dihydroxybenz[a] anthracene and trans-3,4-dihydro-3,4-dihydroxybenz[a] anthracene together with small amounts of the 8,9- and 10,11-dihydrodiols were formed. When oxidised in a similar system, 7,12-dimethylbenz[a] anthracene yielded the K-region dihydrodiol, trans-5,6-dihydro-5,6-dihydroxy-7,12-dimethylbenz[a] anthracene and the non-K-region dihydrodiols, trans-3,4-dihydro-3,4-dihydroxy-7,12-dimethylbenz[a] anthracene, trans-8,9-dihydro-8,9-dihydroxy-7,12-dimethylbenz[a] anthracene, trans-10,11-dihydro-10,11-dihydroxy-7,12-dimethylbenz[a] anthracene and a trace of the 1,2-dihydrodiol. The structures and sterochemistry of the dihydrodiols were established by comparisons of their UV spectra and chromatographic characteristics using HPLC with those of authentic compounds or, when no authentic compounds were available, by UV, NMR and mass spectral analysis. An examination by HPLC of the dihydrodiols formed in the metabolism, by rat-liver microsomal fractions, of benz[a] anthracene and 7,12-dimethylbenz[a] anthracene was carried out. The metabolic dihydriols were identified by comparisons of their chromatographic and UV or fluorescence spectral characteristics with compounds of known structures. The principle metabolic dihydriols formed from both benz[a] anthracene and 7,12-dimethylbenz[a] anthracene were the trans-5,6- and trans-8,9-dihydrodiols. The 1,2- and 10,11-dihydrodiols were identified as minor products of the metabolism of benz [a] anthracene and the tentative identification of the trans-3,4-dihydriol as a metabolite was made from fluorescence and chromatographic data. The minor metabolic dihydriols formed from 7,12-dimethylbenz[a] anthracene were the trans-3,4-dihydrodiol and the trans-10,11-dihydriol but the trans-1,2-dihydrodiol was not detected in the present study.     

Characterization of a photoproduct of 7,12-dimethylbenz[alpha]anthracene and its effects on chick-embryo cells in culture.

A common impurity of 7,12-dimethylbenz[alpha]anthracene was more effective than 7,12-dimethylbenz[alpha]anthracene in inducing morphological alterations, and in causing an increase in glucose uptake, DNA synthesis and cell number in chick-embryo fibroblasts. Gradual morphological transformation follows the increase in DNA synthesis after 2 days when either primary or secondary cultures are treated with 3 microgram of the compound/ml. The compound, isolated from 7,12-dimethylbenz[alpha]anthracene by alumina column chromatography, was characterized by t.l.c., mass spectroscopy, carbon-hydrogen analysis, u.v. and nuclear-magnetic-resonance spectroscopy and thermal decomposition. It was the photo-oxidation product of 7,12-dimethylbenz[alpha]anthracene, 7,12-epidioxy-7,12-dimethylbenz[alpha]anthracene. It is suggested that some of the biological effects observed after treatment of cultures with 7,12-dimethylbenz[alpha]anthracene may be due in part to the presence of the photo-oxidation product.     

Differential excision of carcinogenic hydrocarbon-DNA adducts in mouse embryo cell cultures.

Primary mouse embryo cell cultures efficiently excise DNA damage introduced by the carcinogens 7-bromomethylbenz[$\text{a}$]anthracene and 3-methylcholanthrene but are inefficient in excision of damage introduced by 7,12-dimethylbenz[a]anthracene. Since exposure of the cells to the latter compound does not impair their capacity for excision of adducts introduced by the bromocompound, it is concluded the 7,12-dimethylbenz[$\text{a}$]anthracene-DNA adducts are intrinsically difficult to excise.     

Hepatic DNA and RNA adduct formation from the carcinogen 7-hydroxymethyl-12-methylbenz[a]anthracene and its electrophilic sulfuric acid ester metabolite in preweanling rats and mice

DNA and RNA adducts that were chromatographically identical to those formed in vitro on reaction of 7-sulfooxymethyl-12-methyl-benz[a]anthracene with guanine and adenine nucleosides were formed in the livers of rats and mice given i.p. injections of 7-hydroxymethyl- or 7-sulfooxymethyl-12-methyl-benz[a]anthracene. Considerably higher levels of these hepatic adducts were obtained from the latter short-lived electrophilic ester than from the hydroxymethyl compound. These observations are consistent with the finding of rat liver cytosolic sulfotransferase activity for 7-hydroxymethyl-12-methylbenz[a]anthracene (Watabe et al., Science 215, 403, 1982). Formation of these hepatic adducts from 7-hydroxymethyl-12-methylbenz[a]anthracene was inhibited by prior administration to rats of dehydroepiandrosterone, an inhibitor of the sulfotransferase activity for this hydroxymethyl hydrocarbon.     

Microbial metabolism and detoxification of 7,12-dimethylbenz[a]anthracene

Summary Six strains of fungi grown on Sabouraud dextrose broth in the presence of 7,12-dimethylbenz[a]anthracene (DMBA) were surveyed for their ability to metabolize DMBA. Experiments with [¹⁴C]DMBA indicated that the extent of formation of organic-soluble metabolites ranged from 6 to 28% after 5 days of incubation, depending on the organism tested. The yields of water-soluble metabolites also varied, and ranged from 1 to 33% after 5 days.Cunninghamella elegans ATCC 36112 andSyncephalastrum racemosum UT-70 exhibited the highest DMBA-metabolizing activity among the organisms surveyed.S. racemosum metabolized DMBA primarily to 7-hydroxymethyl-12-methylbenz[a]anthracene (7-OHM-12-MBA)_ and 7,12-dihydroxymethylbenz[a]anthracene (7,12-diOHMBA). Minor metabolites included 7-OHM-12-MBA-trans-5,6-, 8,9- and 10,11-dihydrodiols, and glucuronide and sulfate conjugates of phenolic derivatives of DMBA. In contrast, the major DMBA metabolites produced byC. elegans were water-soluble. The predominant organic-soluble metabolites produced byC. elegans included 7-OHM-12-MBA-trans-5,6-, 8,9- and 10,11-dihydrodiols. DMBA-trans-3,4-dihydrodiol was also detected. Circular dichroism spectral analysis revealed that the major enantiomer of the 7-OHM-12-MBA-trans-8,9-dihydrodiol formed by each organism has anS,S absolute configuration, while the major enantiomers of the 5,6-, 10,11- and 3,4-dihydrodiols had anR,R configuration. The mutagenic activity of extracts fromS. racemosum exposed to DMBA were determined inSalmonella typhimurium TA98. The mutagenicity of DMBA decreased by 36% over a period of 5 days as 33% of the compound was metabolized. Comparison of these results with previously reported results in mammalian systems suggests that there are similarities and differences between the fungal and mammalian oxidation of DMBA and that the overall balance of fungal metabolism is towards a detoxification rather than a bioactivation pathway.