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An increasing amount of Fluoranthene (Fla) and Benz(a)anthracene (BaA) is being produced and used, eventually entering the soil sediments. The accumulation of Fla and BaA will cause poisoning to typical enzymes (α-Amylase) and organisms (Eisenia fetida) in soil. However, the studies about exploring and comparing the different effects of Fla, BaA and their joint effect at different levels are rarely reported. In this paper, the different effects of Fla, BaA and their mixed pollutant on α-Amylase were evaluated and compared at the molecular level, and the effect of Fla-BaA to the antioxidant system of earthworm (Eisenia fetida) was investigated from the aspects of concentration and exposure time at the animal level. The results showed that Fla-BaA had the greatest influence on the skeleton structure and the microenvironment of amino acid residue of α-Amylase compared to Fla and BaA, and in the mixed pollutant system, the joint effect mode was additive mode. The inhibitory effect of Fla-BaA on the activity of α-Amylase was also stronger than that of the system alone. The assays at the animal level showed that low concentrations (below 5 mg/kg) of Fla-BaA increased the activity of GSH-Px and SOD while high concentrations inhibited their activity. The POD that was activated throughout the experiment period suggested its key role in the earthworm antioxidant system. Changes in T-AOC and MDA showed that long-term and high-dose of Fla-BaA exposure inhibited the antioxidant capacity of Eisenia fetida, causing lipid peroxidation and damage to cells.

Not much is known about the biotransformation capability of zebrafish (Danio rerio) embryos. For understanding possible toxicity differences to adult fish, it might be crucial to understand the biotransformation of chemicals in zebrafish embryos i.e. as part of toxicokinetics. The biotransformation capabilities were analysed for two different stages of zebrafish embryos in conjunction with the internal concentrations of a xenobiotic. Zebrafish embryos of the late cleavage/early blastula period (2–26 hpf) and the early pharyngula period (26–50 hpf) were exposed for 24 h to the AhR binding compound benz[a]anthracene (BaA). Time dependent changes in cyp transcription (cyp1a, cyp1b1, cyp1c1 and cyp1c2) as well as concentration & time-dependent courses of BaA in the fish embryo and the exposure medium were analysed. Additionally, the CYP mediated formation of biotransformation products was investigated. We found correlations between transcriptional responses and the internal concentration for both exposure types. These correlations were depending on the start of the exposure i.e. the age of the exposed embryo. While no significant induction of the examined gene transcripts was observed in the first 12 h of exposure beginning in the blastula period a correlation was apparent when exposure started later i.e. in the pharyngula period. A significant induction of cyp1a was detected already after 1.5 h of BaA exposure. Gene transcripts for cyp1b1, cyp1c1 and cyp1c2 showed expressions distinctly different from cyp1a and were, in general, less inducible by BaA in both exposure windows. The toxicokinetic analysis showed that the biotransformation capability was fivefold higher in the older fish embryos. Biotransformation products of phase I reactions were found between 32 hpf and 50 hpf and were tentatively identified as benz[a]anthracene-phenol and benz[a]anthracene-dihydrodiol-epoxide. In conclusion, not only duration but also onset of exposure in relation to the developmental stage of zebrafish embryos is important in the analysis and interpretation of effects due to different biotransformation capabilities.

Benz(a)anthracene (BaA) is a polycyclic aromatic hydrocarbons (PAHs), that belongs to a group of carcinogenic and mutagenic persistent organic pollutants found in a variety of ecological habitats. In this study, the efficient biodegradation of BaA by a green alga Chlamydomonas reinhardtii (C. reinhardtii) CC-503 was investigated. The results showed that the growth of C. reinhardtii was hardly affected with an initial concentration of 10 mg/L, but was inhibited significantly under higher concentrations of BaA (30 mg/L) (p 0.05). We demonstrated that the relatively high concentration of 10 mg/L BaA was degraded completely in 11 days, which indicated that C. reinhardtii had an efficient degradation system. During the degradation, the intermediate metabolites were determined to be isomeric phenanthrene or anthracene, 2,6-diisopropylnaphthalene, 1,3-diisopropylnaphthalene, 1,7-diisopropylnaphthalene, and cyclohexanol. The enzymes involved in the degradation included the homogentisate 1,2-dioxygenase (HGD), the carboxymethylenebutenolidase, the ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) and the ubiquinol oxidase. The respective genes encoding these proteins were significantly up-regulated ranging from 3.17 fold to 13.03 fold and the activity of enzymes, such as HGD and Rubisco, was significantly induced up to 4.53 and 1.46 fold (p 0.05), during the BaA metabolism. This efficient degradation ability suggests that the green alga C. reinhardtii CC-503 may be a sustainable candidate for PAHs remediation.

Cancer is a cellular tumor that unlike benign tumor can metastasize and invade the surrounding tissues. In the present study the anticancer effect of paclitaxel was evaluated on 7, 12 Di Methyl Benz (a) Anthracene induced skin cancer in wistar rats and results were compared with normal, paclitaxel and paclitaxel-Di allyl sulfide combined alternative chemotherapy. By analyzing the various biochemical parameters (lipid Profile and lipid metabolizing enzymes) and Marker enzymes (squamous cell antigen). Skin cancer was induced in rats by 7, 12 Di methyl benz(a) anthracene (DMBA) at the dosage of 5 µg was dissolved in 100µl and administered into experimental animals for 28 weeks. In this study, we demonstrated that combination of paclitaxel and Di allyl sulfide protects the rats from a lethal dose of DMBA for 30 days. Total cholesterol (TC), free cholesterol (FC), phospholipids (PL) and triglycerides (TG) were found to be significantly increased whereas ester cholesterol (EC) and free fatty acids (FFA) were significantly decreased when compared with cancer bearing group II animals. From the present study, the effect of Paclitaxel- Di allyl sulfide combination proved to be a more significant chemotherapeutic agent against DMBA induced skin cancer in wistar rats compared to that of paclitaxel by analyzing the lipid profile and lipid metabolizing enzymes and marker enzymes.

Effect of the black tea extracted from a mangrove plant species, Ceriops decandra (Griff.) was studied on dimethyl benz[a]anthracene (DMBA)-induced changes in blood hematology and plasma non-enzymatic antioxidants in male hamsters. Hamsters were painted with 0.5% solution of DMBA in liquid paraffin on the right buccal pouch three times in a week up to 14 weeks. Each application treated with 0.4mg of DMBA. The mangrove black tea extract (MBTE) was administrated orally with 5mgkg(-1) twice a day and then with DMBA on alternate days. Results showed that the DMBA caused a significant (P

Daucus carota L. ssp. carota (Apiacea) is widely distributed throughout the world and has many uses in traditional medicine.The present study investigates the chemopreventive effects of oil extract of D. carota umbels on 7,12-dimethyl benz(a)anthracene (DMBA)-induced skin cancer in mice.D. carota oil extract (DCOE) was prepared by extracting the dried umbels with 50:50 acetone:methanol. Skin papilloma were initiated by DMBA and promoted by 12-O-tetradecanoyl phorobol-13-acetate (TPA). The extract was administered to animals via gavage (0.02 mL of 100% oil), intraperitoneal (0.3 mL of 2% oil), and topical (0.2 mL of 5, 50, and 100% oil) routes for 20 weeks. Tumor appearance, incidence, yield, and volume were compared with those of a non-treated control group.Topical 100% treatment delayed tumor appearance, and inhibited tumor incidence and yield by 40 and 89%, respectively. Topical 50% treatment inhibited tumor incidence and yield by 30 and 83%, respectively, whereas the 5% treatment inhibited tumor yield by 36%. Tumor volume was decreased by 99, 91, and 70% following topical treatments with 100, 50, and 5% oil, respectively. Intraperitoneal treatment inhibited tumor yield by 43%, and decreased tumor volume by 85%, whereas gavage treatment showed minimal effects on both. Intraperitoneal and topical treatment decreased infiltration and hyperplasia with an increase in the level of hyperkeratosis.These findings demonstrate that DCOE has remarkable antitumor activity against DMBA-induced skin cancer compared with non-treated animals paving the ground for further investigations.

Benz[a]anthracene (BaA) is a ubiquitous polycyclic aromatic hydrocarbon found in numerous aquatic ecosystems. However, ecotoxicological data in aquatic organisms are scarce. To remedy this lack of data, Japanese medaka (Oryzias latipes) embryos were exposed to BaA and toxic effects were investigated at multiple toxicological endpoints. Japanese medaka embryos were incubated onto BaA-spiked artificial sediment for 9 days at low or moderate environmental concentrations ranging from 0.9 to 12 µgg(-1) dw. BaA-exposed embryos exhibited significant tachycardia. BaA exposure was also shown to increase CYP1A activity in the hepato-biliary tissue as well as craniofacial deformities and DNA damage in pro-larvae. The photomotor response of BaA-exposed larvae was reduced in comparison to the control group. According to this set of tests, the lowest tested and observed effect concentration (LOEC) for Japanese medaka early life stages was equivalent to 0.92 µgg(-1) dw of BaA. This concentration fall into the range of concentrations frequently encountered in sediments of polluted aquatic ecosystems. Taking into consideration these results, BaA represents a threat for fish early life stages in particular those developing onto or into contaminated sediments.

Quantitative structure-activity relationships (QSARs) for benz[a]anthracene (BA)mutagens using 73 descriptors were searched. The mutagenicity data was obtained fromAmes assays for Mycobacterium vanbaalenii, Mycobacterium gilvum and Mycobacteriumflavescens strains. These data were fitted using a mutagenicity-cytotoxicity competitionmodel which defines the mutagenic potencies of BA metabolites, and include oxides,phenols, quinones, and dihydrodiols. The QSAR equations were derived using the moleculardescriptor set (charged partial surface area, spatial, thermodynamic and electronicdescriptors) and semi-empirical energetic and charge descriptors. Genetic functionapproximation was used to reduce and fit independent variables, including linear- andquadratic-based functions. Multiple QSAR equations were generated and a separate QSARequation was chosen and evaluated for each strain using conventional r2, F-test, and cross-validated r2. Each strain exhibited its own characteristic descriptors.

Cultures of the ligninolytic fungus Irpex lacteus incubated in a nutrient liquid medium degraded more than 70% of the initially applied benz[a]anthracene within 14 days. At the first step of metabolization, benz[a]anthracene was transformed via a typical pathway of ligninolytic fungi to benz[a]anthracene-7,12-dione (BaAQ). The product was further transformed by at least two ways, whereas one is complied with the anthracene metabolic pathway of I. lacteus. Benz[a]anthracene-7,12-dione was degraded to 1,2-naphthalenedicarboxylic acid and phthalic acid that was followed with production of 2-hydroxymethyl benzoic acid or monomethyl and dimethylesters of phthalic acid. Another degradation product of BaAQ was identified as 1-tetralone. Its transformation via 1,4-naphthalenedione, 1,4-naphthalenediol and 1,2,3,4-tetrahydro-1-hydroxynaphthalene resulted again in phthalic acid. None of the intermediates were identified as dead-end metabolites. Metabolites produced by ring cleavage of benz[a]anthracene using the ligninolytic fungus are firstly presented in this work.

Tobacco smoke and polluted air are risk factors for lung cancer and contain many kinds of polycyclic aromatic hydrocarbons (PAHs) including benzo[a]pyrene (B[a]P) and benz[a]anthracene (BA). BA, as well as B[a]P, is assessed as probably carcinogenic to humans (IARC group 2A). BA is metabolized to several dihydrodiols. Dihydrodiol dehydrogenase (DD), a member of the aldo-keto reductase superfamily, catalyzes NAD(P)+-linked oxidation of dihydrodiols of aromatic hydrocarbons to corresponding catechols. To clarify the role of DD on PAH carcinogenesis, we examined oxidative DNA damage induced by trans-dihydrodiols of BA and B[a]P treated with DD using 32P-5'-end-labeled DNA fragments obtained from the human p53 tumor suppressor gene. In addition, we investigated the formation of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG), an indicator of oxidative DNA damage, in calf thymus DNA by using HPLC with an electrochemical detector. DD-catalyzed BA-1,2-dihydrodiol caused Cu(II)-mediated DNA damage including 8-oxodG formation in the presence of NAD+. BA-1,2-dihydrodiol induced a Fpg sensitive and piperidine labile G lesion at the 5'-ACG-3' sequence complementary to codon 273 of the human p53 tumor suppressor gene, which is known as a hotspot. DNA damage was inhibited by catalase and bathocuproine, suggesting the involvement of H2O2 and Cu(I). The observation of NADH production by UV-visible spectroscopy suggested that DD catalyzed BA-1,2-dihydrodiol most efficiently to the corresponding catechol among the PAH-dihydrodiols tested. A time-of-flight mass spectroscopic study showed that the catechol form of BA-1,2-dihydrodiol formed after DD treatment. In conclusion, BA-1,2-dihydrodiol can induce DNA damage more efficiently than B[a]P-7,8-dihydrodiol and other BA-dihydrodiols in the presence of DD. The reaction mechanism on oxidative DNA damage may be explained by theoretical calculations with an enthalpy change of dihydrodiols and oxidation potential of their catechol forms. DD may play an important role in BA carcinogenesis via oxidative DNA damage.

In groundwater systems, dissolved natural organic matter (NOM) can influence the mobility of organic contaminants by altering the contaminant behavior in water and solid phases. The transport of anthracene and benz(a)anthracene (B(a)A) was studied in the presence and absence of NOM and/or soil organic matter (SOM) in column experiments. The results show that sorption are related to the properties of polycyclic aromatic hydrocarbons (PAHs), NOM and SOM. In the Fe-quartz media, the amount of NOM (20 mg/l) in solution had a little effect on increasing the apparent solubility of anthracene and countering increased anthracene sorption. In the natural (Bemidji) soil, Suwannee river fulvic acid (SRFA, 20 mg/l) and Suwannee river humic acid (SRHA) in water did not compete with SOM for anthracene, indicating that SOM has higher partition efficiency for anthracene. It was also observed that slow diffusion through an organic phase apparently caused most of the observed tailing in column breakthrough curves (BTCs). Even though the fOC of washed Bemidji sediment was very low, the transport of B(a)A was retarded significantly, however, and the transport of B(a)A was shown to be facilitated by dissolved NOM.

Benz[a]anthracene (BA) is one of the most abundant polycyclic aromatic hydrocarbons (PAHs) that are ubiquitous environmental pollutants. PAH carcinogenesis is explained by DNA adduct formation by PAH diol epoxide and oxidative DNA damage by PAH o-quinone. Benz[a]anthracene-trans-3,4-dihydrodiol (BA-3,4-dihydrodiol) is a minor metabolite but shows higher mutagenicity and tumorigenicity than parent BA. We confirmed that a BA o-quinone type metabolite, benz[a]anthracene-3,4-dione (BA-3,4-dione), induced oxidative DNA damage in the presence of cytochrome P450 reductase. Interestingly, we found that BA-3,4-dihydrodiol nonenzymatically caused Cu(II)-mediated DNA damage including 8-oxo-7,8-dihydro-2'-deoxyguanosine formation and the addition of NADH enhanced DNA damage. BA-3,4-dihydrodiol induced a double-base lesion of C and G at the 5'-ACG-3' sequence complementary to codon 273 of the human p53 tumor suppressor gene, which is known as a hotspot. The DNA damage was inhibited by catalase and bathocuproine, indicating the involvement of H(2)O(2) and Cu(I). Time-of-flight mass spectroscopic study suggested that BA-3,4-dihydrodiol undergoes Cu(II)-mediated autoxidation leading to the formation of its hydroxylated form of BA-3,4-dihydrodiol, capable of causing oxidative DNA damage. It is noteworthy that BA-3,4-dihydrodiol can nonenzymatically induce DNA damage more efficiently than BA-3,4-dione with metabolic activation. In conclusion, oxidative DNA damage induced by BA-3,4-dihydrodiol not only via quinone-type redox cycle but also via a new type of redox cycle participates in the expression of carcinogenicity of BA and BA-3,4-dihydrodiol.

Cyclodextrin-modified electrokinetic chromatography (CD-EKC) was investigated for the separation of 12 monomethylbenz[a]anthracene (MBA) isomers. Combined use of a polymeric surfactant, poly(sodium 10-undecenyl sulfate) (poly-SUS), with various types of neutral cyclodextrins (CDs) [beta-CD, gamma-CD, dimethyl-beta-CD (DM-beta-CD), trimethyl-beta-CD (TM-beta-CD) and hydroxypropyl-beta-CD (HP-beta-CD)] were successful in CD-EKC separation of the MBA isomers. Baseline resolution of 10 of the 12 isomers, except for 9-MBA and 2-MBA, was achieved with gamma-CD at pH 9.75. The beta-CD, gamma-CD, and beta-CD derivatives (DM-beta-CD, TM-beta-CD, HP-beta-CD) were found to have different resolution and selectivity. Additionally, the tR/t0 values of isomers were found to be dependent on the type and concentration of the CD additives. In general, tR/t0 values of MBA isomers decrease with an increase in the concentration of beta-CD derivatives, whereas the reversed was true when the concentrations of native beta-CD and gamma-CD were varied. The combination of 5 mM gamma-CD, 0.5% (w/v) poly-SUS, 35% (v/v) acetonitrile at a pH of 9.75 provided the best selectivity and resolution of the MBA isomers with a separation time of 110 min. However, the use of 30 mM DM-beta-CD under similar EKC conditions resulted in much faster separation (ca. 16 min) of 10 MBA isomers.

Microbiological analysis of soils from a polycyclic aromatic hydrocarbon (PAH)-contaminated site resulted in the enrichment of five microbial communities capable of utilizing pyrene as a sole carbon and energy source. Communities 4 and 5 rapidly degraded a number of different PAH compounds. Three pure cultures were isolated from community 5 using a spray plate method with pyrene as the sole carbon source. The cultures were identified as strains of Burkholderia (Pseudomonas) cepacia on the basis of biochemical and growth tests. The pure cultures (VUN 10 001, VUN 10 002 and VUN 10 003) were capable of degrading fluorene, phenanthrene and pyrene (100 mg l−1) to undetectable levels within 7–10 d in standard serum bottle cultures. Pyrene degradation was observed at concentrations up to 1000 mg l−1. The three isolates were also able to degrade other PAHs including fluoranthene, benz[a]anthracene and dibenz[a,h]anthracene as sole carbon and energy sources. Stimulation of dibenz[a,h]anthracene and benzo[a]pyrene degradation was achieved by the addition of small quantities of phenanthrene to cultures containing these compounds. Substrate utilization tests revealed that these micro-organisms could also grow on n-alkanes, chlorinated- and nitro-aromatic compounds.

To evaluate the chemopreventive potential of quercetin in an experimental skin carcinogenesis mouse model. Skin tumor was induced by topical application of 7, 12-dimethyl Benz (a) anthracene (DMBA) and Croton oil in Swiss albino mouse. Quercetin was orally administered at a concentration of 200 mg/kg and 400 mg/kg body weight daily for 16 weeks in mouse to evaluate chemopreventive potential. Skin cancer was assessed by histopathological analysis. We found that quercetin reduced the tumor size and the cumulative number of papillomas. The mean latent period was significantly increased as compared to carcinogen treated controls. Quercetin significantly decreased the serum levels of glutamate oxalate transaminase, glutamate pyruvate transaminase, alkaline phosphatase and bilirubin. It significantly increased the levels of glutathione, superoxide dismutase and catalase. The elevated level of lipid peroxides in the control group was significantly inhibited by quercetin. Futhermore, DNA damage was significantly decreased in quercetin treated mice as compared to DMBA and croton oil treated mice. The results suggest that quercetin exerts chemopreventive effect on DMBA and croton oil induced skin cancer in mice by increasing antioxidant activities.

The three-dimensional structure of 3-methyl-2H-anthra[1,2-b]pyran-2-one, an anticarcinogenic coumarin related to the carcinogen benz[a]anthracene, has been determined by X-ray diffraction techniques. The molecule, apart from hydrogen atoms in the methyl group, is flat, the maximum deviation from its least squares best plane being 0.13 angstroms. The carbonyl C=O bond length is normal [1.206(1) angstroms] and the bonding throughout the molecule indicates localization of double bonds within the coumarin ring, but some delocalization of electrons in the other rings. Molecules pack in planes parallel to each other, the coumarin ring oxygen atom lying between two aromatic rings of other coumarin molecules. The bulky methyl groups are not involved in such stacking, while the carbonyl groups attract C-H groups in neighboring molecules by way of C-H...O interactions. These are the types of interactions that such coumarins could make if they bound to hydrophobic areas in biological macromolecules.

The principal nucleoside-hydrocarbon adducts present in hydrolysates of DNA that had been isolated either from mouse skin or from hamster embryo cells treated with benz[a]anthracene have been examined by chromatography on Sephadex LH20 and by high pressure liquid chromatography on Spherisorb 5 ODS. The results show that one of the major adducts prepared from the DNA of mouse skin and of hamster embryo cells has chromatographic properties similar to those of an adduct formed when anti-BA-8,9-diol 10,11-oxide (r-8,t-9-dihydroxy-t-10,11-oxy-8,9,10,11-tetrahydrobenz[a]anthracene) reacts with DNA, whilst a second major adduct has chromatographic properties similar to those of an adduct prepared by reacting anti-BA-3,4-diol 1,2-oxide (t-3,r-4-dihydroxy-t-1,2-oxy-1,2,3,4-tetrahydrobenz[a]anthracene) with DNA. On the basis of this and other evidence, it appears that both of these diol-epoxides may contribute to the biological activity of benz[a]anthracene.

The rates of reaction and the products formed when two vicinal diol-epoxides derived from benz(a)anthracene, anti-BA-3,4-diol 1,2-oxide (t-3,r-4-dihydroxy-t-1,2-oxy-1,2,3,4-tetrahydrobenz(a) anthracene)* and anti-BA-8,9-diol 10,11-oxide (r-8,t-9-dihydroxy-t-10,11-oxy-8,9,10,11-tetrahydrobenz(a)anthracene) reacted with DNA were studied in vitro and the results were compared with those obtained in similar experiments using anti-BP-7,8-diol 9,10-oxide (r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene). The reactivities appeared to decrease in the order anti-BP-7,8-diol 9,10-oxide > anti-BA-3,4-diol 1,2-oxide > anti-BA-8,9-diol 10,11-oxide. The diol-epoxides reacted to a similar extent with single- and with double-stranded DNA but reactions with dGMP, at equivalent concentrations, were much slower than with DNA. With the diol-epoxides of benz(a)anthracene, two principal adducts were present in DNA hydrolysates and evidence was obtained, based on pK determinations before and after nitrous acid treatment, consistent with their being N2-guanine derivatives, analogous to the known DNA-reaction products of benzo(a)pyrene 7,8-diol 9,10-oxide.

The fluorescence spectral properties of the principal hydrocarbon-nucleoside adducts present in hydrolysates of DNA that had been isolated from hamster embryo cells or from mouse skin treated with benz(a)anthracene have been examined using photon-counting spectrophotofluorimetry and compared with the spectral properties of the products formed when different diol-epoxides derived from benz(a)anthracene react with nucleic acid in solution. The adducts were separated by chromatography on LH20 Sephadex columns and purified by high pressure liquid chromatography. The spectra obtained are phenanthrene-like and not anthracene-like, a result that is consistent with metabolic activation occurring, through the formation of a vicinal diol-epoxide, in the 8,9,10,11-ring, rather than in the 1,2,3,4-ring of this hydrocarbon.

Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants that require metabolic activation to exert their carcinogenic effects. This study investigated the 3,4-diol-1,2-epoxide formation of benz[a]anthracene (BA) and its toxic effects in a human in vitro hepatocyte culture system. Both mRNA and protein expression of metabolic enzymes which can activate PAHs to carcinogenic forms increased after BA exposure in HepG2 cells and our quantitative analysis showed that the formation of BA-3,4-diol-1,2-epoxide in medium extracts increased in a time-dependent manner. We also performed several comparative studies which show that much lower concentrations of BA-3,4-diol-1,2-epoxide had stronger cytotoxicity and genotoxicity than higher doses of BA. These results suggest that BA is activated as the major carcinogenic metabolite 3,4-diol-1,2-epoxide, in human in vitro culture systems by metabolic enzymes and that this metabolite has stronger cytotoxic and genotoxic effects than its parent compound.

Aqueous benz(a)anthracene [B(a)A] solution was treated with chlorine in the absence and presence of bromide ion. Changes in the composition of the halogenated products in water were determined by high performance liquid chromatographic (HPLC) analyses of benzene extracts. Benz(a)anthracene was shown to produce a variety of halogenated compounds, including halogen-substituted B(a)As (BAXs), halogen additive B(a)As (polar compounds) and oxygenated products, B(a)A-7, 12-dione. Production of these compounds depends on the chlorine dose and the reaction time. It was found that oxidation reaction with chlorine of B(a)A in water are accelerated in the presence of bromide ion. Several components obtained from chlorination in water were mutagenic. Some BAXs were mutagenic in TA 100 strain with S-9 mix. The mutagenic responses of BAXs such as 7-chloroB(a)A (BACl) were weaker than that of original B(a)A. The fraction of polar compounds obtained by means of preparative HPLC showed direct mutagenic activities towards both tester strains.

Chlorinated benz[a]anthracenes (Cl-BaA) are halogenated aromatic compounds (typified by dioxins) found in the environment at relatively high concentrations. Fischer 344 rats were intragastrically administered 0, 1, or 10 mg of Cl-BaA or its parent compound benz[a]anthracene (BaA) per kg of body weight for 14 consecutive days. Both chemicals at 10 mg/kg/day inhibited the gain in body weight, and consequent increase in relative liver weight. Hepatic gene expression of cytochrome P450 (CYP) 1A1, 1A2, and 1B1 was significantly stimulated by administration of BaA (10 mg/kg/day) compared with the control. After administration of Cl-BaA, only the CYP1A2 gene was significantly induced, even at the lower dosage; CYP1A1 and 1B1 mRNA levels remained unchanged in Cl-BaA-treated rats compared with controls. To elucidate the role of such Cl-BaA exposure and induced CYPs at toxicity onset, we investigated the mutagenicity of BaA and Cl-BaA using Salmonella typhimurium TA98 and TA100. BaA and Cl-BaA at 10 μg/plate produced positive results in both strains in the presence of rat S-9. Incubation of Cl-BaA with recombinant rat CYP1A2 produced a significantly higher number of revertant colonies in TA98 and TA100 than in controls, but no such change was observed for BaA. In conclusion, BaA changes its own physiological and toxicological actions by its chlorination; (1) daily exposure to Cl-BaA selectively induces hepatic CYP1A2 in rats and (2) Cl-BaA induces frameshift mutations in the presence of CYP1A2, although BaA does not exert mutagenicity. This indicates that CYP1A2 may metabolize Cl-BaA to active forms.

The present study was aimed to evaluate the antigenotoxic effect of Mosinone-A on 7,12-dimethylbenz[a]anthracene induced genotoxicity. The frequency of micronucleated polychromatic erythrocytes [MnPCEs], chromosomal aberrations [CA], DNA damage (comet assay) as cytogenetic markers and the status of lipid peroxidation byproducts, antioxidants and phase II detoxification agents were used as biochemical markers to assess the antigenotoxic effect of Mosinone-A on DMBA induced genotoxicity. A single intraperitoneal injection of DMBA (30 mg/kg b.wt) to golden Syrian hamsters, resulted in marked elevation in the frequency of MnPCEs, aberrations in the chromosomal structure were found in bone marrow and DNA damage (comet assay) was found in blood cells and altered level of lipid peroxidation, antioxidants, and phase II detoxification agents. Oral pretreatment of Mosinone-A (2 mg/kg b.wt) for 5 days to DMBA treated animals significantly reduced the frequency of MnPCEs, chromosomal abnormalities such as chromosomal break, gap, minute, fragment, DNA damage and reversed the status of biochemical variables. Our results thus demonstrated the antigenotoxic effect of Mosinone-A on DMBA induced genotoxicity in male golden Syrian hamsters.

The chromium(0)-carbene complex benzannulation reaction was shown to be applicable to cyclodec-4-yn-1-one. Significant regioselectivity was realized in this reaction with ortho-substituted benzylchromium complexes. Reactions of a novel resultant fused cyclodecenone-naphthoquinone with several bases have been studied. Products apparently arising from either intramolecular Michael addition of a ketone enolate to the quinone or intramolecular aldol condensation of a quinone-stabilized anion with the ketone have been observed. The latter mode constitutes a route to the title substructure and, in principle, a route to reach certain angucycline antibiotics

An investigation was undertaken on the biodegradation of two kinds of polycyclic aromatic hydrocarbons (PAHs), anthracene (ANT) and benz[a]anthracene (BAA), by fungi isolated from PAH-contaminated mangrove sediments environment in Ma Wan, Hong Kong. ANT (50 mg l−1) and BAA (20 mg l−1), respectively, were added to mineral salt medium initially for screening of PAH-degrading fungi, and finally two fungal species capable of using ANT or BAA as the sole carbon source were isolated and identified as Fusarium solani species. Removal of ANT and BAA reached 40% and 60% of the added amount, respectively, after 40 days of incubation. A total of six metabolites were isolated and characterized by solid phase microextraction (SPME) combined with gas chromatography–mass spectrometry (GC/MS), which indicate that F. solani degraded both ANT and BAA via their respective quinone molecules to generate phthalic acid. Free extracellular laccase was detected during the degradation process without detectable lignin peroxidase (LiP) and manganese-dependent peroxidase (MnP), suggesting that laccase might play an important role in the transformation of PAHs compounds.

A palladium-catalyzed tandem C-H activation/biscyclization reaction of propargylic carbonates with terminal alkynes was determined, which allowed the tetracyclic benz[a]anthracene framework to be constructed with high regioselectivity. A possible mechanism for this tandem C-H activation/biscyclization process was discussed.

International audience; Sphingomonas sp. strain LH128 was isolated from a polycyclic aromatic hydrocarbon (PAH)-contaminated soil using phenanthrene as the sole source of carbon and energy. A dioxygenase complex, phnA1fA2f, encoding the alpha and beta subunit of a terminal dioxygenase responsible for the initial attack on PAHs, was identified and isolated from this strain. PhnA1f showed 98%, 78%, and 78% identity to the alpha subunit of PAH dioxygenase from Novosphingobium aromaticivorans strain F199, Sphingomonas sp. strain CHY-1, and Sphingobium yanoikuyae strain B1, respectively. When overexpressed in Escherichia coli, PhnA1fA2f was able to oxidize low-molecular-weight PAHs, chlorinated biphenyls, dibenzo-p-dioxin, and the high-molecular-weight PAHs benz[a]anthracene, chrysene, and pyrene. The action of PhnA1fA2f on benz[a]anthracene produced two benz[a]anthracene dihydrodiols.

A simple and sensitive method for measuring airborne particle-associated benz[a]anthracene-7,12-quinone (BaAQ) based on two-dimensional high-performance liquid chromatography (HPLC) with fluorescence detection was established. The system involves an ODS column for sample clean up, a 6-port switching valve, an ODS column for trapping the fraction containing BaAQ, an ODS column for sample separation, and a Pt-Rh catalytic column for reduction of BaAQ to a corresponding fluorescent compound. The accuracy of the assay, as applied to airborne particulate sample extracts spiked with known amounts of BaAQ was 94–108%. The detection limit was 97 fmol per injection (signal-to-noise ratio = 3), and the calibration range was from 1 to 100 pmol with excellent proportionality (R2 = 0.9995). BaAQ in airborne particles collected in Osaka, Japan every 3 h was successfully measured using the established analytical method with simple sample preparation steps: ultrasonic extraction in organic solvent and concentration under reduced pressure and/or nitrogen stream.

The present study is designed to investigate the effect of piperine in modifying the carcinogenic process, as well as biochemical alterations at the molecular level during DMBA-induced hamster buccal pouch carcinogenesis by FT-IR spectroscopy. Specific changes were noticed in the FT-IR spectral features of DMBA-induced hamster buccal pouch carcinoma. These alterations include structural changes of proteins and possible increase of its content, an increase in the nuclear-to-cytoplasm ratio, an increase in the relative amount of DNA, an enhancement in the phosphorylation of proteins, a loss of hydrogen bonding of the C-OH groups in the amino acid residues of proteins and diminished lipid peroxidation which were accompanied by a significant reduction in the relative amount of lipids compared to untreated control animals. Administration of piperine significantly increased the levels of lipid peroxidation and decreased the levels of proteins and nucleic acid content that were found to increase in oral cancer bearing animals. In conclusion, the results of the present study suggest that piperine may exert its chemopreventive effect by modulating the biochemical changes at the molecular level during DMBA-induced hamster buccal pouch carcinogenesis which can be detected using FT-IR spectroscopic technique.

Product distributions were determined for reactions between 2'-deoxyguanosine, or its anion, and 7-(bromomethyl)benz[a]anthracene in either acetone/H2O (1:1) or 2,2,2-trifluoroethanol at 50 or 70 degrees C. The exocyclic amino-substituted product, N2-(benz[a]anthracen-7-ylmethyl)-2'-deoxyguanosine, was always the major nucleoside product formed in these reactions, although its yield was higher in reactions involving 2'-deoxyguanosine anion than the neutral nucleoside. Reaction with the anion also led to formation of the 1-substituted 2'-deoxyguanosine and a guanidinoimidazole nucleoside resulting from reaction of 2'-deoxyguanosine at carbon 5. Reactions of 2'-deoxyguanosine anion in 2,2,2-trifluoroethanol are shown to produce significant amounts of the N2-substituted product, which is difficult to prepare by other routes.

Polycyclic aromatic hydrocarbon (PAH)-induced C3H/10-T1/2/CL8 mouse embryo fibroblasts (10T1/2) and mouse hepatoma-derived Hepa 1c1c7 cells (Hepa-1), exhibit comparable total cytochrome P450 levels and total PAH-metabolizing activities but very different distributions of PAH metabolites. Based on anti-P450IA1-IgG inhibition data, P450IA1 contributes essentially all PAH metabolism in Hepa-1 microsomes but is not involved in PAH metabolism by 10T1/2 cells. In addition, the microsomal epoxide hydratase (EHm) in Hepa-1 cells is far less effective in dihydrodiol (diol) formation compared to that in 10T1/2 microsomes [Pottenger, L.H. and Jefcoate, C.R. Carcinogenesis, 11, 321-327 (1990)]. In the present study, the levels of expression of P450IA1 and EHm proteins and the corresponding mRNAs, both prior to and following exposure to benz[a]anthracene (BA) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), have been correlated with microsomal PAH metabolism by each cell type. In 10T1/2 cells, P450IA1 protein (56 kd) and mRNA (2.6 kb) were detectable at extremely low levels in only two of five cell preparations and then only after maximum induction by TCDD and BA. Thus although 10T1/2 cells contain functional Ah receptors, their capacity to induce P450IA1 is highly suppressed, representing at most 2% of the total P450. TCDD (10 nM) was 4-fold more effective than BA (10 microM) in inducing P450IA1 mRNA, while the levels of immunodetectable protein were comparable. An even greater discrepancy between P450IA1 mRNA and protein levels was seen in BA-induced Hepa-1 cells, where a 4-fold increase in mRNA was paralleled by a 20-fold increase in protein. This difference is probably due to the greater effect of BA depletion on mRNA compared to protein levels. In 10T1/2 cells, BA and TCDD were equally effective at increasing expression of an unidentified 1.9 kb mRNA sequence that blotted very weakly with the P450IA1 cDNA probe. The expression of this mRNA was independent from that of P450IA1. A similar band was visible in Hepa-1 cells less than 1% of the P450IA1 mRNA. EHm mRNA was almost 3-fold higher in 10T1/2 compared to Hepa-1 cells and was unaffected by cell treatments. In Hepa-1 cells, BA and TCDD elevated EHm protein and hydrating activity to levels comparable to those expressed in 10T1/2 cells. It is, therefore, suggested that the relative ineffectiveness of Hepa-1, compared to 10T1/2 EHm, to hydrate low levels of PAH-epoxides is due to differences between the two proteins or their disposition in the microsomal membrane.

We investigated the effects of postnatal exposure to benz[a]anthracene (B[a]A) and benzo[k]fluoranthene (B[k]F) on the development of the rat uterus. Neonates were injected on each postnatal days 1-14 with B[a]A (0.1, 1.0 or 10.0mg/kg), B[k]F (0.1, 1.0 or 5.0mg/kg), ethynylestradiol (EE; 1.0μg/kg) or a vehicle. The rats were killed on day 23. All doses of B[a]A and B[k]F induced a reduction of the uterine weight, a reduction of the estrogen receptor α expression in the luminal epithelium, glandular epithelium and stromal cells. Moreover, the uterotrophic response to EE (3-day administration of 1.0μg/kg on postnatal days 20-22) in rats exposed to B[a]A, B[k]F and EE was lower than in controls. The study showed that postnatal exposure to B[a]A and B[k]F resultes in morphological and functional disorders of the immature rat uterus.

Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants. Some compounds belonging to this group are considered carcinogenic to people. In order to yield carcinogenic properties, these compounds must be metabolically transformed by enzymes of cytochrome P450 family to oxy-derivatives. In this study, the ability of the following six PAHs: anthracene (Ant), benz(a)anthracene (BA), naphthacene (Nap), benzo(a)pyrene (BaP), dibenz(a,c)anthracene (DB(a,c)A) and dibenz(a,h)anthracene (DB(a,h)A) to induce enzymes of cytochrome P450 (CYP450), in particular CYP1A1 and CYP1A2 in Mcf7 and HepG2 cells was studied. The induction of CYP1A enzymes was assessed at the level of enzymatic protein and enzymatic activity. The change in CYP1A1 and CYP1A2 protein level was assessed by means of confocal microscopy. The ethoxyresorufin-O-deethylase (EROD) and methoxyresorufin-O-deethylase (MROD) assays were applied to determine the CYP1A1 and CYP1A2 activity. The Induction Equivalency Factors (IEFs) were also determined. According to EROD and MROD assay and calculated IEFs the following order of the inducing potency was determined in HepG2 cells: DB(a,h)A > BaP > DB(a,c)A ≈ BA > Nap > Ant, and in Mcf7 cells: DB(a,h)A > DB(a,c)A > BaP > Nap > BA > Ant. The assessment of the protein levels revealed that DB(a,h)A was also the strongest inducer of protein level, however the correlation between enzymatic activity and protein level induction by other PAHs was not always evident. The EROD and MROD activities were higher in Mcf7 than in HepG2 cells, however the CYP1A2 protein level was shown to be higher in HepG2 cells. The results obtained indicate possible catalytic enzymatic activity alterations induced by PAHs.

In order to mitigate the strong microbial resistance of benz(a)anthracene [B(a)A] in soil, a hybrid treatment of Fenton oxidation followed microbial culture was carried out. Based on optimal Fenton oxidation, i.e., 1.0 ml of ethanol, 0.2 ml of 0.5 M Fe2+, and 0.3 ml of 30% H2O2 per 1 g of 500 mg B(a)A/kg soil, about 43% of B(a)A-7,12-dione was generated during oxidation of 97% B(a)A. When the comparative biodegradability between B(a)A-contaminated soil and B(a)A-contaminated soil after Fenton oxidation was examined, it was found that 98% of B(a)A-7,12-dione degraded after 63 d in comparison with only 12% of B(a)A over the same period; results demonstrating that Fenton oxidation enhances biodegradability of B(a)A through B(a)A-7,12-dione.

The cytochrome P450 isoforms responsible for the regio-selective metabolism of benz[a]anthracene (BA) are poorly defined but as with other polycyclic aromatic hydrocarbons (PAHs) may include members of the CYP2C sub-family. Since the expression of some of these is regulated in a gender-specific manner and may be altered by age, rat strain or by phenobarbital treatment, the effects of these variables on metabolism of BA to diols was investigated. These studies used hepatic, microsomal membranes from immature and adult Long-Evans rats and adult Hooded Lister rats. BA-diols were resolved by normal phase HPLC into three discrete peaks identified as benz[a]anthracene-5,6-diol (BA-5,6-diol), benz[a]anthracene-10, 11-diol (BA-10,11-diol) and a mixture of benz[a]anthracene-3,4- and -8,9-diols (BA-3,4-diol and BA-8,9-diol and termed Peak(3/8)). Significant gender-related differences were found in the rates of diol formation in adults of both the Long-Evans and Hooded Lister rat strains. Formation of BA-10,11-diol and to a lesser extent the components of Peak(3/8) were greater in the male compared to female animals by factors of at least 14 and two, respectively. An age-dependent effect is also observed in the Long-Evans rat since these differences are still apparent in prepubertal animals but to a lesser extent (gender ratio male:female BA-10,11-diol 9X; Peak(3/8) 1.4X). In contrast BA-5,6-diol was formed at similar rates by membranes from female and male rats whether mature (Long-Evans and Hooded Lister) or immature (Long-Evans). Phenobarbital treatment of the adult Long-Evans rats resulted in a moderate increase in the formation of each diol other than at the 10,11-position and the induction was not gender specific. The rate of formation of BA-10, 11-diol was decreased in phenobarbital-treated male rats suggesting modulation of a male specific isoform. Measurement of microsomal epoxide hydrolase revealed no gender or age differences and suggests that this enzyme is not rate limiting in BA-diol formation and thus is not responsible for the differences in BA-diol formation observed. The results suggest that CYP2C11 along with a male-specific isoenzyme not regulated by age are important in the formation of BA-10,11-diol and a component(s) of Peak(3/8) in males. CYPs 2B2 and/or 2C6 appear to be involved in formation of BA-5,6-diol in male and female. Identification of the CYPs involved in the regio-selective metabolism of BA may lead to an explanation of the lower carcinogenic potency of this PAH compared to dimethylbenz[a]anthracene and this study provides novel clues concerning the identities of the CYPs, which are important.

Reactive oxygen species and free radicals have been implicated in the multistep cutaneous chemical carcinogenesis. Much of the experimental evidence in this regard is indirect and is based on observations that prooxidant status usually enhances and antioxidant treatments generally inhibit tumor yield. Iron overload is known to enhance peroxidative damage and cause oxidative stress. In this study, we report that iron overload augments 12-O-tetradecanoylphorbol-13-acetate (TPA)-mediated cutaneous tumor promotion. Female Swiss mice were subjected to iron overload by injecting 1 mg iron/mouse/day consecutively for 2 weeks. Tumors were initiated by applying a single dose of 7,12-dimethylbenz(a)anthracene and promoted with twice weekly applications of TPA for 20 weeks and the appearance of first tumor (latency period), percent incidence and number of tumors per mouse were recorded. It has been observed that the level of iron in involved (tumor-bearing) skin was about fourfold higher as compared to uninvolved (non-tumor) skin of iron overload animals and about tenfold higher as compared to the iron level in the skin of normal animals. When compared to the iron-unloaded control group, the iron overload mice showed an increased incidence of tumors. In iron overload animals, the tumors appeared 3 weeks earlier and also the number of tumors per mouse was significantly higher (2.5-fold). These data indicate that iron overload augments TPA-mediated tumor promotion. We propose that oxidative stress generated by iron overload may be responsible for the augmentation of cutaneous tumorigenesis.

Reactions of the benzo[a]pyrene (BP) and benz[a]anthracene (BA) metabolites, (+/-)-trans-7 8-dihydroxy-anti-9, 10-epoxy-7, 8, 9, 10-tetrahydro-BP (BPDE), (+/-)-trans-3, 4-dihydroxy-anti- 1,2-epoxy-1,2,3,4-tetrahydro-BA (BADE), (+/-)-BP-4,5-oxide (BPO), and (+/-)-BA-5, 6-oxide (BAO), were examined under pseudo-first-order conditions at varying Na+ (2.0-100 Mm) and native calf thymus DNA (ctDNA) concentrations. In 0.2 mM ctDNA and 2.0 mM Na+, at a pH of 7.3 most BPDE, BADE, BPO, and BAO (87-95%) undergo DNA catalyzed hydrolysis or rearrangement. For BPDE and BPO, overall, pseudo-first-order rate constants, k, in 2.0 mM Na+ and 0.2 mM ctDNA are 21-72 times larger than values obtained without DNA. For BADE and BAO, the rate constants are less strongly influenced by DNA; k values in 0.2 mM ctDNA are only 9-12 times larger than values obtained without DNA. Kinetic data for BPDE, BPO, BADE, and BAO and DNA intercalation association constants (KA) for BP and BA diols which are model compounds indicate that KA values for BPDE and BPO in 2.0 mM Na+ are 6.6-59 times larger than those of BADE and BAO. The greater DNA enhancement of rate constants for BPDE and BPO, versus BADE and BAO, correlates with the larger KA values of the BP metabolites. DNA adducts, which account for less than 10% of the yields, also form. For BPDE in 0.20 mM ctDNA, k decreases 5.1 times as the Na+ concentration increases from 2.0 to 100 mM. Nevertheless, the DNA adduct level remains constant over the range of Na+ concentrations examined. These results provide evidence that, for BPDE in 0.20 mM DNA and 2.0 mM Na+, ctDNA adduct formation follows a mechanism which is similar to that for DNA catalyzed hydrolysis. The pseudo-first-order rate constant for adduct formation, kAd, given approximately by kAd approximately equal to (kcat,AdKA[DNA])/(1 + KA[DNA]), where kcat,Ad is a catalytic rate constant. for BADE, BPO, and BAO, the influence of varying DNA and Na+ concentrations on k values is similar to that for BPDE, and provides evidence that the formation of adducts follows the same rate law.

Improved technologies for the detection of polycyclic aromatic hydrocarbon adducts are required for human biomonitoring. Therefore, a coupled-column high-performance liquid chromatographic method, with system-integrated sample processing, has been developed and its applicability for determination of tetrahydrotetrols of polycyclic aromatic hydrocarbons in acid hydrolysates of human hemoglobin has been investigated. A novel column-switching technique applying ‘thermotransfer’ is used to separate tetrahydrotetrols of benzo[ a ]pyrene and benz[a]anthracene more efficiently. Derivatives of polycyclic aromatic hydrocarbons from blood hydrolysates are concentrated on a pre-column and then transferred to the analytical column by applying an electrical current to heat the solvent eluting the pre-column. This method allows for rapid and quantitative transfer of the analytes from the pre-column to the analytical column, after HPLC-integrated sample processing.

The metabolism of polycyclic aromatic hydrocarbons by bone marrow, mononuclear cells from normal donors and leukaemia patients in remission has been investigated. When benz[alpha]anthracene (BA) was included with marrow under cell culture conditions, it was converted to materials which were resolved into three peaks by normal phase HPLC, and which had the chromatographic characteristics of BA-dihydrodiols. Formation of hydroxymethyl-or dihydrodiol-derivatives of 7, 12-dimethylbenz[alpha]anthracene were not detected under the same conditions. The BA-metabolites were identified as BA-5,6-dihydrodiol, BA-10,11-dihydrodiol and BA-8,9-dihydrodiol. The identification was based upon chromatographic properties of the metabolites during normal and reverse phase chromatography and on UV spectral and fluorometric characterization. It was not possible to detect the formation of BA-3,4-dihydrodiol since this dihydrodiol co-elutes with BA-8,9-dihydrodiol and BA-10,11-dihydrodiol during normal phase and reverse phase chromatography, respectively. the UV spectra of BA-3,4-dihydrodiol does not have features which enable it to be readily identified in the presence of these other compounds. Formation of the dihydrodiol-metabolites was dependent on cell number and temperature. Two general cytochrome P450 inhibitors, carbon monoxide and piperonyl butoxide, blocked the formation of metabolites but the cyclooxygenase inhibitor, indomethacin had no effect. Large variations were observed in the capacity of marrow from different individuals to form benz[alpha]anthracene-dihydrodiols but, in each sample where dihydrodiols were formed, the relative amount of each metabolite was BA-8,9-dihydrodiol >> BA-5,6-dihydrodiol > BA-10,11-dihydrodiol. Factors which may contribute to this variation, including disease status, genetic and environmental agents, are considered.

Syntheses of benz[e]aceanthrylene, 8-methylbenz[e]aceanthrylene, and their 1,2-dihydro derivatives, as well as 6-methylbenz[j]aceanthrylene and its 1,2-dihydro derivative from benz[a]-anthracene-7,12-dione are described. Compounds containing a methyl group in nonbenzo bay region position, are predicted to be relatively potent carcinogens

Rat liver dihydrodiol dehydrogenase (DDH, EC 1.3.1.20) has been shown to reduce the mutagenicity of benz[ a ]anthracene (BA) in the bacterial Ames test. BA-3,4-dihydrodiol is a highly mutagenic and tumorigenic metabolite of BA. In order to test the hypothesis that this dihydrodiol may be a substrate of DDH, we established two novel assay systems for the NADP + -dependent oxidation of BA-3,4-dihydrodiol by rat liver DDH, an HPLC-based assay procedure and a radiometric assay with specifically labelled [3,4- 3 H]-BA-3,4-dihydrodiol as substrate. With the HPLC-based assay, the kinetic constants of the enzymatic catalysis were as follows: K m (app) = 21 μ M for BA-3,4-dihydrodiol and V max = 20.0 nmol/min · mg enzyme. The reaction product was identified by cochromatography, fluorimetry and mass spectroscopy as BA-3,4-catechol, but interconversions between the catechol and the corresponding o -quinone during the analytical procedures were detected. With the radiolabelled substrate, a linear relationship between substrate concentration and reaction velocity was found. The V/K value for labelled substrate was 0.155 ml/ min · mg enzyme and a ( V/K ) H /( V/K ) T kinetic isotope effect of 6.7 was observed. The non-labelled substrate acted as a competitive inhibitor of the enzymatic oxidation of tritiated BA-3,4-dihydrodiol with a K i value of 56.4 μM. The reaction rates determined in this study suggest an important role of DDH activity in the metabolism of BA.

Polycyclic aromatic hydrocarbons (PAHs) with a methylene-bridge spanning the bay region are often detected in higher concentrations in the environment than PAHs which have a bay-region methyl substituent. The tumor-initiating activity of methylene-bridged bay-region derivatives of chrysene and benz[a]anthracene was evaluated on the skin of female CD-1 mice. 4,5-Methylenechrysene elicited twice as many tumors as chrysene at each of the doses tested. At total initiation doses of 1.5, 0.5 and 0.15 mumol/mouse 4,5-methylenechrysene gave rise to 8.5, 6.8 and 1.1 tumors/animal respectively. There was a 100% incidence of tumor-bearing mice at the two higher doses and a 65% incidence at the lowest dose. This methylene-bridged hydrocarbon was less tumorigenic than 5-methylchrysene at each dose tested. Animals treated with 1,12-methylenebenz[a]anthracene developed 7.6, 4.9 and 1.6 tumors/animal at total initiating doses of 4.0, 2.0 and 0.5 mumol/mouse. The incidence of tumor-bearing mice was 90% at the higher doses and 75% at the lowest dose tested. 12-Methylbenz[a]anthracene was more tumorigenic than the methylene-bridged derivative at each dose. The keto-derivative of 1,12-methylenebenz[a]anthracene did not display significant (P greater than 0.05) tumor-initiating activity at these dose levels. Results from these bioassays indicate that methylene-bridged bay-region derivatives of chrysene and benz[a]anthracene contribute to the overall genotoxicity of environmentally occurring PAHs.