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The potential of polycyclic aromatic hydrocarbons (PAHs) to modulate microsomal epoxide hydrolase activity, determined using benzo[a]pyrene 5-oxide as substrate, in human liver, was evaluated and compared to rat liver. Precision-cut liver slices prepared from fresh human liver were incubated with six structurally diverse PAHs, at a range of concentrations, for 24 h. Of the six PAHs studied, benzo[a]pyrene, dibenzo[a,h]anthracene and fluoranthene gave rise to a statistically significant increase in epoxide hydrolase activity, which was accompanied by a concomitant increase in epoxide hydrolase protein levels determined by immunoblotting. The other PAHs studied, namely dibenzo[a,l]pyrene, benzo[b]fluoranthene and 1-methylphenanthrene, influenced neither activity nor enzyme protein levels. When rat slices were incubated under identical conditions, only benzo[a]pyrene and dibenzo[a,h]anthracene elevated epoxide hydrolase activity, which was, once again accompanied by a rise in protein levels. At the mRNA level, however, all six PAHs caused an increase, albeit to different extent. In rat, epoxide hydroxylase activity in lung slices was much lower than in liver slices. In lung slices, epoxide hydrolase activity was elevated following exposure to benzo[a]pyrene and dibenzo[a,l]pyrene and, to a lesser extent, 1-methylphenanthrene; similar observations were made at the protein level. At both activity and protein levels extent of induction was far more pronounced in the lung compared with the liver. It is concluded that epoxide hydrolase activity is an inducible enzyme by PAHs, in both human and rat liver, but induction potential by individual PAHs varies enormously, depending on the nature of the compound involved. Marked tissue differences in the nature of PAHs stimulating activity in rat lung and liver were noted. Although in the rat basal lung epoxide hydrolase activity is much lower than liver, it is more markedly inducible by PAHs.

The mutagenicity of the commonly used glutathione S-transferase substrates 1-chloro-2,4-dinitrobenzene (CDNB) and 1,2-dichloro-4-nitrobenzene (DCNB) was investigated in the Salmonella mutagenicity assay. CDNB induced a concentration-dependent mutagenic response in Salmonella typhimurium strain TA98. Incorporation of an activation system derived from Aroclor 1254-induced rats did not influence mutagenic response. Under the same conditions DCNB failed to display mutagenic activity. The mutagenic activity of CDNB was attenuated in bacterial strains under-expressing nitroreductase or O-acetylase activity but, in contrast, it was exaggerated in an O-acetylase over-expressing strain. It is inferred that CDNB exhibits a mutagenic response following reduction of the nitro-group to the hydroxylamine, which is further acetylated to form the acetoxy derivative that presumably breaks down spontaneously to generate the nitrenium ion, the likely ultimate mutagen.

The polyphenolic dimers, epicatechin-4β-8-catechin (B1), epicatechin-4β-8-epicatechin (B2), catechin-4β-8-catechin (B3), catechin-4β-8-epicatechin (B4), and the gallate ester epicatechin-4β-8-epicatechin gallate (B′2G) were isolated from grape seeds, and theaflavins and theafulvins from black tea brews. The ability of these naturally-occurring polyphenols to afford protection against the genotoxicity of the heterocyclic amine 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) was compared with that of the monomeric tea flavanols, (+)-catechin (C), (−)-epicatechin (EC), (−)-epicatechin gallate (ECG), (−)-epigallocatechin (EGC) and (−)-epigallocatechin gallate (EGCG). Genotoxic activity was evaluated in human peripheral lymphocytes using the Comet assay. At the concentration range of 1–100 μM, neither the monomeric nor the dimeric flavanols prevented the lymphocyte DNA damage induced by Trp-P-2. In contrast, both of the black tea polyphenols, theafulvins and theaflavins, at a dose range of 0.1–0.5 mg/ml, prevented, in a concentration-dependent manner, the DNA damage elicited by Trp-P-2. Finally, neither the monomeric and dimeric polyphenols (100 μM) nor the theafulvins and theaflavins (0.5 mg/ml) caused any DNA damage in the human lymphocytes. These studies illustrate that black tea theafulvins and theaflavins, if absorbed intact, may contribute to the anticarcinogenic potential associated with black tea intake., The authors wish to thank the Department of Science and Technology, Government of India for a BOYSCAST Fellowship (No. HRU/BYS/L-11/99) to one of us (AD). We also acknowledge gratefully financial support for this study from the European Union under Frameworks 3 (project PL0653) and 4 (POLYBIND project).

The relative mutagenic potentials of 11-amino-16,17-dihydro-15 H -cyclopenta[a]phenanthrene, its 17-keto derivative, and 2- and 5-aminochrysene have been compared in Salmonella typhimurium TA98 and TA100 in the presence of a postmitochondrial liver preparation from Aroclor 1254 induced rats. The 11-amino hydrocarbon is a very weak mutagen (0.27 revertants/nmol), whereas the 11-amino-17-ketone is much more active (129 revertants/nmol). 2-Aminochrysene is the most mutagenic arylamine (∼500 revertants/nmol) among these compounds, but its 5-amino isomer is much less active (0.9 revertants/nmol). Possible reasons for these marked differences are suggested. Use of TA98 with over-expressing O -acetyltransferase (YG 1024) and deficient in this enzyme (TA98/l,8-DNP 6 ) with the 11-amino-17-ketone and with 5-aminochrysene clearly indicates the importance of this enzyme in their bioactivation, implying oxidation of the amino group to the hydroxylamine in both these compounds.

The title compound is a more potent carcinogen than would be anticipated from its simple phenanthrene structure lacking further D-ring conjugation. In vitro it undergoes microsomal metabolism to yield as major metabolites its 15- and 17-alcohols and its 16,17-diol; other minor metabolites are also derived from attack at the 5-membered ring, but no evidence of aromatic oxidation is apparent. The title compound is a weak mutagen in the Ames' test with Salmonella typhimurium TA100, but only with microsomal bio-activation. The 17-ol and 16,17-diol are inactive, with or without biological activation. By contrast the 15-alcohol, a rather reactive compound, is a strong mutagen both in the presence and absence of the bio-activation system. This, therefore, may be the proximate carcinogen, and its structural analogy to the naturally occurring hepato-carcinogen safrole is noted.

The molecular dimensions and electronic structures of 100 chemicals of structural diversity have been determined from molecular orbital calculations and molecular mechanics. From these parameters of molecular structure, those chemicals that are likely substrates of cytochromes P4501 and P4502E have been identified by the computer-optimized molecular parametric analysis of chemical toxicity (COMPACT) programme, and their potential toxicity, mutagenicity and carcinogenicity evaluated. The degree of correlation between COMPACT prediction of toxicity and rodent two species life-span carcinogenicity data is estimated to be 92%, and between COMPACT and Salmonella mutagenicity (Ames test) data is 64%. Anomalous rodent carcinogens are rationalized on the basis of biochemical mechanisms of metabolism, genotoxicity and carcinogenicity. Correlation of the Ames test data with rodent carcinogenicity data was 64%, but correlation of COMPACT plus Ames data versus rodent carcinogenicity data provided the highest correlation of 94%.

The metabolic activation of 2-aminoanthracene to mutagens in the Ames test was investigated using hepatic S9, microsomal and cytosolic fractions from control and Aroclor 1254-treated rats as activation systems. Microsomal and S9 preparations from control animals could activate 2-aminoanthracene, but the efficiency of activation was suppressed by pretreatment of animals with Aroclor 1254. Cytosolic fractions from Aroclor 1254-treated rats could readily activate the promutagen more readily than microsomes. The cytosolic activation of 2-aminoanthracene required NADPH and could not be accounted for by possible microsomal contamination. The molybdenum oxygenases appear not to contribute to the cytosolic activation of this promutagen. It is concluded that (a) the microsomal activation of 2-aminoanthracene is catalysed more effectively by enzyme systems other than the P450 I family and (b) an enzyme system capable of activating this carcinogen in vitro is present in the hepatic cytosol. The implications of these findings in the use of 2-aminoanthracene as a positive control in the Ames test are discussed.

Hepatic microsomal mixed-function oxidase activities were determined in rats pretreated with the aromatic amines 2-aminoanthracene, 2-naphthylamine or 4-aminobiphenyl. All three amines stimulated the O-deethylation of ethoxyresorufin (cytochromes P-448) but none had any effect on the p-hydroxylation of analine. 2-Aminoanthracene and 4-aminobiphenyl also stimulated the NADPH-dependent reduction of cytochrome c and 2-naphthylamine inhibited the N-demethylation of benzphetamine. Hepatic preparations from animals pretreated with 2-aminoanthracene were more efficient in converting this carcinogen to mutagens while in contrast pretreatment with Aroclor 1254 caused a marked decrease in mutagenicity. 4-Aminobiphenyl also enhanced its own activation but Aroclor-pretreated preparations were the most effective. The latter preparations were also more efficient than controls in activating 2-naphthylamine to mutagens. It is concluded that 4-aminobiphenyl and 2-aminoanthracene enhance their own activation at least partly, by inducing the synthesis of cytochromes P-448.