Evaluation of the generation of genotoxic and cytotoxic metabolites of benzo[a]pyrene, aflatoxin B1, naphthalene and tamoxifen using human liver microsomes and human lymphocytes.

1. The ability of model stable epoxides and metabolites generated by human liver microsomes from benzo[a]pyrene, aflatoxin B1, naphthalene and tamoxifen to produce cytotoxicity and genotoxicity in human peripheral lymphocytes has been investigated. 2. The stable epoxides 1,1,1 trichloropropene-2,3-oxide (100 microM) and trans stilbene oxide (100 microM) as well as metabolites generated from aflatoxin B1 (30 microM) and naphthalene (100 microM) by an extracellular metabolising system were toxic to isolated resting mononuclear leucocytes (MNLs), whereas glycidol (100 microM), benzo[a]pyrene (100 microM) and tamoxifen (50 microM) were not. 3. The stable epoxides 1,1,1 trichloropropene-2,3-oxide (100 microM) and trans stilbene oxide (100 microM) but not glycidol (100 microM) were toxic to dividing lymphocytes only after a 72-h exposure. Tamoxifen (30 microM), aflatoxin B1 (30 microM) and their metabolites were also toxic to dividing lymphocytes. Benzo[a]pyrene (100 microM) and naphthalene (100 microM) were not toxic either in the absence or presence of the extracellular metabolising system. 4. Benzo[a]pyrene (100 microM) and aflatoxin B1 (30 microM) were directly genotoxic to lymphocytes, this genotoxicity was significantly enhanced by the presence of the extracellular metabolising system. This indicates that both intracellular and extracellular bioactivation of these two compounds can produce genotoxicity. In contrast, naphthalene and tamoxifen were non-genotoxic.

Characterization of benzo[a]pyrene metabolites formed by 3-methylcholanthrene-induced goldfish, black bullhead and brown bullhead.

Metabolites of BaP formed by 3-MC-induced goldfish (Carassius auratus), black bullhead (Ictalurus melas), brown bullhead (Ictalurus nebulus) and rat have been separated by HPLC. The main metabolites in both bullhead species and the rat were 3-hydroxy-BaP and BaP 9,10- and 7,8-dihydrodiols, whereas BaP 1,6- and 3,6-diones and 9- and 3-hydroxy-BaP predominated in the goldfish. Induced rat formed over 10 times as many total metabolites as the fish. No induction was observed in the 3-MC-treated brown bullheads kept at 7 degrees C. TCPO and alpha-naphthoflavone decreased metabolite production by 50 and 30%, respectively.

Fluorescence study of DNA alkylation by epoxides.

A simple fluorescence assay was devised to measure alkylation of guanine. The assay was tested with simple epoxides: propylene oxide, glycidol, epichlorohydrin, trichloropropylene oxide and styrene oxide, which are known to vary considerably in their mutagenic potency. The order of reactivity parallelled the mutagenic potency, trichloropropylene oxide being the most reactive alkylating agent. Each epoxide alkylated deoxyguanosine faster than single-stranded DNA, at equal concentrations of guanine. Single-stranded DNA was alkylated substantially faster than was double-stranded DNA. The reaction products with each substrate were analysed by thin-layer chromatography and exhibited similar Rf-values. It was concluded that polymers, particularly double-stranded DNA, reacted slower than deoxyguanosine due to the properties of polymers in solution rather than the unavailability of reactive sites for alkylation.