Genotoxicity of intermittent co-exposure to benzene and toluene in male CD-1 mice

Benzene is an important industrial chemical. At certain levels, benzene has been found to produce aplastic anemia, pancytopenia, myeloblastic anemia and genotoxic effects in humans. Metabolism by cytochrome P450 monooxygenases and myeloperoxidase to hydroquinone, phenol, and other metabolites contributes to benzene toxicity. Other xenobiotic substrates for cytochrome P450 can alter benzene metabolism. At high concentrations, toluene has been shown to inhibit benzene metabolism and benzene-induced toxicities. The present study investigated the genotoxicity of exposure to benzene and toluene at lower and intermittent co-exposures. Mice were exposed via whole-body inhalation for 6h/day for 8 days (over a 15-day time period) to air, 50ppm benzene, 100ppm toluene, 50ppm benzene and 50ppm toluene, or 50ppm benzene and 100ppm toluene. Mice exposed to 50ppm benzene exhibited an increased frequency (2.4-fold) of micronucleated polychromatic erythrocytes (PCE) and increased levels of urinary metabolites (t,t-muconic acid, hydroquinone, and s-phenylmercapturic acid) vs. air-exposed controls. Benzene co-exposure with 100ppm toluene resulted in similar urinary metabolite levels but a 3.7-fold increase in frequency of micronucleated PCE. Benzene co-exposure with 50ppm toluene resulted in a similar elevation of micronuclei frequency as with 100ppm toluene which did not differ significantly from 50ppm benzene exposure alone. Both co-exposures – 50ppm benzene with 50 or 100ppm toluene – resulted in significantly elevated CYP2E1 activities that did not occur following benzene or toluene exposure alone. Whole blood glutathione (GSH) levels were similarly decreased following exposure to 50ppm benzene and/or 100ppm toluene, while co-exposure to 50ppm benzene and 100ppm toluene significantly decreased GSSG levels and increased the GSH/GSSG ratio. The higher frequency of micronucleated PCE following benzene and toluene co-exposure when compared with mice exposed to benzene or toluene alone suggests that, at the doses used in this study, toluene can enhance benzene-induced clastogenic or aneugenic bone marrow injury. These findings exemplify the importance of studying the effects of binary chemical interactions in animals exposed to lower exposure concentrations of benzene and toluene on benzene metabolism and clastogenicity. The relevance of these data on interactions for humans exposed at low benzene concentrations can be best assessed only when the mechanism of interaction is understood at a quantitative level and incorporated within a biologically based modeling framework.