Both nitroreduction and ring-oxidation appear to be important pathways for the in vivo metabolic activation of 1-nitropyrene (1-NP), a tumorigenic environmental contaminant. Previous studies, however, suggest that ring-oxidation is primarily responsible for the S9-mediated mutagenicity of 1-NP in Chinese hamster ovary (CHO) cells. In this study, an anaerobic rat liver S9 metabolizing system was used to facilitate the nitroreduction of 1-NP to a mutagen in repair-proficient CHO-K1-BH4 cells and excision-repair-deficient CHO-UV5 cells, and results using this system were compared with those obtained from the aerobic S9 metabolism of 1-NP. Anaerobic S9 metabolism of 2.5-15 micrograms/ml of 1-NP produced 15 +/- 3 mutants/10(6) cells/microgram 1-NP/ml with CHO-UV5 cells and 3 +/- 1 mutants/10(6) cells/microgram 1-NP/ml with CHO-K1-BH4 cells (different at P less than 0.001). When the assays were conducted with CHO-K1-BH4 cells, the number of mutants produced by 1-NP using aerobic treatment conditions was similar to that found using anaerobic conditions. In contrast, the aerobic incubations resulted in significantly fewer 1-NP-induced mutants than the anaerobic treatments when the assays were conducted with CHO-UV5 cells. Examination of the metabolites produced during these incubations indicated that under anaerobic conditions 1-NP was efficiently converted to 1-aminopyrene, while aerobic metabolism resulted in the formation of 1-NP phenols and dihydrodiols. DNA adduct analysis by 32P-postlabeling revealed that 1-NP treatment using the anaerobic procedure produced CHO-cell adducts by the reduction of 1-NP to N-hydroxy-1-aminopyrene, while aerobic incubations resulted in adducts produced by other metabolic pathways, probably involving ring-oxidation. These findings indicate that the S9-mediated metabolism of 1-NP under anaerobic conditions produces mutations and DNA adducts in CHO cells that are the result of nitroreductive metabolism. The results with aerobic S9 metabolism were consistent with the previous conclusion that this system mediated the mutagenicity of 1-NP in CHO cells mainly through the generation of ring-oxidized metabolites. The combination of the anaerobic and aerobic S9 metabolism procedures provides a new approach for evaluating the mutagenicity of nitropolycyclic aromatic hydrocarbons in mammalian cells.