Nitric oxide (NO) is both an endogenously generated species and the active species released from a variety of important drugs. Due to its endogenous generation and use as a therapeutic agent, the metabolism and fate of NO is of interest and concern. To date, most attention regarding the metabolism and fate of NO has been paid to its oxidized metabolites. Due to the reducing environment of cells, we considered that NO may also undergo reductive metabolism as well. Therefore, we have examined the reductive metabolism of NO by hepatocytes. Generation of nitrous oxide (N(2)O) was used as an indication of NO reduction. Indeed, we observed that NO could be reduced to N(2)O by the cytosolic fraction of hepatocytes. The N(2)O production was partially inhibited by the thiol modifying agent, N-ethylmaleimide and thiol consumption was observed during N(2)O formation. Thus, our results indicate that NO reduction is feasible and likely occurs via a thiol-dependent process.