In the present study, the metabolism of NF has been examined in detail in both rat lung and liver 9000 g supernatants using a specific radiometric HPLC assay. Over 92% of the total radioactivity chromatographed with authentic NF after incubations from either organ were carried out under oxygen for 60 min. Under anaerobic conditions, only 19% and 5% of the total unbound radioactivity corresponded to unchanged NF in lung and liver respectively. At least 4 metabolites were evident from the HPLC trace (M1, M2, M3, M4 according to increasing retention times). In the absence of oxygen, liver 9000 g supernatants generated 65% more M1 and 260% more M3 than did lung 9000 g supernatants, but the lung produced significantly more M4. Covalent binding to tissue macromolecules was similar in both tissues under oxygen but was 7 times greater in lung than in liver in the absence of oxygen (compared per unit protein). Neither piperonyl butoxide nor indomethacin affected NF metabolism. However, allopurinol almost completely inhibited the anaerobic and aerobic (superoxide generation measured by the rate of acetylated cytochrome c reduction) metabolism in the lung with little or no effect in the liver. The data indicate a quantitative difference in NF metabolism between the two tissues that may be related to the organ-selective toxicity of the drug.