The effects of chronic ethanol ingestion on NADPH-oxidase and on the NADPH-catalyzed peroxidation of lipids in rat liver microsomes have been studied. It was demonstrated that the rates of NADPH oxidation, of oxygen consumption, and of malondialdehyde formation increased significantly above control values after one month of ethanol ingestion. Further, the fatty acid composition of these microsomes revealed a decrease in arachidonate and in the C22 polyenes. Also, the energies of activation for the formation of malondialdehyde increased in the microsomes from the ethanol-treated animals. These results were interpreted to mean that ethanol ingestion had induced changes in the microsomal membranes such that additional or alternate, possibly abnormal, pathways for lipid peroxidation were functional. Finally, these data suggest a mechanism whereby chronic ethanol ingestion inhances the production of lipid peroxides via the microsomal-catalyzed oxidation of NADPH.