The results of these experiments indicate that thein vitro metabolism of diazinon is accomplished by enzyme preparations from fish liver homogenates, with the microsomal fraction being most active. The enzyme system responsible for diazinon metabolism in channel catfish liver microsomes required NADPH and oxygen for the oxidative desulfuration of diazinon to diazoxon, and probably for cleavage of diazinon and diazoxon to diethyl phosphorothioic acid and diethyl phosphoric acid, respectively. Thus, these reactions are presumed to be catalyzed by the NADPH-cytochrome P450 mixed function oxidase system. The only slight increase in diazinon metabolism by channel catfish hepatic microsomal or soluble enzymes with exogenous GSH suggests that GSH-dependent enzymes were not very active.