The rate of metabolism of [14C]isofenphos (IFP) to isofenphos oxon (IFP-oxon), des N-isofenphos (d-N-IFP), and des N-isofenphos oxon (d-N-IFP-oxon) by rat, guinea pig, monkey, dog, and human liver microsomal P-450 enzymes was studied to obtain Vmax and Km values for Michaelis-Menten kinetics. The monkey had the highest Vmax value for the conversion of IFP to IFP-oxon (desulfuration), 162 nmol isofenphos hr-1 per 1.3 nanomoles P-450, followed by guinea pig (98), rat (66), dog (43), and human (14). The Km values for the desulfuration of isofenphos were 19.2, 7.4, 14.1, 23.3, and 18.4 microM, respectively, for the monkey, guinea pig, rat, dog, and human. The Vmax values for the dealkylation process (conversion of IFP to d-N-IFP) were 64.6, 17.2, 9.7, and 7.3 nmol isofenphos hr-1 per 1.3 nanomoles P-450 for the monkey, rat, dog, and human, respectively. For the dealkylation process, monkey had the highest Km value, 16.3 microM IFP, followed by human (11.2), rat (9.9), and dog (9.3). The rate of metabolism of IFP-oxon and d-N-IFP to d-N-IFP-oxon were separately studied. The Vmax and Km values obtained in this study for animal and human liver P-450 enzymes will be used to develop a PB-PK/PB-PD model to predict the fate and toxicity of isofenphos in animals and man.