An increase in folate catabolism has been suggested as the cause of the folate deficiency observed in many clinical conditions, including chronic anticonvulsant therapy. Previous studies have shown that the radioactive catabolites, excreted after an equilibration period of 3 d, consisted exclusively of folates that had been cleaved to produce pteridines and p-aminobenzoylglutamate, most of which was excreted as acetamidobenzoylglutamate. We have developed an experimental animal model using mice to determine the rate of catabolism of [3H]pteroylglutamate (folic acid) by the quantitative estimation of [3H]p-aminobenzoylglutamate and [3H]acetamidobenzoylglutamate in urine. Administration of diphenylhydantoin at three different doses (0.5, 20, and 50 mg/kg) significantly increased the rate of catabolism as measured by an increase in both the mean daily excretion and the cumulative excretion of these catabolites. Administration of intramuscular phenobarbitone on the other hand, did not affect the rate of catabolism, when compared with controls.