Indomethacin is now widely used during the neonatal period, but few data exist concerning its metabolism during this period. Thus, we studied the maturational changes in the metabolism of indomethacin into desmethylindomethacin by isolated liver microsomes from 1- to 60-day-old rats. From 8 days of age on, there was a progressive increase in enzymatic activity which was inversely correlated with the change in Michaelis-Menten constant. However, the maximal velocity was not significantly affected by the age. These findings suggest that the deficient metabolism of indomethacin in the newborn rat is due to decreased affinity between enzyme and substrate, or presence of competitive inhibitors. To determine the substrate specificity of the monooxygenase-catalyzing inducers, sodium phenobarbital and beta-naphthoflavone given during 3 days in neonatal rats were also studied. Although beta-naphthoflavone did not exhibit any significant effect, phenobarbital significantly increased the activity and maximal velocity of indomethacin O-demethylation.