The addition for 6 days of 1 % orotic acid to the diet of rats reduced the activity of aniline hydroxylase in the 9000 g supernatant from 1683 ± 111 (S. E.) to 640 ± 96 n-moles/liver/10 min in the male, but had no effect in the female. Further, there was no significant change in microsomal protein. The addition of 0.25 % adenine sulfate along with the orotic acid gave the intermediate value of 1244 ± 116 nmoles/liver/10 min. On the other hand, the small decrease after orotic acid in ethylmorphine N-demethylase activity of microsomes was not statistically significant. Orotic acid decreased the level of cytochrome P-450 from 0.093 ± 0.004 to 0.058 ± 0.005 E (450–490)/ mg of microsomal protein in male rats without affecting the concentration of this cytochrome in females. Further, in male and female rats, the activity per unit of cytochrome P-450 decreased from 76.6 ± 5.6 and 67.5 ± 2.4 nmoles p-aminophenol formed/10 min/unit of cytoehrome P-450, respectively, to 60.3 ± 5.3 and 52.2 ± 1.5. The soluble fraction from orotic acid-fed animals increased aniline hydroxylase by 30–50 per cent in microsomes from control and treated animals.