The level of N-acetyl-L-glutamate showed a diurnal rhythm, which was associated primarily with dietary ingestion. Since the hepatic level of arginine, activator for acetylglutamate, is one of the plausible principal factors responsible for the regulation of acetylglutamate level, the effect of arginine was further studied. The intraperitoneal administration of a large amount of arginine increased the level of acetylglutamate and the effect was augmented by the combined administration of glutamine. The arginine administration stimulated the hepatic synthesis of acetylglutamate as measured with [14C]glutamate as tracer, but did not affect the half-life (about 20 min) of the synthesized 14C-labeled acetylglutamate.The effect of arginine in raising the acetylglutamate level was also demonstrated in vitvo in isolated rat liver cells. On the other hand, pent-4-enoic acid, known to inhibit fatty acid oxidation and to decrease acetyl-CoA production in the liver, lowered the level of acetylglutamate in the rat liver cells. It was shown that the cellular ureogenic capacity, as measured with NH4Cl as nitrogen source, showed a close correlation with the varying levels of acetylglutamate, brought about by arginine or pent-4-enoic acid.The carbamoyl phosphate level, which may reflect the balance between its synthesis and utilization for citrulline synthesis, was low (about 1 nmol/g) and remained relatively constant in the livers of intact mice even when the rate of urea synthesis was expected to vary with changes in the dietary protein content. A transient increase in the level was brought about by agents expected to increase the synthesis or to inhibit the utilization, such as ammonia, N-acetylglutamate diethyl ester, or norvaline. These results, together with others, suggest that in intact animals the production of carbamoyl phosphate is coupled to its utilization for citrulline synthesis.