Maturation of hepatic bile formation and lipid secretion were examined in neonatal guinea pigs in an attempt to explain the earlier observation that the neonate (15 days old) is less susceptible to lithocholate-induced cholestasis than the adult. Bile flow and bile acid secretion were lowest at 12 h postpartum and increased to attain a maximum at 15 days. Thereafter, values decreased to a level which was not significantly different from that at 3 days of age. Bile acid analysis indicated the presence of chenodeoxycholic acid, 7-ketolithocholic acid and ursodeoxycholic acid in bile at all ages. But, at 15 days ursodeoxycholic acid was a major contributor to total bile acid secretion. Phospholipid and cholesterol secretion increased between the 1st and the 3rd day of age but decreased markedly at 15 days. This indicated an uncoupling of biliary lipid secretion and endogenous bile acids which was related in part to the nature of the bile acids secreted. Serum bile acid levels were markedly increased at 15 days of age. This hypercholanemia may reflect a rapid expansion of the bile acid pool including an enhanced bile acid secretion which could explain in part the resistance to the lithocholate cholestasis by increasing the secretory capacity for the toxic bile acid.