We studied the postnatal development of hepatic bile acid conjugation in the rat. Overall conjugating activity (homogenate assay) and the activity of the individual enzymes involved in the two step conjugation reaction (coenzyme A-bile acid thioester formation by microsomal cholyl-CoA ligase and amino acid transfer to this intermediate mediated by cytosolic cholyl-CoA:taurine N-acyltransferase were measured on days 1, 7, 14, 21, and 56 of life. Cholic acid conjugation was significantly lower in the suckling rat on days 1, 7, and 14 compared to the adult rat on day 56. At weaning (21 days) there was a marked increase in homogenate activity to a level higher than at 56 days. This peak in conjugating activity could be precociously induced on day 14 by cortisone acetate injection on days 10 through 13. Both cholyl-CoA ligase and cholyl- CoA:taurine N-acyltransferase activities were significantly lower during the first 14 days of life in comparison with 56 days. At 21 days of age there was a sharp increase in cholyl-CoA ligase activity to 1.5 times the activity in the adult; however, cholyl-CoA:taurine N-acyltransferase activity remained less than half of that found in the adult. We conclude that 1) cholic acid conjugation, as assessed by determination of either total conjugating activity or by the individual enzymatic steps, is decreased in the suckling rat; 2) the peak in conjugating activity noted at weaning may be mediated by the hormonal milieu, and 3) cholyl- CoA ligase activity closely parallels the increase in total activity and appears to be rate-limiting during development.