The kinetics of disappearance of radioactivity from plasma after intravenous injection of radiolabeled bile acids was characterized in healthy subjects. Cholic and chenodeoxycholic acids and their N-glycine and N-taurine conjugates were studied along with the secondary bile acid, deoxycholic acid, and a synthetic conjugated bile acid, dehydrocholyltaurine. Disappearance of each primary bile acid was rapid and could be described by a biexponential curve (t 1/2 the first component, 1.7 to 3 min; second component, 7 to 16 min). Cholyglycine and cholyltaurine had identical disappearance rates which were significantly faster than the disappearance rates of chenodeoxycholylglycine and chenodeoxycholyltaurine, which were also identical. Radioactivity of a conjugated bile acid disappeared more rapidly than that of its unconjugated form, but all of these differences, although consistent, were small. Deoxycholic acid radioactivity disappeared slightly more slowly than that of the primary bile acids. In vitro, dihydroxy bile acids were bound by serum proteins more completely (96 to 98%) than trihydroxy bile acids (83 to 91%), whether free or conjugated, and binding was unrelated to bile acid concentration. Thus, plasma disappearance rates were related to the degree of protein binding to some degree in that those bile acids which were more tightly bound had the longer t 1/2 values. However, dehydrocholyltaurine showed little protein binding (42%), yet had the slowest disappearance rates; t 1/2 values averaged 4 and 36 min for the two components of the disappearance curve. The results suggest that the residence time of any bile acid in blood is extremely short, and that measurable concentrations of serum bile acids persist only because of continuous intestinal absorption. It is speculated that the reported difference between the pattern of fasting state serum bile acids and biliary bile acids is explained by differences in intestinal absorption and hepatic uptake of individual bile acids.