Experiments were performed using isolated mucosal cells from the rat jejunum or using the perfused jejunum in the anesthetized rat to test whether lipophilic unconjugated dihydroxy bile acids are absorbed from the proximal small intestine via the same carrier mechanism involved in the uptake of long chain fatty acids. With isolated jejunal mucosal cells, the cellular uptake rate of deoxycholic acid or chenodeoxycholic acid increased linearly with time, showed no evidence of saturation, and was not decreased by the presence of a monospecific antibody to the membrane fatty acid binding protein. In contrast, oleate uptake was saturable, was inhibited by the same antibody, but was not affected by the presence of chenodeoxycholic acid or deoxycholic acid. Bile acid uptake by isolated enterocytes occurred at one-eighth the rate of fatty acid uptake if expressed in relation to total solute concentration; if expressed in relation to monomeric concentration, initial bile acid uptake was four orders of magnitude slower than fatty acid uptake. In the isolated perfused jejunal segment, chenodeoxycholic acid and deoxycholic acid uptake was not influenced by the presence of the antibody to membrane fatty acid binding protein, whereas absorption of oleate was inhibited by more than 70%. These experiments indicate that absorption of unconjugated lipophilic dihydroxy bile acids in the rodent jejunum does not involve the carrier mediated uptake mechanism involved in the absorption of long chain fatty acids--the mechanism is likely to be passive nonionic diffusion.