To study the fate of oleate in the colon, oleate labeled with carbon 14 was instilled into the cecum of four rats through chronically implanted cecal cannulas. Fecal recovery of 14C and 14CO2 excretion were measured over a 3-day period. A mean of only 57% +/- 6% of the dose of [14C(U)]oleate was recovered as 14C in fecal lipid. About 8% was recovered in the aqueous phase of feces and 4% was recovered as 14CO2. Occlusion of the terminal ileum did not diminish 14CO2 excretion, excluding ileal reflux with small bowel absorption. Studies in two germ-free rats showed no conversion of [14C(U)]oleate to fecal water-soluble compounds, indicating that [14C]oleate is converted into water-soluble compounds by bacterial metabolism. The metabolism of [14C]oleate to 14CO2 was also observed in germ-free rats, indicating oxidation in the colon or other host tissues. We conclude that the colonic absorption of lipid or lipid metabolites plus conversion to water-soluble fecal compounds or volatile metabolites results in a fecal fat measurement that appreciably overestimates small bowel absorption of lipid.