Effects of pectin, a soluble dietary fiber, on fatty acid and glucose absorption were studied in vivo in rats and humans by perfusing the intestine with linoleic acid and glucose solutions with and without pectin. Linoleic acid and glucose absorption decreased with increasing concentrations of pectin. The reduction in linoleic acid absorption was not caused by binding of linoleic acids by pectin or impaired micelle formation due to binding of bile acid by pectin. The unstirred water layer expanded with increasing concentrations of pectin. These results suggest that enlargement of the unstirred water layer is closely associated with the reduction of absorption of fatty acid and glucose ingested with pectin.