The role of folate deficiency in the pathogenesis of the abnormalities of structure and transport of the small intestine that are often present in certain disorders such as alcoholism is unclear. One reason for this is that the folate status of the intestinal mucosa has never been ascertained in humans. Previous investigations have shown that a deficient diet supplemented with sulfa does produce folate deficiency of the intestinal mucosa in weanling rats and that this is associated with diarrhea and structural abnormalities of the jejunum. In the present study, we used then in vivo marker perfusion technique to examine the jejunal transport of water, sodium, xylose, glucose, and L-leucine in such folate-deficient rats. Supplementation of the normal diet with sulfa did not produce folate deficiency of the intestinal mucosa, but it was associated with a moderate reduction in water absorption; this was attributed to cecal enlargement in these rats. Rats placed on a folate-deficient diet plus sulfa developed markedly depressed serum and jejunal folate concentrations, net secretion of water and sodium, and persistent diarrhea; absorption of xylose, glucose, and L-leucine remained normal, however. Folate repletion of these rats by water and sodium transport to absorption ano cessation of diarrhea. These observations indicate that folate deficiency of the intestinal mucosa alters the transport of water and electrolytes but not that of such solutes as xylose, glucose, and L-leucine. They suggest that when transport abnormalities of these solutes occur in folate-deficient alcoholics, this in not due to the folate deficiency but rather to other pathogenic factors such as ethanol.