After oral d-xylose ingestion, cystic fibrosis patients have significantly higher blood levels of xylose than controls. The aim of this study was to examine whether nutrient absorption at the mucosal level is altered in cystic fibrosis. Steady-state perfusion experiments using isotonic test solutions were performed in 11 healthy controls and 10 cystic fibrosis patients. Net d-glucose absorption was higher in cystic fibrosis when the perfusate contained a glucose concentration of ≤ 50 mM. Kinetic analysis by three different methods, including Lineweaver-Burk analysis, revealed a lower apparent K_m as well as a lower apparent V_max in cystic fibrosis as compared with healthy controls (33.9 mM and 52.5 mmol/20 cm · h vs. 81.8 mM and 68.3 mmol/20 cm · h, respectively, p < 0.01). Absorption of d-fructose and glycine demonstrated a tendency for increased net absorption in cystic fibrosis but the results were not significantly different. l-Xylose absorption and electrolyte movement were not altered in cystic fibrosis. Among several possible mechanisms investigated, a decrease in the apparent K_m for glucose absorption would be consistent with a decrease in diffusion barriers overlying the jejunal mucosa in cystic fibrosis. Using an electrical method, the unstirred water layer thickness was significantly decreased in cystic fibrosis (546 ± 41 μm in cystic fibrosis vs. 780 ± 110 μm in controls, p < 0.05). A decrease in the mucosal surface area in the cystic fibrosis group or an intrinsic defect in the mucosal glucose transport system could account for differences in the apparent V_max values. We suggest, however, that enhanced absorption in cystic fibrosis is most likely due to a decrease in intestinal diffusion barriers possibly due to abnormal mucus overlying the intestinal mucosa.