Transport of enterostatin (VPDPR) across rat jejunum was investigated using Grass-Sweetana diffusion chambers. The rate of absorption of enterostatin and its metabolites were studied in absence and in presence of diisopropylfluorophosphate (DFP), a serine protease inhibitor. An extensive hydrolysis of enterostatin was observed during incubation with brush border membranes and when exposed to the mucosal side of the jejunal epithelium. No accumulation of enterostatin occurred in serosal tissue. Addition of DFP delayed enterostatin disappearance and abolished des-arg-enterostatin degradation. Under these conditions, a low amount of enterostatin was able to cross the epithelium intact. Moreover, a substantial amount of des-arg-enterostatin is absorbed across the jejunal epithelium, probably through passive diffusion. Thus, a decreased metabolic activity increased the absorption of a tetrapeptide (VPDP). Dipeptidylaminopeptidase IV, remains a limiting step in transfer of intact enterostatin and its metabolite des-arg-enterostatin across intestinal wall.