The intestinal uptake of phenytoin was studied as a function of concentration, intestinal region, coperfused glucose, and calcium chloride in rat intestinal perfusions and everted intestinal rings. Steady-state intestinal membrane permeabilities were obtained in an in situ perfusion system and initial rates of intestinal tissue uptake were obtained in an in vitro everted ring system as rate of absorption parameters. Steady-state membrane permeabilities were independent of phenytoin perfusion concentration and decreased from duodenum to ileum. Coperfusion of glucose increased, and high calcium chloride concentrations decreased phenytoin permeabilities. While phenytoin uptake in the in vitro ring system was also concentration-independent and depressed by high calcium concentrations, regional variations and glucose enhancement were not observed. Thus, drug-nutrient interactions involved in intestinal absorption from phenytoin solutions are a function of the isolation procedure.