The effect of chemically-induced acute diabetes mellitus on the intestinal transport of 5-methyltetrahydrofolate, the predominate dietary form of folate, was examined using an in vivo perfusion technique in unanesthetized restrained rats and the in vitro everted sac technique. Diabetes was induced by an intraperitoneal injection of 70 mg/kg of streptozotocin. Transport experiments were performed 4 days after the induction of diabetes and results were compared to appropriate controls. The amount of 5-methyltetrahydrofolate transported in vivo and in vitro was significantly higher in the diabetic group as compared to controls. The kinetic parameters of the in vitro transport of 5-methyltetrahydrofolate showed an increase in the Vmax value for the diabetic rats as compared to controls (1.52 and 0.80 nmol/g initial tissue wet wt/30 min, respectively) with no change in apparent Kt (1.62 and 1.80 microM, respectively). These observations suggest that the increase in the transport of 5-methyltetrahydrofolate observed in the diabetic rats is due to an increase in the activity and/or the number, but not the affinity, of the transport carriers involved. We conclude that chemically-induced acute diabetes mellitus is associated with an enhancement in folate absorption.