The effect of bovine milk beta-lactoglobulin (BLG) on intestinal uptake of retinol was examined in suckling rats with the everted sac technique. Uptake of 0.06 mumol retinol/L bound to BLG (BLG-retinol) was significantly (p less than 0.01) higher than that of 0.06 mumol free retinol/L both in the jejunum and the ileum. The enhancing effect of BLG on retinol uptake was specific because equimolar concentrations of bovine serum albumin and lactoferrin had no effect on retinol uptake. However, serum retinol-binding protein (RBP), which shares structural and conformational similarities with BLG, also enhanced retinol uptake. BLG, BLG-retinol, and RBP-retinol all inhibited the uptake of retinol from BLG-[3H]retinol in a concentration-dependent manner. Uptake of retinol from BLG-retinol was saturable (apparent Km = 5.6 mumol/L, Vmax = 22.7 nmol.g-1.5 min-1), not affected by metabolic inhibitors, and partially temperature dependent (Q10 = 2.77). BLG also significantly (p less than 0.01) enhanced retinol uptake in the intestine of adult rats. These results demonstrate that BLG specifically enhances intestinal uptake of retinol and suggest the possibility of a receptor for BLG-like proteins at the brush border membrane of the enterocyte.