Changes in concentrations of milk and serum carotenoids, retinol, and alpha-tocopherol of five healthy, well-nourished, lactating women were measured over a 28-d supplementation trial with 30 mg beta-carotene and for 4 wk thereafter. beta-Carotene supplementation increased mean beta-carotene concentrations in milk and serum 6.4- and 7.4-fold, respectively. Concentrations of other major carotenoids, retinol, and alpha-tocopherol did not change substantially in either milli. or serum. Uptake of beta-carotene into both serum and milk followed apparent first-order kinetics, occurring more rapidly into serum (t(1/2),,, = 5.5 d) than into milk (t(1/2),, = 9 d). After supplementation, milk and serum beta-carotene concentrations decayed slowly, reaching approximately twofold initial concentrations by 4 wk. Kinetics of uptake and decay in milk beta-carotene concentrations paralleled those in serum, The data show that short-term supplementation of healthy, lactating mothers with purified beta-carotene at approximately fivefold the average daily dietary intake substantially increased milk and serum beta-carotene concentrations while not interfering with concentrations of other carotenoids, retinol, or alpha-tocopherol in milk or serum. Thus, an increased intake of beta-carotene by healthy, lactating women increases the supply of milk beta-carotene available to their breast-feeding infants.