We tested the hypothesis that preweaning diet alters cholesterol metabolism in adult baboons. Eighty baboons, progeny of 6 sires and 80 dams, were either breast-fed or fed one of three infant formulas containing 2, 30 or 60 mg cholesterol/100 mL. At 16 wk of age the baboons were weaned to one of four diets containing 1.0 or 0.01 mg cholesterol/kcal with 40% of energy from saturated [polyunsaturated fat/saturated fat [P/S) = 0.37] or unsaturated fat [P/S = 2.1] and maintained on these diets until they were necropsied as young adults at 7-8 yr of age. We observed no significant effects of formula cholesterol content on serum lipid or lipoprotein concentrations measured at 6-8 yr of age, but formula cholesterol intake influenced the cholesterol turnover rate and several variables of cholesterol metabolism in the adult. At 6-8 yr, baboons that were breast-fed during infancy, compared with those that were formula-fed, had lower high-density lipoprotein cholesterol (HDL-C) concentrations and higher ratios of very-low-density plus low-density lipoprotein cholesterol (VLDL + LDL-C) to HDL-C. Breast-fed baboons, as adults, had lower cholesterol production rates, masses of the rapidly exchanging cholesterol compartment (pool A) and neutral steroid excretion rates than did those fed formula as infants. Breast and formula feeding differentially influenced the adult metabolic responses to dietary cholesterol or fat saturation. These results demonstrate that breast vs. formula feeding in infancy alters cholesterol metabolism and serum lipoprotein concentrations in adult baboons.