Several methods were evaluated for the solubilization of supraphysiological concentrations of β-carotene in culture media. The addition of β-carotene in ethanol as 0.1% (vol/vol) of culture medium that contained 10% fetal bovine serum and incubation at 37°C for 30 minutes solubilized approximately 50% of the added carotenoid. Solubilized β- carotene concentrations from 3.06 to 36.4 μmol/l were obtained by this method. Ultracentrifugation studies localized the solubilized β-carotene to low-density lipoproteins (67.0%) and high-density lipoproteins (24.9%). Other methods utilizing various amounts of fetal bovine serum, ethanol, and hexane as solvents did not significantly improve the solubilization of β-carotene compared with the use of ethanol as 0.1% (vol/vol) of culture medium that contained 10% fetal bovine serum. Nonetheless, solubilization of supraphysiological concentrations of β-carotene indicated the feasibility of preparing culture media containing physiological concentrations (≃0.5 μmol/l) of β-carotene.