An isocratic high-performance liquid chromatographic method specifically developed to allow simple and rapid determination of β-carotene concentrations in serum and plasma is reported. Using a method modified from a previously published technique, serum and plasma proteins are denatured by exposure to perchloric acid, and β-carotene is subsequently extracted into an organic matrix consisting of ethyl acetate—tetrahydrofuran (1:1); no evaporation step is required. Separation is achieved using isocratic elution from a reversed-phase C 18, column with UV detection at 436 nm. Recovery of β-carotene from water and plasma was greater than 98.1%; β-carotene was stable in the extraction matrix for at least 4 h. Three anticoagulants (oxalate, citrate, and EDTA) caused losses of β-carotene; perchloric acid and tetrahydrofuran could also destroy β-carotene under certain conditions. Each run required less than 15 min ; within-day coefficient of variation for identical samples averaged 2.3%, between-day coefficient of variation was 4.4% and sensitivity was better than 10 ng/ml. Stability of β-carotene in plasma was also examined. This method permits a simple, rapid, sensitive, precise, and accurate determination of β-carotene using 0,5 ml of serum or heparinized plasma.