The uptakes of albumin-bound nonesterified fatty acids and of [1-14C]palmitic acid complexed to albumin by the isolated perfused rat liver were compared. During perfusion, the rate of uptake of nonesterified fatty acids decreased and became zero when the fatty acid:albumin molar ratio reached 0.3, but the rate of uptake of radioactive palmitic acid remained constant. This finding suggests the existence of two pools of fatty acids bound to albumin with different fractional turnover rates. This view was supported by the fact that when delipidated albumin complexed in vitro to radioactive and nonradioactive fatty acids was used no difference was observed between the uptakes of nonesterified fatty acids and radioactive fatty acids by perfused liver. Similar results were found with albumin-bound radioactive fatty acid in vivo (obtained from rats fed radioactive palmitic acid), showing a homogeneous distribution of the label in both pools. The existence of two nonesterified fatty acid pools in plasma would arise from the differences in the nature of bonds between fatty acid and albumin molecules, which could determine the rate of exchange of fatty acids between the albumin-bound and soluble forms preceding their uptake by the cells.