Binding of laurate and myristate anions to human serum albumin has been studied over a range of temperatures, 5-37'C, at pH 7.4. The binding curves indicate that the strength of binding of the first few molecules of fatty acid to albumin ( r < 5) decreases with increasing temperature, whereas binding of the following molecules seems to proceed independently of temperature.Binding data were analyzed according to the general binding equation yielding several sets of acceptable binding constants within a probability limit of 0.75. From the temperature dependence of the first step constant, it was possible to calculate values for the changes in enthalpy and entropy during the initial binding step. For the medium-chain fatty acids, laurate and myristate, binding of the first molecule to albumin appeared to be enthalpic, with a tendency to an increasing contribution of entropy to binding energy with increasing chain length of the fatty acid.Serum albumin, being synthesized in the liver, is the most abundant protein in the extracellular fluid. The concentration in human plasma is about 0.6 mM, whereas in the interstitial fluids it varies in the different tissues between 0.2 -0.5 mM [l].Albumin is known to bind a plethora of different compounds including inorganic ions and a multitude of metabolites, hormones, and drugs [2]. These very flexible binding properties constitute the transport function of albumin.Albumin binding of fatty acids which have a low water solubility is important for the energy-metabolizing muscles. The fatty acids are released into the plasma as a result of mobilization of fat from adipose tissue during fasting, stress or exercise, and in plasma the concentration of fatty acids can reach 2.5 mM, i.e. four times the concentration of albumin Estimation of biological responses to varying concentrations of fatty acids therefore necessitates evaluation of multiple equilibria between fatty acids and albumin. Results, as well as difficulties, from this type of investigation have recently been reviewed by Spector [3, 41. We have introduced a dialysis method allowing determination of the concentration of unbound fatty acid anion by measuring the rate of exchange of labelled fatty acid across a dialysis membrane under conditions of equilibrium [5, 61. With this method we have obtained solubility data for different fatty acids and established binding isotherms for multiple equilibria between albumin and lauric acid [6] as well as myristic acid [7].In the present paper we extend this investigation by studying the effect of temperature on the binding curves in an attempt to estimate changes of enthalpy and entropy on binding of the anions of lauric and myristic acid to human serum albumin.131.