Ferric human serum heme-albumin (heme-HSA) shows a peculiar nuclear magnetic relaxation dispersion (NMRD) behavior that allows to investigate structural and functional properties. Here, we report a thermodynamic analysis of NMRD profiles of heme-HSA between 20 and 60 degrees C to characterize its hydration. NMRD profiles, all showing two Lorentzian dispersions at 0.3 and 60 MHz, were analyzed in terms of modulation of the zero field splitting tensor for the S = 5/2 manifold. Values of correlation times for tensor fluctuation (tau(nu)) and chemical exchange of water molecules (tau(M)) show the expected temperature dependence, with activation enthalpies of -1.94 and -2.46 +/- 0.2 kJ mol (1), respectively. The cluster of water molecules located in the close proximity of the heme is progressively reduced in size by increasing the temperature, with Delta H = 68 +/- 28 kJ mol(-1) and Delta S = 200 +/- 80 J mol (1) K (1). These results highlight the role of the water solvent in heme-HSA structure-function relationships. (C) 2009 Elsevier Inc. All riglits reserved.