Affinity chromatography study of magnesium and calcium binding to human serum albumin: pH and temperature variations

Abstract

The magnesium and calcium binding on human serum albumin (HSA) was studied using an affinity chromatography approach. The effects of the mobile phase pH, its ionic strength and column temperature on the transfer equilibrium constants were studied. The thermodynamic data corresponding to the electrostatic interactions occurring during the HSA-ion binding were determined. Enthalpy–entropy compensation revealed that the ion binding mechanism at HSA was independent of the ionic strength, the same at four pH values (6.5, 8, 8.5 and 9), but presented a weak change at physiological pH around 7–7.5 due to a HSA phase transition. A theoretical model based on the Gouy–Chapman theory allows to determine the relative charge density of the HSA surface implied in the binding process and the variation of the number of ions bound to one albumin molecule with the pH.