Study of Binding of Benzyl-Thiouracil to Human Serum Albumin by Gel Filtration Chromatography

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Abstract
The binding of benzyl-thiouracil to human serum albumin was studied at 37°C and pH 7.4 by Sephadex filtration chromatography based upon Hummel and Dreyer's method. As the benzyl-thiouracil (ligand) was adsorbed on to the gel matrix, the free ligand concentrations had to be corrected through the ligand distribution between the stationary and mobile phases. A good agreement was found between binding parameters—calculated by this method and by the classical method (equilibrium dialysis). Binding is characterized by one binding site with a moderate association constant (K1 = 5.7 × 104 M-1) and two binding sites with a low association constant (K2 = 7.8 × 103 M-1).