Reactive dyes human serum albumin (HSA) binding by frontal analysis affinity chromatography

The binding with human serum albumin (HSA) of Cibacron Blue F3GA, Levafix Brilliant Blue EB, the structure of which has been carefully investigated, Reactive Orange 046, and Reactive Red 022 dyes, immobilised on sepharose Ct 6B, has been quantitatively investigated by frontal affinity chromatography. This technique allowed the calculation of the association constants of The ligand-HSA complexes, on the basis of the dependence of the HSA elution volume on its initial concentration. The values obtained were high and comparable for Cibacron and Levafix dyes (5.4 and 5.7x10(6) M-1, respectively) and significantly smaller for Reactive Orange 046 and Reactive Red 022 (0.23 and 0.4x10(6) M-1). A possible interpretation of these data, based on the different structures of the bonded dyes, is proposed.