Study of affinity supports based on reactive polymers immobilized on silica: affinity constant determination from...

Polymers bearing benzamidine moieties have been prepared from reactive copolymer containing chloroformate functions and deposited on porous silica matrices. These high-performance affinity chromatography supports were characterized by quantitative methods, which analyse the zonal elution behaviour of trypsin in the presence of a soluble competitor (L-arginine). The column loading capacity for trypsin was measured by the zonal elution method in mass overload conditions. On the basis of a Langmuir isotherm, the influence of the protein capacity and the concentration of the soluble ligand on the elution volume was studied for the determination of the binding constants. The plate heights determined for silica supports of various porosities and particle diameters show that the strong affinity interactions between trypsin and p-aminobenzamidine are mainly responsible for the low efficiencies observed.