Influence of water-miscible aprotic solvents on alpha-chymotrypsin stability

The influence of five different water-miscible aprotic solvents (dimethyl sulfoxide, dimethylformamide, acetonitrile, acetone, and tetrahydrofuran) on the stability of immobilized alpha-chymotrypsin adsorbed onto Celite, has been studied. In all cases, alpha-chymotrypsin exhibited non-first-order deactivation kinetics, which were adequately analyzed by a two-step series-type deactivation model. The main effects of solvents were observed in the first-step of the kinetic mechanism. The most hydrophilic solvent (DMSO) enhanced greatly the enzyme stability, while the increase in solvent hydrophobicity determines a loss in their protective effect. For the most assayed hydrophobic solvent (THF) a denaturative effect was always showed. These facts were also observed from the analysis of the evolution of the half-live of the enzyme, as a function of the solvent concentrations and the water distribution between both the macro- and microenvironment of the enzyme.