Improvement of activity and stability of soluble and sol–gel immobilized naringinase in co-solvent systems

The hydrolysis of naringin, a bitter flavonone glycoside, with naringinase leads to reducing sugars (rhamnose and glucose), to prunin and to the aglycone, naringenin. To overcome the low solubility of naringin in the enzymatic reaction media, the effect of different solvents was studied, in order to improve the productivity and yield of the system. The effect of increasing concentration of co-solvents on the stability of both soluble and immobilized naringinase expressing α-l-rhamnosidase and β-d-glucosidase activities was evaluated. The enzyme was immobilized onto sol–gel matrices of tetramethoxysilane and glycerol. Combining the higher naringin solubility, and the higher residual activity of both α-l-rhamnosidase and β-d-glucosidase expressed by naringinase, eight solvents were chosen for stability and activity studies: dimethyl sulfoxide, N,N-dimethylmethanamide, methanol, ethanol, acetone, tetrahydrofurane, 1,2-dimethoxyethane and 1,4-dioxane.Deactivation of soluble naringinase was analyzed according to a first-order kinetic model. For the sol–gel immobilized enzyme the two-step deactivation model, of Henley and Sadana, was adjusted.Sol–gel immobilization stables naringinase in all tested co-solvents systems. This effect was specially pronounced at higher co-solvent concentration (10%). The half-life of α-l-rhamnosidase and β-d-glucosidase expressed by naringinase even increased 21- and 59-fold, respectively, in aqueous co-solvation with tetrahydrofurane.These are high innovative and sounding results showing the protective effect of immobilization onto sol–gel (tetramethoxysilane + glycerol) matrices with naringinase in co-solvent systems, which is a great advantage for non-conventional biocatalysis.