Modification of β-galactosidase for use in organic mono-phase hexanol system

Abstract

Different methods for preparation of the Aspergillus oryzae β-galactosidase were evaluated and the total enzyme activity was determined in hexanol mono-phase system using p-nitrophenyl-β-D-galactoside as a substrate. Several supports such as Acurell EP-100, Amberlite IRC (50), Celite, Dowex, Eupergit C and silica gel were tested in order to select the most suitable matrix for immobilization of the β-galactosidase for performing transgalactosylation reactions in hexanol. Celite and Amberlite IRC (50) were selected as the most appropriate carriers. Albumin, PEG 6000, starch and glycine, added prior to the immobilization procedure, acted as stabilizers of the galactosidase. By adding albumin on Celite, a 3.3-fold increase of enzyme activity was achieved. Sodium dodecyl sulphate (SDS), dioctyl sulfosuccinate (AOT), Tween 65 and crown ether were used directly in the reaction medium in order to increase its homogeneity. Addition of SDS to the medium resulted in a 3.65-fold increased activity of the β-galactosidase deposited on Celite. A micro-emulsion system created by addition of AOT resulted in an increased catalytic activity. The β-galactosidase showed enhanced total activity with increased water activity in the system having the highest value for the water activity close to the saturation level (0.92).