A novel chitosan derivative to immobilize alpha-L-rhamnopyranosidase from Aspergillus niger for application in beverage technologies

alpha -L-rhamnopyranosidase (Rha, EC 3.2.1.40) is an enzyme of considerable importance to food technology in increasing the aroma of wines, musts, fru it juices and other beverages. The aim of this research is the immobilizati on of the Rha contained in a commercial preparation already used in the win emaking industry and purified in the manner described in a previous study [ 1]. The immobilization supports tested were chitin, chitosan and derivatize d chitosan, diethylaminoethyl chitosan (DE-chitosan) never previously used for this type of application. Particularly, on DE-chitosan, the Rha was ads orbed and cross-linked with various bifunctional agents (glutaraldehyde, di epoxyoctane, suberimidate and carbodiimide), whose best results (immobiliza tion yields and activity) were obtained with carbodiimide (EDC) that allowe d a reduction in the involvement of the enzyme amine groups that are probab ly important in catalytic mechanism. In addition, the use of rhamnose and a succinimide (NHS) during cross-linking enhanced the action of the EDC and so increased the immobilization yield and activity. The immobilized Rha ret ained the kinetic parameters (K-m and V-max) of the free enzyme and increas ed stability. Moreover, this biocatalyst allowed an increase in the aroma i n a model wine solution containing glicosidic precursors with a marked redu ction in specificity toward tertiary monoterpenols as compared to the free enzyme. (C) 2001 Elsevier Science Inc. All rights reserved.