Immobilization of β‐Galactosidase on Fibrous Matrix by Polyethyleneimine for Production of Galacto‐Oligosaccharides from Lactose

Abstract

The production of galacto-oligosaccharides (GOS) from lactose by Aspergillus oryzae beta-galactosidase immobilized on cotton cloth was studied. A novel method of enzyme immobilization involving PEI-enzyme aggregate formation and growth of aggregates on individual fibrils of cotton cloth leading to multilayer immobilization of the enzyme was developed. A large amount of enzyme was immobilized (250 mg/g support) with about 90-95% efficiency. A maximum GOS production of 25-26% (w/w) was achieved at near 50% lactose conversion from 400 g/L of lactose at pH 4.5 and 40 degrees C. Tri- and tetrasaccharides were the major types of GOS formed, accounting for about 70% and 25% of the total GOS produced in the reactions, respectively. Temperature and pH affected not only the reaction rate but also GOS yield to some extend. A reaction pH of 6.0 increased GOS yield by as much as 10% compared with that of pH 4.5 while decreased the reaction rate of immobilized enzyme. The cotton cloth as the support matrix for enzyme immobilization did not affect the GOS formation characteristics of the enzyme under the same reaction conditions, suggesting diffusion limitation was negligible in the packed bed reactor and the enzyme carrier. Increase in the thermal stability of PEI-immobilized enzyme was also observed. The half-life for the immobilized enzyme on cotton cloth was close to 1 year at 40 degrees C and 21 days at 50 degrees C. Stable, continuous operation in a plug-flow reactor was demonstrated for about 3 days without any apparent problem. A maximum GOS production of 26% (w/w) of total sugars was attained at 50% lactose conversion with a feed containing 400 g/L of lactose at pH 4.5 and 40 degrees C. The corresponding reactor productivity was 6 kg/L/h, which is several-hundred-fold higher than those previously reported.

Immobilization of β-Galactosidase in Adsorptive Membranes for the Continuous Production of Galacto-Oligosaccharides from Lactose

“…The corresponding reactor productivity for the production of GOS from lactose with Mustang TM Q in a Continuous Membrane-Chromatography-Reactor-System (CMCRS) was 98.7 grams GOS per hour and cubic centimeter membrane volume, which significantly exceeds previously reported results [12,13,20].…”

“…Some techniques have been developed for immobilization of β-galactosidase including non-covalent adsorption, covalent binding, entrapment and encapsulation [1][2][3][10][11][12][13][14][15].…”

“…Some of the advantages of immobilizing enzymes via physical adsorption on an activated support are: the activated support is chemically inert, good stability during storage, controlled residence time on the enzyme, simple recovery of the enzyme after usage, the possibility of a continuous process in enzyme reactors, the elimination of carryover of the enzyme to the final product [2,12,14].…”

Membrane chromatography reactor system for the continuous synthesis of galactosyl-oligosaccharides

“…The enzyme was reported to have stronger hydrolytic activity than transferase activity and produced a high proportion of trisaccharide in the synthetic GOS mixtures [1,2,7]. Some techniques have been developed for immobilization of ß-galactosidase including non-covalent adsorption, covalent binding, entrapment and encapsulation [1][2][3][8][9][10][11]. The newly developed synthetic micro porous membrane adsorbers as chromatographic media are an attractive alternative to traditionally used packed bed chromatography, which has several limitations [12].…”

“…In our study, the membrane adsorbers have been investigated as an alternative support for immobilization of enzyme to produce GOS. Some of the advantages of immobilized enzymes via physical adsorption of enzyme on activated support are: the chemically inertness of activated supports, good stability during storage, controlled residence time on the enzyme, the recovery of the enzyme after usage, the possibility of a continuous process in enzyme reactors and the elimination of the enzyme from the final product [3,10]. This present study focuses on developing of a method for immobilization of ß-galactosidase in chromatography membranes, investigating the properties of immobilized enzyme and the possibility of using the immobilized enzyme system for the synthesis of GOS from lactose.…”