Synthesis of a novel prebiotic trisaccharide by a type I α-glucosidase from B. licheniformis strain TH4-2

The use of alpha-glucosidase from Bacillus licheniformis TH4-2 in the glucosyl transfer reaction for the synthesis of oligosaccharides (OS) was evaluated. The conditions for alpha-glucosidase production were optimized. The enzyme was 112-fold purified with a 28% yield. The molecular mass was 64 kDa, and the optimum pH and temperature were 6.0 and 45 degrees C. The highest hydrolytic activity was observed towards p-nitrophenyl alpha-D-glucopyranoside followed by isomaltose, sucrose and maltose, supporting that it is a type I alpha-glucosidase. The enzyme could synthesize OS by transglucosylation from sucrose donor using various saccharides as acceptor. Melibiose, an alpha-galactoside, was selected as an efficient and interesting acceptor. TLC and HPLC analysis of the products revealed that the optimum condition for OS production was pH 6.0 with 15% melibiose, 5% sucrose and 5 U/ml enzyme at 45 degrees C for 24 h. Under these conditions two product peaks were observed in the HPLC profile with yields of 17.2% and 3.3%. The main product was isolated by Sephadex LH-20 column and analyzed by MS and NMR as the novel 504 Daltons trisaccharide, alpha-D-glucopyranosyl-(1 -> 6)-alpha-D-galactopyranosyl-(1 -> 6)-D-glucopyranose. The prebiotic nature of this product was suggested from its resistance to hydrolysis by rat intestinal acetone powder containing digestible enzymes of rat intestine. (C) 2010 Elsevier Ltd. All rights reserved.