Enzymatic synthesis and identification of oligosaccharides obtained by transgalactosylation of lactose in the presence of fructose using β-galactosidase from Kluyveromyces lactis

摘要

The enzymatic transgalactosylation of lactose in the presence of fructose using β-galactosidase from Kluyveromyces lactis (KIβGal) leading to the formation of oligosaccharides was investigated in detail. The reaction mixture was analyzed by high performance liquid chromatography with differential refraction detector (HPLC-RI) and two main transgalactosylation products were discovered. To elucidate their overall structures, the products were isolated and purified using preparative liquid chromatography and analyzed by LC/MS, one-dimensional (ID) and two-dimensional (2D) NMR studies. Allo-lactu-lose((5-D-galactopyranosyl-(l→ 1 )-D-fructose) with two main isomers in D_2O was identified to be the major transgalactosylation product while lactulose(p-D-galactopyranosyl-(l → 4)-D-fructose) turned out to be the minor one, indicating that K/βGal was regioselective with respect to the primary C-l hydroxyl group of fructose. The maximum yields of allo-lactulose and lactulose were 47.5 and 15.4 g/l, respectively, at 66.5% lactose conversion (200 g/l initial lactose concentration).