Synthesis of potentially-bioactive lactosyl-oligofructosides by a novel bi-enzymatic system using bacterial fructansucrases

Abstract Efficient enzymatic synthesis of lactosyl-oligofructosides (LFOS) with a degree of polymerization from 4 to 8 was achieved in the presence of sucrose:lactosucrose and sucrose:lactose mixtures by transfructosylation reaction. The main synthesized LFOS which consist of β-2,1-linked fructose to lactosucrose: β- d -galactopyranosyl-(1 → 4)-α- d -glucopyranosyl-[(1 → 2)-β- d -fructofuranosyl]n-(1 → 2)-β- d -fructofuranoside (where n refers to the number of transferred fructose moieties) was structurally characterized by nuclear magnetic resonance (NMR). The maximum formation of LFOS was 81% (in weight with respect to the initial amount of lactosucrose) and was obtained after 24 h of transfructosylation reaction based on sucrose:lactosucrose (250 g L − 1 each) catalyzed by an inulosucrase from Lactobacillus gasseri DSM 20604 (IS). The production of LFOS in the presence of sucrose:lactose mixtures required a previous high-yield lactosucrose synthesis step catalyzed by using a levansucrase from Bacillus subtilis CECT 39 (LS) before the inulosucrase-catalyzed reaction. This novel one-pot bi-enzymatic system led to the synthesis of about 22% LFOS in weight, with respect to the initial amount of lactose (250 g L − 1 ). The results revealed a high specificity for the substrate involved in the inulosucrase-catalyzed reaction given that, although lactosucrose ( O -β- d -galactopyranosyl-(1 → 4)- O -α- d -glucopyranosyl-(1 → 2)-β- d -fructofuranoside) acted as a strong acceptor of β-2,1-linked fructose, lactose (β- d -galactopyranosyl-(1 → 4)-α- d -glucose) was found to be an extremely weak acceptor.