Efficient Synthesis and Characterization of Lactulosucrose by <i>Leuconostoc mesenteroides</i> B‑512F Dextransucrase

Abstract

This work describes an efficient enzymatic synthesis and NMR structural characterization of the trisaccharide β-d-galactopyranosyl-(1→4)-β-d-fructofuranosyl-(2→1)-α-d-glucopyranoside, also termed as lactulosucrose. This oligosaccharide was formed by the <i>Leuconostoc mesenteroides</i> B-512F dextransucrase-catalyzed transfer of the glucosyl residue from sucrose to the 2-hydroxyl group of the reducing unit of lactulose. The enzymatic reaction was carried out under optimal conditions, i.e., at 30 °C in 20 mM sodium acetate buffer with 0.34 mM CaCl₂ at pH 5.2, and the effect of factors such as reaction time (0–48 h), enzyme charge (0.8, 1.6, and 2.4 U mL^–1), and sucrose:lactulose concentration ratios (20:40, 30:30, and 40:20, expressed in g/100 mL) on the formation of transfer products were studied. The highest formation in lactulosucrose was attained at 8 and 24–32 h by using 20%:40% and 30%:30% sucrose:lactulose mixtures, respectively, with 1.6 or 2.4 U mL^–1 dextransucrase, leading to lactulosucrose yields of 27–35% in weight respect to the initial amount of lactulose. Furthermore, minor tetra- and pentasaccharide, both probably derived from lactulose, were also detected and quantified. Likewise, the capacity of lactulosucrose to act as d-glucosyl donor once the sucrose was consumed, could explain its decrease from 16 to 24 h when the highest charge of dextransucrase was used. Considering the chemical structure of the synthesized oligosaccharides, lactulosucrose and its derivatives could potentially be excellent candidates for an emerging prebiotic ingredient