High-yield production of α-glucosylglycerol using Xanthomonas campestris WU-9701 derived XgtA

α-Glucosylglycerol (αGG), an α-glucoside found in alcoholic beverages, exhibits beneficial physiological effects. However, high costs and low yields limit its availability in the market. Therefore, we aimed to optimize the conditions for high-yield αGG production and establish a method for effective αGG separation. Accordingly, maltose and glycerol were used as the α-glucosyl donor and acceptor, respectively. Enzymatic production was executed using XgtA, an α-glucosidase derived from Xanthomonas campestris WU-9701, and glucose isomerase (GI). The products were αGG comprising 1-O-αGG and 2-O-αGG. αGG was separated from the reaction solution using activated carbon, followed by high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry. Under optimal conditions, the concentrations of αGG and 1-O-αGG reached 1141 mM and 1072 mM, respectively, within 48 h. These are the highest values obtained by enzymatic production to date. 1-O-αGG and 2-O-αGG were separated using an HPLC system equipped with the DCpak PMPC column. The area ratio of the two separated peaks of the products was 94:6 for the enzymatic reaction solution containing 1141 mM αGG. The products corresponding to these peaks were isolated for nuclear magnetic resonance analysis. The main product was confirmed to be 1-O-αGG, whereas the other was 2-O-αGG. Furthermore, the melting point of 1-O-αGG was 126 °C. Finally, the enzymatic products were confirmed to be 1-R-O-αGG, 1-S-O-αGG, and 2-O-αGG. These findings confirmed the successful high-yield and enzymatic production of αGG, and the first isolation and chemical identification of 1-O-αGG. Our method can potentially be implemented in industrial processes, owing to its high productivity. KEY POINTS: • We enzymatically produced the highest 1-O-α-GG yield reported to date • We developed methods for 1-O-αGG and 2-O-αGG detection and characterization • The 1-O-αGG:2-O-αGG ratio changed according to the reaction temperature.