α-anomer-selective glucosylation of menthol with high yield through a crystal accumulation reaction using lyophilized cells of Xanthomonas campestris WU-9701

l-Menthyl α-D-glucopyranoside (α-MenG) is a desirable derivative of l- menthol with useful properties for the production of new flavors and novel food additives. Bacteria were screened for α-anomer-selective glucosylation activity toward l-menthol, resulting in the isolation of two strains, Xanthomonas campestris WU-9701 and Stenotrophomonas maltophilia WU-9702, from independent soil samples. Since the safety of X. campestris for use in the food industry is well established, WU-9701 was selected as the more suitable strain for further study. When 50 mg X. campestris WU-9701 lyophilized cells as a biocatalyst were incubated with 1.0 M maltose and 100 mg l-menthol in 10 ml of 10 mM H₃BO₃ - NaOH - KCl buffer (pH 8.0) at 40°C, α-MenG was accumulated, mainly in a crystalline form, through the anomer-selective synthesis reaction without any by-product formation. Under the optimal conditions, 202 mg α-MenG was obtained over 48 h with a highest conversion yield of 99.1% based on the supplied l-menthol. Crude α-MenG formed through this 'crystal accumulation reaction' was easily collected from the reaction mixture by separation on filter paper. Plank-like crystals of purified α- MenG were subsequently obtained by recrystallization in ethyl acetate solution.