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

L-Menthyl alpha-D-glucopyranoside (alpha-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 alpha-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 H3BO3NaOHKCl buffer (pH 8.0) at 40 degrees C, alpha-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 alpha-MenG was obtained over 48 h with a highest conversion yield of 99.1% based on the supplied L-menthol. Crude alpha-MenG formed through this "crystal accumulation reaction" was easily collected from the reaction mixture by separation on filter paper. Plank-like crystals of purified alpha-MenG were subsequently obtained by recrystallization in ethyl acetate solution.