The β-glucosidase-catalyzed synthesis of n-alkyl-β-d-glucosides through the condensation of glucose and n-alcohols with alkyl chain lengths of 6 to 12 was carried out in a biphasic system, where the organic phase consisted only of an alcohol substrate. All the alcohols tested were condensed with glucose to give the corresponding n-alkyl-β-d-glucosides. An equation was formulated to predict the equilibrium yields of n-alkyl-β-d-glucosides under various conditions, and its validity was experimentally verified. The equilibrium constant for the formation of n-alkyl-β-d-glucosides was independent of the alkyl chain length and was 1.90 at 60°C. The volume ratio of the organic to the aqueous phase was a factor significantly affecting the equilibrium yield. It was also shown that the synthesis at lower initial glucose concentrations effectively yielded alkylglucosides with less β-glucobiose formation.