Enterodiol (END) possesses the estrogenic and antiestrogenic activities, which could prevent the development of breast cancer and prostate cancer, as well as menopausal syndrome. Previous studies in our laboratory set up a bio-transformation method for largely yielding secoisolariciresinol (SECO) from the substrate of defatted flaxseeds by strain Bacteroides umformis ZL-I. In this research, another two single colonies, designated as strain ZL-II and strain ZL-III, were isolated, which were closely related to Eubacterium limosum species (ZL-II), and Eggerthella lento species (ZL-III) on the basis of 16S rRNA gene sequence data. Under the combining actions of strains ZL-(I+II+III), END could be produced from defatted flaxseed directly, ZL-II was proved to possess the activities of demethylation, while ZL-III had the activities of dehydroxylation. Secoisolariciresinol diglucoside (SDG) existed in the form of oligomeric with 3-hydroxy-3-methyl glutaric acid in flaxseed could be efficiently transformed into END under the co-culture of strains ZL-(I+II+III), with the conversion rate of more than 90%. The method for mass-producing END from defatted flaxseed reported here is meaningful not only for the medicinal values of END but also for the resource utilization of flaxseed materials. (C) 2013 Published by Elsevier Ltd.