In order to develop a clean and effective approach for producing the valuable drug diosgenin from Dioscorea zingiberensis tubers, two successive processes, enzymatic saccharification and microbial transformation, were used. With enzymatic saccharification, 98.0% of starch was excluded from the raw herb, releasing saponins from the network structure of starch. Subsequently, the treated tubers were fermented with Trichoderma reesei under optimal conditions for 156 h. During microbial transformation, glycosidic bonds, which link beta-D-glucose or alpha-L-rhamnose with aglycone at the C-3 position in saponins, were broken down effectively to give a diosgenin yield of 90.6+/-2.45%, 42.4% higher than that obtained from bioconversion of raw tubers directly. Scaled up fermentation was conducted in a 5.0-l bioreactor and gave a diosgenin yield of 91.2+/-3.21%. This is the first report on the preparation of diosgenin from herbs through microbial transformation as well as utilizing other available components in the raw material, providing an environmentally friendly alternative to diosgenin production.