Abstract: OBJECTIVE Two species of C21 steroidal aglycones of Caudatin steroidal aglycones and Kaider steroidal aglycones, which were extracted from Cynanchum auriculatum, could be C11 α-hydroxylated with low by-product content and high-yield objectives. METHODS Rhizopus nigricans and Absidia sp. Were used to synchronized transformation and by shaking cultivation for 70 hours at 30 ℃ in water/butanol two-phase system with pH=5.0-6.0. RESULTS The products were confirmed to be C11 α-hydroxylated Caudatin steroidal aglycones and Kaider steroidal aglycones by IR, MS, and NMR. The results also indicated that by-product content was low and the yield of objectives was up to 70.1%. CONCLUSIONS The method was high effective for C11 α-hydroxylation of C21 steroidal aglycones in water/butanol two-phase system by Rhizopus nigricans and Absidia sp