Contents of epimedin C and icariin in 3 Guizhou mainstream varieties of the medicinal material epimedium from different harvesting season were compared.The high performance liquid chromatog. (HPLC) method was adopted to determine the epimedin C and icariin content in Epimedium acuminatum, Epimedium wushanense T. and Epimedium leptorrhizum Stearn.The samples were separated on the Elite SinoChrom ODS-AP column (250 mm × 4.6 mm, 5.0μm); the mobile phase was acetonitrile (A)-water (B) by the gradient elution (0-22 min, 27%→29% A; 22-23 min, 29%→100% A; 23-34 min, 100% A; 34-36 min, 100%→27% A; 36-50 min, 27%).The linear ranges of epimedin C and icariin were 13.22-396.6μg (r=0.999 9) and 3.026-151.3μg (r=0.999 9) resp.The average recovery rates of epimedin C and icariin were 98.0%-105.0%; the contents of epimedin C and icariin in Epimedium acuminatum were 0.516%-2.973% and 0.096%-0.216%, resp.The contents of epimedin C and icariin in Epimedium leptorrhizum Stearn were 0.071-0.185% and 0.081%-0.164% resp.The contents of epimedin C and icariin in Epimedium wushanense T. were 1.015%-4.219% and 0.080%-0.190% resp.Epimedium wushanense T. has the highest content of epimedin C and icariin, followed by Epimedium acuminatum and Epimedium leptorrhizum Stearn is in the end, which indicates that different producing areas and different varieties of epimedium medicinal material have quite different contents of epimedin C and icariin.