The active flavonoids in silymarine extract prominently consist of silychristin, silydianin, silybin (A and B) and isosilybin (A and B). Among these active flavonoids, silybin is the most important one. It is often used to treat liver disorders. An HPLC method for determining thg active flavonoids is described in this paper. The components in the extract can be separated by using a reversed-phase system with a C18 column, eluted with methanol and phosphate buffer under gradient conditions, and detected at 280 nm. In comparing with the method of derivatization with 2,4-dinitrophenylhydrazine and detection with UV spectrometry which can only determine the total active flavonoids. The HPLC method not only can separate taxifolin from the flavolignans but also permit individual estimations of silychristin, silydianin, silybin and isosilybin. It has better repeatability. The relative standard deviation is below 2%(n = 5). It can be used for quality control of the crude extracts and the pharmaceutical preparations.