OBJECTIVETo establish HPLC characteristic fingerprints of the saponins in Pulsatilla medicinal plants, and provide the basis for authentication and classification of Pulsatilla species.

METHODThe HPLC profiles were determined at 35 degrees C on a Symmetry C18 column (4.6 mm x 250 mm,5 microm) eluted with water (A) and acetonitrile (B) as mobile phases in a linear gradient elution with the flowrate of 0.5 mL x min(-1). The elution program was as follows: 0-8 min, 90% A to 77% A, 8-25 min, changed to 71% A, 25-40 min, to 60% A, 40-50 min, to 50% A, 50-75 min, to 10% A, 75-80 min, to 0% A. The detection wavelength was set at 210 nm.

RESULTThe different species of Pulsatilla showed different HPLC fingerprints, but with 10 common peaks. A cluster analysis of 14 accessions indicated that they were divided into four groups: all accessions from P. koreana were classified into group I, P. ambigua in group II, P. dahurica and P. turczaninovii in group III, and P. chinensis in group IV, respectively. The significant differences between P. koreana and P. dahurica, and between P. turczaninovii and P. ambigua were observed.

CONCLUSIONThe results obtained were in agreement with the traditional taxonomic study. The method was rapid and precise, not only can be used to classify and authenticate Pulsatilla species, but also provides important references for HPLC fingerprints and quality control of Pulsatilla medicinal plants.