A high-performance anion-exchange chromatography coupled with diode array detection method was developed for the determination of dencichine in Panax notoginseng and related species. The analysis was performed on an Eprogen Synchropak WAX column (4.6 × 250 mm, 6 μm) with 50 mM NaH PO aqueous solution isocratic elution. The method was validated in terms of linearity, sensitivity, precision, stability, and accuracy. It was found that the calibration curve for dencichine showed good linearity (R = 0.9999) within the test range. The LOD and LOQ were 0.77 and 3.06 ng, respectively. The RSD for intra- and interday repeatability was 0.2 and 0.5%, respectively. The test solution of dencichine is stable at least for three days at room temperature and for seven days at 4°C. The mean recovery of dencichine was 102.0%. The established method was successfully applied to determine dencichine in the raw root of P. nogoginseng, P. ginseng, and P. quinquefolium as well as the steamed root of P. notoginseng. Compared with previous reports, this method is sensitive, selective, and accurate, which is helpful to evaluate the quality of P. notoginseng and related species. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.