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In our continuous study on the quality assurance of Eleutherococcus Senticosus Rhizome (ESR), we investigated the botanical origin of the commercial ESR obtained in Heilongjian, China using ITS sequence analysis of nuclear rDNA. Furthermore, we established a simple and rapid authentication method of ESR based on the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and estimated the detection limit of the method in preparation for its application as a purity test. As a result, two ITS genotypes were observed in the commercial ESR and they were supposed to be the inherent origin, Eleutherococcus senticosus and the related plants such as E. sessiliflorus. These data are in accord with our previous study. The authentication method based on the PCR-RFLP method could clearly discriminate the inherent material from the counterfeits. In addition, the related plants were detected in all 11 samples when artificial 5% adulterant samples, which consisted of 95% E. senticosus and 5% related plant in weight, were assayed by the authentication method. Therefore, it was found that the purity test of ESR utilizing the PCR-RFLP method can detect contaminants at the 5% level.