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A novel strategy was developed to assess the quality of TCM (traditional Chinese medicine) based on the correlation analysis of the chemical fingerprint and biological effect. Using Rehmanniae glutinosa (RG) to treat the kidney yin deficiency as an example, chemical fingerprints of 27 RG samples were analyzed by liquid chromatography–mass spectrometry (LC–MS), and urinary metabolic profiling of RG treatment of kidney yin deficiency in rats was explored by using LC–MS. A correlation analysis between the chemical fingerprints and efficacy evaluation was developed to identify quality marker components to assess TCM quality. Thirty-four variables in chemical fingerprints were successfully defined to have a close relationship with the efficacy of RG. The validation test with a new RG sample indicated that these efficacy-related components could be used to evaluate the integral quality of RG accurately. Compared with conventional chemical fingerprint methodology, not only is the proposed approach a powerful tool to identify efficacy-related components for the quality evaluation of TCM, but the approach can also be used to predict the therapeutic efficacy of TCMs.The bubble chart indicated that chemical fingerprints of Rehmanniae glutinosa (RG) from different origins had a significant difference. The larger the diameter of the bubble, the worse the therapeutic effects of RG treating kidney yin deficiency. Efficacy-related components could be used to evaluate the quality and predict therapeutic effects of RG. PH: processed RG samples from Henan province. PS: processed RG samples from Shanxi province. RH: raw RG samples from Henan province. RS: raw RG samples from Shanxi province. PS4: processed RG sample from Shanxi province, used for the quality verification test as a new RG sample.