Abstract: OBJECTIVE To establish HPLC fingerprinting and multi-indicator quantitative analysis methods for Rhei Radix et Rhizoma and to predict potential quality markers of it in combination with network pharmacology.METHODS The content of 13 index components, including gallic acid, catechin and epicatechin, was determined in 20 batches of Rheum palmatum L. samples by HPLC, and the similarity analysis was performed by using the Chinese Medicine Chromatographic Fingerprint Similarity Evaluation System. The network pharmacology was used to predict the targets and pathways of Rhei Radix et Rhizoma-related components, construct a network diagram of components-targets-pathways and perform organ localization analysis on the core targets to predict the potential quality markers of Rhei Radix et Rhizoma.RESULTS The similarity of fingerprint profiles ranged from 0.882 to 0.998, and a total of 15 common peaks were identified. Emodin-3-methyl ether,catechin, emodin-1-O-glucoside, physcion-8-O-glucoside and aloe-emodin-8-O-glucoside were the differential compounds,which could be used to identify and differentiate Rhei Radix et Rhizome quality. The screening yielded 15 compounds with 8core targets including AKT1, TP53, JUN, HSP90AA1, CASP3, EGF, TNF, IL-6, involved in response to oxidative stress,response to lipopolysaccharide, endopeptidase activity, involved in lipid and atherosclerosis, MAPK signaling pathway,AGE-RAGE signaling pathway in diabetic complications pathways. Non-alcoholic fatty liver disease, and the construction of a component-target-pathway map. Prediction of gallic acid, catechin, aloe-emodin, rhein acid and emodin as potential quality markers of Rhei Radix et Rhizoma.CONCLUSION The established fingerprint profile of Rhei Radix et Rhizoma is highly characterized and can be used to control its quality. The characterization profile and network pharmacological analysis predict the quality markers of Rhei Radix et Rhizoma, which provide a reference for further research on the mechanism of action of Rhei Radix et Rhizoma.