In this study,the ultra performance liquid chromatography(UPLC) was used to establish fingerprint and multi-component determination method of Herba Serissae,and provide a comprehensive evaluation of the drug from different habitats.The further quality assessment of the drug were carried out by chemical identification of reference substance combined with similarity evaluation,principal component analysis (PCA),and orthogonal partial least squares discriminant analysis (OPLS-DA).The UPLC fingerprint of Herba Serissae was established and fourteen common peaks were designated. Six common peaks were identified,namely deacetyl asperulosidic acid,neochlorogenic acid,geniposide,asperulosidic acid,vicenin Ⅱ and quercetin-3-O-α-L-rhamnopyranosyl(1-2)-β-D-glucopyranoside-7-O-α-L-rhamnopyranoside by comparing with the reference substances and UPLC-ESI-QE-Orbitrap-MS technical analysis.Multivariate statistical analysis showed that 16 batches of Herba Serissae were divided into four categories,and eight quality differential markers were determined.The content of deacetyl asperulosidic acid,asperulosidic acid and vicenin Ⅱ in 16 batches of Herba Serissae standard were decoction ranged from 2.035 to 4.422 mg/g,1.210 to 3.279 mg/g and 0.087 to 0.288 mg/g,respectively.The contents of deacetyl asperulosidic acid and asperulosidic acid ether in Herba Serissae of Henan province were the highest,while the content of vicenin Ⅱ was low.The method established in this study is simple and accurate,which can provide an quality control and evaluation for the Herba Serissae from different producing areas and provide reference.