Previous studies reported that Agrimonia pilosa (AP) Ledeb possessed diverse biological activi-ties,including anti-inflammatory,antioxidant,and anti-tumor activities.However,the effect of AP on ulcerative colitis (UC) remains unclear.In this study,we investigated the therapeutic effect and mechanisms of AP on dextran sodium sulfate (DSS)-induced colitis.The potential constituents of AP were investigated by ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS).A total of 13 compounds were recognized by UPLC-Q-TOF/MS chromatogram.Furthermore,a network pharmacology approach revealed that there are 297 candidate targets of UC and 549 common targets for the 13 active ingredients of AP.GO enrichment and KEGG pathway analysis indicated that AP might have a protective effect on UC through the nuclear factor κB (NF-κB) and nucleotide-binding oligomerization domain (NOD)-like receptor signaling pathways.Subsequent experimental validation in a DSS-induced colitis model revealed that AP alleviated the severity of DSS-induced colitis,reduced the production of proin-flammatory factors,and protected against the loss of intestinal integrity.Moreover,AP inhibited the phosphorylation of NF-κB p65 and the activation of the NLRP3 inflammasome.In conclusion,AP ameliorated DSS-induced colitis through suppressing the activation of the NLRP3 inflammasome and NF-κB signaling pathways.