BACKGROUND: Ulcerative colitis (UC) is a stubborn disease that occurs globally. SSP-TXYF is a traditional Chinese medicine (TCM) compound, which is widely used in the treatment of UC, but its picking and preparation is complicated. Therefore, this work analyzes the components of SSP-TXYF to find an natural compound that can treat UC, and verifies its efficacy and mechanism, aiming to explore new natural compound with the same efficacy as SSP-TXYF. METHODS AND RESULTS: In this study, we used the quality markers (Q-marker) strategy to analyze the SSP-TXYF. We obtained that angelicin (Ang) is the main active ingredient. Combined with reductive dimethyl labeling and tandem orthogonal proteolysis-activity-based protein profiling (rdTOP-ABPP) and network pharmacology, we found that ERK1/2 and HDAC1 may be the binding proteins. Then, we demonstrated that Ang not only suppresses inflammation in lipopolysaccharide-induced IEC-6 but also effectively inhibits 2,4,6-trinitrobenzenesulfonic acid-induced UC in vivo. Increasing the concentration of propionate and butyrate, activating GPR43 and GPR109A receptors, increasing ERK1/2 activity, promoting Sp1 phosphorylation, thus competitively inhibiting the combination of HIF-1alpha and HDAC1, and finally increasing the acetylation of HIF-1alpha is the molecular signaling mechanism for Ang. CONCLUSIONS: In conclusion, Ang can improve the symptoms of UC in rats and activate the GPR/ERK/SP1/HDAC1/HIF-1alpha pathway in colonic epithelial cells to repair the damaged intestinal mucosal barrier (IMB). The key pharmacodynamic mechanism of Ang is consistent with SSP-TXYF, and it is preliminarily believed that Ang can be used as a new natural medicine for treating UC.