Identifying Active Compounds in Cnidii Fructus Against Ulcerative Colitis: Integration of Chromatographic Analysis, Spectrum‐Effect Relationship, and In Vitro/In Vivo Experiments

Cnidii Fructus (CF) is a Chinese herbal medicine with bioactive immune properties and potential applications in treating ulcerative colitis (UC), but its well-defined bioactive constituents are understudied. The current study aimed to investigate CF's anti-UC bioactivity and elucidate its bioactive constituents via chromatographic analysis, in vitro/in vivo tests, and spectrum-effect analysis. Results showed 10 batches of CF inhibited the inflammatory response induced by lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. High-performance liquid chromatography (HPLC) fingerprint profiles of the 10 batches were established, with 16 common peaks identified. Subsequently, 15 of these peaks (including eight reference-validated compounds) were characterized by ultraperformance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS). Four anti-UC-related compounds were identified via spectrum-effect relationships, namely, diosmin, xanthotoxol, imperatorin, and osthole. Experimental verification confirmed these as key components potentially linked to alleviating inflammatory responses in UC. This study is the first to establish a spectrum-effect relationship for CF's anti-UC effects, identify its critical active constituents, and provide a scientific basis for CF quality control and UC therapeutic development.