Diverse thymol analogs from Eupatorium fortunei: Their anti-inflammatory activity and mechanism.

Twenty-five thymol analogs including six new ones (1-6), along with a new phenylpropanoid (7), were isolated from Eupatorium fortunei Turcz. The structures of the new compounds were elucidated through comprehensive spectroscopic data analyses (HRESIMS, 1D and 2D NMR, and UV). Compound 7 exists as a pair of inseparable isomers at C-10 or C-11, due to the possible intramolecular transesterification. Five thymol derivatives (5, 12, 15, 17, and 22) significantly inhibited nitric oxide (NO) production in LPS-induced RAW264.7 macrophages, with IC50 values of 8.1 ± 0.4, 14.1 ± 0.1, 11.9 ± 0.5, 13.0 ± 0.3 and 30.4 ± 0.9 μM, respectively. Integrated network pharmacology prediction and experimental validation revealed that compound 5 exerts anti-inflammatory effects primarily via inhibiting the p38 MAPK/NF-κB signaling axis, thereby leading to downregulation of iNOS and the pro-inflammatory cytokines Il6, Il1b and Tnfa. In conclusion, E. fortunei is rich in thymol derivatives, which may collectively contribute to the anti-inflammatory properties of this herb.