Abstract: OBJECTIVE To study on the terpenoids and their pharmacological activities in Eupatorium adenophorum Spreng. METHODS D101, HP20 macroporous resin, silica gel column chromatography, dextran gel chromatography, high performance liquid chromatography and other methods were used for separation analysis. Their structures were determined by MS, ¹H-NMR and ¹³C-NMR and other spectrographic techniques, as well as ECD calculation. The cytotoxic activity of these compounds were screened by the MTT method. RESULTS Ten compounds were isolated from the petroleum ether and ethyl acetate extraction fractions of 95% ethanolic extracts of Eupatorium adenophorum Spreng, and they were identified as eupatorinone D(1), thymoquinol 5-O-β-glucopyranoside(2), thymoquinol 2-O-β-glucopyranoside(3), 2R^*,3S^*-toxol-7- O-β-D-glucopyranoside(4), 6-methoxy-kaempferol-7-O-β-D-glucoside(5), patuletin-4'-methoxyl-7-O-β-D-glucopyranoside(6), 6-hydroxykaempferol-7-O-β-D-glucopyranoside(7), eupatonriochromene(8), demethoxyencecalin(9) and encecalin(10). Among them, the IC₅₀ value of compound 10 against esophageal cancer cells was 178.4 μmol·L^-1. CONCLUSION Compound 1 is a new cadinane-type sesquiterpene, and compound 6 was isolated from genus Eupatorium for the first time, compounds 2, 3 and 7 were isolated from Eupatorium adenophorum Spreng for the first time. Compound 10 has certain cytotoxic activity against esophageal cancer cells Eca-109.