Study On Separation Of Anti?tumor Components And Pharmacodynamics Of Euphoorbia Kansui

The dried roots of Euphorbia kansui L.(Euphorbiaceae)are known as “Gan Sui” in Traditional Chinese Medicine.It is abundantly distributed in northwestern China.“Gan Sui” was firstly recorded in Sheng Nung's Herbal as a low grade drug and it is bitter,cold and poisonous.Modern researches indicated that it had numbers of activities,such as purgative,abortive,analgesic,immunosuppressive and anti-tumor activity.Previous investigations have shown that the ethyl acetate extract of Euphorbia kansui showed strongest anti-tumor activity in all kinds of commonly used solvent extracts.However the research of monomers was still in blank.In this study,a new strategy of the convergence of two-dimensional preparative high performance liquid chromatography(HPLC)and real-time cell analysis(RTCA)was developed for bioassay-guided separation of anti-tumor components from the ethyl acetate extract of Euphorbia kansui.The method might lay the material foundation for the anti-tumor mechanism of this medicine.The first-dimensional separation was conducted on silica column in industrial preparative high performance liquid chromatography and seven fractions were gatherd.After anti-tumor screening by iCELLigence real-time cell analysis system,fraction 4 and 6 were selected to sepatare by the second-dimensional preparation.Fraction 4 was prepared by 2D-NPLC/RPLC chromatography system with silica column and C18 column and fraction 6 was prepared by 2D-NPLC/NPLC chromatography preparation system with silica column and amide column.In this method,eight compounds esulone A(1),kansuinin A(2),(3?,11?)-3,11-dihydroxylanosta-8,24-dien-7-one(3),kansuinin E(4),kansuinin B(5),isoscopoletin(6),kansuinin D(7)and kansuinin G(8)were efficiently isolated in purity above 97%.Of all the compounds,esulone A has been isolated from this plant for the first time.The structures were identified by NMR spectroscopy and comparison with the previous literatures.The bioassay revealed that compounds 1,2,3,7 and 8 almost entirely inhibited the proliferation of A549 cells and that compound 8 was also thought to be the most active compound against Hep-G2 cells.The result of primary pharmacodynamics revealed that kansuinin A and kansuinin B showed obvious concentration-dependent effect on lewis lung cancer mice.The inhibitory ratios of kansuinin A and kansuinin B were similar to cyclophosphamide with the concentrations of 11.11mg/kg and 99.99mg/kg,respectively.The experiment might provide the reference for further research of its action mechanism.The convergence of 2D-HPLC and RTCA system had distinctly shorten the experiental period.It also prepared considerable assist for separation and anti-tumor screening of complicated compounds of Traditional Chinise Medicine.