Studies On The Chemical Constituents Of Metaplexis Japonica And Their Effects On Multi-drug Resistance Of Cancer Cell And Lipid Accumulation In Vitro

Background & ObjectiveMetaplexis japonica(Thunb.)Makino is a plant of Asclepiadaceae.This plant is mainly grown in Henan,Gansu,Shanxi,Guizhou Provinces.Numbers of bioactive constituent have been isolated and identified from the plants of Asclepiadaceae.C21-steroids,flavonoids,cardiac glycosides,fatty acids,fatty acid esters and so on are found.Among them C21-steroids or pregnanes are representative characteristic components.While only a few chemical and pharmacological researches about the plant Metaplexis japonica were reported.Our research group reported that four tenacigenin B derivatives were identified from this plant and two ester derivatives of tenacigenin B showed activity in reversing MDR in P-gp overexpressed human cancer cells.Based on literature M.japonica has effects of tonifying the kidney and improving the milk-secretion for lactation woman.To learn more of the C21 steroidal compounds with new structure and their prospect of drug discovery,this thesis focuses on the chemical constituents isolated from the total aglycone of this plant,and the effects of the isolates on the reversal of multi-drug resistance in P-gp overexpressed cancer cells and the lipid accumulation in liver cell line.Content and Method1.Separation and identification of the chemical constituents from plant Metaplexis japonica(Luo-Mo).The chemical constituents were isolated from the total aglycones of the ethanol extract of Metaplexis japonica(Thunb.)Makino through repeated silica gel,ODS,Sephadex LH-20,HPLC column chromatographies.2.Reversal effect of the pregnane compounds on P-pg mediated multi-drug resistance in cervical cancer cells He La/Tax.3.Regulatory effect of of the isolated compounds on lipid production in liver cell line HepG2 cells induced by oleic acid,which were detected by Oil red O staining.Results1)On the basis of spectroscopic data,including NMR,UV,MS,IR,the chemical structures of the six isolated compounds(VI-8,A2,C1,C3,D1,and V-6)(1-6)were respectively identified as: 3β,12β,17β,14β,20-pentahydroxy-5α-pregnan-12β(E)-O-(2-methyl-2-butenoate),i.e.,12β-O-tigloyl-tomentogenin)](1);12β-O-(E)-cinnamoyl-5α-pregnan-3β,12β,14β,17β,20(S)-pentol(2);5α-pregnan-3β,12β,14β,17β,20(S)-pentol-20(S)-O-(E)-cinnamate(3);5α-pregnan-3β,12β,14β,17β,20(S)-pentol-12-O-nicotinate(4);5α-pregnan-3β,12β,14β,17β,20(S)-pentol-20-O-nicotinate(5);and ethyl-β-Dthevetopyranosyl-(1→4)-α-D-oleandropyranoside(6).2)Our in vitro anticancer experiment showed that compounds 2 – 4,at a non-cytotoxic concemtration of 5 and 10 μM,increased the cytotoxicity of paclitaxel(Taxol)in a P-gp overexpressed cervical cancer cell line He La/Tax,among these three compounds the reversal fold of 2 for taxol against P-gp mediated He La/Tax cells was 12 and 20,similar to those of MT2,a positive MDR reversal agent found by our research team.3)Compound 2,at a concentration of 2.5 ～ 10 μg/m L,obviously inhibited the production of lipid in HepG2 cells.ConclusionCompounds 1-6 were isolated and identified from the total aglycones manufactured by mild acidic hydrolysis on the ethanolic extract of plant Metaplexis japonica for the first time.Among them compounds 3,4,and 5 are new compounds.Compounds 2 – 4,at a noncytotoxic concentration may sensitize the cytotoxicity of paclitaxel against overexpressed Pgp mediated MDR cervical cancer cells.The inhibitory effect of compound 2 on lipid production in HepG2 cells were reported for the first time.