Preparation Of Single Angeloyl Oleanane Theasapogenol,Synthesis Of Galactose Glycosylation And Structure-activity Relationship Against Magnaporthe Oryzeae

This laboratory preliminary study found oleanane-type saponins of Schima superba leaf have strong inhibition activity of rice blast fungus.Structure-activity relationship shows that it is close to restrain activity of rice blast fungus of C21,C22 angelica acyl and C3 carbohydrate chain.Considering low contents of Schima superba leaf saponins and more difficult to get enough sapogenin,the same family plant camellia is expected to provide alternative angelica acyl oleanane-type saponins.Therefore,this study attempts to use tea saponin as starting material to achieve the diversification of molecular design synthesis and structure-activity relationship study.In this paper,tea saponins were obtained using water bath extraction,resin adsorption and precipitation.The theasapogenols were prepared using acid hydrolysis,solvent extraction,column chromatography and HPLC.Qualitative and quantitative analysis of theasapogenol was performed by chromatography and spectrum.The results showed that the conditions for the water extraction of teasaponins were ratio of material to liquid 1:25(m/V)under water 80 ? for 2 h,triplicate.Teasaponins were purified by AB-8 resin with elution H2O,0.1%NaOH,H2O and 90%ethanol successively,then precipitated 3 h,centrifuged and obtained with purity 96%.Two theasapogenols were prepared by the process of acid hydrolysis in 3 mol/L 60%HCl-Methanol aqueous solution with the ratio of material to liquid 1:15(m/V),reflux at 85 ? for 5 h;extract with ethyl acetate,elution through silica gel column with petroleum ether-ethyl acetate,and semi-preparative HPLC.The theasapogenol was identified as 21?-O-angeloyloxy-15a,16a,23,28-tetrahydroxy-22?-O-(2-methylbutanoyloxy)theasapogenol according to their spectrum data and compared with literatures(abbreviation:TS).Catalyzed by racemic camphor sulfonic acid,using PhCH(OMe)2 for ortho primary?secondary hydroxyl protection of TS,obtain glycosylacceptors 1 and 2(named respectively referred to as:TS1 and TS2),galactose trichloroacetimidates as donors,TMSOTf as catalyst,respectively glycosylation reaction with TS1 and TS2,received eight galactosyl oleanane-type saponins.Through separation and purification,1H NMR and13C NMR identification,their chemical structures,for respectively:3,15-dihydroxy-21-O-angel oyloxy-22-O-(2-methylbutanoyloxy)-16,28-O-benzal-ole an-12-ene23-O-?-D-Gal(TSl-Gal-1);15-hydroxy-21-O-angeloyloxy-22-O-(2-Methylbutan oyloxy)-16,28-O-benzal-olean-12-ene-3-O-?-D-Gal-23-O-p-D-Gal(TS 1-Gal-2);15,16-dih ydroxy-21-O-angeloyloxy-22-0-(2-methylbutanoyloxy)-3,23-O-benzal-olean-12-ene28-O-?-D-Gal(TS2-Gal-1);16-hydroxy-21-0-angel oyloxy-22-O-(2-methylbutanoyloxy)-3,23-0-benzal-olean-12-ene-15-O-?-D-Gal-28-O-?-D-Gal(TS2-Gal-2);21-O-angeloyloxy-22-O-(2-methylbutanoyloxy)-3,15,16,28-tetrahydroxy-olean-12-ene23-O-?-D-Gal(TS1-Gal-1");21-O-angel oyloxy-22-O-(2-Methylbutanoyloxy)-15,16,28-trihydroxy-olean-12-ene-3-O-?-D-Gal-23-O-?-D-Gal(TS1-Gal-2");21-O-angeloyloxy-22-O-(2-methylbutanoyloxy)-3,15,16,23-tetrahy droxy-olean-12-ene28-O-?-D-Gal(TS2-Gal-1");21-O-angeloyloxy-22-O-(2-methylbutanoyloxy)-3,16,23-trihydroxy-olean-12-ene-15-O-?-D-Gal-28-O-?-D-Gal(TS2-Gal-2").Inhibitory activity was determined of tea saponin,teasapogenin and galactosyl oleanane-type saponins on Magnaporthe oryzae by mycelium growth rate method,their antimicrobial activity respectively is 79.61%.42.45%.11.19%.30.98%.38.33%.23.87%?29.58%?4.91%?48.38%?29.64%?35.83%?26.16%,their structure-activity relationship:(1)C3 polysaccharide side chain groups of tea saponin against rice blast fungus a ctivity has the very important role,bacteriostatic activity reduced a lot when the h ydrolysis of the polysaccharide chain;(2)Embedding the hydroxyl groups on steroids skeleton of teasapogenin,bacteriost atic activity reduced in different degree,especially embedding C3 with C23 hydrox y1 at the same time;(3)When C16 and C28 hydroxyl condensation,connect one galactose in C23 or C3,C23 separately,active increased in varying degrees,but not more than teasapo genin;(4)When C3 and C23 hydroxyl condensation,connect one galactose in C28,Acti vity decreased but the change is not obvious,while connect one galactose in C16 and C28 separately,active direct almost lost;(5)Remove benzylidene groups,connect one galactose in C23,the active increase is the largest and more than raw material activity on rice blast fungus;Therefore,make the teasapogenin C16 and C28 hydroxyl bare,at the same time increase sugar base types of C3 and C23,and C3 sugar base number,effective optimization screening molecular design direction of oleanane-type saponins activities on Magnaporthe oryzae,to increase the number and types of glycosylation need to further study.