Extraction Purification And Structural Indentification Of Flavonoids From Hyperrcum Perforatum L.

The hypericum perforatum L.is distributed widely in ourcountry.Currently,the artificial cultivation area of hypericumperforatum L.is expanding constantly. The hypericum perforatumL.was recorded in the Chinese dictionary of drug in2005.From thenon,the studies about it become increasing. Traditional chinesemedicine reported that hypericum perforatum L.is moderateherbs,and it has the effectiveness of fever reduction, detoxification,meridians regulation, promoting blood circulatin, hemostatic andmuscel regeneration. Modern phamaacology proved that hypericumperforatum L. has the effectiveness of antidepressant, anti-virus,antibacterial and analgesic.The flavonoids of hypericum perforatumL. have multiple biological activity, such as anti-aging, free radicalscavenging, anti-diabetic and antioxidant.The experiment used hypericum perforatum L. as material, andexplored the process of extraction, preliminary purification,thencontinued to purificate, until getting the flavonoids monomer fromhypericum perforatum L.Finally, the experiment identificated thestructural of the flavonoids monomer.This experiment providetheoretical basis for development and utilization of hypericumperforatum L.herbs efficiently and increased the herbs’ medicinalvalue.The UV-visible spectrophotometry method,was established forthe quantitative determination of the flanovoids in the hypericumperforatum L.The standard solution of80%alcohol was scanned inUV district,and the result showed that the UV maximum absorption wavelength was360nm.Added color reagent to the standard solutionand scanned in visible district,the result showed that the maximumabsorption wavelength of visible was510nm. When the concent ofstandard solution was between0.015mg/mL and0.075mg/mL,therutin concentration and A510have good linear relationship.In orderto extract flavonoids from hypericum perforatum L., the experimentused of ultrasonic assisted organic solvents method,and investigatedthe impaction of differences organic solvents, the concentration ofsolvent,solid-liquid ratio,time, temperature, frequency and times ofextraction to the yield of flavonoids. The experiment used the yieldas an indicator,then designed orthogonal experiment by some factorswhich have greater impaction. Finally, the optimal extraction craftwas that crude drug was infused by60%alcohol,liquid ratio1:30,twice in extraction,and each extraction last45minutes,and thecontent of flavonoids was maximum which reached7.003%.The preliminary purification experiment of flavonoids fromhypericum perforatum L. selected3types of macroporous adsorptionresin,which were suitable for purificating flavonoids, and they areD101,D4020and AB-8. By static absorption and static desorptionexperiment,the result showed that the static absorption of D101wasmaximum which reached78.767mg/g,and the static desorption ratiowas90.2%.So,in three types macroporous resins, D101macroporousadsorption resin was the optimum resin which can be used forpreliminary purification experiment.Then used total flavonoidscontent as an indicator,the experiment investigated the impaction ofthe optimum flavonoids concentration of the original solution, thevolume of extract solution, the volume fraction of eluant and theconsumption of eluant by dynamic adsorption and elution tests.Theresult showed that the optimum purification process was theconcentration of original solution was3.202mg/ml, the volume ofextract solution was8times of resin,the eluant was40%alcohol andthe consumption of eluant was3BV. At the same time, the experiment used stratified extractionmethod to purificate flavonoids from hypericum perforatum L.,andreagents were followed by petroleum ether(Boiling range60℃-90℃),ethyl acetate and butanol.The flavonoids extraction samples which getted by stratifiedextraction and D101macroporous adsorption resin,were purificatedby silica gel column chromatography repeatedly. The eluant wereselected the gradient system of petroleum ether(boiling range60℃-90℃)-ethyl acetate and ethyl acetate-methanol.The eluant whichwere collectted and concentrated were appraisal qualitatively by thinlayer chromatography.The developing agent of thin layerchromatography was selected Chloroform: Methanol: Formicacid(17:2:1).After two or three times silica gel columnchromatography,three flavonoids monomer were getted frompreliminary purification samples. By NMR structural identification,They are quercitrin,isoquercetin and rutin respectively.