The Research On The Extraction, Purification, Isolation And Identification Technology Of Flavonoids From Licorice Residue And The Active Antibacterial Property

Historically,liquorice is widely medicinal. From the 1950s,the researchers in countries have systematically studied different kinds of licorice of chemical composition. Currently 170 various chemical composition have been isolated from different licorice. Flavonoids,triterpene compounds and Glycyrrhiza Polysaechiad are the main ingredients in licorice. People have been more concerned with terpene saponins compounds in licorice,neglecting the research of the development of licorice flavonoids.According to reports,after extracting glycyrrhizic acid, licoric residue still contains a lot of flavonoids compounds. Previously,most licorice residue was abandoned what made licorice lost part value and failed to fully develop licorice resources. In recent years,with licorice flavonoids in pharmacological research, people find licorice flavonoids in with antibacterial,anti-inflammatory,anti-cancer,etc.Thus licorice flavonoids increasingly become pharmacological research hotspot. With the expansion of licorice exploitation degree,liquorice demand is increasing rapidly,and people no-limits unreasonable abuse, making limited resource of wild licorice threatened.Today natural resources is increasingly nervous,and we should seek a comprehensive utilization way of limited resources. Therefore this paper selects licorice residue as experimental materials which has been extracted glycyrrhizic acid by laboratory personnel ,and explores the best technology of extraction of licorice flavonoids. With the help of modern instrument testing and analysis means ,the chemical composition of licorice residue were separated and identified.While chemical constituents of bacteriostasis were tracked and selected. The experimental study provide a theory basis for the further application of flavonoid compounds in licorice residueFirst, establish the method of quantitative determination of flavonoids in licorice residue:The lutin as standard sample and NaNO₂,Al(NO₃)₃,NaOH reagent as the indicator were used to determined the content of flavonoids at the wavelength of 510 nm,to establish the linear regression equation which is about absorbency and concentration. The method is simple and feasible,which can rapidly determine the content of flavonoids in liquorice residue.Second,the optimal extraction technology and parameters of flavonoids in licorice residue are selected with single factor and orthogonal test.The result showed the optimal extracting condition were:using Microwave extracting technology,the volume ratio of ethyl and ethanol ,1:4; Microwave power,500W; ratio of material to liquid,1:10; extracting temperature ,70℃; extracting time,90min; extracting times,2.At last,the content 18.65% and the yield 1.41% of flavonoids licorice residue from can be realized.Third,D101 macroporous resin is a suitable route to purify flavonoids from licorice residue. The process conditions are as follows:choose pH=7,40% ethanol solution as sample solution for licorice flavonoids, concentration of sample solution 0.9 mg/mL,with a flow rate 2 BV/h; gradient elution method was used, remove impurities with 40 % ethanol solution,use 80%of ethanol and 5BV ethanol as eluant,with a flow rate of 4 BV/h.The content of licorice flavonoids was 40.3% after purification by D101 macroporous resin.The fourth,three kinds of compounds were isolated from extraction of licorice residue.By using the modern spectral methods, three kinds of compounds were identified.They are liquiritin,formononetin,β-Sitosterol.The fifth,through ethanol extraction and ethyl acetate extraction of licorice residue compared with ethanol extraction of licorice and crude products of glycrrhizin,the ethyl bacteriostasis of acetate extraction of licorice residue is only weaker than ethanol extraction of licorice ;and ethyl acetate extraction of licorice residue has stronger bacteriostatic ability on bacteria than fungus. MIC of ethyl acetate extraction of licorice residue on staphylococcus aureus,pseudomonas aeruginosa, escherichia coli,candida albicans are 1.77mg/mL,3.53mg/mL,1.77mg/mL and 7.06mg/mL.The sixth,liquirtin and formononetin separated from ethyl acetate extraction of licorice residue was studied ,tracked and screen about bacteriostatic research. Eventually found formononetin has stronger antibacterial activity than glycrrhizin and glycyrrhetinic acid and liquiritin has no bacteriostatic effect. Further research suggests that mononetin has better effect against staphylococcus aureus and escherichia col than pseudomonas aeruginosa and candida albicans.And MIC of formononetin on staphylococcus aureus, pseudomonas aeruginosa, escherichia coli, candida albicans are 78.2μg/mL,156.3μg/mL,78.2μg/mL and 156.3μg/mL.