Study On Extraction And Separation And Biological Activities Of Flavonoids From Aeschynomene Indica L.

Aeschynomene indica L.is the dry whole plant of Aeschynomene indica Linn..It taste sweet and bitter.It has some functions of heat dampness,dieretic and promoting lactation.At the same times,the present research have showed that the extract of Aeschynomene indica L.has antioxidant,anti-bacteria,anti-inflammatory and antiallergic activities,but now there are few literature reports about its effective active site.In addition,the modern pharmacological studies have shown that flavonoids is one of the main active ingredients of natural plants.It has biological activities such as antioxidant,anti-inflammatory,etc.But The research about flavonoids of Aeschynomene indica L.has rarely reported and this affected the effective utilization of Aeschynomene indica L.Therefore this paper using Aeschynomene indica L.as the research object provided a theoretical basis for the effective utilization and comprehensive development of Aeschynomene indica L.and the active ingredients through the researches of the extraction technology,separation and purification,structure identification,content determination of multiple index component and biological activity in Aeschynomene indica L.of flavonoids.The main contents are as following:1.Research on the extraction technology of flavonoids from Aeschynomenene indica L.The extraction process of flavonoids from Aeschynomene indica L.was optimized by single-factor tests and RSM.According to the actual situation,the optimum extraction conditions were as follows: extraction temperature of 90 ℃,extraction twice and 60 min each time,ethanol concentration of 65% and liquid-solid ratio(g/m L)of 25:1.Under the optimum conditions,the yield of total flavonoids higher to 4.08 mg/g.2.Separation,purification and structure identification of flavonoids from Aeschynomene indica L.This experiment had a system research of flavonoids from Aeschynomene indica L.for the first time.Four flavone components were separated from Aeschynomene indica L.by using organic solvent extraction,polyamide column chromatography,silica gel column chromatography,gel column chromatography,ODS reversed-phase column chromatography and recrystallization methods.It has been indentified as quercetin,avicularin,isoquercitrin and rutin,respectively,by physicochemical properties,TLC,MS and NMR.3.Simultaneous determination of the contents of rutin,isoquercitrin,avicularin,quercetin in Aeschynomene indica L.by HPLC Simultaneous determination of the contents of rutin,isoquercitrin,avicularin,quercetin of crude extract and total flavonoids in Aeschynomene indica L.was established.The results showed that the compositions of purified flavonoids basically remained unchange.And the contents of four flavone conponents in purified total flavonoids were enriched,which the purified avicularin is up to 54.312 mg/g.This method is rapid,simple and high repeatability.It can provide certain reference basis for the control of inner quality in medicinal materials and lay a theoretical foundation for the research of subsequent pharmacodynamics.4.Research on the antioxidant activity of flavonoids of Aeschynomene indica L.The antioxidant activity of crude extract,total flavonoids and flavone components of Aeschynomene indica L.has been studied by four types of external antioxidant indexs.The research showed that the antioxidant activity of total flavonoids was higher than crude extract in concentration range of 25 μg/m L to 200 μg/m L.Furthermore,quercetin showed highest antioxidant activity,which associated with the structure of components.The sticking point of antioxidant activity in quercetin depends on the existence of catechol in the B ring,a double bond of 2,3 post,3-OH and 5-OH while antioxidant activity in rutin,isoquercitrin,avicularin weaken obviously because of glycosylation in 3-OH.5.Research on the anti-inflammatory activity of flavonoids of Aeschynomene indica L.The influence of flavonoids of Aeschynomene indica L.on the activity of macrophages through cck-8.The result showed that there is no obvious side effects on macrophages in the range of the samples concentrations of crude extract,total flavonoids and flavone components.The appropriate concentrations of samples was selected and the inflammatory model was setted by using LPS stimulate macrophages to study the influence of flavonoids of Aeschynomene indica L.on NO secretion capacity of macrophages.The result showed that the anti-inflammatory activity of total flavonoids higher than crude extract in concentration range of 50 μg/mL to 200 μg/mL.Furthermore,in concentration range of 10 μmol/L to 40 μmol/L,quercetin showed higher anti-inflammatory activity than avicularin.While rutin and isoquercitrin has no significant anti-inflammatory activity,which was owing to glycosidic replaced the hydroxyl which reduces the free hydroxyl combined with the site to make the anti-inflammatory activity of the greatly reduced or even missing.