Flavonoids In Puerariae Lobata Flos Quality Control And Protect Liver Therapy Of Pharmacodynamics And Mechanism Research

Objective : For leguminous plants flower of Pueraria lobata(Willd.)Ohwi or P.t homsoii Beeth.the unopened bud,synonym Puerariae lobatal article flowers,is the commonly used traditional Chinese medicine(TCM)therapy in our country.In recent years,the study reported the main effective components in Puerariae lobatale flos solution wine to protect liver isoflavones,consistent with this experimental pilot study;Experimental flavonoids site of Puerariae lobatal flos partsl parts for the main chemical composition,efficacy and mechanism of research,clinical application and quality standards,specification to provide a theoretical basis for the development and utilization of resources.Methods:After extraction and purification,the Puerariae lobatale flos herbs come flavonoids site of Puerariae lobatal flos partsl parts,respectively by using thin layer chromatography(TLC)and ultraviolet-visible spectrophotometry(UV)to flavonoids site of Puerariae lobatal flos partsl parts of main ingredients identification and content determination of total flavonoids;Method using high performance liquid chromatography(HPLC)determination of iris in flavonoids site of Puerariae lobatal flos partsl parts Tectoridin,Tectorigenin-7-O-xylosylglucosideose glycosidase prosthetic,Tectorigeninan,the content of and establish flavonoids site of Puerariae lobatal flos partsl parts fingerprint.Experiment choose ICR mice model of acute alcohol intoxication,with flavonoids site of Puerariae lobatal flos partsl parts of high,medium and low dose preventive intervention,by observing the flavonoids site of Puerariae lobatal flos partsl parts of drunk turn positive reflection,the ability of learning and memory in mice and the influence of the movement coordination and in the determination of ethanol concentration in blood of mice and activity of alcohol dehydrogenase(ADH),analysis of flavonoids site of Puerariae lobatal flos partsl parts of acute alcohol intoxication protection;Choose Wistar rats with acute or chronic alcoholic liver damage model is established,with flavonoids site of Puerariae lobatal flos partsl parts of high,medium and low dose preventive intervention,the detection of acute or chronic alcoholic liver injury in rats serum and liver in the influence of the biochemical indexes and liver pathology structure,analysis of flavonoids site of Puerariae lobatal flos partsl parts of acute or chronic alcoholic liver injury of rat liverprotection;Using enzyme-linked immune ELISA method and fluorescence quantitative PCR method to detect chronic alcoholic liver injury in rats serum tumor necrosis factor alpha(TNF alpha),transforming growth factor beta 1(TGF-beta 1),interleukin 6(IL 6)and interleukin 8(IL-8)levels and liver of nuclear factor related factor-2(NRF-2),nuclear transcription factors-kappa B(nf-kappa B)predominate,TGF-beta 1,m RNA expression of CD68,of chronic alcoholic liver injury rat liver protection mechanism,on the basis of the research.Results:Flavonoids site of Puerariae lobatal flos partsl parts after extraction and purification,qualitative identification by TLC,strong specificity,obvious spots were clear,separating degree is better;Establish a UV method of determination of total content of flavonoids in the flavonoids site of Puerariae lobatal flos partsl parts,Tectoridin as the comparison of regression equation Y = 0.0742 + 0.0123 X,correlation coefficient r =0.9995.Methodology validation results: precision,repeatability,stability,all conform to the rules;The recovery rate is 97.24 ~ 102.32%,RSD was 1.80%.Flavonoids site of Puerariae lobatal flos partsl parts of the flavonoids content of 93.24%.Establish HPLC and determination of three kinds of component content in flavonoids site of Puerariae lobatal flos partsl parts,the method of Tectorigenin-7-O-xylosylglucosideose glycosidase prosthetic regression equation y = 2840528.18 x-29185.93,correlation coefficient r = 0.9999;Tectoridin of regression equation y =4444648.34 x-139596.80,correlation coefficient r = 0.9999;Tectorigeninan equation y =7328041.12 x-61890.82,correlation coefficient r = 0.9998.Methodology validation results: precision,repeatability,stability,all conform to the rules;Tectorigenin-7-O-xylosylglucosideose glycosidase prosthetic prosthetic sample recovery rate was 98.88 ~ 103.44%,RSD was 1.80%;Tectoridin sample recovery rate was 97.26 ~101.19%,RSD was 1.56%;Tectorigeninan sample recovery rate was 98.80 ~ 102.11%,RSD was 1.40%.10 average three components in a batch of flavonoids site of Puerariae lobatal flos partsl parts content: Tectorigenin-7-O-xylosylglucosideose glycosidase prosthetic was 15.78%,Tectoridin was 45.08%,Tectorigeninan was 13.05%.Through the analysis of 10 batches of flavonoids site of Puerariae lobatal flos partsl parts HPLC,established the flavonoids site of Puerariae lobatal flos partsl parts fingerprint,calibration14 common peak,similarity of 0.952 to 0.952.Acute alcohol poisoning experiment results show that: compared with model group,flavonoids site of Puerariae lobatal flos partsl parts drunk prolonged the incubation period of mice(P < 0.05),the sleep time(P < 0.05)and sober time(P < 0.05),and with a certain amount of activity relationship.In a drunken excitement period,flavonoids site of Puerariae lobatal flos partsl parts can strengthen the ability of learning and memory,to extend the time of incubation period to get an electric shock(P < 0.01),reduce the number of electric shock(P < 0.05),and enhance the coordination of the movement of a drunken mice(P < 0.05).Flavonoids site of Puerariae lobatal flos partsl parts ethanol concentration in serum of mice(P < 0.05),but has no effect on alcohol dehydrogenase activity in the liver.Acute alcoholic liver injury pharmacological experimental results show that:compared with model group,flavonoids site of Puerariae lobatal flos partsl parts can effectively reduce the acute alcoholic liver injury in the rat liver index(P < 0.01),lower serum ALT(P < 0.01),the activity of AST(P < 0.01)and liver MDA(P < 0.01)content,make liver damage conditions have different degrees of improvement,that flavonoids site of Puerariae lobatal flos partsl parts protective effect of acute alcoholic liver injury.Chronic alcoholic liver damage the pharmacological experiment results show that the flavonoids site of Puerariae lobatal flos partsl parts can effectively reduce the liver index of chronic alcoholic liver injury in rats(P < 0.01),lower serum ALT(P < 0.01),AST(P <0.01),AKP activity(P < 0.05)and liver(P < 0.01)the content of MDA increased liver SOD(P < 0.01),GSH(P < 0.05)content,reduce the degree of liver injury,that flavonoids site of Puerariae lobatal flos partsl parts can effectively protect caused by chronic alcoholic liver damage.Chronic alcohol liver injury mechanism,the experimental results show that:Flavonoids site of Puerariae lobatal flos partsl parts can significantly inhibit the rat liver mRNA expression of CD68(P < 0.01),reduce the liver of macrophage activation,cut the nf-kappa B mRNA expression(P < 0.01),inhibit inflammation,reduce the inflammatory factors and inflammatory mediators(TNF alpha(P < 0.01),the TGF-beta 1(P < 0.01),IL-6(P < 0.01),IL-8(P < 0.05),the release of inflammatory cells of normal liver cells;reduction Can cut mRNA expression of TGF-beta 1(P < 0.05),the mechanism of the formation of liver cells,to reduce the degree of liver fibrosis,inhibit liver injury aggravated;At the same time increase the NRF-2 mRNA expression(P < 0.01),activation of downstream oxidation,give play to the role of liver protection.Conclusion : This study established a Puerariae lobatal flos partsl parts quality control method,the solution wine to protect liver pharmacodynamics and mechanism were studied,To establish the HPLC fingerprint and the three kinds of chemical composition of Tectoridin,Tectorigenin-7-O-xylosylglucosideose glycosidase prosthetic,Tectorigeninan content has carried on the determination,,through acute alcohol poisoning,acute or chronic liver injury of experimental study the flavonoids site of Puerariae lobatal flos partsl parts protect liver function,by ELISA method and fluorescence quantitative PCR method for flavonoids site of Puerariae lobatal flos partsl parts of the liver protective effect mechanism carries on the preliminary research,to develop leguminous plants flower of the clinical application and development of traditional Chinese medicine(TCM)therapy research provides theory basis.