Extraction And Purification And Biological Activities Of Alkaloid From Thermopsis Lanceolate

According to the study,there are alkaloids,organicacids,flavone,phenolic compounds,polysaccharide and other components of Thermopsis lanceolate.Alkaloids are the main bioactive and toxiferous chemicals.This aspect of the story is rarely covered the extraction process and pharmacology of total alkaloids of Thermopsis lanceolate until now.Accordingly,this artic le not only make a study on alkaloids of extraction separation and identification of total alkaloids of Thermopsis lanceolate,but also accessed to the higher purity of a single alkaloids.The dissertation studies the alkaloids and total alkaloids of Thermopsis lanceolate on the pharmacology.The obtained main contents are as follows:1.Pre-experiment of Chemical Components for Thermopsis lancenolate.To explore the active constituents of Thermopsis lancenolate by preexperiment.The extractives from the different parts of Thermopsis lancenolate in different(purified water,solvent ac idic ethanol,thin layer chromatographic method and diethyl ether)solvent was investigated by the law of test tube and chromatography inc luding various indicators.The results indicate that Thermopsis lancenolate contains alkaloids,glycosides,polysaccharide,flavonoids,sitostanol,triterpenoids and so on.Active ingredients of Thermopsis lancenolate mainly contains alkaloids and flavonoids which existed in root and leaf,and stems.2.Extraction technology of total alkaloids from Thermopsis lancenolate.Total alkaloid is extracted by the hot reflux extraction and utrasonic assisted extraction method of Thermopsis lancenolate with methanol.The results show that the extractives were extracted by 70% hot methanol and the extracting temperature 65?and the extracting time 2h from Thermopsis lanceolata,acidified with 1%Hcl and filtrated to give ac id liquid.After the acid liquid was extracted with chloroform toremoveimpurity,its p H was alkalization by strong ammonia the basic liquid was extracted with methanol-chloroform,ethyl acetate and n-butanol in due order.Results showed that the extraction rate of total alkaloids was 0.66%.The utrasonic assisted extraction method is the ratio of material to solvent 1:20?the extracting temperature 60??the extracting time 1h?the utrasonic power 500 W.Under the condition mentioned-above,the extraction rate of total alkaloids of utrasonic-assisted extraction method was 0.76%.The result of TLC analys is indicated that the methanol-chloroform contained 7 kind of alkaloids,n-butanol part contained 4 kind of alkloids and ethyl acetate part does not contain alkaloids.N-butanol part does not contain minimum polarity alkaloids.3.The isolation by silica gelcolumn chromatography of alldoids from Thermopsis lanceolata.By applying the silica gelcolumn chromatography separation technologies to separate different polarity alkaloids from Thermopsis lanceolata.It is to obtain 2 monomer compound(crysta1? and ?)and alkaloid mixture of A,E,G,Jphase.Crystal?is yellow needle.The result of TLC analysis showed 2 orange spots and was not fluorescent.Crystal?is schistose structure.The result of TLC analys is showed 5 red purple spots and was not fluorescent.4.Analysis of alkaloid from Thermopsis lanceolata By GC-MS.The extractives from the different parts of Thermopsis lancenolate was acidified with 1%HCl and filtrated to give acid liquid.After the ac id liquid was extracted with chloroform toremoveimpurity,its p H was alkalization by strong ammonia the basic liquid was extracted with methanol.Finally,five kinds of alkaloid was retrieved in the maximum polarity from Thermopsis lanceolate were identified by GC-MS.They are Sparteine,Dodecahydro-7,14-methanodipyrido[1.2-a:1,2 '-E] [1,5]Diazocine Argentamin,Cytis ine,Thermopsine.The Dodecahydro-7,14-methanodipyrido[1.2-a:1,2'-E][1,5]Diazocine of Thermopsis lanceolate was very few,occupied only 0.26%.The Sparteine had the highest contents,accounted for 55.61%.5.Purification and Characterization of alkaloid from Thermopsis lanceolata.Crystallization obtained by colum chromatography was collected.Two crytal with high purity were obtained.The chemical structures of the monomer compound were identified by means of GC-MS and TLC analys is.The result showed that the yellow needle of Crystal I is Thermopsine and the schistose structure of Crystal II is Cytisine.In the same conditions of GC-MC analysis tests,Thermopsine and Crystal I's time of maximum concentration are 12.2m.However,Cytis ine 's time of maximum concentration is different from Crystal II.6.The study on toxicity of alkaloids of Thermopsis lanceolataTo study the toxicity of alkaloids of Thermopsis lanceolata and Copy out here in Quinolizidine poisoning pathology model.The results showed that pour mice were died with 200mg/kg of alkaloid from Thermopsilanceolata.As results,LD50 was less than 200mg/kg.Blood biochemical examination of chronic intoxication in mice found that alkaloids of Thermopsis lanceolata make mice's HGB and RBC and WBC and ALT and AST and CK were higher,however mice's BUN were lower.The heart,liver,kidney were appeared degeneration and they were arget organ.7.Effect of Thermopsis lanceolata alkaloids on Pathogenic bacteria of dairycow endometritisTo understand effect of Thermopsis lanceolatas alkaloid on pathogens in vitro of dairy cow endometritis,from dairy cows with clinical puerperal metritis were adopted to isolate and identify pathogenic bacteria.The results showed that the mixed infection of the Esche-richiacoli and hemolytic Streptococcuswere uniquely found.Total alkaloids of Thermopsis lanceolatas had similar antibacterial activity,especially non-hemolytic streptococcus was most sensitive,followed by Salmonella,Esche-richiacoli,hemolytic streptococcus.The Sparteine has no antibacterial activity.8.Effects of Thermopsis lanceolata alkaloid on immune function of miceThe research ami ed to study the effect of Thermopsis lanceolata alkaloid on immune function of mice.the icr mice were divided into three experimental groups given medicament and were managed with same amount of normal saline for 20 days.Effects of Thermopsis lanceolata alkaloid on mice bodyweight and organweight,organ indices,the phagocytosis of enterocoelia macrophage,T lymphocyte counts were observed.Compared with controlgroup,those items of the medicated groups decreased.Under the above conditions,meso and high concentration of concentration Thermopsis lanceolata alkaloid has inhibitory effect on the mi mune function of mice.9.Cytotoxicity experimental study of alkaloids from Thermopsis lanceolataThe camptothecin was used as as a positive control to determinate the cytotoxity of Thermopsis lanceolata and Cytis ine on the normal BHK cells and SW480 cancer cells by using MTT colorimetry.The results showed that Thermopsis lanceolata has inhibition of SW480 to the growth activity.As the dose increased,inhibition rate also increased significantly.Toxicity of camptothecin was rather obvious and both BHK and SW480 in the group had better inhibition.Thermopsis lanceolata and Cytisine had low toxicity to BHK cells and had a certent inhibition to SW480.s...