Study On Extraction, Abstraction, Purification And Bacteriostatic Effect In Vitro Of Alkaloids From Chelidonium

Greater celandine(Chelidonium majusL) is the Papaveraceae chelidonium plant,the alias is wintergreen barberry root,the mountain watermelon.It is grown in the mountain valley moist region,nearby the edge of forest drainage,widely distributed in the Eurasia,places such as North America,mainly distributes in our country in Northeast,North China numerous provinces and so on,the natural resource is extremely rich and abundance.It is first recorded for medicinal purposes in jiu huang ben cao.The crude drug with cool-natured, bitter in taste,slightly poisonous,enters into heart,kidney,liver and lung meridian.The main effective component in Greater celandine is the remarkable biological activity alkaloids material,it including the alkaloid 0.97～1.87%in the entire grass,containing alkaloid 1.9～4.14%in the root. So far it is already accumulated the reported that Greater celandine included twenty-nine kinds of alkaloids belonged to type of benzophenanthrene,gen tropine,bisisoquinoline and aporphines.According to modern pharmacological research,the important active constituent is the type of isoquinoline alkaloid,mainly for chelidonine,chelerythrine,sanguinarin and so on.So far,study on monomer abstraction from Greater celandine deeply,discovered that until now the alkaloid extraction process and the content determination method study report are few about Greater celandine. It is really micro studying on the common bacterial disease of livestock and fowl among the numerous pharmacologicalactions of different extraction from Greater celandine.According to the above,this article study on each committed step for new veterinary medicine,including the alkaloid extraction,concentrates,the purification and the pharmacological action research,to provide some basic theory for a large-scale industry.First,research on the technical of alkaloid extraction,this thesis the assay method of assay method with RP-HPLC determination.Using the orthogonal experimental designed method,taken the evaluating target by the extracting weight and the chelidonine content,analyzed by software SPSS14.0,inspected two kind of alcohol extraction process which are the hot dipping and the ultrasonic wave method When take the extracting weight as goal material,the extraction efficiency of hot lixiviated method surpasses the ultrasonic wave extraction process,and the difference achieves extremely obviously.The condition is 65%alcohol,ratio dosage liquor 1:20,extract temperature 80℃,extract time 2h.When take chelidonine as goal material,the extraction efficiency of the ultrasonic wave extraction process surpasses the hot lixiviated method,and the difference achieves extremely obviously.The condition is 95%alcohol,the ratio of dosage liquor 1:10,the extraction temperature 50℃,extraction time 0.5h,the extraction frequency were 85Hz.Second,study on the method of Concentration.The thesis investigated the adsorption effect of chelidonine with six kind of common maeroporous resin.Respectively type of D101 and AB-8 which are made in china,type of HP-20,HP-2MG,sp-700,sp-850 which are made in Japan.Bolting the most suitable type macroporous resin for purifying chelidonine sp-700.It is prominent of this kind of macroporous resin,first processing and the regeneration are easy.The new resin uses 30BV95%alcohol processing can be used,the extract after crossed the resin can dry easily.After the HPLC examination,mixed peak have been reduced obviously.Third,study on separation and purification.we applied with the classic the traditional Chinese medicine chemistry means to reserach the chemical constituents of Greater celandine.The method of silica gel normal phase column chromatography has been taken,elution system of petroleum benzin and acetone, trichlormethane and MeOH are be used.Obtained two monomer compound,one is accredited chelidonine by MS,¹HNMR,¹³C NMR,IR chromatograph appraisal.Fourth,study on antibacterial effect in vitro of alkaloids from Chelidonium.Choosing the six common pathogenic bacterium,which are Escherichia coli ATCC25922,Staphylococcus aureus ATCC25923, Staphylococcus aureus persister,streptoc ATCC55121 salmonella,Escherichia coli persister,to obtain the six kinds of extraction MIC value.Method of the six different extraction MIC value are:90%alcohol elute the AB-8,60%alcohol to elute the AB-8,the chloroform extraction,the water decoctum extraction,the water level extraction,the n-butanol level extraction.By flat plate double dilution to determin the MIC,the 90% alcohol to elute the AB-8 to Escherichia coli ATCC25922 is 7.8125 mg/mL,Escherichia coli persister 15.625mg/mL.The 60%alcohol to elute the AB-8 to Escherichia coli ATCC25922 is 7.8125mg/mL,Staphylococcus aureus ATCC25923 is 15.625mg/mL,Staphylococcus aureus persister is 31.25mg/mL.Escherichia coli persister is 15.625mg/mL.The medicine MIC value of the chloroform extraction to Escherichia coli ATCC25922,Staphylococcus aureus ATCC25923,Escherichia coli persister is 7.8125 mg/mL,15.625mg/mL,31.25mg/mL,15.625 mg/mL.The medicine MIC value of the water decoctum extraction to Staphyloco ccus aureus ATCC25923,Staphylococcus aureus persister is 7.8125 mg/mL,31.25 mg/mL respectively.