Study On Extraction Process Of Main Alkaloids And Enrichment Process Of Cephalotaxine From Cephalotaxus Hainanensis Li

Hainan is known as the "natural medicine library".Cephalotaxus hainanensis Li is a rare medicinal plant to tropical zones in China and is a endemic species in Hainan.The number of cephalotaxus esters from Cephalotaxus hainanensis Li is the largest(11 species in total)and the content of cephalotaxus esters is the highest in the same genus,especially four kinds of cephalotaxus esters with significant anti-tumor effect on animals and clinics.These alkaloids are extremely scarce and expensive in the market as active pharmaceutical ingredients(APIs)or important API intermediates.It is urgent to resolve the contradiction between supply and demand of APIs.Therefore,it is of great practical significance to obtain high-content Cephalotaxine,and to enrich them,and then to obtain first-line anti-tumor drugs with high activity such as Homoharringtonine by semi-synthesis,which can fundamentally solve the shortage of APIs.Based on its significant medicinal and economic value as a natural source of anti-tumor drugs,this thesis focused on the extraction and enrichment process and separation process of alkaloids from Cephalotaxus hainanensis Li.In this thesis,based on the establishment of a method for determining the content of three alkaloids,the extraction process of main alkaloids from Cephalotaxus hainanensis Li was studied,and the enrichment process of Cephalotaxine was studied by single factor method.An isolation and purification method of Cephalotaxine was established by preparative high performance liquid chromatography and the cephalotaxine monomer was separated and prepared by this method.The main research contents were as follows:1.A method for the determination of three alkaloids from Cephalotaxus hainanensis Li was developed by ultra performance liquid chromatography-mass spectrometry technique(UPLC-MS).The linear relationship and methodological test showed that the method was accurate and reliable,which could be used for the determination of the content of three alkaloids.2.The main alkaloids from Cephalotaxus hainanensis Li were extracted by ultrasonic-assisted method.The extraction process of main alkaloids from Cephalotaxus hainanensis Li was optimized and determined by single-step factor screening and response surface optimization.Single-step factor screening was carried out on 8 factors that affected the extraction effect of main alkaloids from Cephalotaxus hainanensis Li,including the type of solvents,the changes in p H,the particle size of raw materials,the extraction times,the extraction time,the extraction temperature,the material to liquor ratio,and the ultrasonic power.After investigation,the optimal value range of each factor was determined.Taking the material to liquor ratio,the extraction time,and the extraction temperature as the investigation factors,and taking the content of three alkaloids as the evaluation indicator,the optimal extraction process parameters were determined by response surface optimization.The optimal extraction process parameters were as follows:the extraction solvent was sulfuric acid with p H 1;the particle size of raw materials was 10 mesh;the material to liquor ratio was 1:70(g:m L);the extraction time was60 minutes;the extraction temperature was 50 degrees Celsius;the ultrasonic power was 480 watts;and the number of extraction was four times.3.Based on the consensus that some of the cores of cephalotaxus esters are the same structure as Cephalotaxine,the ester cleavage of carbon-oxygen single bond was carried out under different alkaline conditions for the enrichment of Cephalotaxine.The four factors were investigated by single-step factor screening,including the type of bases,the reaction time,the concentration of the base,and the reaction temperature which affected the enrichment efficiency.And the optimal enrichment process parameters of Cephalotaxine were as follows: the reaction time was 10 minutes;the reaction temperature was the room temperature(28 ℃);the base used in the reaction was KOH;and the concentration of potassium hydroxide solution was 0.1 mol/L.4.An isolation and purification method of Cephalotaxine was established by preparative high performance liquid chromatography.And the cephalotaxine monomer was separated and purified by this method.