不同道地产区木瓜指标成份分析及川木瓜总黄酮提取纯化工艺研究

【作者】 俞佳；

【导师】 刘友平；

【作者基本信息】 成都中医药大学 ， 药物化学， 2007， 硕士

【摘要】 木瓜为《中国药典》2005年版（一部）收载品种,是蔷薇科植物贴梗海棠Chaenomeles speciosa（Sweet）Nakai的干燥近成熟果实。味酸、温,归肝、脾经。具有平肝舒筋,和胃化湿的功效。传统认为安徽宣城、浙江、湖北所产木瓜为道地药材,质量最好。川木瓜在全国产量最大,关于川木瓜质量研究的报道较少,为了评价川木瓜药材质量,研究其与道地产地药材质量的区别,本研究对不同产地木瓜药材指标成份进行分析,为其开发、利用提供一定的依据;木瓜含有多种黄酮类化合物,提取川木瓜总黄酮,不仅为黄酮类物质的提取提供新的材料,也可为川木瓜产业寻找到新的开发点,为进一步进行其药理研究奠定基础。为此本课题以三萜酸和有机酸作为指标成份对川木瓜及其他产地的木瓜进行了分析比较研究;同时对川木瓜总黄酮的提取、纯化工艺进行了研究主要研究。结果如下:一、不同产地木瓜指标成份比较分析1.有机酸的含量比较研究采用电位滴定法测定了不同产地木瓜中总有机酸的含量,结果除产自广元的含量较低外,其余川木瓜总酸含量都在6%以上,与宣木瓜含量接近,说明川木瓜中有机酸的含量较高。不同产地木瓜药材中苹果酸和柠檬酸含量的测定。首次建立了RP-HPLC法同时测定木瓜中苹果酸和柠檬酸含量的方法。苹果酸的进样量在0.426～2.982ug范围内峰面积值与进样量之间线性关系良好,回归方程为Y=18*10⁶X+65895,相关系数为r=0.9995,平均回收率为99.84%;柠檬酸的进样量在0.164～1.148μg范围内与峰面积积分值呈良好的线性关系,回归方程为Y=918764X-59076,相关系数为0.9992,平均回收率为99.71%。该方法具有良好的稳定性、重现性和准确性,可作为检测木瓜中苹果酸和柠檬酸含量的方法。采用此法对收集样品的含量进行测定,结果川木瓜和宣木瓜中这两种成份的含量总和都在11mg/g以上。2.三萜酸的含量测定采用RP-HPLC法对齐墩果酸、熊果酸含量进了测定,结果表明不同产地样品的含量差异较大。3.总黄酮的含量测定采用比色法测定了不同产地药材中总黄酮的含量。以NaNO₂-AlCl₃显色,检测波长为510nm,以芦丁为对照品,在4.64～27.84ug/ml范围内对照品的量与吸收度呈良好的线性关系。回归方程为Y=1.0998X-0.0073,相关系数为0.9991,回收率为97.84%。该方法具有较高的回收率,稳定性、精密度和重复性较好。应用所建立的方法对不同产地木瓜药材中总黄酮含量进行测定,得出目前市售不同产地木瓜中黄酮的含量有一定的差异。其中川木瓜和宣木瓜的含量相近。4.对不同产地的药材进行了一般检查,结果表明药材都符合药典规定标准。二、川木瓜总黄酮的提取纯化研究1.优化了川木瓜总黄酮提取工艺。提取工艺为:采用回流提取法,乙醇浓度为80%、固液比1:10、提取3次、每次1h。2.研究了大孔吸附树脂纯化川木瓜总黄酮的工艺参数。选用D₁₄₀大孔吸附树脂,吸附条件为:药液浓度0.1g/mL、药液pH值为5、上样速度为1BV/h;洗脱条件为:蒸馏水、10%乙醇、50%乙醇溶液各3BV,以2BV/h的流速依次洗脱,收集50%洗脱部位,回收乙醇,浓缩,真空减压干燥得精制木瓜总黄酮。采用该工艺,产品纯度可达到52.13%。更多还原

【Abstract】 Mugua is the dry and mature fruits of the Rose family plant chaenomeles（Sweet） Nakai,recorded in the China pharmacopoeia（2005 edition）.It is a common Rheumatism Chinese medicine,which is sour in flavor,warm in nature,and attributive to the liver and spleen meridians.It is thought that the quality of mugua which from Anhui,Zhejiang,Hubei are the best.Sichuan has it largest production,but the quality research reports about Chuan mugua are very few.In order to appraise the quality of Chuan mugua,this research carries on the analysis to the different habitat mugua raw material for medicine target ingredient.Mugua includes the many kinds of flavonoids,extract total flavanone from it,not only for the flavonoids matter provides the new material,also may seek for the Chinese flowering quince industry to the new developmentspot.For further conducts its pharmacology research to lay the foundation.The comparison of organic acid and the triterpenoids acid between Chuan mugua and other different soursed mugua were researched.The extraction,purification of flavonoids in Chuan mugua was researched too.The main results are as follows:1.Different sourced mugua ingredients comparative analysis.The content of total organic acid of different sourse mugua were researched.The result is that the content of different Chuan mugua samples were all above 6%,and the result is to closede to Xuan mugua,except one sample which from Guanyuan.For the first time,The RP-HPLC was established for determinating the content of malic acid and citric acid in mugua simultaneously.There was good linearity between peak areas and injection quantity of malic acid range from 0.426～2.982ug and citric acid range from 0.164～1.148ug.The average recovery rate was 99.84%for malic Acid and 99.71%for citric acid.The meathod was reasonable,stable and accurate. The result of determination can be applied to judge the quality of mugua.The two kind of ingredients content of eight different sourced mugua were determined.The result is that the kind of ingredients content sum total all above 11mg/g. Ursolic acid and oleanolic acid as reference substance were determined by RP-HPLC method.The result is that the content of different samples are different.A spectrophotometric method was established for quantitative determination the total content of flavonoids in chaenomeles（Sweet） Nakai.The result showed that the optimal determination wavelength was 510nm.There was a good linear relationship between the absorbance and the quantity of rutin when the concentration of rutin（contrast） within the range of 4.64ug～27.84ug/ml.Regression equation was Y=1.0998X-0.0073 with the correlation coeffcient of 0.9991.The average recovery rates were 97.84%.This method is stable and precise as well as high recovery rates. We determine the total flavonoids content of eight different sourced mugua.The result is that the content of total flavonoids from Sichuan is as much as Anhui.4.The general inspection to the different habitat raw material for medicine has been carried,finally indicated raw material for medicine all conforms to the pharmacopoeia stipulation standard.2.The extraction,purification of flavonoids in Chuan mugua.The extraction process of the total flavonoids in Chuan mugua was investigated by single factor and orthogonal experiment.The optimum condition for the extraction of the total flavonids in it was that alcohol concentration is 80%;temperature is 60C;extraction time is three and one hour of each and the ratio of solid to liquid is 1:10.The column Chromatography was employed to enhance the content of the total flavonoids.In terms of the result of static adsorption and static de-adsorption test, D₁₄₀ resin was the best one for isolation flavonoids.The optimal adsorption parameters were feeding rate 1 BV/h,pH value 5～6,medicine liquid concentration 0.1mg/ml.The optimal de-adsorption parameters were eluted with distilled water, 3BV 10%ethanol and 3BV 50%ethanol in turn,flow rate 2BV/h.The purified total flavonoids was collected in 50%ethanol eluting solvent.更多还原