【作者】 刘婷；

【导师】 梁宗锁；

【作者基本信息】 西北农林科技大学 ， 药用植物学， 2010， 硕士

【摘要】 以采自镇平县农科所五年生味连上的叶片与须根为试验材料,以紫外分光光度法测得的总生物碱含量作为考察指标,通过单因素试验和正交试验设计对影响乙醇回流提取法和纤维素酶解提取法提取效率的因素及水平进行了研究,同时考察了D101大孔树脂吸附法富集纯化黄连总生物碱的工艺条件。本试验旨在在试验室水平范围内找出从黄连叶片、须根中提取小檗碱的最佳条件及使用大孔树脂纯化黄连生物碱的工艺条件,为开发黄连叶片与须根、提高黄连综合利用水平提供参考。取得主要结论如下:1.传统上利用很少但生物量比例很大的黄连叶片及须根中含有相当含量的生物碱,在最佳提取条件下黄连叶片和须根中总生物碱得率分别为5.48%和7.1%,其中小檗碱含量占黄连总生物碱的一半左右,因此其具有潜在的开发价值。2.通过单因素试验和正交试验设计确定了黄连叶片的最佳回流提取条件为加入10X体积的70%乙醇回流提取3次,每次1.5h,生物碱得率为5.48%;而须根的最佳回流提取条件为加入12X体积的80%乙醇,回流提取3次,每次2.0h,生物碱得率为7.1%。3.通过单因素试验和正交试验设计确定了黄连叶片的最佳纤维素酶酶解条件为加入样品质量2.0%的纤维素酶粉和10X体积的pH=2.6的缓冲液后在50℃水浴中反应150min;须根的最佳纤维素酶解条件为加入样品质量1.5%的纤维素酶粉和10X体积的pH=2.6的缓冲液后在55℃水浴中反应120min。从中得到的黄连生物碱得率分别为4.83%和6.01%。4.无论用哪种方法提取,叶片与须根比较生物碱含量都较低,说明该种黄连的叶片中所含生物碱比须根中生物碱少。两种提取方法比较,无论材料是叶片还是须根都表现出乙醇回流法提取更彻底,生物碱得率更高,但是该法耗时长,且耗用了更多的试剂;而酶法省时、消耗少,生物碱得率相对较低,但这并不能完全说明酶解法不适应于黄连叶片及须根中生物碱的提取。5.考察了非极性大孔树脂D101富集纯化黄连生物总碱的条件。试验结果表明,用D101大孔树脂纯化黄连生物碱的最佳条件为:0.0555g生物碱/g湿树脂的上样量,上样流速为1BV/h时具有最好的吸附效果,而用pH>12的30%乙醇以2BV/h的流速冲洗柱子时,能将更多的生物碱冲洗下来,在该条件下,黄连总生物碱回收率达到83.72%,结合重结晶法可以得到更高纯度的盐酸小檗碱。更多还原

【Abstract】 Single factor experiments and orthogonal tests were used to secelect the optimum conditions of ethanol extraction and enzyme extraction from the leaves and fibrous roots of Coptis chinensis which had lived for five years.The test materials were from Zhenping agricultural science research institute and the extraction amount of alkaloid which detected by UV spectrophotometry was used as a marker.At the same time ,the process conditions of enrichment of coptis alkaloids by D101macroporous resin were studied.The purpose of the study was to find the best conditions of extraction and purification of coptis alkaloids from the leaves and fibrous roots of Coptis chinensis in laboratory and the data should be guidance for enhancing the level of comprehensive utilization of Coptis chinensis. Obtains the main conclusion to be as follows:1.The leaves and fibrous roots of Coptis chinensis which were unutilized have large proportion of biomass.And the yield rate of alkaloids from the leaves and fibrous roots of Coptis chinensis were 5.48% and 7.1% under optimum extraction conditions.More than 50% of the alkaloid was berberine,so the leaves and fibrous roots of Coptis chinensis should be exploited.2.The best extraction conditions of reflux extraction for leaves was: the solid-liquid ratio is 10,the ethanol concentration is 70%, the extraction time was 1.5 hours, and the extraction times were 3 times;and the fibrous roots’was: the solid-liquid ratio is 12,the ethanol concentration is 80%, the extraction time was 2 hours, and the extraction times were 3 times.The yield rates of coptis alkaloids were respectively 5.48% and 7.1%.3.The best extraction conditions of enzyme extraction for leaves was:enzyme amount is 2.0%,the pH of buffer is 2.6 ,enzymolysis time was 150min in water bath which temperature was 50℃;and the fibrous roots’was: enzyme amount is 1.5%,the pH of buffer is 2.6 ,enzymolysis time was 120min in 55℃water bath . The yield rates of coptis alkaloids were respectively 4.83% and 6.01%.4.Compared with leaves ,the fibrous roots of Coptis chinensis has more alkaloid no matter which extraction method was used.Ethanol reflux extraction has higher extraction efficiency but need more time and solvent.Enzymatic hydrolysis used less time but has lower extraction efficiency. Ethanol reflux extraction was better than enzymatic hydrolysis in extracting alkaloid from the leaves and fibrous roots of Coptis chinensis can not concluded by these experiments at all.5.The enrichment conditions of coptis alkaloids by D101 macroporous resin were studied,the result showed that the best accumulation conditions were:the adsorption valus is 0.055526g/g(alkaloid/macroporous resin), sample flow rate is 1BV/h,the eluent is pH>12 30% ethanol with a 2BV/h elution speed.83.72% alkaloid was recovered under these conditions.Highly purified berberine should obtained by recrystallizing after enrichment by macroporous resin.更多还原