Box-Behnken响应面法优化女贞子总三萜纯化工艺
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篇名: Box-Behnken响应面法优化女贞子总三萜纯化工艺
TITLE: Optimization of the Purification Technology of Total Triterpenoids of Ligustrum lucidum by Box-Behnken Re- sponse Surface Methodology
摘要: 目的:提高女贞子总三萜中齐墩果酸和熊果酸的转移率和纯度,优化该类成分的纯化工艺。方法:以齐墩果酸和熊果酸作为女贞子药材中总三萜类代表成分,采用高效液相色谱法测定两者含量。色谱柱为ThermoBDSHypersilC18,流动相为甲醇-0.02%乙酸铵溶液(80∶20,V/V),检测波长为210nm,柱温为30℃,流速为1.0mL/min,进样量为20μL。采用单因素试验,以齐墩果酸、熊果酸的转移率为指标,考察工艺中的水沉温度、水沉时间和复溶乙醇用量对纯化工艺的影响;以两种成分的转移率和纯度为指标,考察活性炭用量、析晶乙醇体积分数对纯化工艺的影响。在此基础上,以复溶乙醇用量、活性炭用量、析晶乙醇体积分数为因素,采用Box-Behnken响应面法进一步优化该纯化工艺并验证。结果:最佳纯化工艺为女贞子药材提取浓缩液加4倍量(mL/g,下同)水,在0℃静置2h(水沉);然后加入药材1倍量的乙醇溶解(复溶);再加入4%的活性炭(除杂);最后加水调整乙醇体积分数为80%,在4℃下静置12h(析晶),离心,干燥。3次验证试验结果显示,采用该优选纯化工艺所制得的总三萜中齐墩果酸、熊果酸的转移率分别为61.11%、65.78%,纯度分别为53.44%、19.79%,RSD均小于3%。结论:优化所得纯化工艺提取效率高、操作简便,可用于女贞子总三萜类成分纯化的工业化生产以及相关制剂的开发。
ABSTRACT: OBJECTIVE:To i mprove the transfer rate and purity of oleanolic acid and ursolic acid in total triterpenoids from Ligustrum lucidum ,so as to optimize the purification technology. METHODS :Oleanolic acid and ursolic acid were used as representative components of total triterpenoids ,and their contents were determined by HPLC. The determination was performed on Thermo BDS Hypersil C 18 column with mobile phase consisted of methanol- 0.02% ammonium acetate solution (80∶20,V/V)at the flow rate of 1.0 mL/min. The detection wavelength was set at 210 nm,and column temperature was 30 ℃. The sample size was 20 μ L. In single factor tests,using transfer rate of oleanolic acid and ursolic acid as index ,the effects of water precipitation temperature and time ,the amount of redissolved ethanol on the purification technology was investigated ;using transfer rate and purity of two components as indexes ,the effects of the amount of activated carbon and volume fraction of crystallization ethanol were investigated. Based on it ,using the amount of redissolved ethanol and activated carbon ,volume fraction of crystallization ethanol as factors ,Box-Behnken response surface methodology was used to optimize the purification technology ,and validation tests were performed. RESULTS :The optimal purification technology was adding 4-fold(mL/g,the same below )water in L. lucidum concentrated solution ,placing for 2 hours at 0 ℃(water precipitation );adding 1-fold ethanol to dissolve (redissolution); adding 4% activated carbon (edulcoration);finally adding water to adjust the volume fraction of ethanol to 80%,placing at 4 ℃ for 12 hours(crystallization),centrifuging and drying. The results of 3 times of validation tests showed that the transfer rates of oleanolic acid and ursolic acid in total triterpenoids prepared by optimized technology were 61.11% and 65.78%,the purities of them were 53.44% and 19.79%,and RSDs were both lower than 3%. CONCLUSIONS :The optimized purification technology has high extraction efficiency and simple operation ,which can be used for industrial production of purification of total triterpenoids from L. lucidum and the development of corresponding preparations.
期刊: 2020年第31卷第18期
作者: 高爽,王玉真,薄福民,张丽,孙越,杨山景,封安杰,李凌军
AUTHORS: GAO Shuang ,WANG Yuzhen ,BO Fumin ,ZHANG Li,SUN Yue,YANG Shanjing ,FENG Anjie,LI Lingjun
关键字: 女贞子;总三萜;齐墩果酸;熊果酸;纯化工艺
KEYWORDS: Ligustrum lucidum ;Total triterpenoids ;Oleanolic acid ;Ursolic acid ;Purification technology
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