响应面法优化救必应提取纯化工艺及其治疗肝炎的研究

Optimization of Extraction and Purification Process of Ovateleaf Holly Bark by Response Surface Methodology and Its Hepatoprotective Effects on CCl_4-induced Liver Injury

以救必应(植物铁冬青Ilex rotunda Thunb的干燥树皮)为原料,研究了救必应的乙酸乙酯与乙酸丁酯提取及聚酰胺树脂纯化工艺。以救必应提取物重量得率为评价指标,对救必应提取时间、固液比、乙酸乙酯与乙酸丁酯提取液的混合比例、生药量与聚酰胺的混合比例、拌药与未拌药的聚酰胺混合比例、洗脱液浓度共6个因素进行单因素试验,并结合响应面分析对提取纯化工艺进行优化。结果表明:乙酸乙酯-乙酸丁酯混合液的体积比为1∶2,药材生药量∶聚酰胺重量为3∶1,提取时间为124 min、固液比为1∶21(g∶m L)、洗脱液浓度为39%、拌药的聚酰胺∶未拌药的聚酰胺为1∶3时,救必应提取物的得率达到最大,为95. 41%,达到预测值的99. 63%。采用小鼠急性肝损伤模型对救必应提取物的保肝、抗炎作用进行研究。结果显示,救必应提取物对CCl_4诱导的肝脏肿大有明显地改善效果;救必应提取物能降低小鼠血清中谷丙转氨酶(ALT)和谷草转氨酶(AST)的活性(P<0. 01),明显抑制丙二醛(P<0. 05)的生成,同时降低炎性介质关键酶环氧化酶-2(P<0. 05)的表达,从而抑制炎性介质的合成。因此认为救必应提取物具有保肝、抗炎的作用,具有较好的开发应用前景。

The ethyl acetate-butyl acetate extraction and polyamide resin purification of ovateleaf holly bark were investigated in this work. The independently investigated parameters of the extraction process included extraction time,solid-to-liquid ratio,mixing ratio of extract,mixing ratio of polyamide and crude drug,polyamide ratio of mixed medicine and non-mixed medicine,and eluent concentration. The optimized extraction conditions were determined with the aid of response surface analysis. Results indicated that the extraction yield of ovateleaf holly bark was up to 95. 41% under the optimized conditions: extraction time 124 min,solid-to-liquid ratio at 1 ∶ 21 ( g ∶ mL) ,mixing ratio of extract at 1 ∶ 2,mixing ratio of polyamide and crude drug at 3 ∶ 1,polyamide ratio of mixed medicine and non mixed medicine at 1 ∶ 3. To determine the liver protection and anti-inflammation of ovateleaf holly bark extract,we used the established acute liver injury mouse mode. Results showed that significant inhibitory effect on liver injury was observed,and the inhibition rate was up to 86. 09%. The changes in tissue sections also indicated protective effect of liver. Results also showed a significant reduction of ALT and AST activity ( P<0. 01) and the formation of MDA and the expression of COX-2 were significantly suppressed. These data suggest that the ovateleaf holly bark extract has the functions of liver protection and anti-inflammation,and good prospects for development and application.