超声-酶辅助双水相提取枳壳黄烷酮工艺的建立与优化

Establishment and optimization of the ultrasonic-enzyme assisted aqueous two-phase extraction for obtaining the flavanones from Zhiqiao(Fructus aurantii)

枳壳是我国柑橘属常见的药食同源植物,所含丰富的黄烷酮即新橙皮苷和柚皮苷,是其主要的药理成分。为提高枳壳新橙皮苷和柚皮苷的提取分离效率,该研究采用响应面法优化了超声辅助双水相提取的最佳条件,并进一步建立超声-酶辅助双水相法(ultrasonic-enzyme assisted aqueous two-phase extraction,UEA-ATPE)提取工艺和制备液相色谱分离纯化方法。结果表明,(NH_(4))_(2)SO_(4)/乙醇为最佳双水相体系,建立并优化的UEA-ATPE最佳工艺条件为乙醇质量分数27%,(NH_(4))_(2)SO_(4)质量分数21%,超声功率137 W,复合酶质量比为中性蛋白酶∶果胶酶=1∶1。在此条件下,新橙皮苷和柚皮苷的得率分别为(92.27±2.13)、(72.24±1.56)mg/g,优于水煎煮法、50%乙醇浸提法、超声辅助95%乙醇提取法和超声辅助双水相法,同时此新方法提取液中杂质的种类和含量更少。提取液经制备型液相色谱进一步分离纯化后,得到了纯度为99.37%的新橙皮苷。该方法具有提取效率高、绿色环保的优点,有助于枳壳作为保健食品的精深开发利用,提升其经济附加值。

Zhiqiao(Fructus aurantii)is a common medicinal and edible plant of citrus species in China.It contains abundant flavanones,namely neohesperidin and naringin,which are the major pharmacological components.To improve the extraction and separation efficiency of neohesperidin and naringin from Zhiqiao,the optimal conditions of the ultrasonic-assisted aqueous two-phase extraction using response surface methodology were explored,and a further ultrasonic-enzyme assisted aqueous two-phase extraction(UEA-ATPE)technique and a preparative liquid chromatography were established.The results showed that the optimal conditions of UEA-ATPE to obtain flavanones from Zhiqiao were as follows:mass fraction of ethanol,27%;mass fraction of(NH_(4))_(2)SO_(4),21%;ultrasonic power,137 W;the ratio of neutral protease:pectinase=1∶1(mass ratio)in the complex enzymes.The yield of neohesperidin and naringin reached(92.27±2.13),(72.24±1.56)mg/g,respectively,which was better than that of water decoction,extraction using 50%ethanol,ultrasonic-assisted extraction using 95%ethanol and ultrasonic-assisted aqueous two-phase extraction,while the composition and content of unwanted extracts by the new method were less.The extracts were finally separated and purified using a preparative liquid chromatography and the purity of neohesperidin was 99.37%.The method has the advantages of high extraction efficiency and greenness,which is helpful for the deep development and utilization of Zhiqiao as a health food and improving its economic added value.