柱层析法分离纯化血橙花色苷  

Isolation and Purification of Anthocyanins from Blood Oranges by Column Chromatography

作　　者：曹少谦[1] 潘思轶[1] 姚晓琳[1] 傅虹飞[1] 

机构地区：[1]华中农业大学食品科技学院,武汉430070

出　　处：《中国农业科学》2009年第5期1728-1736,共9页Scientia Agricultura Sinica

基　　金：农业部农业结构调整重大技术研究专项(04-09-03B);湖北省自然科学基金(2006ABA168);公益性行业(农业)科研专项(nyhyzx07-023)

摘　　要：【目的】研究适合分离纯化血橙花色苷的柱层析法,并对血橙中的花色苷进行初步鉴定。【方法】分别采用12种不同类型树脂对血橙花色苷静态和动态吸附解吸试验进行比较,优化了大孔树脂提取分离血橙花色苷的工艺,采用Toyopearl TSK HW-40S柱层析对血橙花色苷提取物进一步分离纯化。同时,利用高效液相色谱-电喷雾质谱联用(HPLC-ESI/MS)对血橙花色苷进行定性分析。【结果】大孔树脂NKA-9对血橙花色苷的分离效果最好,最佳工艺条件为:50%乙醇用柠檬酸调pH为2.5时洗脱效果最好;ToyopearlTSKHW-40S柱层析分离纯化条件为:35%甲醇(经2%甲酸酸化)为洗脱液,流速0.6ml·min-1,可得到3个单一花色苷组分和1个混合花色苷组分。HPLC-ESI/MS分析表明,血橙花色苷主要是为矢车菊素-3(3″-丙二酰)葡萄糖苷和矢车菊素-3(6″-二乙二酰)葡萄糖苷(组分1),矢车菊素-3-葡萄糖苷(组分2),矢车菊素-3(6″-丙二酰)葡萄糖苷(组分3)及矢车菊素-3-葡萄糖苷加成物4-乙烯基儿茶酚(组分4)。【结论】NKA-9大孔树脂与ToyopearlTSKHW-40S凝胶柱层析相结合,可以有效分离纯化血橙中花色苷,HPLC-ESI/MS分析可以方便、快捷、有效地为花色苷的鉴定提供依据。[ Objective ] Isolation and purification of anthocyanins from blood oranges by column chromatography were made, and then the anthocyanins of blood orange were identified. [ Method] The behavior of static adsorption and desorption, dynamic adsorption and desorption of twelve kinds of resins was compared. Toyopearl TSK HW-40S column was employed to separate and purify the anthocyanin extracts from blood orange. Then, the anthocyanins of blood orange were identified by HPLC-ESI/MS analysis. [Result] The results indicated that NKA-9 macroporous resin was the optimum for the anthocyanins of blood orange purification, and the optimum elution reagent was 50% ethanol with citric acid （pH 2.5）. The best separation of Toyopearl TSK HW-40S column was obtained using a mobile phase of 35% methanol with 2% formic acid at a flow-rate of 0.6 ml·min^-1. Three major anthocyanins of blood orange were isolated. HPLC-ESI/MS analysis results show that the anthocyanins of blood orange are mainly cyanidin-3-glucoside （35.2%） and cyaniding-3-（6＂-malonyl） glucoside （42.9%）. Furthermore, cyanidin-3-（3＂-malonyl） glucoside, cyanidin 3-（6′′-dioxalyl） glucoside and cyanidin-3-glucoside adduct： 4- vinylcatechol were identified in blood orange. [Conclusion] The combination ofNKA-9 macroporous resin and Toyopearl TSK HW-40S column chromatography for separation and purification of blood orange anthocyanins is an effective method, and HPLC-ESI/MS analysis is a convenient, rapid and effective method for identification of anthocyanins from blood orange.

关 键 词：血橙 色素 花色苷 纯化 鉴定 

分 类 号：TQ464.3[化学工程—制药化工]