纯橙汁中类黄酮的RP-HPLC-DAD-ESI-MS^2分离鉴定

Separation and Identification of Flavonoids in Orange Juice by RP-HPLC-DAD-ESI-MS^2

为了建立纯橙汁中类黄酮的快速、灵敏的反相高效液相色谱-电喷雾电离质谱法(RP-HPLCDAD-ESI-MS2)分离鉴定方法。本试验通过对反相高效液相色谱法(RP-HPLC)分离条件进行优化,建立了纯橙汁中类黄酮的RP-HPLC-DAD分离方法。结果显示:6种类黄酮对照品可以在40 min内全部检出,其检出限达到0.02~0.17μg·m L-1;方法的精密度和稳定性检验显示6种类黄酮对照品保留时间和峰面积的相对标准偏差(RSD)均≤4.23%,加标回收率为89.90%~105.90%,且RSD均小于5%。在此基础上,采用RP-HPLC-DAD-ESI-MS2法对纯橙汁样品进行分析,并与对照品进行比较,结果显示:纯橙汁样品中除含有柚皮素-7-β-芸香糖苷、橙皮苷、香蜂草苷、甜橙黄酮、川皮苷、橘皮素外,可能还含有槲皮素-3,7-三葡糖甙、新橙皮苷、芹菜素-6,8-二-C-吡喃葡萄糖苷、香叶木素-6,8-二-C-葡萄糖苷、二-C-葡萄糖基黄酮、香叶木素-6-C-β-葡萄糖苷、柚皮素、橙皮素等8种类黄酮成分。运用此方法可以建立橙汁类黄酮指纹图谱,并为橙汁鉴伪提供借鉴。

The purpose of this study is to establish a rapid and sensitive RP-HPLC-DAD-ESI-MS2 method to separate and identify the flavonoids in orange juice. By optimizing the condition of RP-HPLC,established a RP-HPLC-DAD method for separating flavonoids in orange juice. The results showed that： six flavonoids reference substance could be detected totally in 40 minutes and the detection limits were 0. 02 to 0. 17 μg·m L^-1. Precision and stability test of the method showed relative standard deviation（RSD） of retained time and peak area of six flavonoids reference substance were both below 4. 23%. Recoveries were 89. 90% to 105. 90% and RSD were below 5%. Based on this,the orange juice was analysed by RP-HPLC-DAD-ESI-MS^2 and compared with the reference substance,the result showed the orange juice may contain another eight kinds of flavonoids which included quercetin 3,7-triglucoside,neohesperidin,6,8-di-Cglucopyrano sylapigenin,6,8-di-C-β-glucosyl diosmin,di-C-glucosylflavone,6-C-β-glucosyl diosmin,naringenin,hesperitin besides naringenin-7-β-rutinoside,hesperidin,didymin,sinensetin,nobiletin,tangeretin.We can build orange juice flavonoids fingerprints through this method and provide an experimental basis for the orange juice discrimination