花椒游离脂肪酸的微波快速提取与衍生化HPLC荧光检测方法研究

建立了花椒中游离脂肪酸的微波快速提取技术,并采用苯并[b]吖啶酮-5-乙基对甲苯磺酸酯（BAETS）作为柱前衍生试剂,建立了19种长链饱和与不饱和游离脂肪酸的衍生化HPLC荧光检测分析方法。微波提取功率700W,溶剂为体积比1∶1的氯仿/甲醇,70℃提取10min,可达近100%提取率。采用色谱柱Eclipse XDB-C8（4.6mm×150mm,5μm）,酸性乙腈/水流动相进行梯度洗脱,HPLC荧光激发和发射波长分别为273nm和505nm。同时进行在线质谱辅助鉴定（APCI-MS）,更提高了HPLC荧光定量的准确度。结果表明,微波提取快速高效,分析方法线性良好,线性范围14.286～0.0139μmol/L,最低检出限为0.636nmol/L,回收率在87.60%～106.59%之间。本方法快速、准确、高灵敏,可作为花椒相关样品的标准化检测方法。 

Microwave rapid extraction of free fatty acids from Zanthoxylum bungeanum Maxim and their HPLC fluorescence detection by pre-column derivatization

Free fatty acids (FFAs) in Zanthoxylum bungeanum Maxim were extracted by microwave-assisted extraction and derivatized using 2- (12-oxobenzo[b]acridin-5 (12H) -yl) ethyl-4-toluenesulfonate (BAETS) as derivatization reagent.This method was proposed to detect 19 long-chain saturated and unsaturated FFA derivatives by HPLC fluorescence detection using pre-column derivatization.Microwave-assisted extract percentages almost reached 100% when FFAs were extracted at microwave power 700W, 70℃ and 10min using chloroform/methanol (1∶1, v∶v) .Good chromatographic separation was achieved using acidic acetonitrile/water as mobile phase on Eclipse XDB-C 8 column (4.6mm×150mm, 5μm) by gradient elution.On-line APCIMS identification was used for producing more accurate HPLC fluorescence quantitation.Fluorescence excitation and emission wavelengths for HPLC detection were 273nm and 505nm, respectively.The results indicated that microwave-assisted extraction was fast and efficient, and the developed analytical method had good linearity.Linear range was from 14.286 to 0.0139μmol/L, the lowest detection limits was 0.636nmol/L, the recovery was in the range of 87.60% ～106.59% .The established method was rapid, accurate, and highly sensitive, it can be used as standardized detection method for free fatty acids.