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建立固相萃取(solid phase extraction,SPE)前处理-高效液相色谱串联二极管阵列和荧光检测器(high performance liquid chromatography with diode array detector/fluorescence detector,HPLC-DAD/FLD)快速检测甜橙汁中9种多甲氧基黄酮(polymethoxyflavones,PMFs)的方法。橙汁经粗滤后过C18固相萃取柱,依次以超纯水、30%乙腈清洗除杂,乙酸乙酯洗脱,氮气吹干后用甲醇溶解,制得橙汁PMFs样品。以0.05%磷酸、甲醇、50%四氢呋喃为HPLC流动相进行反相梯度洗脱,利用DAD、FLD同时获得各物质的紫外和荧光光谱及峰面积,从而对样品PMFs做定性和定量检测。该条件下,9种PMFs在10 min内实现快速分离。采用外标法定量,紫外检测波长330 nm,荧光激发和发射波长分别为340 nm和450/500 nm,各物质线性关系良好(R2>0.999),紫外和荧光定量限分别为4.00~61.2μg/L和0.024~0.61 mg/L(对原果汁),橙汁PMFs加标回收率为96.4%~104.2%(紫外)和94.2%~102.1%(荧光),两种检测器定量结果具有良好的一致性。该方法前处理简便、耗时短、成本低,适合橙汁中多甲氧基黄酮成分的快速检测。按本方法对11个汁用甜橙品系鲜榨汁进行了分析,结果显示“渝早橙”PMFs含量最高,具备进一步开发的优良潜力。
A rapid solid phase extraction (SPE) was developed for the rapid determination of 9 in sweet orange juice by HPLC-DAD / FLD (high performance liquid chromatography with diode array detector / fluorescence detector) A method of polymethoxyflavones (PMFs). The orange juice was coarsely filtered and then passed through a C18 solid phase extraction column, followed by ultrapure water, 30% acetonitrile for impurity removal, ethyl acetate elution, and dried in nitrogen and then dissolved in methanol to prepare a sample of orange juice PMFs. Reversed phase gradient elution was carried out with 0.05% phosphoric acid, methanol and 50% tetrahydrofuran as mobile phase of HPLC. The UV and fluorescence spectra and peak area were obtained simultaneously by DAD and FLD. Qualitative and quantitative detection of PMFs was performed. Under these conditions, nine PMFs were rapidly separated within 10 min. The UV detection wavelength was 330 nm. The fluorescence excitation and emission wavelengths were 340 nm and 450/500 nm respectively. The linearity of each material was good (R2> 0.999). The UV and fluorescence quantification limits were 4.00 ~ 61.2 μg / The recoveries of spiked PMFs ranged from 96.4% to 104.2% (UV) and from 94.2% to 102.1% (fluorescence) in the range of 0.024-0.61 mg / L for the original juice. The quantitative results of the two detectors were in good agreement . The method has the advantages of simple pretreatment, short time-consuming and low cost, and is suitable for the rapid detection of polymethoxyflavone in orange juice. According to this method, the fresh juices of 11 juices with sweet orange were analyzed. The results showed that the content of PMFs of Yu Ju Ju Orange was the highest, which possessed the excellent potential for further development.