高效液相色谱-荧光检测器法同时测定小麦粉中L-抗坏血酸和D-异抗坏血酸

Simultaneous Determination of L-ascorbic Acid and D-isoascorbic Acid in Wheat Flour by High Performance Liquid Chromatography-Fluorescence Detector

建立了灵敏快速的高效液相色谱-荧光检测器法(High Performance Liquid Chromatography-Fluorescence Detection,HPLC-FLD),检测小麦粉中L-抗坏血酸与D-异抗坏血酸含量。样品中的L-抗坏血酸和D-异抗坏血酸采用偏磷酸-乙酸溶液提取,经活性炭催化脱氢后,与邻苯二胺衍生成具有强荧光性的物质。以20 mmoL/L磷酸氢二钠水溶液和乙腈体积比90︰10为流动相,经亲水性Waters T;色谱柱分离,用配备荧光检测器的高效色谱仪进行测定。结果表明,L-抗坏血酸与D-异抗坏血酸在0.1~10.0 μg/mL线性良好(R;≥0.999),检出限和定量限分别为0.7 mg/kg和2.0 mg/kg。以空白样品为基体,在2.22 mg/kg、22.20 mg/kg和222.00 mg/kg这3个浓度添加水平下进行加标回收实验,平均添加回收率为93.5%~101.0%,相对标准偏差≤1.73%(n=6)。该方法具有操作简便、灵敏度高、精密度好及准确性高的优点,成功用于市售11种小麦粉中L-抗坏血酸和D-异抗坏血酸含量的测定。

A sensitive and rapid High Performance Liquid Chromatography-Fluorescence Detector (HPLC-FLD) method was established to detect the content of L-ascorbic acid and D-isoascorbic acid in wheat flour.The L-ascorbic acid and D-isoascorbic acid in the sample were extracted with metaphosphoric acid-acetic acid solution,and then catalyzed dehydrogenation by activated carbon,and then derivatized with o-phenylenediamine into substances with strong fluorescence.Under the condition that the volume ratio of 20 mmoL/L disodium hydrogen phosphate aqueous solution and acetonitrile is 90∶10 as the mobile phase,it was separated on a hydrophilic Waters T3 chromatographic column,and measured with a high-performance chromatograph equipped with a fluorescence detector.The developed HPLC-FLD method exhibited good linearity (R;≥0.999) over the concentration range of 0.1~10.0 μg/mL,low limits of detection (LOD) and limits of quantification (LOQ) of 0.7 mg/kg and 2.0 mg/kg,respectively.The average recovery rate of L-ascorbic acid and D-isoascorbic acid under three addition levels of 2.22,22.20,and 222.00 mg/kg ranged between 93.5%~101.0%,satisfactory precision with relative standard deviations ≤1.73%(n=6).The method has the advantages of simple operation,high sensitivity,good precision and high accuracy,it has been successfully used for the determination of L-ascorbic acid and D-isoascorbic acid in 11 kinds of wheat flour on the market.