两种同时检测茶叶中赭曲霉毒素A与桔霉素方法的比对

Comparison of 2 methods for the simultaneous detection of ochratoxin A and citrinin in tea

中文关键词:  茶叶  赭曲霉毒素A  桔霉素  适配体

英文关键词:tea  ochratoxin A  citrinin  aptamer  ultrafiltration

中文摘要:

      目的 对比两种同时检测茶叶中赭曲霉毒素A (ochratoxin A, OTA)与桔霉素(citrinin, CIT)的方法。方法 考察不同提取方法、溶剂、料液比的提取效果, 比较适配体结合超滤法与免疫亲和柱纯化效果; 对比高效液相色谱-荧光检测器(high performance liquid chromatography-fluorescence detector, HPLC-FLD)与超高效液相色谱-三重四极杆质谱法(ultra performance liquid chromatography-tandem mass spectrometry, UPLC-MS/MS)测定OTA与CIT的检出限、精密度、准确度等差异, 评价方法的适用性。结果 采用70%甲醇超声与振荡的提取方法, 在料液比为1:3的条件下, OTA、CIT的提取率最佳, 回收率分别为99.49%的92.77%; 经适配体结合超滤纯化后, CIT与OTA的回收率为82.07%~92.95%, 与免疫亲和柱纯化方法效果类似; UPLC-MS/MS测定CIT和OTA的基质效应(20.0%、?20.0%)小于HPLC-FLD测定CIT和OTA的基质效应(48.8%、50.3%), 二者测定CIT与OTA均具有良好线性度, 相关性系数均不小于0.9989; HPLC-FLD测定CIT与OTA的检出限分别为0.09 μg/kg、0.12 μg/kg; UPLC-MS/MS测定CIT和OTA的检出限分别为0.12 μg/kg、0.09 μg/kg, 两种方法的加标回收率分别为84.20%~100.70%与89.22%~97.88%, 相对标准偏差分别为1.30%~4.31%与0.74%~4.04%。结论 适配体结合超滤纯化方法操作简单, 成本低廉, 免疫亲和柱纯化方法成本较高且低浓度下的准确度高; HPLC-FLD和UPLC-MS/MS均适用于茶叶基质中CIT与OTA的提取与检测, 但后者基质效应更低, 定性更准确。

英文摘要:

      Objective To compare 2 methods for the simultaneous detection of ochratoxin A (OTA) and citrinin (CIT) in tea. Methods The effects of different extraction methods, solvents, and sample-to-solvent ratios on extraction efficiency were investigated, the binding of molecularly imprinted superfiltration was compared with immunoaffinity column purification. The limits of detection, precision, accuracy and other differences between high performance liquid chromatography-fluorescence detector (HPLC-FLD) and ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) in determining OTA and CIT were evaluated to assess the applicability of the detection methods. Results When the ratio of solid to liquid was 1:3, the best extraction rates of OTA and CIT were obtained by 70% methanol ultrasonic and oscillating extraction method, and the recoveries were 99.49% and 92.77%, respectively. After purification by aptamer-binding ultrafiltration, the recoveries of CIT and OTA were 82.07%?92.95%, which were similar to the results obtained using immunoaffinity column purification alone. The matrix effects of UPLC-MS/MS for quantitation of CIT and OTA (20.0%, ?20.0%) were lower than HPLC-FLD (48.8%, 50.3%). Both methods showed good linearity for CIT and OTA, with correlation coefficients not less than 0.9989. The limits of detection for CIT and OTA by HPLC-FLD were 0.09 μg/kg and 0.12 μg/kg, respectively. The limits of detection for CIT and OTA by UPLC-MS/MS were 0.12 μg/kg and 0.09 μg/kg, respectively. Spike recoveries ranged from 84.20%?100.70% with relative standard deviations of 1.30%?4.31% for HPLC-FLD and 89.22%?97.88% with relative standard deviations of 0.74%?4.04% for UPLC-MS/MS. Conclusion Aptamer combined with ultrafiltration purification method is simple and cost-effective, immunoaffinity column purification is more costly and has high accuracy at low concentrations. Both HPLC-FLD and UPLC-MS/MS are suitable for the extraction and detection of CIT and OTA in tea matrix, but the latter has lower matrix effect and more accurate characterization.