Lhotská Ivona, Šatínský Dalibor, Havlíková Lucie, Solich Petr
Department of Analytical Chemistry, Faculty of Pharmacy, Charles University, Heyrovského 1203, Hradec Králové, 500 05, Czech Republic.
Anal Bioanal Chem. 2016 May;408(12):3319-29. doi: 10.1007/s00216-016-9402-6. Epub 2016 Mar 18.
A new fast and sensitive method based on on-line solid-phase extraction on a fused-core precolumn coupled to liquid chromatography with fluorescence detection has been developed for ochratoxin A (OTA) and citrinin (CIT) determination in lager beer samples. Direct injection of 100 μL filtered beer samples into an on-line SPE-HPLC system enabled fast and effective sample extraction including separation in less than 6 min. Preconcentration of OTA and CIT from beer samples was performed on an Ascentis Express RP C18 guard column (5 × 4.6 mm), particle size 2.7 μm, with a mobile phase of methanol/0.5% aqueous acetic acid pH 2.8 (30:70, v/v) at a flow rate of 2.0 mL min(-1). The flow switch from extraction column to analytical column in back-flush mode was set at 2.0 min and the separation was performed on the fused-core column Ascentis Express Phenyl-Hexyl (100 × 4.6 mm), particle size 2.7 μm, with a mobile phase acetonitrile/0.5% aqueous acetic acid pH 2.8 in a gradient elution at a flow rate of 1.0 mL min(-1) and temperature of 50 °C. Fluorescence excitation/emission detection wavelengths were set at 335/497 nm. The accuracy of the method, defined as the mean recoveries of OTA and CIT from light and dark beer samples, was in the range 98.3-102.1%. The method showed high sensitivity owing to on-line preconcentration; LOQ values were found to be 10 and 20 ng L(-1) for OTA and CIT, respectively. The found values of OTA and CIT in all tested light, dark and wheat beer samples were significantly below the maximum tolerable limits (3.0 μg kg(-1) for OTA and 2000 μg kg(-1) for CIT) set by the European Union.
已开发出一种基于在线固相萃取的快速灵敏方法，该方法采用熔融核预柱与液相色谱联用荧光检测，用于测定贮藏啤酒样品中的赭曲霉毒素A（OTA）和桔霉素（CIT）。将100 μL过滤后的啤酒样品直接注入在线固相萃取-高效液相色谱系统，可在不到6分钟的时间内实现快速有效的样品萃取，包括分离。在Ascentis Express RP C18保护柱（5×4.6 mm，粒径2.7 μm）上对啤酒样品中的OTA和CIT进行预富集，流动相为甲醇/0.5%乙酸水溶液（pH 2.8，30:70，v/v），流速为2.0 mL min⁻¹。在反冲模式下，从萃取柱到分析柱的流动切换时间设定为2.0分钟，分离在熔融核柱Ascentis Express苯基己基柱（100×4.6 mm，粒径2.7 μm）上进行，流动相为乙腈/0.5%乙酸水溶液（pH 2.8），采用梯度洗脱，流速为1.0 mL min⁻¹，温度为50℃。荧光激发/发射检测波长设定为335/497 nm。该方法的准确度（定义为OTA和CIT在淡色和深色啤酒样品中的平均回收率）在98.3% - 102.1%范围内。由于在线预富集，该方法具有高灵敏度；OTA和CIT的定量限分别为10和20 ng L⁻¹。在所有测试的淡色、深色和小麦啤酒样品中发现的OTA和CIT值均显著低于欧盟设定的最大耐受限量（OTA为3.0 μg kg⁻¹，CIT为2000 μg kg⁻¹）。
