免疫亲和柱净化/柱前衍生化-高效液相色谱荧光检测法测定粮谷中的T-2毒素

Determination of T-2 Toxin in Cereal Grains by High Performance Liquid Chromatography with Fluorescence Detection after Immunoaffinity Column Clean-Up and Precolumn Derivatization

建立了免疫亲和柱净化／柱前衍生化-高效液相色谱荧光检测器测定粮谷中T-2毒素含量的方法。样品经甲醇-水（体积比为80：20）混合溶剂提取，通过免疫亲和柱（IAC）净化，以氰酸蒽（1-AN）为衍生化试剂、4-二甲基氨基吡啶（DMAP）为催化剂进行衍生，以ZORBAX Eclipse XDB-C18柱为分离柱，乙腈-水（体积比为80：20）为流动相进行高效液相色谱分离及荧光检测，荧光检测的激发波长为381nm，发射波长为470nm。T-2毒素的质量浓度为0．01～1．5mg／L时与峰高呈良好的线性，相关系数为0．9985。在0．01～1．5μg／g添加水平下，回收率为79．7％～94．5％，相对标准偏差小于7％；检出限（S/N=3）为0．01μg／g。该方法净化效果好，灵敏度高，操作简便快速。

A method has been developed for the determination of T-2 toxin in cereal grains by high performance liquid chromatography with fluorescence detection after immunoaffinity column clean-up and precolumn derivatization. The derivatization reaction was used to develop a sensitive, reproducible and accurate method for the determination of T-2 toxin in wheat, corn, barley and rice. T-2 toxin was extracted with methanol-water （80： 20, v/v） , purified by immunoaffinity columns containing antibodies specific for T-2 toxin, and quantified by reversedphase high performance liquid chromatography with fluorescence detection （excitation wavelength, 381 nm; emission wavelength, 470 nm） after derivatization with 1-anthroylnitrile （1- AN） and 4-dimethylaminopyridine （DMAP）. ZORBAX Eclipse XDB-C18 column and mobile phase of acetonitrile-water （80： 20, v/v） were used for the analysis. Recoveries from the different cereals spiked with T-2 toxin at levels ranging from 0. 01 to 1.5 wg/g were from 79. 7% to 94. 5%, the relative standard deviations were lower than 7% and the limit of detection was 0.01 wg/g based on a signal-to-noise ratio of 3： 1.