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免疫亲和柱净化并经1-蒽腈衍生化后,采用液相色谱-荧光检测法测定谷物中的T-2毒素。

Determination of T-2 toxin in cereal grains by liquid chromatography with fluorescence detection after immunoaffinity column clean-up and derivatization with 1-anthroylnitrile.

作者信息

Pascale Michelangelo, Haidukowski Miriam, Visconti Angelo

机构信息

Institute of Sciences of Food Production, C.N.R., Viale L. Einaudi 51, 70125 Bari, Italy.

出版信息

J Chromatogr A. 2003 Mar 14;989(2):257-64. doi: 10.1016/s0021-9673(03)00081-5.

Abstract

1-Anthroylnitrile (1-AN) has been shown to be an efficient labelling reagent for the determination of T-2 toxin (T-2) by high-performance liquid chromatography (HPLC)-fluorescence detection. This reaction has been used to develop a sensitive, reproducible and accurate method for the determination of T-2 in wheat, corn, barley, oats, rice and sorghum. The method uses immunoaffinity columns containing antibodies specific for T-2 for extract clean-up, pre-column derivatization with 1-AN and HPLC with fluorescence detection for toxin determination. Ground cereal samples were extracted with methanol-water (80:20, v/v), the extracts were purified by immunoaffinity columns and the toxin was quantified by reversed-phase HPLC with fluorometric detection (excitation wavelength 381 nm, emission wavelength 470 nm) after derivatization with 1-AN. Recoveries from the different cereals spiked with T-2 at levels ranging from 0.05 to 1.5 microg/g were from 80 to 99%, with relative standard deviations of less than 6%. The limit of detection was 0.005 microg/g, based on a signal-to-noise ratio of 3:1.

摘要

1-蒽腈(1-AN)已被证明是一种通过高效液相色谱(HPLC)-荧光检测法测定T-2毒素(T-2)的有效标记试剂。该反应已被用于开发一种灵敏、可重现且准确的方法,用于测定小麦、玉米、大麦、燕麦、水稻和高粱中的T-2。该方法使用含有对T-2具有特异性抗体的免疫亲和柱进行提取物净化,用1-AN进行柱前衍生化,并通过HPLC结合荧光检测来测定毒素。将谷物磨碎样品用甲醇-水(80:20,v/v)提取,提取物通过免疫亲和柱纯化,毒素在与1-AN衍生化后,通过反相HPLC结合荧光检测(激发波长381nm,发射波长470nm)进行定量。在添加了浓度范围为0.05至1.5μg/g的T-2的不同谷物中的回收率为80%至99%,相对标准偏差小于6%。基于3:1的信噪比,检测限为0.005μg/g。

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