Heidari Ghasem, Hashemi Hazaveh Seyed Jamal, Daraei Bahram, Bayat Mansour
Department of Pathobiology, Science and Research Branch, Islamic Azad University, Tehran, Iran.
Department of Medical Parasitology and Mycology, School of Public Health, Food Microbiology Research Center, Tehran University of Medical Sciences, Tehran, Iran.
Iran J Pharm Res. 2019 Fall;18(Suppl1):182-189. doi: 10.22037/ijpr.2019.112398.13735.
This study focuses on optimization and validation of an Ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method for simultaneous analysis of 11mycotoxins: Aflatoxins (B, B, G, and G), Ochratoxin A, Deoxynivalenol, Fumonisins (B and B), Zearalenone, T-2, and HT-2toxin, in wheat matrix. Sample extraction and cleanup procedure is based on a single extraction step using acetonitrile/water/acetic acid mixture (79.5/20/0.5 v/v/v) and rapid clean-up of samples were performed with the Myco6in1 Immunoaffinity column. Electrospray ionization at positive mode was operated to the simultaneously analysis of selected mycotoxins in a single run time of 15 min. Multiple Reaction Monitoring (MRM) mode was selected for quantification and detection of the mycotoxins. The analysis method was validated for selected mycotoxins at different spike levels (2-150 ngg for AFs, T-2, OTA; 20-1500 ngg for ZER, HT-2 toxin; and 100-1500 ngg for DON and FB+B) in wheat. Calibration curves were plotted based on the area of peak analyte in spike samples. Limits of detection (LOD) ranged from 0.7 to 33.3 ngg and limits of quantification (LOQ) ranged from 2 to 100 ngg. Recovery values were between 70 and 120% for all the mycotoxins, except for AFG (72-123%) and T-2 toxin (77-122%) with good repeatability. The recoveries and repeatabilities were in accordance with the criteria determined by European Union (EU) Recommendation 519/2014.
本研究聚焦于超高效液相色谱串联质谱法（UHPLC-MS/MS）的优化与验证，该方法用于同时分析小麦基质中的11种霉菌毒素：黄曲霉毒素（B₁、B₂、G₁和G₂）、赭曲霉毒素A、脱氧雪腐镰刀菌烯醇、伏马毒素（B₁和B₂）、玉米赤霉烯酮、T-2毒素和HT-2毒素。样品提取和净化程序基于使用乙腈/水/乙酸混合物（79.5/20/0.5 v/v/v）的单一提取步骤，并采用Myco6in1免疫亲和柱对样品进行快速净化。在正模式下进行电喷雾电离，以便在15分钟的单次运行时间内同时分析选定的霉菌毒素。选择多反应监测（MRM）模式对霉菌毒素进行定量和检测。该分析方法针对小麦中不同加标水平（黄曲霉毒素、T-2毒素、赭曲霉毒素A为2-150 ng/g；玉米赤霉烯酮、HT-2毒素为20-1500 ng/g；脱氧雪腐镰刀菌烯醇和伏马毒素B₁+B₂为100-1500 ng/g）的选定霉菌毒素进行了验证。根据加标样品中分析物峰面积绘制校准曲线。检测限（LOD）范围为0.7至33.3 ng/g，定量限（LOQ）范围为2至100 ng/g。除AFG₂（72-123%）和T-2毒素（77-122%）外，所有霉菌毒素的回收率在70%至120%之间，重复性良好。回收率和重复性符合欧盟（EU）第519/2014号建议确定的标准。
