基于UPLC同时测定三种小麦赤霉病毒素方法的构建与应用

Construction and Application of Simultaneous Determination of Three Wheat Fusarium Mycotoxins Based on UPLC

为测定小麦籽粒中因赤霉病产生的毒素脱氧雪腐镰刀菌烯醇(deoxynivalenol,DON)、雪腐镰刀菌烯醇(nivalenol,NIV)和玉米赤霉烯酮(zearalenone,ZEN),本研究构建了一种多功能净化柱-超高效液相色谱-二极管阵列检测器(multifunctional column-ultra performance liquid chromatography-diode array detector,MFC-UPLC-DAD)方法,可同时测定上述三种毒素,对该方法与多功能净化柱-高效液相色谱-二极管阵列检测器(multifunctional column-high performance liquid chromatography-diode array detector,MFC-HPLC-DAD)法测定9个小麦品种中上述3种毒素含量的结果进行比较;并用MFC-UPLC-DAD法检测了129份小麦品种(系)的毒素含量。结果显示,MFC-UPLC-DAD法检测三种毒素在检测范围内(0.01~10.00μg·mL^(-1))标准曲线的相关系数均大于0.994;三种毒素的检出限为15~78μg·kg^(-1);回收率在80.25%~118.35%之间,相对标准偏差为0.20%~1.92%;9份小麦品种中,3种毒素含量的测定结果在两种方法间差异均不显著。利用MFC-UPLC-DAD法测定自然发病的129份材料发现,DON毒素与NIV毒素检出率较高,分别为99%和92%;符合限量标准且毒素含量较低的材料共80份。本研究构建的MFC-UPLC-DAD法可同时测定小麦种DON、NIV和ZEN含量,测定结果重复性好,准确度高,可对抗赤霉病育种提供方便。

To determine the mycotoxins caused by Fusarium head blight in wheat,an MFC-UPLC-DAD(multifunctional column-ultra performance liquid chromatography-diode array detector)method was constructed for simultaneous determination of deoxynivalenol(DON),nivalenol(NIV)and zearalenone(ZEN)in grains by UPLC(ultra-performance liquid chromatography).This method and MFC-HPLC-DAD(multifunctional column-ultra performance liquid chromatography-diode array detector)were used to determine the accumulation of DON,NIV and ZEN in nine wheat varieties.After comparison,MFC-UPLC-DAD was used to detect and analyze the mycotoxin accumulation in 129 varieties after natural infection.The results of MFC-UPLC-DAD showed that a good linear relationship in the detection range(0.01-10.00μg·mL^(-1))in determination and the correlation coefficients of the standard curves were all greater than 0.994.The limits of detection(LOD)of DON,NIV and ZEN were 15-78μg·kg^(-1).The recovery rate was in the range of 80.25%-118.35%,and the relative standard deviation(RSD)was in the range of 0.20%-1.92%.It showed that MFC-UPLC-DAD and MFC-HPLC-DAD have no significant difference in comparing the determination of nine varieties.The detection rate of DON and NIV was 99%and 92%by MFC-UPLC-DAD,respectively.A total of 80 varieties with low toxin content were in line with the limit standard.The MFC-UPLC-DAD constructed in this study for simultaneous determination of DON,NIV and ZEN showed good repeatability,high accuracy,which can be used for simultaneous determination of the three mycotoxins in the grains of different wheat varieties,providing a reference for anti-toxin accumulation breeding.