高效液相色谱-串联质谱法测定玉米及其制品中隐蔽型伏马毒素

Determination of hidden fumonisins in maize and maize products by high performance liquid chromatography-tandem mass spectrometry

目的建立高效液相色谱-串联质谱法(high performance liquid chromatography-tandem mass spectrometry, HPLC-MS/MS)检测玉米及玉米制品中隐蔽型伏马毒素的方法。方法样品中加入伏马毒素B1和B_2的稳定同位素内标,用2 mol/L氢氧化钠溶液进行水解,使隐蔽型及游离型伏马毒素一并转变为水解型伏马毒素,用乙酸乙酯萃取水解型伏马毒素。色谱分离采用0.1%甲酸(A)和甲醇(B)作为流动相进行梯度洗脱,质谱采用正离子扫描(positive ion electrospray ionization,ESI4)、多反应监测模式(multiple reaction monitoring,MRM)对水解型伏马毒素B_1和B_2(hydrolyzed fumonisin B_1 and B_2, HFB_1和HFB_2)的定量离子和定性离子进行监测。另取一份平行样品,根据国家标准GB 5009.240-2016第二法测定其中的游离型伏马毒素含量。样品中隐蔽型伏马毒素的含量(按伏马毒素当量计)通过水解型伏马毒素减去游离型伏马毒素含量计算而得。结果本方法在15 min内完成HFB1和HFB2及其稳定同位素内标的高效液相色谱-串联质谱分离分析。HFB1和HFB2在50、100和1000μg/kg添加水平的回收率为89.1%～109.2%,相对标准偏差为3.3%～12.2%(n=6),方法定量限为15～20μg/kg。结论该方法准确、灵敏,前处理简单,适合测定玉米及其制品中的隐蔽型伏马毒素。

Objective To establish a method for the determination of hidden fumonisins B_1 and B_2 in maize and maize products by high performance liquid chromatography-tandem mass spectrometry（HPLC–MS/MS）. Methods Stable isotope internal standards of fumonisin B_1 and B_2 were added to the sample, 2 mol/L sodium hydroxide solution was used for hydrolysis, which transformed the hidden and free types of fumonisins into hydrolyzed fumonisins, and the hydrolyzed fumonisins was extracted with ethyl acetate. Chromatographic separation was performed using 0.1% formic acid（A） and methanol（B） as mobile phases for gradient elution. Quantitative and qualitative ions of hydrolyzed fumonisins B_1 and B_2（HFB_1 and HFB_2） were monitored by mass spectrometry with positive ions（electrospray ionization, ESI＋） and multiple reactions monitoring（MRM） mode. The free fumonisins in a parallel sample were determined according to method 2 of GB 5009.240-2016. The hidden fumonisins（measured by the equivalent of fumonisins） were calculated by subtracting free fumonisins from total fumonisins. Results HFB_1 and HFB_2 and their stable isotopes were separated and analyzed by high performance liquid chromatography-tandem mass spectrometry within 15 min. The recoveries of hydrolyzed fumonisins B_1 and B_2 at spiking levels of 50, 100 and 1000 μg/kg ranged from 89.1% to 109.2%, with the relative standard deviations of 3.3%-12.2%（n=6）, and the limits of quantitation were 15-20 μg/kg. Conclusion The method is accurate, sensitive and simple, which is suitable for the determination of hidden fumonisins in maize and maize products.