超高效液相色谱-串联质谱法测定鱼丸中的河鲀毒素

Determination of tetrodotoxin in fishballs by ultra-liquid chromatography-tandem mass spectrometry

目的建立鱼丸中河鲀毒素(TTX)的超高效液相色谱-串联四级杆质谱联用测定方法。方法鱼丸样品中的TTX经酸化甲醇提取,免疫亲和柱净化,氮气吹干后用流动相定容,经Hilic Amide(2.1 mm×100 mm,1.8μm)色谱柱分离,甲醇-0.1%甲酸(含5 mmol/L乙酸铵)溶液作为流动相梯度洗脱,选择电喷雾离子源正离子多反应监测(MRM)模式进行监测,外标法基质标准曲线定量。结果 TTX在HILIC Amide柱上能与杂质良好分离,在2 ng/ml^200 ng/ml时,线性关系良好,相关系数为0.999 1。在10μg/kg、50μg/kg和100μg/kg的加标水平下,回收率为72.9%~95.2%,相对标准偏差为2.32%~5.62%,方法的定量限为10μg/kg。应用该方法对市售的30份鱼丸样品进行检测,均未检出TTX。结论该法操作简单快速,准确度和灵敏度高,适合于鱼丸中TTX的检测,具有较好的应用价值。

Objective To establish a method for the determination of tetrodotoxin（ TTX） in fishball by ultra- high performance liquid chromatography- tandem mass spectrometry（ UPLC- MS）. Methods TTX in fishball samples was extracted with acetic acid- methanol,purified with immunoaffinity column. The separation was performed by ultimate Hilic Amide column（ 2. 1 mm ×100 mm,1. 8 μm） with the solution of methanol- 0. 1% formic acid- 5 mmol / L ammonium acetate as mobile phase for gradient elution. The detection was performed by mass spectrometer in the positive electrospray ionization（ ESI） under multiple monitoring mode（ MRM）,and quantification was based on external standard method with the matrix- matched calibration curve. Results TTX was well separated with impurity,and there was a good linearity for TTX in the range of 2 ng / ml- 200 ng / ml（ r =0. 999 1）. The recoveries were within 72. 9%- 95. 2% with the relative standard deviations（ RSD） within 2. 32%- 5. 62% at spiked concentrations of 10 μg / kg,50 μg / kg and 100 μg / kg,respectively. The limit of quantification was 10 μg / kg. With the application of this method to detect 30 commercially available fish samples,TTX was not detected. Conclusion This method is simple,rapid,accurate and sensitive. It is suitable for the detection of TTX in fish ball samples,with higher application value.