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目的:建立干酪中6种生物胺的高效液相色谱-柱后衍生检测方法。方法:采用高氯酸提取干酪中生物胺,以邻苯二甲醛(OPA)为柱后衍生试剂,高效液相色谱-柱后衍生-荧光法检测。采用Agilent Eclipse XDBC18色谱柱;流动相A为乙酸钠-辛烷磺酸钠溶液,流动相B为乙酸钠-辛烷磺酸钠和乙腈混合溶液,流速1.0m L/min,梯度与等度相结合洗脱方式;柱温40℃;荧光探测器激发波长330 nm,发射波长465 nm;衍生剂流量0.3 m L/min,衍生池温度45℃。结果 :采用本洗脱程序,40 min内6种生物胺完全分离;衍生剂OPA质量浓度0.4 mg/m L,流速0.3 m L/min时衍生效果最佳;6种生物胺标准曲线具有良好的线性关系(R2>0.9982),平行样品相对标准偏差(RSD)<3.8%,方法稳定性良好,6种生物胺样品平均回收率89.25%~103.09%。结论:本法样品预处理简便,灵敏度高,重复性好,准确性高,适合检测干酪中的生物胺。
Objective: To establish a method for the determination of six biogenic amines in cheese by post-column derivatization. Methods: Biogenic amines were extracted with perchloric acid, OPA was used as post - column derivatization reagent, and high performance liquid chromatography - post column derivatization - fluorescence method was used. Using Agilent Eclipse XDBC18 column; mobile phase A sodium acetate - sodium octane sulfonate solution, mobile phase B sodium acetate - sodium octane sulfonate and acetonitrile mixed solution, flow rate 1.0m L / min, gradient and isocratic phase The column temperature was 40 ℃. The excitation wavelength of the fluorescence detector was 330 nm and the emission wavelength was 465 nm. The flow rate of the derivative was 0.3 m L / min and the temperature of the derivatization cell was 45 ℃. Results: Six kinds of biogenic amines were completely separated within 40 min. The best derivatization efficiency was obtained when the concentration of OPA was 0.4 mg / m L and the flow rate was 0.3 m L / min. The standard curves of six biogenic amines had good The linear relationship (R2> 0.9982) and the relative standard deviation (RSD) of parallel samples were less than 3.8%. The stability of the method was good. The average recoveries of six biogenic amines were 89.25% ~ 103.09%. Conclusion: This method sample pretreatment is simple, high sensitivity, good repeatability, high accuracy, suitable for the detection of biogenic amines in cheese.