高效液相色谱—柱后衍生法测定黄曲霉毒素的方法改进

An improved method for the determination of aflatoxins by high performance liquid chromatography and post-column derivatization

目的改进GB 5009.22—2016高效液相色谱—柱后衍生法测定不同种类样品中4种黄曲霉毒素的样品前处理和色谱条件,以简化分析步骤,提高回收率。方法样品经甲醇—水混合溶液提取,免疫亲和柱净化,不氮吹,50%甲醇溶液介质中直接进样。以Acclaim C;色谱柱(4.6 mm×150 mm,3μm)分离,流动相为甲醇—水(43∶57,V/V),经光化学柱后衍生器衍生、荧光检测器检测,外标法定量。结果4种黄曲霉毒素在一定浓度范围内呈良好线性,其相关系数均为1.0000。以3倍信噪比计算得出4种组分的检出限,依次为0.008μg/kg、0.03μg/kg、0.01μg/kg、0.03μg/kg。以空白样品做基体进行加标回收试验,回收率在98.2%~100.8%,相对标准偏差(n=6)在0.72%~0.83%。结论本方法简便、准确、灵敏,可用于不同样品中4种黄曲霉毒素的测定,尤其适合测定批量样品。

Objective To improve the sample pretreatment conditions and chromatographic conditions for determining four aflatoxins in different kinds of sample by GB 5009.22—2016 post-column derivatization coupled with high performance liquid chromatography(HPLC).Methods The sample was extracted with methanol-water,cleared up by the immune affinity column;then pretreated with no nitrogen blowing and directly injected to HPLC with the medium of 50%methanol solution.Acclaim C_(18)(4.6 mm×150 mm,3μm)was used as the chromatographic column,the methanol-water(43∶57,V/V)was used as mobile phase.After the post column derivatization and fluorescence detection,external standard method was used for quantification.Results The four aflatoxins had a good linear relationship with correlation coefficients(r)of 1.0000.Detection limits(3S/N)for four aflatoxins were 0.008μg/kg,0.03μg/kg,0.01μg/kg,0.03μg/kg,respectively.The recoveries from spiked blank sample were in the range of 98.2%-100.8%,with RSDs(n=6)of 0.72%-0.83%.Conclusion The method is simple,accurate,and sensitive.Therefore,the method can be successfully used for the determination of four aflatoxins in different kinds of samples,which is especially suitable for analysis of large batch of samples.