高效液相色谱法测定粮食及其制品中赭曲霉毒素A提取条件的优化

Optimizion of extraction conditions for the determination of ochratoxin A in grain and its products by different extraction methods

中文关键词:  粮食及其制品  赭曲霉毒素A  高效液相色谱法  方法优化  国际实验室间验证

英文关键词:grain and its products  ochratoxin A  high performance liquid chromatography  method optimization  the international lab validation

中文摘要:

      目的 优化高效液相色谱法测定粮食及其制品中赭曲霉毒素A(ochratoxin A, OTA)的提取条件。 方法 样品通过乙腈-水(60:40, V:V)提取, 高速均质2 min或振荡提取20 min, 采用赭曲霉毒素A免疫亲和柱浓缩净化、高效液相色谱-荧光检测器定量检测。结果 选用OTA自然污染的粮食样品优化方法更符合实际检测。60%乙腈-水的提取效率优于80%的甲醇-水, 且振荡提取20 min, 一次提取即可提取样品中99%以上的OTA。4种粮食及其制品中OTA在0.2~50 ?g/kg范围内线性关系良好, 相关系数为r2=0.9997。加标回收率为80.7%~108.0%, 变异系数为1.6%~7.5%, 检出限为0.06 μg/kg, 定量限为0.2 μg/kg。采用此方法参加比利时国际能力验证, |Z|<2, 结果满意。同时, 基于本方法组织的国际实验室环形验证中, 粮食及其制品中OTA测定值的统计结果符合《食品法典委员会程序手册》(第二十版)中关于联合验证中HorRat在2之内的规定。 结论 优化后的方法灵敏度高、准确性好, 适合粮食及其制品中赭曲霉毒素A的精准测定。

英文摘要:

      Objective To optimize the extraction conditions for determination of ochratoxin A (OTA) in grain and its products by high performance liquid chromatography (HPLC). Methods The samples were extracted by acetonitrile-water (60:40, V:V), the mixture was homogenised at high speed for 2 min or shaking for 30 min. Their concentration and purification were achieved with the immunoaffinity column (IAC), then the samples were quantitatively detected by high performance liquid chromatography with fluorescence detector (HPLC-FLD). Results The optimizing method using grain samples with OTA natural contamination was more suitable for actual detection. The extraction efficiency of 60% acetonitrile-water was better than 80% methanol-water, and the oscillation extraction time was 20 min, and more than 99% OTA in the sample could be extracted after one extraction. The OTA in grain and its products had good linear relationships in the range of 0.2-50 ?g/kg, and the correlation coefficients (r2) was 0.9997. The recoveries were in the range of 80.7%-108.0%, with the variable coefficient of 1.6%-7.5%. The limit of detection was 0.06 μg/kg, and the limit of quantitation was 0.2 μg/kg. Using this method to participate in Belgium international capacity verification, the results were satisfactory (|Z|<2). At the same time, the statistical results of OTA measurement values in food and its products in the international laboratory circular verification organized by this method conformed to the provisions of the Codex alimentarius commission procedures manual (20th edition) on HorRat within 2 in the joint verification. Conclusion The optimized method has high sensitivity and good accuracy, which is suitable for accurate determination of ochratoxin A in grain and its products.