基于超高效液相色谱-四极杆/静电场轨道阱高分辨质谱系统,建立了食品中牛奶过敏原酪蛋白的快速筛查和定量检测方法.样品经缓冲液提取后,采用5 kD超滤膜去除小分子杂质,得到蛋白质提取液.以数据依赖采集(data-dependent acquisition, DDA)方式获得全扫描质谱图,进行蛋白质定性确证,以平行反应监测(parallel reaction monitoring, PRM)技术对目标特征肽段进行定量分析.针对特征肽段,设计并合成了内标肽和内标物质,以降低基质效应和抵消处理过程中的损失.该方法应用于食品中的α-酪蛋白、β-酪蛋白和κ-酪蛋白的快速筛查和定量检测.结果表明,该方法在5~250 μg/L范围内线性关系良好,定量限为0.2~5.5 μg/kg,平均回收率在68.8%~104.4%之间,RSD<6%.该方法可用于果汁饮料、果酱、面包、早餐谷物中牛奶过敏原酪蛋白的快速筛查和定量分析.
Based on ultra performance liquid chromatography-quadrupole/electrostatic field orbitrap high resolution mass spectrometry (UPLC-Q/Orbitrap MS) system, a rapid screening and quantitative detection method for milk allergen caseins was developed.After the sample was extracted with protein extraction buffer, a 5 kD ultrafiltration membrane was used to remove small molecule impurities.The extracted protein was then digested by trypsin.The full-scan mass spectra obtained by the data-dependent acquisition (DDA) mode were used for protein identification, and parallel reaction monitoring (PRM) technique was used for quantitative analysis of the target signature peptide.The internal standard peptide and the internal standard substances for every signature peptide were designed and synthesized to reduce the matrix effect and eliminate the loss in sample pretreatment.The proposed method showed a good linear relationship in the range of 5-250 μg/L.The limits of quantitation were in the range of 0.2-5.5 μg/kg.The observed recoveries ranged from 68.8% to 104.4%, and the RSDs were lower than 6%.This method can be used for rapid screening and quantitative analysis of milk allergen casein in juice, jam, bread and breakfast cereals.
参考文献
- 下载量()
- 访问量()
- 您的评分：
-
10%
-
20%
-
30%
-
40%
-
50%