液相色谱-串联质谱法测定牛排产品中大豆蛋白含量

Determination of Soybean Protein in Steak Products by Liquid Chromatography-Tandem Mass Spectrometry

建立液相色谱-串联质谱技术测定牛排产品中大豆蛋白含量的方法。首先利用高分辨质谱结合Unitprot数据库,对大豆特异性多肽进行鉴别及筛选,获得可用于大豆准确定性的多肽序列信息,并利用skyline软件将多肽序列转换为离子对信息;其次利用液相色谱-串联质谱(liquid chromatography-tandem mass spectrometry,LC-MS/MS)对大豆蛋白特异性多肽进行确证,构建大豆蛋白LC-MS/MS定性判别方法;进一步开展用于定量多肽的筛选研究,最终筛选到线性和回收率较好的定量多肽共3条,分别为GSDLVNVR、VSDDEFNNYK、LVFCPQQAEDDK,线性相关系数均达到0.99以上,加标回收率为81.7%~115.8%,精密度小于13%,检出限为0.15%,定量限为0.5%;同时对市售28个牛排进行大豆蛋白含量的测定,构建LC-MS/MS方法测定牛排中大豆蛋白含量的方法。结果表明,该方法特异性好,准确度高,同时可推广应用至其他肉制品。

A method for the determination of soybean protein in steak products by liquid chromatography-tandem mass spectrometry(LC-MS/MS)was developed.Firstly,the specific soybean peptides were identified and screened by highresolution mass spectrometry based on the Unitprot database to obtain the peptide sequences for accurate identification of soybean protein.Then,the soybean peptide sequences were converted into ion pair information using the skyline software.Next,the specific soybean peptides were confirmed by LC-MS/MS,and further quantified.Finally,3 peptides for quantitative analysis of soybean protein with good linearity and recovery rate were selected,which were GSDLVNVR,VSDDENNNYK,and LVCCPQQAEDDK.For all three peptides,the correlation coefficients were above 0.99,the spiked recoveries were 81.7%–115.8%,and the precision expressed as relative standard deviation(RSD)was less than 13%.The limit of detection(LOD)was 0.15%,and the limit of quantification(LOQ)was 0.5%.The soybean protein contents of 28 commercially available steaks were determined successfully using the proposed method.The results showed that the method was highly specific and accurate,and could be applied to other meat products.