基于液相色谱-串联质谱技术的羊牛乳特征肽段鉴别及测定

Identification and detection of goat and bovine milk peptide marker based on liquid chromatography-tandem mass spectrometry

中文关键词:  高效液相色谱-串联质谱法  特征肽段  羊乳  牛乳  鉴别

英文关键词:liquid chromatography-tandem mass spectrometry  peptide marker  goat milk  bovine milk  identification

中文摘要:

      目的 建立液相色谱-串联质谱法(liquid chromatography-tandem mass spectrometry, LC-MS/MS)鉴别羊牛乳及定量检测羊乳中牛乳的方法。方法 样品经胰蛋白酶水解后, 采用纳升液相色谱-串联飞行时间质谱仪(nano liquid chromatography-tandem time-of-flight mass spectrometer, nano LC-TOF-MS)检测, 经ProteinPilotTM软件、UniProt数据库和blast分析, 筛选出特征肽段。然后基于高效液相色谱-串联质谱法(high performance liquid chromatography-tandem mass spectrometry, HPLC-MS/MS), 应用牛乳、羊乳的特征肽段对样品进行鉴别和定量测定。结果 本方法在羊乳中牛乳含量为5%~50%范围内, 线性关系良好, 相关性系数大于0.99, 检出限为1.5%, 定量限为5.0%, 在5%、20%和40%添加水平的回收率为85.2%~114.0%, 相对标准偏差小于10% (n=6)。结论 该方法快速、准确, 适合应用于羊乳中牛乳的鉴别及定量分析。

英文摘要:

      Objective To establish a method for the identification and detection of goat and bovine milk peptide marker and quantitation of bovine milk in goat milk based on liquid chromatography-tandem mass spectrometry (LC-MS/MS). Methods Samples were determined by nano liquid chromatography-tandem time-of-flight mass spectrometer (nano LC-TOF-MS) after tryptic digestion. Peptide markers were screened by data analysis with ProteinPilotTM software and UniProt protein database, and searched with blast. The samples were identified and quantified based on high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) with peptide markers. Results The proposed method had good linear dependence for bovine milk in goat milk from 5% to 50% with correlation coefficient r2＞0.99, the limit of detection (LOD) was 1.5% and the limit of quantitation (LOQ) was 5.0%. The recoveries were ranged from 85.2% to 114.0% at 5%, 20% and 40% spiked levels, and the relative standard deviations (RSDs) were less than 10% (n=6). Conclusion The proposed method is fast and accurate, which is suitable for the identification and quantitative detection of bovine milk in goat milk.