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食品与发酵工业  2021, Vol. 47 Issue (18): 258-266    DOI: 10.13995/j.cnki.11-1802/ts.028030
  分析与检测 本期目录 | 过刊浏览 | 高级检索 |
超高效液相色谱-高分辨质谱法测定肉类特征肽
王忠合, 李晓婷, 胡文梅, 王军*
(韩山师范学院 食品工程与生物科技学院,广东 潮州,521041)
Determination of peptide markers of meat species by ultra-high pressure liquid chromatography coupled with high resolution mass spectrometry
WANG Zhonghe, LI Xiaoting, HU Wenmei, WANG Jun*
(College of Food Engineering and Biotechnology, Chaozhou 521041, China)
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摘要 采用超高效液相色谱串联四极杆飞行时间质谱法测定8条肉类特征肽,探讨色谱分离条件、净化方法和质谱参数对特征肽含量测定的影响。采用C18柱分离,以体积分数0.1%的甲酸水溶液和乙腈(含体积分数0.1%甲酸)为流动相梯度洗脱,电喷雾正离子化检测、采用飞行时间质谱的准多反应监测模式(time-of-flight mass spectrometry-pseudo-multiple response monitoring,TOF-MRM)采集方法定量测定。结果表明,目标特征肽的母离子带2个或3个电荷的信号强度高、稳定性好,二级质谱图中各肽段的碎片离子主要是y离子,且母离子带3个电荷的肽段中也有带2个电荷的碎片离子,使用TOF-MRM采集模式和目标增益采集数据可增加目标离子的灵敏度。优化后的梯度洗脱条件分离效果非常好,质谱信号强,8条合成肽标准溶液的质量浓度和检测响应的线性关系良好,其相关系数均大于0.98;检出限在0.19~1.50 μg/L,TOF-MRM法准确度高,精密度和重复性均可满足要求。对牛肉丸制品中特征肽含量测定及鉴别发现,七类样品中共检测出5条特征肽,其种属来源分别为牛肉2条、猪肉1条、鸡肉2条,这表明测定的牛肉丸样品中不同程度地混有鸡肉或猪肉,且其混入量不等,与牛肉丸价格高低无关,而种属来源为鸭肉的2条特征肽未检出,这表明测定的牛肉丸样品中未混入鸭肉。该文建立了一种肉类特征肽的检测方法,为肉类蛋白质来源鉴定提供了有力的技术支持。
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王忠合
李晓婷
胡文梅
王军
关键词:  肉种属  肽标志物  高分辨质谱  准多反应监测  确证    
Abstract: Eight marker peptides in meat were identified and quantitatively determined by ultra-high pressure liquid chromatography coupled with quadrupole time-of-flight mass spectrometry,and the effects of chromatographic separation conditions and mass spectrometry parameters on the determination of marker peptides in meat were investigated. The samples were separated by a C18 column, and gradient elution was performed with 0.1% formic acid in water and acetonitrile (containing 0.1% formic acid) as mobile phase. Then, the electrospray positive ion mode (ESI+) was used to detect the marker peptides, and quantitative analysis and data collection were performed by pseudo-multiple reaction monitoring by time-of-flight mass spectrometry (TOF-MRM). Results showed that the parent ions of the targeted marker peptides had a 2+ or 3+ charge, which showed high signal intensity and good stability. And the fragment ions of each marker peptide in the mass spectrometry was mainly y ion. Moreover, the fragment ions with 3+ charge were accompanied by the formation of 2+ charges. The TOF-MRM acquisition mode and target enhancement increased the detection sensitivity of target ions. Under the optimized conditions, the separation of 8 marker peptides was good and the mass signals were strong. There was a good linear relationship between the concentration of 8 marker peptides standards and detection response, and the correlation coefficients were greater than 0.98. The limit of detection (LOD) for analysts was in the range of 0.19 - 1.50 μg/L, and the optimized TOF-MRM method was successfully applied to the analysis of marker peptides in meat samples due to the high accuracy, precision and repeatability. The application of marker peptides on 7 class beef meatball products showed that five marker peptides could be detected including 2 beef, 1 pork and 2 chicken meat, which indicated that cheaper chicken or pork was spiked in beef meatball samples. The abundance of marker peptides had no correlation with the price. However, no duck-specific marker peptides were found. The established species-specific peptide detection method provided powerful technical support for the identification of protein species origin.
Key words:  meat species    marker peptides    high resolution mass spectrometry    pseudo-multiple reaction monitoring    authentication
收稿日期:  2021-05-18      修回日期:  2021-06-03                发布日期:  2021-10-18      期的出版日期:  2021-09-25
基金资助: 广东省基础与应用基础研究项目(2018A0303070006);广东省教育厅特色创新项目(2015KTSCX088);广东省科技发展专项资金项目(2016A020210135);2020年度烹饪科学四川省高等学校重点实验室开放基金项目(PRKX2020Z04)
作者简介:  博士,副教授(王军副教授为通讯作者,E-mail:wangjun19811210@163.com)
引用本文:    
王忠合,李晓婷,胡文梅,等. 超高效液相色谱-高分辨质谱法测定肉类特征肽[J]. 食品与发酵工业, 2021, 47(18): 258-266.
WANG Zhonghe,LI Xiaoting,HU Wenmei,et al. Determination of peptide markers of meat species by ultra-high pressure liquid chromatography coupled with high resolution mass spectrometry[J]. Food and Fermentation Industries, 2021, 47(18): 258-266.
链接本文:  
http://sf1970.cnif.cn/CN/10.13995/j.cnki.11-1802/ts.028030  或          http://sf1970.cnif.cn/CN/Y2021/V47/I18/258
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