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A combined tryptic peptide and winged peptide internal standard approach for the determination of α‐lactalbumin in dairy products by ultra high performance liquid chromatography with tandem mass spectrometry
Author(s) -
Lai Shiyun,
Zhang Jingshun,
Zhang Yu,
Chen Qi,
Huang Baifen,
Ren Yiping
Publication year - 2015
Publication title -
journal of separation science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.72
H-Index - 102
eISSN - 1615-9314
pISSN - 1615-9306
DOI - 10.1002/jssc.201401279
Subject(s) - peptide , chemistry , chromatography , lactalbumin , mass spectrometry , tandem mass spectrometry , bottom up proteomics , digestion (alchemy) , sample preparation in mass spectrometry , peptide mass fingerprinting , whey protein , protein mass spectrometry , biochemistry , proteomics , electrospray ionization , gene
A robust ultra high performance liquid chromatography with tandem mass spectrometry method at peptide level was established for measuring α‐lactalbumin in various dairy products. An isotope‐labeled winged peptide (VKKILDKVG *I NYW *L AHKALCSEKL) with extra amino acids of the sequence of signature peptide concatenated at each end as the internal standard was spiked in samples to participate in the whole tryptic digestion process. The peptide VG *I NYW *L AHK that resulted from the isotope‐labeled winged peptide was used as the final isotopically labeled internal standard of the α‐lactalbumin signature peptide (VGINYWLAHK) during the quantitative analysis. The contents of α‐lactalbumin in samples were calculated based on the equimolar relationship between the α‐lactalbumin protein and signature peptide. The optimized molar ratio of trypsin to protein (1:60) and enzymatic digestion time (5 h) could not only improve the digestion efficiency and reduce the cost, but also minimize the period of sample pretreatment. Considering the robustness of the current method using the isotopically labeled internal standard and acceptable measurement cost, its application may promote the development of nutrient investigation and quality control of α‐lactalbumin in dairy products. This protein analysis method might provide a new reference strategy for food analysis and quantitative protein analysis.