title: 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.
abstract: 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*INYW*LAHKALCSEKL) 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*INYW*LAHK 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.
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China.|Zhejiang University of Technology, Hangzhou, China.
authors.forename: Shiyun
authors.initials: S
authors.lastname: Lai
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China.
authors.forename: Jingshun
authors.initials: J
authors.lastname: Zhang
authors.affiliation: College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Chen
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China.
authors.forename: Baifen
authors.initials: B
authors.lastname: Huang
authors.affiliation: Zhejiang Provincial Center for Disease Control and Prevention, Hangzhou, China.
authors.forename: Yiping
authors.initials: Y
authors.lastname: Ren
mesh_terms: D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003611:Dairy Products / Q000032:analysis*; D007768:Lactalbumin / Q000737:chemistry*; D057230:Limit of Detection; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D012015:Reference Standards; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014357:Trypsin / Q000378:metabolism*
chemical_list: D007768:Lactalbumin; D014357:Trypsin
keywords: Bovine α-lactalbumin; Isotopically labeled peptides; Signature peptide; Tryptic digestion; Ultra high performance liquid chromatography with tandem mass spectrometry