Characterization and determination of bovine immunoglobulin G subtypes in milk and dairy products by UPLC-MS

•Ultra-high performance liquid chromatography mass spectrometry was applied to detect IgG in milk.•Characteristic peptides were qualitatively confirmed by high-resolution mass spectrometry.•Isotope dimethylation can be the internal standard method to correct for matrix effects.•The validated method is suitable for bovine immunoglobulin G subtypes detection. In this study, ultra-high performance liquid chromatography mass spectrometry (UPLC-MS) method was established for the characterization and quantitative determination of immunoglobulin G (IgG) subtypes (IgG1, IgG2, IgG3) in bovine dairy products. High-resolution mass spectrometry (HRMS) was applied to qualitatively confirm the theoretical peptides with specificity, enzymatic hydrolysis curve and stability among in heavy chain constant (CH1, CH2 and CH3) regions. The characteristic peptides VHNEGLPAPIVR, EPSVFIFPPKPK, GLPAPIVR, VVSALR were screened to quantitative analysis bovine IgG1, IgG2, IgG3 and the total amount of bovine IgG1 and IgG3, respectively. Isotope-labeled peptides were obtained by isotope dimethylation reaction, which aimed to correct the matrix effects. The results showed that the recovery was between 98.7% and 103.5%, and the precision of inter-day and intra-day was less than 6.8%. Moreover, this method had good linearity (R2 ≥ 0.999). Therefore, this research provided an effective method for quantitatively detecting bovine IgG subtypes in milk and dairy products.