Simultaneous determination of bovine α-lactalbumin and β-lactoglobulin in infant formulae by ultra-high-performance liquid chromatography–mass spectrometry

A reliable ultra-high-performance liquid chromatography–mass spectrometry method for simultaneous determination of bovine α-lactalbumin (α-La) and β-lactoglobulin (β-Lg) was developed. Compared to the previous methods, the developed approach with mass spectrometer operated in selected area monitoring mode offered increased speed and enhanced lower detection limit. A linear gradient mobile phase, consisting of (A) water containing 0.1% trifluoroacetic acid (TFA) and (B) acetonitrile containing 0.1% TFA, and an Acquity UPLC BEH300 C18 column (150 mm × 2.1 mm, 1.7 μm) were employed to obtain the best resolution of the target analytes. The accurate quantitation was achieved by employing human α-lactalbumin as the internal standard. The established method was extensively validated by determining the linearity (R2 ≥ 0.9991), sensitivity (limit of quantitation, 0.15–0.19 μg mL−1), recovery (94.0–98.7%), precision (relative standard deviation ≤ 11.1%) and repeatability (relative standard deviation ≤ 5.7%). It was shown to be a suitable method for simultaneous determination of the major whey proteins in biological samples. Current validated method was successfully applied to the nutrient investigation of infant formulae.