Absolute Quantitation of β-Lactoglobulin by Protein Liquid Chromatography−Mass Spectrometry and Its Application to Different Milk Products

The absolute quantitation of proteins in biological matrixes is of great interest in many fields and can be accomplished by different methodologies. Here, a method for the absolute quantitation of the whey protein β-lactoglobulin using protein liquid chromatography coupled to mass spectrometry is reported. The developed approach was characterized in detail and applied to the determination of β-lactoglobulin contents in various milk products. A special focus was placed on the recovery rates of the isolation procedure and on robust quantitation by LC−MS. For these purposes protein internal standards were employed. The observed recovery rates of β-lactoglobulin from various samples ranged from 100% for whole milk to just over 50% for a strongly processed yogurt-based baby food product. The influence of processing was investigated in greater detail, showing that an increasing intensity of the applied heat treatment resulted in an increasing loss of β-lactoglobulin. LC−MS quantitation at the protein level proved to be highly suitable, avoiding a potentially problematic digestion step. The use of an appropriate internal standard to compensate for sample losses during sample workup was shown to be essential for obtaining accurate results.