HPLC separation of casein components on a diol-bonded silica column with MALDI TOF/TOF MS identification

A comparative study has been made of three home-made bonded silica columns for the separation of the components of bovine milk casein (alpha-, beta- and kappa-casein). A gradient elution HPLC procedure is now proposed for separation of the protein components, and the tests conducted have confirmed the superiority of columns based on diol-bonded silica over octadecyl- or amine-bonded silica. A matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometer (MALDI-TOF MS) has been utilized as a detector for off-line identification of the components of the casein fractions. The method selected enables effective and rapid separation of the casein components, together with their detailed MS characterization.