Analysis of tryptic digests of bovine beta-casein by reversed-phase high-performance liquid chromatography

Reversed-phase high-performance liquid chromatography (RP-HPLC) was used to separate the tryptic peptides of beta-casein. A gradient of 0-50% acetonitrile in 0.1% trifluoroacetic acid at a flow-rate of 0.8 ml/min resolved ten of the thirteen peptides. A modified gradient resolved the three peptides eluted at ca. 25% acetonitrile. RP-HPLC proved superior to high-voltage paper electrophoresis in analysis time, resolution and flexibility. The methods developed for the analysis of proteolysis of the milk protein, beta-casein, are now being applied to study the action of extracellular proteases from dairy bacteria on milk proteins.