A Simple Method to Separate the Antimicrobial Peptides from Complex Peptic Casein Hydrolysate and Identification of a Novel Antibacterial Domains within the Sequence of Bovine αs-Casein

A simple method to separate antimicrobial peptides from complex peptic hydrolysate of bovine casein is proposed. It consists to precipitation of the active peptides under ionic strength and pH conditions. The peptide extract obtained was fractionated using reverse phase high performance liquid chromatography and the actives fractions are analyzed by mass spectrometry (LC-ESI-MS and MADI-TOF). Several peptides derived mostly from αs-casein are identified. The major component in active fractions are αs2-CN f(164-179), αs2-CN f(148-166), αs2-CN  f(180-207), αs2-CN  f(183-207), αs2-CN f(131-174), αs1-CN f(153-196), αs1-CN f(141-187), αs2-CN f(176-207), αs2-CN f(164-207), αs2-CN f(99-163). The minimum inhibitory concentration and mode of action of the peptide extract and purified peptide αs2-CN f(164-207) were studied against several Gram-positive and Gram negative Bacteria