Bovine lactoferrin was hydrolysed with a range of proteolytic enzymes including calf rennet, fungal
rennin, and porcine pepsin. Lactoferrin hydrolysates were assessed for their antibacterial activities
against Escherichia coli and Bacillus subtilis. At pH 3, calf rennet lactoferrin hydrolysate before (LFH)
showed the highest antimicrobial activity, then pepsin LFH, while fungal rennin LFH was the least active.
The calf rennet and pepsin LFH were fractionated using autofocusing and chromatographic techniques.
The activity-guided fractionation of calf rennet LFH identified a potent antimicrobial peptide of 11-
residues, lactoferricin B (Lf-cin B), and three other novel antibacterial peptides. The 11-residues Lf-cin B
was the most potent antibacterial peptide and was isolated from both rennet and pepsin LFH. Pepsin LFH
had a main antimicrobial peptide of 25-residues, which was not detected in calf rennet LFH. It could be
concluded that calf rennet LFH had stronger antibacterial properties than porcine pepsin LFH. Besides,
autofocusing could be used for scaling up the isolation of the potent rennet LFH peptides that would have
a widespread commercial use as a natural food preservative substituting porcine pepsin digest, especially
in Islamic communities.