Cathelicidin antimicrobial peptides BuMAP-28 and (BuMAP-28)18 were synthesized and used for the present study. Aminoacid sequences of the antimicrobial domain were deduced from the gene sequence of myeloid antimicrobial peptide of buffalo. Peptides are highly cationic, amphipathic showed a net charge of +11 for both BuMAP-28 and its analogue and a predicted hydrophobic ratio of 39% for BuMAP-28 and 33% for (BuMAP-28)18. Ramachandran plot analysis indicating a high structural stability for the peptides. In silico structural analysis of the peptides revealed a helix-turn helix which is later confirmed by CD analysis. Biological activity testing of the peptides revealed that the peptide has got a broad spectrum antimicrobial and anticancerous activity. Peptides showed wide spectrum of activity against both Gram- positive, Gram- negative bacteria, fungi, spirochetes and virus. Peptides are active against even methicillin resistant S.aureus with lower MIC values. In vitro antibacterial and antifungal activities were later confirmed by morphological testing and SEM. Peptides are also active against HeLa cell lines. Cytotoxic studies revealed that the truncation of BuMAP-28 could reduce the haemolytic activity when compared to the parent peptide. Murine models injected with Duck pasteurella (DP1), when treated with the peptides protected 100% of the animals at 12.5µM doses. Studies revealed that the truncation of the peptide could reduce the haemolytic activity without a considerable change in antimicrobial activity.