In this study, porcine lactoferricin LFP-20 and its analogs were prepared by chemical synthesis with an aim to understand structure-function relationships of these peptides and thereby to obtain improved analogs. The minimum inhibitory concentration (MIC) measured by broth microdilution method and hemolytic analysis showed that the 20-residue porcine lactoferricin (LFP-20) and its analogs LF2A, LF-1, and LF-3 displayed the antimicrobial activity against Escherichia coli, Salmonella choleraesuis, Salmonella typhimurium, Staphylococcus aureus and Staphylococcus epidermidis. The minimum inhibitory concentrations of LFP-20 ranged from 64 to 128 μg/mL, and LF-1 and LF-3 were 2 to 4 times more effective than LFP-20. The studies demonstrated that the analog LF2A, replacing the 2- and 17- Cys of LFP-20 with Ala, did not show increased activities against bacteria, but exhibited decreased hemolytic activity. The analog LF-1, replacing the 9-and 18- Ile of LFP-20 with Trp, showed improved antimicrobial activity. But the hemolytic activity of LF-1 was also increased at 4, 32, 64, 128, and 256 μg/mL (P<0.05). The cytotoxic potential of LFP-20 analogs was quantified by colorimetric WST-1 and LDH assays in peripheral blood mononuclear cell （PBMC）. LF2A, LF-1 and LF-3 increased cell proliferation and viability in a dose dependent fashion. Compared with LFP-20, 25 to 200 μg/mL LF-1 improved significantly cell proliferation (P<0.05), while 400 μg/mL LF-1 decreased cell proliferation (P<0.05). Both 200 and 400 μg/mL LF-1 induced an increase in lactate dehydrogenase (LDH) release from PBMC (P<0.05) whereas 25 to 50 μg/mL decreased the LDH release (P<0.05). Moreover, LF-3 exhibited obviously enhanced potential to depolarize the cytoplasmic membranes at relatively low concentrations (4 μg/mL). In contrast, LFP-20 and LF2A had more modest antibacterial activities, and a weaker ability to depolarize the cytoplasmic membrane. In conclusion, the antimicrobial activity of LF-3 is found to be 2 to 4 folds higher than that of LFP-20, which dose not couple with increased heomolysis and cytotoxicity to PBMCs. Moreover, LF-3 can disrupt the membrane potential by depolarizing the bacterial membrane, which is proposed to be one of the mechanisms of action of LF-3.［Chinese Journal of Animal Nutrition，2011，23（2）：241-249］