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To compare amidation and acylation of lactoferrin (LF) from bovine milk, as a means of enhancing its antimicrobial and antiviral properties.LF was chemically modified by amidation with a 1-ethyl-3-[3-(dimethylamino) propyl] carbodiimide (EDC) in the presence of ammonium ions or by acylation with either succinic or acetic anhydride. In the test systems used, amidation substantially enhanced the activity of LF against Pseudomonas fluorescens in comparison with native LF. However, increasing the net negative charge of LF by acylation had no effect on the activity of LF against P. fluorescens, and abrogated the antimicrobial activity of LF against Bacillus subtilis and Saccharomyces cerevisiae. Increasing the net negative charges of LF by acylation eliminated its antimicrobial and antiviral effects against poliovirus and feline calicivirus (nonenveloped viruses).The addition of positive charges to LF via amidation enhanced antimicrobial properties in contrast to increasing the negative charges by acylation, which abolished both the antimicrobial and antiviral properties of LF.The effects of charge alteration of LF determined in this study provides a basis for further development of LF formulations with enhanced antimicrobial effectiveness for use in food process hygiene, veterinary and health-care applications.