Lactoperoxidase is a peroxidase enzyme found in milk. This enzyme has antimicrobial and antioxidant properties. It is fairly heat resistant and was widely used as an indicator of over pasteurization of milk. In the present study, bovine lactoperoxidase (LPO) was purified from skimmed milk using amberlite-CG-50-H+ resin, CM-sephadex-C-50 ion-exchange chromatography and sephadex-G-100 gel filtration chromatography. Enzyme activity was determined using 2, 2-azino-bis-diammonium salt as a chromogenic substrate at pH 6, and purification degree was controlled by means of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Antibacterial and antifungal activity of bovine lactoperoxidase was determined by disk diffusion method. Bovine LPO exhibited high antifungal and antibacterial activity in 100 mM thiocynate-100 mM hydrogen peroxide (H2O2) medium on some fungi (Candida albicans, Candida glabrata, Candida krusei, Candida parapsilosis and Saccharomyces boulardii) and bacteria (Citrobacter freundii, Escherichia coli, Streptococcus pneumoniae, Staphylococcus epidermidis and Staphylococcus intermedius), respectively. Antibacterial and antifungal activity of LPO system was compared to those of well known antibacterial and antifungal substances such as cefaclor, erythromycin, tetracycline and fluconazole. This study has investigated for the first time the antifungal and antibacterial effects of bovine LPO system on C. albicans, C. glabrata, C. krusei, C. parapsilosis, S. boulardii, S. pneumoniae, S. epidermidis and S. intermedius.