We report on the synthesis and the characterization of an azobenzene-based compound. The molecule shows high antimicrobial activity against Staphilococcus aureus and Candida albicans, as demonstrated by in vitro microbiological test in solution. Two antimicrobial polymeric systems were realized introducing this antimicrobial molecule in two polyolefin matrices. The polymeric materials containing different percentages of azo-compound were mold-casted and the obtained films were tested in vitro against the same Gram-positive bacterium and fungus. The films show biocide activity at low percentage of azo dye (up to 0.01%), holding promise for the fabrication of bacteria-resistant polymer films by means of simple melt processing.