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Biodegradable polymers for localized delivery of antibiotics play an important role in treatment of wound infections. This paper describes the in vitro evaluation of biodegradable ofloxacin loaded chitosan-based microspheres and the relation between the release of ofloxacin and the biological activity. The amount of ofloxacin released was determined with UV/VIS spectroscopy. Biological tests on cytotoxicity were performed to prove the possibility of the ofloxacin-releasing chitosan-based microspheres for the treatment of infections associated to wounds.