In the present study, polyurethane materials were obtained from castor oil, polycaprolactone and isophorone diisocyanate by incorporating different concentrations of chitosan (0.5, 1.0 and 2.0% w/w) as an additive to improve the mechanical properties and the biological activity of polyurethanes. The polyurethanes were characterized by Fourier transform infrared spectroscopy, thermogravimetric analysis, scanning electron microscopy, stress/strain fracture tests and swelling analysis, and the hydrophilic character of the surface was determined by contact angle trials. The objectives of the study were to evaluate the effect of the incorporation of chitosan on the changes of the physico-chemical and mechanical properties and the in vitro biological activity of the polyurethanes. It was found that the incorporation of chitosan enhances the ultimate tensile strength of the polyurethanes and does not affect the strain at fracture in polyurethanes with 5% w/w of polycaprolactone and concentrations of chitosan ranging from 0 to 2% w/w. In addition, PCL5-Q-PU formulations and their degradation products did not affect cell viability of L929 mouse fibroblast and 3T3, respectively. Polyurethane formulations showed antibacterial activities against Staphylococcus aureus and Escherichia coli bacteria. The results of this study have highlighted the potential biomedical application of this polyurethanes related to soft and cardiovascular tissues.