Novel antibacterial and biocompatible transparent films based on chitosan or pullulan and two bioactive ionic liquids (ILs), cholinium hexanoate and cholinium citrate, were prepared. These ILs were selected based on their MIC values against several microbial strains, film-forming ability when blended with the polysaccharides and biocompatibility against designated human cell lines. The films were obtained through simple casting of polysaccharide aqueous solutions containing different amounts of the ILs (20 and 40 wt% with respect to the amount of polysaccharide). The physical properties of the films were investigated using transmittance measurements, thermal analysis, mechanical testing and antibacterial assays. In general, the addition of both ILs does not affect the optical transparency (up to 80% transmittance within 400–700 nm) of the films but decreased their stiffness (acting as plasticizers) and thermal stability. All chitosan-based films showed antibacterial activity against S. aureus and K. pneumoniae but for pullulan only those with cholinium citrate were bioactive.