Carbon quantum dots as a novel type of carbon nanomaterials have attracted the attention of many researchers because of their unique optical, antibacterial, and anticancer properties as well as their biocompatibility. In this study, for the first time, carbon quantum dots were prepared from -phenylenediamine dissolved in toluene by a solvothermal route. Subsequently, the prepared carbon quantum dots were encapsulated into polyurethane films by a swelling-encapsulation-shrink method. Analyses of the results obtained by different characterization methods (AFM, TEM, EDS, FTIR, photoluminescence, and EPR) indicate the significant influence of the precursor on structural, chemical, and optical properties. Antibacterial and cytotoxicity tests showed that these dots did not have any antibacterial potential, because of the low extent of reactive oxygen species production, and showed low dark cytotoxicity. By investigating the cellular uptake, it was established that these dots penetrated the HeLa cells and could be used as probes for bioimaging.