In this study, cellulose acetate (CA)/polyvinylpyrrolidone (PVP) and CA/PVP/Mn 2+ nanofibers were produced by the electrospinning method, and these cellulose-derived membranes were used as carriers for arginase immobilization for the first time. The structural and morphological analysis of these cellulose-derived nanofibers were determined by attenuated total reflection-Fourier to transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA), X-ray diffraction (XRD), and scanning electron microscopy (SEM). After the immobilization process, it was observed that the thermal properties of the cellulose-derived nanofibers especially improved. The optimum temperature value for free arginase was found to be 35 °C, and this value was found to be 40 °C for arginase immobilized cellulose-derived nanofibers. When the free arginase retained only about 35% of its activity at 50 °C and 60 °C after 60 min, arginase immobilized nanofibers protected 65% of their activity under the same conditions. In addition, arginase immobilized CA/PVP and CA/PVP/Mn 2+ nanofibers managed to retain 50% of their activity even after 9 and 12 reuses, respectively.