In recent years, there is great attention on the L-Arginine (Arg) which is a unique, non-toxic, and biocompatible green material because it can be utilized as an agent for the functionalization then stabilization of magnetite [Fe3O4] nanoparticles (MNPs) against oxidation and aggregation during or after synthesis procedure. Our studies demonstrate that Arg has two great impacts on the MNPs which include increasing and decreasing its stability and particle size respectively. Besides, saturation magnetization and electrostatic interactions of Arg-coated magnetic nanoparticles (RMNP) have a direct impact on the biological molecules such as proteins and genes. Therefore, controlling Arg concentration gives a great chance to accurate control above mentioned characterizations which is a useful tool for applications such as antibody connecting, catalyst, drug loading, and MNPs stability modifier. In the current study, RMNPs with different arginine concentrations aka 0.42μg, 1.62μg, and 2.29μg per mg in three synthesized RMNPs (as named 푝0.5, 푝1, and 푝1.5) on the surface of MNPs based on a colorimetric determination, has been successfully synthesized through the simple co-precipitation method. Besides, as-synthesized RMNP powders were characterized by XRD, SEM/EDAX, FT-IR, VSM, and Zeta Potential analyser. The effect of these nanoparticles on the stability of lysozyme protein as a model protein was investigated; also the secondary structure and stability of lysozyme along with these nanoparticles were detected by Ultraviolet (UV) spectroscopy, circular dichroism (CD), fluorescence spectroscopy and enzymatic activity measurements. The results show that magnetic nanoparticles capped with arginine, improve the properties of magnetite (biocompatibility, prevent aggregation of nanoparticles, reduce the size of nanoparticles) and improve the secondary structure of the protein, as well as increase the stability and protein activity. Results from XRD, SEM, and FT-IR was argued that arginine was completely capped to magnetite nanoparticles. Also, the results of VSM showed that, with increasing amount of arginine, magnetite saturation magnetization decreased due to the increase in the diameter of the coating. Finally, the results of bioassay tests proven stability and protein efficiency.