In biomedical applications, biocompatible and nontoxic compounds are the most critical types of cytotoxic effects. In this work, the nanobiomaterial MCC/IL/Ag-AgCl NPs were fabricated by immobilizing Ag-AgCl nanoparticles on the surface of organic biopolymer [microcrystalline cellulose (MCC)] without using any reducing agent. The physical and chemical properties of nanobiomaterial were characterized by techniques including transmission electron microscopy (TEM), scanning electron microscopy (SEM), energy-dispersive X-ray (EDX) analysis, X-ray diffraction (XRD), FOurier transform infrared (FT-IR) spectroscopy, and atomic absorption spectroscopy (AAS). The in vitro antibacterial activity of MCC/IL/Ag-AgCl NPs was investigated on methicillin-resistant Staphylococcus aureus (ATCC-43300) and multidrug-resistant Pseudomonas aeruginosa (ATCC-27853). The results such as MIC and MBC illustrated that the antimicrobial effect of MCC/IL/Ag-AgCl NPs against Pseudomonas aeruginosa (1−2 μg/mL of MIC and MBC value) was higher than that of Staphylococcus aureus (4 μg/mL of MIC and MBC value) and also by using the microtiter plate method, the antibiofilm effect of MCC/IL/Ag-AgClNPs against Pseudomonas aeruginosa was remarkable in a variety of low concentrations (32, 16, 8, 4, 2, and 1 μg/mL). Besides, the cytotoxicity study (cell death induction) by MCC/IL/Ag-AgCl NPs in Caco-2 cells revealed low toxic effects in different doses of MCC/IL/Ag-AgCl NPs up to 400 ppm for Ag-AgCl nanoparticles and MCC/IL support, which were further confirmed in biomedical products. It is expected that using these biocompatible compounds can inhibit the biofilm formation of bacteria on surfaces and equipment, to reduce nosocomial infections, in hospital environments, especially in the intensive care unit, and surgery unit/room.