Functionalize nano-architecture CaO/Ag-chitosan bio-nanoparticles were prepared by the acetic acid sol-gel process and were loaded into the chitosan nanocomposite. The so prepared bioactive nano-CaO/Ag-chitosan nanocomposite was characterized using XRD, TEM, FE-SEM, optical, and the antimicrobial activity was evaluated. The antibacterial evaluations implied that chitosan, nano-CaO, and nano-CaO/Ag-chitosan are exhibited subtle antimicrobial activities against both Gram-positive (Staphylococcus aureus) microbes and Gram-negative (Escherichia coli).Ag doping of CaO produces an oxygen vacancy, defects, and improved light scattering and so leads to transmission increase and absorption increase which in turn lead to increase of the bandgap energy. The optical conductivity of the nanocomposites decreased with Ag addition as a direct result of the bandgap increase. These prefatory investigations of the bio nanocomposites will open a good platform for their use in various bio-industries.