The evolution of eco-friendly, green route and cheap technology for synthesizing nanostructured zinc oxide (ZnO) thin films using plant extracts is a promising choice because such materials present a widespread potential for numerous technological applications. This study proposes the green and cost-effective technique to synthesize stable ZnO thin films using a good reducing agent and facilitating many natural L-ascorbic acids (AA) metabolic reactions capacity. The influence of AA concentrations in the starting bath solution on ZnO samples' structural, morphological, electrical and antibacterial performances has been reported in detail. The main physical characteristics of the ZnO materials were improved by supplementing of reducing and capping agents AA. Average particle size varies with the adding AA from 58.29 to 48.68 nm and also thickness of these films was decreased from 0.82 to 0.44 μm. Also, it was seen that, the presence of AA in the bath solution significantly affected the absorption process and causes a morphological alteration due to the reaction between Zn2+ and AA during the deposition process. FTIR transmittance spectra of bare ZnO presented that a transmittance peak about 886 cm−1 and 748 cm−1 was created by the characteristic stretching vibration mode of the Zn–O. The resistivity of the produced films significantly changed with AA concentration in the bath solution. Antibacterial potentials of bare ZnO and ascorbic acid added ZnO films were examined against economically important Staphylococcus aureus (ATCC 25923) gram-positive bacteria and Escherichia coli (ATCC 35218) gram-negative bacterial disease agents via handling paper disc diffusion assay. The obtained diameter of the zones of inhibition was 20.1 mm for E. coli and 28.1 mm for S. aureus at the dose of ZnO+AA 8.0%. These inhibition diameters were larger than the diameter of ampicillin as our positive control alone. This proves that the newly synthesized compound is a powerful antibacterial agent.