Nosocomial infections (NIs) are prevalent in intensive care units due to antibiotic overuse. Metal oxide nanoparticles (NPs), like ZnO, offer potential solutions, yet understanding how NPs size impacts their antibacterial efficacy are lacking. This study focuses on the effect of nanoparticle size on kinetics of bacterial strains growth. NPs were synthesized using a sol-gel process with monoethanolamine (MEA) and water, characterized using X-ray diffraction (XRD), transmission electron microscopy (TEM), and Raman spectroscopy, confirming crystallization and size variations. ZnO NPs with mean size of 22, 35 and 66 nm were used against the most common nosocomial bacteria strains Escherichia coli (Gram-negative), Pseudomonas aeruginosa (Gram-negative), and Staphylococcus aureus (Gram-positive). The evaluation of NPs minimal inhibitory concentration (MIC) and bactericidal concentration (MBC) revealed superior antibacterial activity in smaller NPs. The bacterial population was monitored via optical absorbance, showing reduced specific growth rate, prolonged latency period, and increased inhibition percentage with smaller NPs, indicating a substantial deceleration in the growth of microorganisms. Pseudomonas aeruginosa exhibited the smallest sensitivity to ZnO NPs, attributed to its environmental stress resistance. Furthermore, the antibacterial efficacy of paint containing 1 wt% of 22 nm ZnO NPs was assessed and displayed activity against E. coli and S. aureus.