Ozone, a gas composed of three oxygen atoms, is notable for its oxidative and antimicrobial properties, showing potential to inhibit microbial growth. Its effectiveness depends on concentration and exposure time. The bactericidal action of ozone against medically relevant pathogens motivates studies exploring its therapeutic applications as an alternative against antimicrobial resistance.

To evaluate the efficacy of ozone as an inhibitor of bacterial growth in clinically relevant species, investigate its antimicrobial mechanisms, and provide scientific evidence for potential applications in the control of microbial resistance.

A bacterial killing curve was constructed using clinically relevant microorganisms standardized to 0.5 McFarland in saline solution. Ozonized serum was applied at varying concentrations and exposure times (0, 5, and 10 minutes), followed by serial dilution and plating. Identical control cultures were processed without ozone exposure. All samples were incubated at 37°C for 18 hours. After incubation, colony-forming units (CFUs) were counted and compared (test vs. control) to evaluate the bacterial killing curve under different ozone concentrations and exposure times. Statistical analysis will be performed using the Student’s t-test.

In the first experiment with Staphylococcus aureus, using 25 µg/mL of ozone, a control culture was compared with an ozone-treated culture at 0, 5, and 10 minutes. After dilution, incubation, and colony counting, the control showed 5 × 10⁹ CFU, while the ozone-exposed culture showed 3 × 10⁶ CFU (0 min), 1.2 × 10³ CFU (5 min), and 1 × 10² CFU (10 min). An immediate reduction in CFUs was observed at time 0 compared with the control, with a progressive decline as exposure time increased, demonstrating a time-dependent bactericidal effect.

The initial experiment demonstrated a marked reduction in S. aureus CFUs compared with control and across increasing ozone exposure times, confirming its antimicrobial action against this species. Preliminary results suggest the need for further experiments to validate these findings.