The current study aimed to assess the efficacy of ozone therapy in the treatment of stage II and stage III periodontitis. This prospective split-mouth study selected patients who were diagnosed with either stage II or stage III periodontitis. All patients were treated with scaling and root-planing (SRP) on the control side and SRP with ozone therapy on the test side. Probing depth (PD), clinical attachment loss (CAL), O’Leary plaque index (PI), and bleeding on probing (BOP) scores were recorded at baseline and six weeks after the SRP treatment. A total of 46 patients were selected for this study, including 31 males and 15 females. All periodontal variables (PD, CAL, PI, and BOP) showed significant changes (p < 0.0001) from baseline to six weeks. Moreover, significant changes (PD = 0.0001, CAL = 0.0001, PI = 0.042 and BOP = 0.0001) were also observed between the control and test sides. Gender showed no significance on periodontal variables (p > 0.05) except PD on the test side (p = 0.030). In addition, periodontal stages and grades showed no significant changes (p > 0.05) in any periodontal variables on both sides. Ozone therapy significantly improves the periodontal condition compared to SRP treatment alone. However, the stages and grades of periodontitis do not influence the outcome of ozone therapy.