Background The gold standard in treatment of periodontitis is mechanical removing of dental biofilm but using local delivery drugs as adjunctive to SRP is widely used to modulate inflammatory host and eradicate microbes. Tea tree oil (TTO) has a broad-spectrum antimicrobial, anti-inflammatory, antifungal, antiviral, antioxidant effect. This study aimed to assess clinically and biochemically the effect of intrapocket application of TTO (Melaleuca alternifolia) gel adjunctive to scaling and root planing (SRP) in the treatment of stage 2 (moderate) periodontitis and to correlate the biochemical levels with clinical response. Methods A randomized, controlled clinical trial was conducted on thirty patients with stage 2 periodontitis. Patients were equally divided into two groups: Control Group treated with (SRP) alone and Test Group treated with SRP and locally delivered 5% TTO gel. Clinical assessment included pocket probing depth (PPD), clinical attachment loss (CAL), gingival index (GI) and bleeding on probing (BOP) measured at baseline and after 3 and 6 months. The level of matrix metalloproteinase-8 (MMP-8), in the gingival crevicular fluid (GCF) was also assessed at baseline and after1, 3 and 6 months by Enzyme-linked immunosorbent assay (ELISA) kit. Chi-square, Student t- tests, Mann–Whitney U test and Spearman correlation were the statistical tests used in the study. Results An improvement of all clinical and biochemical parameters was observed (at p < 0.001) in both groups. A significant difference between the two groups was found in both clinical and biochemical parameters. Conclusion The local delivery of TTO gel adjunctive to SRP proved to be effective in the treatment of stage II periodontitis. Trial registration The study was retrospectively registered at clinicaltrials.gov NCT04769271, on 24/2/2021.