Assad Assad Amara Matouga¹, Una Mohamed El-Shinnawi², Hesham AL-Sharkawy³, Reham Mohammed El-Farahaty^{4 1} BDS, Faculty of Dentistry, West Mountain University, Libya. ² Professor of Oral medicine, Periodontology, X-Ray & Diagnosis, Faculty of Dentistry, Mansoura University, Mansoura, Egypt. ³ Associate Professor of Oral medicine, Periodontology, X-Ray & Diagnosis, Faculty of Dentistry, Mansoura University, Mansoura, Egypt. ⁴ Associate Professor of Clinical Pathology, Faculty of Medicine, Mansoura University, Mansoura, Egypt. Abstract: Objectives: Evaluate the level of 25-Hydroxy vitamin D3 and osteocalcin in GCF and serum befor and after scaling and root planning in chronic periodontitis patients. Methods: A total of forty patients (20 with moderate to severe chronic periodontitis (study group) and 20 chronic gingivitis patients were chosen as a control group) were selectively collected for contribution in the present study. The diseased patients (study group) received initial periodontal therapy (SRP). Gingival crevicular fluid (GCF) and serum sample were collected at baseline and six weeks after therapy for study group and at baseline for control group. Systemic and local levels of 25-hydroxy vitamin D3, osteocalcin were measured using radioimmunoassay or enzyme-linked immunosorbent assay kits and compared. Results: The respective local osteocalcin level are significantly dropped from baseline to six weeks after (SRP) (9.56ng/ml versus 7.38ng/ml, P=0.001). The respective systemic osteocalcin level significantly increased after six weeks from SRP (10.85ng/ml versus 17.74 ng/ml, P=0.001) .The respective local 25-Hydroxy vitamin D3 level are significantly increased from baseline to six weeks after (SRP) (3.41ng/ml versus 4.57ng/ml, P=0.001). The respective systemic 25-Hydroxy vitamin D3 level significantly increased after six weeks from SRP (39.88ng/ml versus 41.48 ng/ml , P=0.001). Conclusions: Scaling and root planinig (SRP) is the mainstay of treatment of periodontal diseases as SRP was effective in improving clinical parameters in patients with chronic periodontitis. 25-hydroxy vitamin D3 might have an important role in the pathogenesis of periodontal disease and could be used as adjunctive therapeutic modality for the prevention and treatment of different types of periodontitis. Osteocalcin could be used as a potential diagnostic marker for periodontal disease activity in both serum and gingival crevicular fluid. Keywords: 25-Hydroxy Vitamin D3, Osteocalcin, Chronic Periodontitis.