I) Patients:

The present study will be carried on periodontitis patients. The patients will be informed about the treatment steps. This includes the possible effects or risks, and other treatment options according to the rules of the Ethical Committee of Faculty of dentistry, Mansoura University.

Groups:

Group I (negative control): 15 periodontally healthy subjects. Group II (positive control): 15 patients will be treated with mechanical debridement only.

Group III (study group A): 15 patients will be treated with mechanical debridement followed by delivery of sunflower seed oil gel for each pocket. (J MAC BOTANICALS, organic cold pressed helianthus annuus sunflower seed oil, Waterglider LLC. Spokane Valley, WA 99216, USA).

Group IV (study group B): 15 patients will be treated with mechanical debridement followed by delivery of ozonated sunflower seed oil gel for each pocket.( PurO3, Ozonated Sunflower Oil combines organic sunflower oil and ozone, giving you an oxygen-rich, soothing gel with high amounts of ozone, 1220 S Happy Hollow Rd Fayetteville, AR 72701, USA).

Periodontal evaluation will be performed for all patients at baseline and after three months including the following parameters:

• Plaque index (PI).(15)
• Gingival index.(16)
• Probing pocket depth (PPD).(17)
• Clinical attachment level (CAL).(18) B) Gingival crevicular fluid sampling. C) Local drug delivery Adequate amount of prepared ozonated sunflower and sunflower oil gel will be delivered into the selected periodontal pockets.

B) Immunological analysis of TNF-α in GCF

I) Patients:

The present study will be carried on periodontitis patients. The patient will be selected from Out Patient Clinic, Department of Oral Medicine, Periodontology, Diagnosis and Oral Radiology, Faculty of Dentistry, Mansoura University. The patients will be informed about the treatment steps. This includes the possible effects or risks, and other treatment options according to the rules of the Ethical Committee of Faculty of dentistry, Mansoura University. The patients must be legally competent to provide written consent before performing any required steps.

A)Inclusion Criteria

1. Patients with Stage II Periodontitis grade B periodontitis (horizontal bone loss with maximum probing depth ≤ 5 mm and with 3-4 mm clinical attachment loss).
2. Minimum of 20 remaining teeth.
3. Age range (25 - 50) years.
4. Good compliance with the plaque control instructions following initial therapy.
5. Availability for follow up and maintenance program. B) Exclusion Criteria

1. Systemic diseases / drugs which could influence the outcome of the therapy (According to Cornell Medical Index-Health Questionnaire).

2. Receiving periodontal therapy in past 6 month. 3. Pregnant and lactating females. 4. Vulnerable groups of patients (e.g handicapped patients and decisionally impaired individuals).

5. Smoking.

D) Groups:

Group I (negative control): 15 periodontally healthy subjects. Group II (positive control): 15 patients will be treated with mechanical debridement only.

Group III (study group A): 15 patients will be treated with mechanical debridement followed by delivery of sunflower seed oil gel for each pocket. (J MAC BOTANICALS, organic cold pressed helianthus annuus sunflower seed oil, Waterglider LLC. Spokane Valley, WA 99216, USA).

Group IV (study group B): 15 patients will be treated with mechanical debridement followed by delivery of ozonated sunflower seed oil gel for each pocket.( PurO3, Ozonated Sunflower Oil combines organic sunflower oil and ozone, giving you an oxygen-rich, soothing gel with high amounts of ozone, 1220 S Happy Hollow Rd Fayetteville, AR 72701, USA).

II) Clinical Procedures:

• Medical and dental histories will be taken from patients.
• Mechanical debridement will be performed using suitable ultrasonic and hand instruments for positive control and study groups.

A) Periodontal assessment:

Periodontal evaluation will be performed for all patients at baseline and after three months including the following parameters:

• Plaque index (PI).(15)
• Gingival index.(16)
• Probing pocket depth (PPD).(17)
• Clinical attachment level (CAL).(18)

B) Gingival crevicular fluid sampling. The GCF samples will be collected from all subjects at baseline and three months after treatment.(19)

C) Local drug delivery Adequate amount of prepared ozonated sunflower and sunflower oil gel will be delivered into the selected periodontal pockets.

III) Laboratory procedures

A) Pharmaceutics Section: Formulation development:

1. Sunflower oil and derevatives:

   The study will involve using sunflower oil as pure form and/or ozonated using controlled conditions. The oils will be used in preparation of gel formulation for applications in the mouth cavity using different formulation techniques.

2. Development of Formulations:

Based on the oil formulation, various gels will be developed with a focus on creating formulations suitable for clinical application.

• Different gel bases will be evaluated to optimize their physicochemical characteristics.
• Physicochemical Parameters of the gel formulations will be assessed to ensure the desired properties.
• The developed gels will undergo in-vitro testing under different conditions.
• Once optimized, the selected gel formulation(s) will be prepared for testing in clinical trials.

B) Immunological analysis of TNF-α in GCF

Assessment of crevicular fluid TNF-α will be performed using enzyme-linked immunosorbent assay technique

Inclusion Criteria:

1. Patients with Stage II Periodontitis grade B periodontitis (horizontal bone loss with maximum probing depth ≤ 5 mm and with 3-4 mm clinical attachment loss).
2. Minimum of 20 remaining teeth.
3. Age range (25 - 50) years.
4. Good compliance with the plaque control instructions following initial therapy.
5. Availability for follow up and maintenance program.

Exclusion Criteria:

1. Systemic diseases / drugs which could influence the outcome of the therapy (According to Cornell Medical Index-Health Questionnaire).
2. Receiving periodontal therapy in past 6 month.
3. Pregnant and lactating females.
4. Vulnerable groups of patients (e.g handicapped patients and decisionally impaired individuals).
5. Smoking.