Dental disease prevention is commonly associated with a reduction of some gram positive and gram negative microorganism. Presently no such study is available regarding the in vivo effect of star anise.

The primary outcome measure used for this study will be to determine anti-inflammatory property and astringent property by Loe and Silness Gingival Index and papillary bleeding index respectively. The indices will be measured at baseline, 14^th day and after 28 days of administration of the mouthwash. After evaluating the findings in 28 days, they will be made to undergo complete ultrasonic scaling.

Antimicrobial activity: Subjects will be made comfortable and asked to rinse mouth with normal tap water. They will be asked to spit their saliva into the sterile plastic containers. About 3–4 ml of whole unstimulated saliva will be collected over 2 min. All the saliva samples of participants will be coded during the period of sample collection and processing. The samples collected will be handed over to the pathology laboratory for analysis on the same day. In the laboratory, samples will be stored at room temperature (17°C–25°C) prior to the analysis.  

All samples will be sonicated (Vibra Cell 400 W, Sonics & Materials Inc. – 5% amplitude, 9.9 s cycle, six pulses) and diluted in a saline solution (100 and 1000 times). The diluted samples (5 ul) will then spread on blood agar. The Petri dishes will be placed in an incubator (Jovan IG 150) with 10% CO₂ at 37°C for 48 hours. Subsequently, the dishes with blood agar will be placed in an aerobic incubator at 37°C, for an additional 24 hours. Total oral microorganism counts will be performed in blood agar. Bacterial colonies will be enumerated and the number of viable bacteria reported as colony forming units per ml (CFU/ml).

Thus, the aim of this study is to observe the antimicrobial activity of star anise against oral microorganism and to test the efficacy of star anise as astringent and anti-inflammatory agent.