Introduction: Calcium hydroxide has been used with various vehicles in the treatment modalities of Endodontics. Owing to the increased antibacterial resistance, Ozone is currently being discussed as a possible alternative vehicle because of its reported high antimicrobial property without the development of drug resistance. Aim: The purpose of this study was to investigate and compare the effectiveness of calcium Hydroxide in combination with 2% chlorhexidine and ozonated chlorhexidine for elimination of E.faecalis bacteria. Materials and Methods: In this study aqueous ozone was used, it was prepared by passing gaseous ozone through CHX in a glass beaker. Minimal inhibitory concentration of ozonated CHX was obtained by serial dilution method. The samples were assigned to 3 groups: Group I- Calcium hydroxide with saline; Group II- Calcium hydroxide with 2% Chlorhexidine; Group IIICalcium hydroxide with ozonated chlorhexidine. Efficacy of intracanal medicaments against E.faecalis was carried out by disc diffusion method. Results were statistically analysed by one way ANOVA. Results: The antimicrobial effectiveness of the samples in Group II where ozone was ozonated with reduced percentage i.e. 0.12% was around 21mm which is almost equal to that of 2% CHX and the results were found to be statistically significant at 0.5. Conclusion: Ozonated CHX with reduced concentration possessed similar antimicrobial effect as conventional CHX in combination with calcium hydroxide.