Background: Chemo-mechanical preparation is essential for the debridement of root canal, aiming to remove microorganisms, pulpal tissues and smear layer. Conventional techniques are not enough, incorporation of nanoparticles with ultrasonic activation is suggested to improve the antimicrobial properties of an endodontic irrigant. Aim: The present study was done to compare the efficiency of magnesium oxide in the nano form with NaOCL when used as an endodontic irrigants against E. Faecalis biofilm. Furthermore, the influence of ultrasonic activation on the irrigant's action was evaluated. Materials and methods: Sixty intact human permanent single rooted teeth with mature apices were decoronated, instrumented to size #50, sterilized and incubated with culture suspension of Enterococcus Faecalis. Samples were divided into three groups equally according to the type of irrigant used. Group 1 were irrigated by MgO nanoparticles, group 2 were irrigated by NaOCL 5.25% and group 3 were irrigated by saline. Each group were subdivided into two groups equally A and B. Subgroups A were irrigated passively, subgroups B the irrigant was ultrasonically activated 5 min, 25 HZ. Antimicrobial efficacy was evaluated by counting colony forming units. Results: Ultrasonic activation of nano MgO was the best in bacterial reduction followed by activated NaOCL. Conclusions: MgO in the nano form exhibit superior antibacterial action and its action was inhanced by ultrasonic activation.