Aim: Evaluation and comparison of the antimicrobial effi ciency of the medicinal plant extracts, when used as root canal irrigants, on single species of Enterococcus faecalis. Materials and Methods: A comparative bioactivity-guided fractionation of four medicinal plants was carried out using solvents with diff erent polarities (ethanol, hexane, ethyl acetate, butanol, and water). The organism was freshly inoculated in brain-heart infusion broth for 24 h prior to the test, and the inoculum was standardized. Cultures were spread over the surface of the plate, and wells were punched out in every plate. Each well was inoculated by each extract and incubated for 24 h and the zone of inhibition around each well was recorded. A total of 50 extracted human single rooted teeth were disinfected decapitated and prepared, then divided into six groups: Group I: Neem leaf extract, Group II: Ginger extract, Group III: Miswak extract, Group IV: Lemon solution, Group V: NaOCl 5.25% (+ve control), and Group IV: Saline (−ve control). Each root canal was completely fi lled with bacterial suspension and incubated at 37°C for 24 h, then completely fi lled with one of the medicinal plants extract according to each group. Root specimens were then incubated for 24 h at 37°C. The root canals were sampled, and the samples were contained and incubated for 24 h at 37°C. The colony forming units (CFU) were calculated. Data presented as mean and standard deviation values. One-way analysis of variance (ANOVA)was used for comparisons between more than two groups. Tukey's post-hoc test was used for pair-wise comparison between the groups when ANOVA test was signifi cant. Results: NaOCl 5.25% showed signifi cantly less CFU/ml count, when compared with all tested plant extracts (P < 0.001). This was followed by fresh lemon solution which showed signifi cantly less CFU/ml count when compared with all of the other tested plant extracts. Butanol fraction of ginger showed signifi cantly less CFU/ml count than ethyl acetate fraction of miswak and neem. Conclusions: Medicinal herbs may off er a new source of antibacterial agents for use; medicinal plants are natural antimicrobial agents and might be used in the development of a promising irrigants, which might be safer than other chemical compounds used in the endodontic treatment process.