To evaluate the effect of GSE, NaOCl, CHX and QMix as an antimicrobial agents against Enterococcus faecalis and their influence on flexural and ultimate tensile strength of root canal dentine.Root canals were divided into five groups (n=10) according to the substances used: 2.5% NaOCl, 2% CHX, 6.5% GSE, Qmix and control group (distilled water) (DW). Final irrigation was done with 17% EDTA in all groups, except when DW was used. The number of colony-forming units was used to evaluate the antimicrobial activity. Dentine beams were used to assess the flexural strength after treatment with substances as described before (n=10). The UTS was evaluated after the treatment of root dentine hourglass shape sections with the same substances (n=30).The lowest bacteria contamination was observed for CHX and GSE, while NaOCl and QMix showed an intermediate antimicrobial activity (p>0.05). NaOCl and QMix significantly reduced the mechanical properties of dentine (flexural strength and UTS) (p<0.05) and no statistical difference was found among CHX, GSE and DW (p>0.05).CHX and GSE were more effective than NaOCl and QMix against E. faecalis. Furthermore, they did not harm dentine mechanical properties as observed for NaOCl and QMix.The use of GSE can be recommended for endodontic procedures since it has good antimicrobial activity and does not interfere in the mechanical properties of dentine; similarly to CHX.