OBJECTIVE The purpose of this study was to evaluate the effects of endodontic irrigants on the microhardness of root canal dentin. METHODS Thirty extracted single-rooted human teeth were used. The crowns were sectioned at the cementoenamel junction. Each root was transversely sectioned into cervical, middle, and apical segments, resulting in 90 specimens. The 3 sections of each root were separately mounted in an individual silicon device with acrylic resin. The specimens were randomly divided into the following 3 groups (n = 30), according to the irrigant solution used: (1) group 1, control (saline solution); (2) group 2, 2% chlorhexidine gluconate solution; and (3) group 3, 1% sodium hypochlorite (NaOCl). After 15 minutes of irrigation, dentin microhardness was measured on each section at 500 microm and 1000 microm from the pulp-dentin interface with a Vickers diamond microhardness tester in Vickers hardness number (VHN). RESULTS Data obtained were analyzed using analysis of variance and the Tukey test (5%). Specimens irrigated with 2% chlorhexidine (group 2) or 1% NaOCl (group 3) presented lower values of dentin microhardness, with significant difference in relation to the control group (P < .05). CONCLUSIONS It could be concluded that chlorhexidine and NaOCl solutions significantly reduced the microhardness of root canal dentin at 500 microm and 1000 microm from the pulp-dentin interface.