Purpose: This study compared the effects of different irrigation solutions on the microhardness of root dentin. Methods: Fifty single-rooted human teeth were chosen, and the roots were randomly divided into five groups (n = 10) according to the irrigant used for 5 min: 17% ethylenediaminetetraacetic acid (EDTA), 2% boric acid (BA), 5% BA, 10% BA, and distilled water (DW). After the irrigation procedure, dentin surface microhardness was calculated using a Vickers indenter 100 μm from the root canal lumen. Comparisons between the groups were performed with a two-way ANOVA test and and Tukey’s multiple comparison test (p = 0.05).