The purpose of this study was to evaluate the effect of 17 % ethylenediaminetetraacitic acid (EDTA) at pH 7.21 and 10 % citric acid at pH 1.55 on the microhardness of root dentin. Thirty human mandibular premolars were split longitudinally and randomly divided into six groups. Specimens were treated as follows: Gr1: 17 % EDTA for 1 min, Gr2: 17 % EDTA for 3 min, Gr3: 17 % EDTA for 5 min, Gr4: 10 % citric acid for 1 min, Gr5: 10 % citric acid for 3 min and Gr6: 10 % citric acid for 5 min. Dentin microhardness was measured with a Knoop indenter under a 50-g load and a 15-seconds dwell time. Data were statistically analyzed using Paired sample t-test, One-way ANOVA and Independent samples t-test at 5 % significance level. The results indicated that both solutions significantly decreased the microhardness of root dentin when contacted at 1, 3 and 5 minutes. The reductions of the microhardness values of root dentins that were exposed to 10 % citric acid at 3 and 5 minutes were significantly higher than those of EDTA groups. Under the conditions of this study, 17 % EDTA at pH 7.21 could be safely used as an irrigating solution without a change of dentin surface microhardness when contact time is 1-5 minutes.