Aim: The aim of present study is to compare and evaluate the changes in microhardness of root canal dentin following application of different irrigating solutuions. Materials and methods: A total of 50 freshly extracted single rooted teeth were selected and decoronated at 15 mm from root apex. Root canal instrumentation was done using step back technique and roots were sectioned longitudinally using diamond disc dividing into two halves; which were then polished and embedded into acrylic blocks separately. Half section of each specimen was sent for pretreatment microhardness testing whereas other half was immersed in different irrigating solution as per group in closed glass plate for 5 minutes. 50 specimens were divided into 5 groups of 10 teeth according to the irrigating solution used. Group A : 3% Sodium hypochlorite, Group B : 17% EDTA (Ethylene diamine Tetraacetic acid), Group C : 2% Chlorhexidine, Group D : Chloroquick solution, Group E : Saline (Control group). All the samples were placed in respective irrigant for 5 minutes at 37°C. All the tested specimens were then subjected to microhardness testing using Vickers hardness tester. Readings were recorded and statistical analysis was done using One–way ANOVA and Tukey’s Post Hoc test. Results: 1) The result of present study showed that microhardness of root canal dentin decreases following application of different irrigating solutions. 2) The microhardness of root canal dentin was found to be comparatively least affected by using Chloroquick irrigating solution followed by 2% Chlorhexidine, 3% Sodium hypochlorite and 17% EDTA. Conclusion: Chloroquick is an efficient root canal irrigant, as it was found to be least detrimental to root dentin microhardness. Besides, it has additional advantages like antimicrobial properties and efficient in smear layer removal