Inhibition of carbonic anhydrase (CA) has emerged as a promising approach for the treatment of a variety of diseases such as glaucoma, epilepsy, obesity and cancer. As a result, the design of CA inhibitors (CAIs) is an outstanding field of medicinal chemistry. Due to the therapeutic potential of isoindoline-1,3-diones as CAIs, herein hCA I and hCA II isozymes were purified from human erythrocytes using affinity chromatography and the inhibitory effects of a series of isoindoline-1,3-diones on hydratase activities of these isozymes were investigated. Among these compounds, compound 3a was found to be the most effective compound on hCA I with an IC50 value of 7.02 mu M, whereas compound 3c was the most potent compound on hCA II with an IC50 value of 6.39 mu M. Moreover, molecular docking studies were carried out for all compounds and acetazolamide (AAZ), the reference agent, in the active sites of hCA I and hCA II. Generally, the compounds showed high affinity through salt bridge formation and metal coordination with Zn2+ ion and pi-stacking interaction with His94 residue. According to in silico Absorption, Distribution, Metabolism and Excretion (ADME) studies, the pharmacokinetic parameters of all compounds were within the acceptable range. (C) 2019 Elsevier B.V. All rights reserved.