The in vitro investigation of antiglaucoma effect of new acridine derivatives

Carbonic anhydrase (CA) (E.C. 4.2.1.1.) is a metalloenzyme that contains zinc ion in its active site. It catalyses that the reversible hydration of carbon dioxide to yield bicarbonate and proton in a two-step reaction. Human eye tissue contains CA I, CA II and CA IV isoenzymes. CA I and CA II isoenzymes are cytosolic, but CA IV isoenzyme is membrane-bound. Carbonic anhydrase inhibitors have been used for treatment of glaucoma, because of their decrease higher intraocular pressure which occurs with excessive secretion of aqueous humor.In this study, the inhibitory effects of new carbonic anhydrase inhibitors, which candidates for treatment of glaucoma, on this enzyme were investigated in vitro.Firstly, hCA I and hCA II isoenzymes were purified from human erythrocytes by using affinity gel (Sepharose-4B-L-tirosine-sulfanilamide). The qualitative and quantitative protein assay was made and the purity of the enzymes was checked with SDS-PAGE electrophoresis. Enzyme activities were determined with kinetic studies.Later, the inhibition effects of newly synthesized compounds (1-9) on hCA I and hCA II isoenzymes were determined. To determine inhibitory effects of the compounds, the hydratase and esterase activities of carbonic anhydrase enzyme were measured. %Activity vs. [I] graphics were drawn and the IC50 values were calculated for potential inhibitory compounds. The Ki constants were calculated from Lineweaver-Burk graphics.According to in vitro studies, any inhibition effects of 1-9 compounds were not observed on hydratase activity of hCA I and hCA II isoenzymes. These new compounds inhibited the esterase activities of hCA I and hCA II isoenzymes. The IC50 values of compounds for esterase activity are 0,71-0,27µM for hCA I and 0,52-0,21µM for hCA II, respectively. The Ki values of these inhibitors are 0,47-0,19µM for hCA I and 0,34-0,032µM for hCA II, respectively.Keywords: Glaucoma, Carbonic Anhydrase, Sulfonamides, Acridine Compounds, IC50 Values.