| Assay Method Information | |
| | AChE and BChE Inhibition Assay |
| Description: | To evaluate in vitro AChE activity, modified Ellman's method was performed using a 96-well plate reader (BioTek ELx808, Highland Park, IL, USA). Each well contained 50 µL potassium phosphate buffer (KH2PO4/K2HPO4, 0.1 M, pH 8), 25 µL sample dissolved in 50% methanol and 50% DMSO, and 25 µL enzyme (final concentration 0.22 U/mL in buffer). They were preincubated for 20 min at room temperature, and then, 125 µL DTNB (3 mM in buffer) was added. Characterization of the hydrolysis of ATCI catalyzed by AChE was performed spectrometrically at 405 nm followed by the addition of substrate (ATCI 3 mM in water). The change in absorbance was measured at 405 nm after 20 min. The IC50 values were determined graphically from inhibition curves (log inhibitor concentration versus percent of inhibition). A control experiment was performed under the same conditions without inhibitor and the blank contained buffer, DMSO, DTNB, and substrate. The described method was also used for BChE inhibition assa |
| Affinity data for this assay | |
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