4.5.2. Inhibition of Acetylcholinesterase

The acetylcholinesterase (AChE) inhibitory activity assay was measured by the Ellman colorimetric method [53]. A 50-µL appropriate diluted sample solution, 20 μL of acetylcholine (0.2 U/mL), and 90 µL of Ellman’s reagent (containing 15 µL of 15 mM ATCI and 75 µL of 3 mM DTNB) were added to a 96-well plate in darkness for 10 min. Finally, 20 μL of 0.2 U/mL AChE was added and kept in darkness for 5 min, and the absorbance was read at 405 nm (As). Methanol in place of the sample was used as a blank (Ab), phosphate buffer in place of acetylcholine was used as a negative control (Ac), and galanthamine was used as a positive control. The inhibition rate calculated was similar to Equation (2).