A colorimetric assay for acetylcholinesterase activity and inhibitor screening based on the thiocholine–induced inhibition of the oxidative power of MnO₂ nanosheets on 3,3′,5,5′–tetramethylbenzidine

The authors describe a colorimetric method for the determination of the activity of acetylcholinesterase (AChE). Manganese dioxide (MnO₂) nanosheets directly reacts with 3,3′,5,5′–tetramethylbenzidine (TMB) in the absence of hydrogen peroxide (H₂O₂). This leads to the formation of a blue product (oxTMB) with an absorption peak at 652 nm. If AChE hydrolyzes its substrate acetylthiocholine chloride, thiocholine is formed which blocks the oxidative power of the MnO₂ nanosheets. Hence, oxTMB will not be formed. The decreased absorbance is directly related to the AChE activity in the 0.01–1.0 mU·mL⁻¹ range. The detection limit is 0.01 mU·mL⁻¹ and the relative standard deviation is 1.2% (for n = 11 at 0.5 mU·mL⁻¹). The method was also applied to screen for inhibitors of AChE. .