Enzyme colorimetric cellulose membrane bioactivity strips based on acetylcholinesterase immobilization for inhibitors preliminary screening

Abstract

To quickly screen the active pharmaceutical ingredient that can be used as acetylcholinesterase inhibitors (AChEIs) to treat Alzheimer's disease, an enzyme colorimetric cellulose membrane bioactivity strip (CBS) was developed for simple and rapid screening of AChEIs. The amino group of acetylcholinesterase (AChE) undergoes Schiff base reaction with the aldehyde group on the oxidized cellulose membranes, then the AChE was covalently cross-linking on the surface of cellulose membranes, enabling the screening based on Ellman's enzyme colorimetric method. When the enzyme activity of AChE was inhibited after incubation with inhibitors, the hydrolysis of S-Acetylthiocholine iodide decreased, consequently, the 5-thio-2-nitrobenzoic acid generated by the reaction with 5,5'-dithiobis (2-nitrobenzoic acid) also decreased, leading to a decreased color intensity. In addition, CBSs had fast chromogenic time, excellent specificity, and extraordinary storage stability. Tacrine and Donepezil were used as representative inhibitors during the detection, while their IC₅₀ and limit of detection were determined. Therefore, our work not only established a platform for effective preliminary screening of AChEIs but also inspired the further development of other cellulose membrane-based biosensors.

Keywords: Acetylcholinesterase immobilization; Cellulose membrane bioactivity strips; Enzyme colorimetric detection; Inhibitors screening.