Superficially Bound Acetylcholinesterase Based on a Chitosan Matrix for Neurotoxic Compound Assay by a Photographic Technique

• Published in: Analytical letters 2018-07, Vol.51 (10), p.1622-1632
• Main Authors: Kostelnik, Adam, Pohanka, Miroslav
• Format: Article
• Language: English
• Subjects: Acetic acid; Acetylcholinesterase; Analytical chemistry; Assaying; Chitosan; Choline; colorimetry; Enzymes; immobilization; indoxylacetate; inhibitor; Inhibitors; Laboratory equipment; Model testing; Neurotoxicity; Photography; Smartphones; Spectrophotometry; Three dimensional printing
• ISSN: 0003-2719; 1532-236X
• DOI: 10.1080/00032719.2017.1381846

Smartphones have become popular in the last decade and they have been used in analytical chemistry as easy detection systems with comparable parameters to standard laboratory equipment. This work focuses on the attachment of enzyme acetylcholinesterase on the previously activated chitosan. Acetylcholine splits the neurotransmitter acetylcholine, as a natural substrate, into choline and acetic acid. However, this compound can cleave alternative substrates, e.g., indoxyl acetate, which was used in this work. The conversion of the substrate is blocked or slowed down by inhibitors from which galantamine and tacrine were tested as model inhibitors with detection limits of 1.1 and 0.18 µM, respectively. The measurement procedure was performed on a three-dimensional printed holder and red-green-blue channels were used for digital photography evaluation. The method was validated using a standard Ellman's spectrophotometric assay. We successfully attached acetylcholinesterase on the chitosan surface that was used for the inhibitor assay. The long-term stability of the immobilized enzyme as well as the sensitivity to organic solvents were also tested. The proposed method appeared to be suitable for the rapid and inexpensive assay of neurotoxic compounds.