A rapid TLC bioautographic method for the detection of acetylcholinesterase and butyrylcholinesterase inhibitors in plants

Abstract

A simple and rapid bioautographic enzyme assay on TLC plates has been developed for the screening of acetylcholinesterase and butyrylcholinesterase inhibition by plant extracts. Enzyme activity was detected by the conversion of naphthyl acetate into naphthol and the formation of the corresponding purple-coloured diazonium dye with Fast Blue B salt. Inhibitors of cholinesterases produced white spots on the dye-coloured background of the TLC plates. The alkaloids galanthamine and physostigmine, which are known inhibitors of acetylcholinesterase, were used to determine the sensitivity of the assay. Various plant extracts were tested using the bioassay.

Modified TLC bioautographic method for screening acetylcholinesterase inhibitors from plant extracts

“…1), while the least amount of physostigmine required was 1610 -5 lg (Fig. 2) which was far lower than published values [7].…”

“…However, the results will be uncertain when testing colored samples or hydrophobic samples. TLC bioautographic method has arisen in late years [6,7]. As a simple and rapid method, it has been used to screen and bioassay-guidedly isolate acetylcholinesterase inhibitors from plant extracts.…”

“…To overcome the disadvantages, we modified Marston's method [7] by reducing the concentration of the enzyme to 1.0 U/mL and changing the operations, the concentration of the substrate and the chromogenic reagent, and the time of the reaction correspondingly. As a result, the consumption of the enzyme was reduced by 85% and the detection limits were also decreased remarkably.…”

In vitroacetylcholinesterase activity of peptide derivatives isolated from two species of Leguminosae

“…The isolated compounds 1 and 2 were evaluated by TLC, which showed positive spots on the bioautographic TLC test for AChE activity using suitable Marston et al (2002) methodology. The in vitro inhibition of AChE was determined by the spectrophotometric method of Ellman et al (1961).…”

Determination of organophosphorus pesticide residues in vegetables by an enzyme inhibition method using α-naphthyl acetate esterase extracted from wheat flour

“…1) (Marston et al, 2002). The enzyme's activity was determined by the linear relationship between absorbance of purple-colored diazonium dye and time.…”