Electrochemical Determination of Activity of Acetylcholinesterase Immobilized on Magnetic Particles

Pohanka, Miroslav; Kostelník, Adam; Cegan, Alexander

doi:10.20964/2016.06.39

Cited by 11 publications

(7 citation statements)

References 33 publications

(25 reference statements)

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“…Despite broad use, this method has some drawbacks like instability of Ellman's reagent and hemoglobin interference [ 2 , 4 , 5 ]. Furthermore, there is possibility of measuring enzyme activity electrochemically [ 6 , 7 ]. Colorimetric detection can be also based on pH measurement.…”

Section: Introductionmentioning

confidence: 99%

Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone

Kostelník

Cegan

Pohanka

2017

International Journal of Analytical Chemistry

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Smartphones are widely spread and their usage does not require any trained personnel. Recently, smartphones were successfully used in analytical chemistry as a simple detection tool in some applications. This paper focuses on immobilization of acetylcholinesterase (AChE) onto commercially available pH strips with stabilization in the gelatin membrane. AChE degrades acetylcholine into choline and acetic acid which causes color change of acid-base indicator. Smartphone served as a tool for measurement of indicator color change from red to orange while inhibitors blocked this process. AChE inhibitors were measured with limits of detection, 149 nM and 22.3 nM for galanthamine and donepezil, respectively. Organic solvents were measured for method interferences. Measurement procedure was performed on 3D printed holder and digital photography was evaluated using red-green-blue (RGB) channels. The invented assay was validated to the standard Ellman's test and verified on murine plasma samples spiked with inhibitors. We consider that the assay is fully suitable for practical performance.

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Section: Introductionmentioning

confidence: 99%

Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone

Kostelník

Cegan

Pohanka

2017

International Journal of Analytical Chemistry

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“…Mediator transfers the electron to the electrode via redox reactions. Oxidation and reduction process on cyclic voltammogram are referred to solid and dashed lines in Figure 3, respectively [18,19]. Electrons on the electrode are detected by external electrical circuit (potentiostat).…”

Section: Measurement Proceduresmentioning

confidence: 99%

Electrochemical acetylcholinesterase biosensor for detection of cholinesterase inhibitors: study with eserine

Lokar¹,

Kononenko²,

Drobne³

et al. 2019

Informacije MIDEM

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Cholinesterase inhibitors are widely used as pesticides, as chemical warfare agents and as drugs to treat symptoms of Alzheimer’s disease. Therefore, it is a high need to develop methods for their detection which are fast, sensitive, and reliable. This paper reports a preliminary work in the development of an electrochemical biosensor based on acetylcholinesterase (AChE) which is constructed by immobilization layers – cysteamine/glutaraldehyde/AChE on thin layer gold electrode for detection of cholinesterase inhibitors. Eserine (physostigmine) was used as a test inhibitor. The enzyme immobilization efficacy was evaluated by measuring activity of immobilized enzyme via Ellman’s method. The enzyme activity of the initial reduction of 33% in five days remained after that stable for at least one week. Chronoamperometric response to substrate acetylthiocholine chloride (ATCl) was assumed to follow Michaelis-Menten kinetics. After exposure biosensor to 25 mM eserine for 10 min, 70% inhibition of enzyme was detected. Reactivation factor of inhibited AChE was determined as 0.016 min-1.

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“…Measuring of AChE activity is important in diagnostics or serves as a tool in analytical chemistry in inhibitor determination. A commonly acknowledged method for activity determination is Ellman’s reaction but there is potential for pH or electrochemical detection [ 2 , 3 , 4 , 5 , 6 , 7 ]. There is a demand for the determination of AChE inhibitors in a wide spectrum of used compounds like Alzheimer’s disease drugs (donepezil, rivastigmine, huperzine, galantamine), pesticides (carbofuran, malaoxon, malathion), and chemical warfare agents (sarin, soman, tabun, VX) [ 8 ].…”

Section: Introductionmentioning

confidence: 99%

“…MPs have been used for many years in protein immobilization as a good platform for the attachment of proteins through free carboxyl, hydroxyl, thio, or amino groups on their surface [ 16 , 17 , 18 ]. Many protocols for enzyme immobilization on MPs have already been reported [ 4 , 19 , 20 , 21 ]. Silane-coupling reagents (like 3-aminopropyltrimethoxysilane or 3-aminopropyltriethoxysilane) are easily used for surface modification of synthetized MPs plus provide free amino groups which can be used for enzyme immobilization [ 22 , 23 , 24 ].…”

Section: Introductionmentioning

confidence: 99%

Construction of an Acetylcholinesterase Sensor Based on Synthesized Paramagnetic Nanoparticles, a Simple Tool for Neurotoxic Compounds Assay

Kostelník

Kopel

Cegan

et al. 2017

Sensors

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Magnetic particles (MPs) have been widely used in biological applications in recent years as a carrier for various molecules. Their big advantage is in repeated use of immobilized molecules including enzymes. Acetylcholinesterase (AChE) is an enzyme playing crucial role in neurotransmission and the enzyme is targeted by various molecules like Alzheimer’s drugs, pesticides and warfare agents. In this work, an electrochemical biosensor having AChE immobilized onto MPs and stabilized through glutaraldehyde (GA) molecule was proposed for assay of the neurotoxic compounds. The prepared nanoparticles were modified by pure AChE and they were used for the measurement anti-Alzheimer’s drug galantamine and carbamate pesticide carbofuran with limit of detection 1.5 µM and 20 nM, respectively. All measurements were carried out using screen-printed sensor with carbon working, silver reference, and carbon auxiliary electrode. Standard Ellman’s assay was used for validation measurement of both inhibitors. Part of this work was the elimination of reversible inhibitors represented by galantamine from the active site of AChE. For this purpose, we used a lower pH to get the original activity of AChE after inhibition by galantamine. We also observed decarbamylation of the AChE-carbofuran adduct. Influence of organic solvents to AChE as well as repeatability of measurement with MPs with AChE was also established.

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Electrochemical Determination of Activity of Acetylcholinesterase Immobilized on Magnetic Particles

Cited by 11 publications

References 33 publications

Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone

Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone

Electrochemical acetylcholinesterase biosensor for detection of cholinesterase inhibitors: study with eserine

Construction of an Acetylcholinesterase Sensor Based on Synthesized Paramagnetic Nanoparticles, a Simple Tool for Neurotoxic Compounds Assay

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