Pohanka M


Full name : Pohanka Miroslav

First name : Miroslav

Mail : Centre of advanced studies\; Faculty of Military Healths Sciences\; University of Defense\; 50002 Hradec Kralove

Zip Code :

City :

Country : Czech Republic

Email : rau@atlas.cz

Phone : +420720427263

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References (83)

Title : Open Meter Duo: Low-Cost Instrument for Fluorimetric Determination of Cholinesterase Activity - Kerestes_2024_Sensors.(Basel)_24_
Author(s) : Kerestes O , Mozo JD , Pohanka M
Ref : Sensors (Basel) , 24 : , 2024
Abstract : Environmental screening is essential due to the increased occurrence of harmful substances in the environment. Open Meter Duo (OMD) is an open-source field photo/fluorimeter that uses an RGB diode that imitates a color according to the selected wavelength and uses a UV LED from the security kit diode as an excitation light source. The prepared PCB shield with a 3D-printed aperture was connected to Arduino UNO R4 WiFi. This system was used for the fluorescent detection of cholinesterase activity with the indoxyl acetate method. Carbofuran-a toxic pesticide-and donepezil-a drug used to treat Alzheimer's disease-were tested as model inhibitors of cholinesterase activity. The limit of detection of indoxyl acetate was 11.6 micromol/L, and the IC50 values of the inhibitors were evaluated. This system is optimized for wireless use in field analysis with added cloud support and power source. The time of analysis was 5 min for the fluorimetric assay and 20 min for the optional photometric assay. The time of field operation was approximately 4 h of continuous measurement. This system is ready to be used as a cheap and easy control platform for portable use in drug control and point-of-care testing.
ESTHER : Kerestes_2024_Sensors.(Basel)_24_
PubMedSearch : Kerestes_2024_Sensors.(Basel)_24_
PubMedID: 38544037

Title : Affordable Portable Platform for Classic Photometry and Low-Cost Determination of Cholinesterase Activity - Kerestes_2023_Biosensors.(Basel)_13_
Author(s) : Kerestes O , Pohanka M
Ref : Biosensors (Basel) , 13 : , 2023
Abstract : Excessive use of pesticides could potentially harm the environment for a long time. The reason for this is that the banned pesticide is still likely to be used incorrectly. Carbofuran and other banned pesticides that remain in the environment may also have a negative effect on human beings. In order to provide a better chance for effective environmental screening, this thesis describes a prototype of a photometer tested with cholinesterase to potentially detect pesticides in the environment. The open-source portable photodetection platform uses a color-programmable red, green and blue light-emitting diode (RGB LED) as a light source and a TSL230R light frequency sensor. Acetylcholinesterase from Electrophorus electricus (AChE) with high similarity to human AChE was used for biorecognition. The Ellman method was selected as a standard method. Two analytical approaches were applied: (1) subtraction of the output values after a certain period of time and (2) comparison of the slope values of the linear trend. The optimal preincubation time for carbofuran with AChE was 7 min. The limits of detection for carbofuran were 6.3 nmol/L for the kinetic assay and 13.5 nmol/L for the endpoint assay. The paper demonstrates that the open alternative for commercial photometry is equivalent. The concept based on the OS3P/OS3P could be used as a large-scale screening system.
ESTHER : Kerestes_2023_Biosensors.(Basel)_13_
PubMedSearch : Kerestes_2023_Biosensors.(Basel)_13_
PubMedID: 37366964

Title : Caffeine Role in the Age-Related Neurodegenerative Diseases, A Review - Pohanka_2022_Mini.Rev.Med.Chem__
Author(s) : Pohanka M
Ref : Mini Rev Med Chem , : , 2022
Abstract : Caffeine, a simple purine alkaloid with the proper chemical name 1,3,7-trimethylpurine-2,6-dione, is an abundant compound contained in beverages like coffee, food and drugs. It interacts with various pathways of which antagonism of adenosine receptors is the most significant but the other physiological pathways can be influenced by caffeine as well. Interaction with glutamate and dopamine neurotransmission pathways, competition with other substrates on cytochrome P450, non-competitive inhibition of acetylcholinesterase, blocking of nicotinic acetylcholine receptor and competitive inhibition of cyclic nucleotide phosphodiesterase can be mentioned. Because of caffeine availability as a part of foods, beverages and drugs, it has practical relevance even if the effect is weak. Intake of coffee containing edibles for a long period or even for a substantial part of life makes caffeines impact significant. Low acute and chronic toxicity of caffeine is another important specification. The discoveries from the last few years point to the fact that caffeine would interfere with the progression of some age-related neurodegenerative disorders like Alzheimer and Parkinson diseases and dementia with Lewy bodies. In this review article, the recent findings about caffeines impact on neurodegenerative diseases are presented and important facts about the caffeine effect including the substantial discoveries are described.
ESTHER : Pohanka_2022_Mini.Rev.Med.Chem__
PubMedSearch : Pohanka_2022_Mini.Rev.Med.Chem__
PubMedID: 35422212

Title : A Butyrylcholinesterase Camera Biosensor Tested for Carbofuran and Paraoxon Assay - Pohanka_2022_Int.J.Anal.Chem_2022_2623155
Author(s) : Pohanka M , Zakova J
Ref : Int J Analytical Chemistry , 2022 :2623155 , 2022
Abstract : Biosensors containing cholinesterase are analytical devices suitable for the assay of neurotoxic compounds. In the research on biosensors, a new platform has appeared some years ago. It is the digital photography and scoring of coloration (photogrammetry). In this paper, a colorimetric biosensor is constructed using 3D-printed multiwell pads treated with indoxylacetate as a chromogenic substrate and gold nanoparticles with the immobilized enzyme butyrylcholinesterase. A smartphone camera served for photogrammetry. The biosensor was tested for the assay of carbofuran and paraoxon ethyl as two types of covalently binding inhibitors: irreversible and pseudoirreversible. The biosensor exerted good sensitivity to the inhibitors and was able to detect carbofuran with a limit of detection for carbofuran 7.7 nmol/l and 17.6 nmol/l for paraoxon ethyl. A sample sized 25 microl was suitable for the assay lasting approximately 70 minutes. Up to 121 samples can be measured contemporary using one multiwell pad. The received data fully correlated with the standard spectrophotometry. The colorimetric biosensor exerts promising specifications and appears to be competitive to the other analytical procedures working on the principle of cholinesterase inhibition. Low-cost, simple, and portable design represent an advantage of the assay of the biosensor. Despite the overall simplicity, the biosensor can fully replace the standard spectroscopic methods.
ESTHER : Pohanka_2022_Int.J.Anal.Chem_2022_2623155
PubMedSearch : Pohanka_2022_Int.J.Anal.Chem_2022_2623155
PubMedID: 35432544

Title : Pharmacological Influencing of The Cholinergic Anti-inflammatory Pathway in Infectious Diseases and Inflammatory Pathologies - Pohanka_2021_Mini.Rev.Med.Chem_21_660
Author(s) : Pohanka M
Ref : Mini Rev Med Chem , 21 :660 , 2021
Abstract : The cholinergic anti-inflammatory pathway is a part of the parasympathetic nervous system and it can also be entitled as an anti-inflammatory reflex. It consists of terminations of the vagal nerve into blood, acetylcholine released from the terminations, macrophages and other cells having alpha7 nicotinic acetylcholine receptor (alpha7 nAChR), calcium ions crossing through the receptor and interacting with nuclear factors, and erythrocytes with acetylcholinesterase (AChE) terminating the neurotransmission. Stopping of inflammatory cytokines production is the major task for the cholinergic antiinflammatory pathway. The cholinergic anti-inflammatory pathway can be stimulated or suppressed by agonizing or antagonizing alpha7 nAChR or by inhibition of AChE. This review is focused on cholinergic anti-inflammatory pathway regulation by drugs. Compounds that inhibit cholinesterases (for instance, huperzine, rivastigmine, galantamine), and their impact on the cholinergic anti-inflammatory pathway are discussed here and a survey of actual literature is provided.
ESTHER : Pohanka_2021_Mini.Rev.Med.Chem_21_660
PubMedSearch : Pohanka_2021_Mini.Rev.Med.Chem_21_660
PubMedID: 33208075

Title : A Smartphone Camera Colorimetric Assay of Acetylcholinesterase and Butyrylcholinesterase Activity - Pohanka_2021_Sensors.(Basel)_21_
Author(s) : Pohanka M , Zakova J
Ref : Sensors (Basel) , 21 : , 2021
Abstract : Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) can serve as biochemical markers of various pathologies like liver disfunction and poisonings by nerve agents. Ellman's assay is the standard spectrophotometric method to measure cholinesterase activity in clinical laboratories. The authors present a new colorimetric test to assess AChE and BChE activity in biological samples using chromogenic reagents, treated 3D-printed measuring pads and a smartphone camera as a signal detector. Multiwell pads treated with reagent substrates 2,6-dichlorophenolindophenyl acetate, indoxylacetate, ethoxyresorufin and methoxyresorufin were prepared and tested for AChE and BChE. In the experiments, 3D-printed pads containing indoxylacetate as a chromogenic substrate were optimal for analytical purposes. The best results were achieved using the red (R) channel, where the limit of detection was 4.05 microkat/mL for BChE and 4.38 microkat/mL for AChE using a 40 microL sample and a 60 min assay. The major advantage of this method is its overall simplicity, as samples are applied directly without any specific treatment or added reagents. The assay was also validated to the standard Ellman's assay using human plasma samples. In conclusion, this smartphone camera-based colorimetric assay appears to have practical applicability and to be a suitable method for point-of-care testing because it does not require specific manipulation, additional education of staff or use of sophisticated analytical instruments.
ESTHER : Pohanka_2021_Sensors.(Basel)_21_
PubMedSearch : Pohanka_2021_Sensors.(Basel)_21_
PubMedID: 33807562

Title : Diagnoses of Pathological States Based on Acetylcholinesterase and Butyrylcholinesterase - Pohanka_2020_Curr.Med.Chem_27_2994
Author(s) : Pohanka M
Ref : Curr Med Chem , 27 :2994 , 2020
Abstract : Two cholinesterases exist: Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). While AChE plays a crucial role in neurotransmissions, BChE has no specific function apart from the detoxification of some drugs and secondary metabolites from plants. Thus, both AChE and BChE can serve as biochemical markers of various pathologies. Poisoning by nerve agents like sarin, soman, tabun, VX, novichok and overdosing by drugs used in some neurodegenerative disorders like Alzheimers disease and myasthenia gravis, as well as poisoning by organophosphorus pesticides are relevant to this issue. But it appears that changes in these enzymes take place in other processes including oxidative stress, inflammation, some types of cancer and genetically conditioned diseases. In this review, the cholinesterases are introduced, the mechanism of inhibitors action is explained and the relations between the cholinesterases and pathologies are explained.
ESTHER : Pohanka_2020_Curr.Med.Chem_27_2994
PubMedSearch : Pohanka_2020_Curr.Med.Chem_27_2994
PubMedID: 30706778

Title : Inhibitors of Cholinesterases in Pharmacology: the Current Trends - Pohanka_2020_Mini.Rev.Med.Chem_20_1532
Author(s) : Pohanka M
Ref : Mini Rev Med Chem , 20 :1532 , 2020
Abstract : Inhibitors of cholinesterases are a wide group of low molecular weight compounds with a significant role in the current pharmacology. Besides the pharmacological importance, they are also known as toxic compounds like military nerve agents. In the pharmacology, drugs for Alzheimer disease, myasthenia gravis and prophylaxis of poisoning by nerve agents can be mentioned as the relevant applications. Besides this, anti-inflammation and antiphrastic drugs are other pharmacological applications of these inhibitors. This review is focused on a survey of cholinesterase inhibitors with known or expected pharmacological impact and indications of their use. Recent literature with comments is provided here as well.
ESTHER : Pohanka_2020_Mini.Rev.Med.Chem_20_1532
PubMedSearch : Pohanka_2020_Mini.Rev.Med.Chem_20_1532
PubMedID: 31656151

Title : Indoxyl Acetate as a Substrate for Analysis of Lipase Activity - Valek_2019_Int.J.Anal.Chem_2019_8538340
Author(s) : Valek T , Kostelnik A , Valkova P , Pohanka M
Ref : Int J Analytical Chemistry , 2019 :8538340 , 2019
Abstract : Lipases play a crucial role in metabolism of microbes, fungi, plants, and animals, and in analytical chemistry, they are often used in detection of fats and triglycerides. Determination of lipase activity is also important in toxicology, when lipase activity can be both increased and decreased by organophosphates and other pesticides and in medicine for diagnosis of heart diseases. The standard method for lipase activity determination is based on cleaving ester bonds in lipase buffer containing Tween. Our aim was to find a method with faster and more sensitive response. It is known that acetylcholinesterase belongs to the same group of hydrolases enzymes as lipases and it cleaves indoxyl acetate, so we assume indoxyl acetate could report a similar reaction with lipase. Our method is based on indoxyl acetate as a substrate for lipase, where indoxyl acetate is cleaved by lipase to indoxyl and acetate moiety and blue indigo is created. The method was optimized for different times and amount of enzyme and compared with the standard Tween assay. The calibration curve measured in reaction time 20 minutes with 10 mul of lipase exhibited the best analytical parameters, and it showed Michaelis-Menten response with the Michaelis-Menten constant equal to 8.72 mmol/l. The indoxyl acetate-based method showed faster and more sensitive response than the standard method for lipase activity determination, so it has great potential in biosensor construction and it could be used in industry, medicine, toxicology, and common practice where the activity of lipases is need to be measured.
ESTHER : Valek_2019_Int.J.Anal.Chem_2019_8538340
PubMedSearch : Valek_2019_Int.J.Anal.Chem_2019_8538340
PubMedID: 31885593

Title : Biosensors and Bioassays Based on Lipases, Principles and Applications, a Review - Pohanka_2019_Molecules_24_
Author(s) : Pohanka M
Ref : Molecules , 24 : , 2019
Abstract : Lipases are enzymes responsible for the conversion of triglycerides and other esterified substrates, they are involved in the basic metabolism of a wide number of organisms, from a simple microorganism and to mammals. They also have broad applicability in many fields from which industrial biotechnology, the production of cleaning agents, and pharmacy are the most important. The use of lipases in analytical chemistry where it can serve as a part of biosensors or bioassays is an application of growing interest and has become another important use. This review is focused on the description of lipases chemistry, their current applications and the methods for their assay measurement. Examples of bioassays and biosensors, including their physical and chemical principles, performance for specific substrates, and discussion of their relevance, are given in this work.
ESTHER : Pohanka_2019_Molecules_24_
PubMedSearch : Pohanka_2019_Molecules_24_
PubMedID: 30744203

Title : Copper and copper nanoparticles toxicity and their impact on basic functions in the body - Pohanka_2019_Bratisl.Lek.Listy_120_397
Author(s) : Pohanka M
Ref : Bratislavske Lekarske Listy , 120 :397 , 2019
Abstract : Copper is a biogenic metal having multiple functions in basic processes in organisms and it is common in all kingdoms of life. Limited intake of copper is a problem; however, doses of copper exceeding the recommended alimentary source are problematic as well and toxicity is soon manifested. Impact of copper nanoparticles on human health is another serious issue taken into consideration in this review. Regarding to copper toxicity, neurodegenerative disorders including Alzheimer and Parkinson diseases are suspected to be linked to copper toxicity or copper can contribute to their progression. Wilson and Menke diseases are also described as examples of copper intolerance. This paper is focused on the description, literature survey and discussion of the current knowledge about copper and copper nanoparticles toxicity and their involvement in various pathological processes (Tab. 6, Fig. 2, Ref. 171). Keywords: reactive oxygen species; Alzheimer disease; acetylcholinesterase; copper; Fenton reaction; disorder; heavy metal; pollution; nanoparticle.
ESTHER : Pohanka_2019_Bratisl.Lek.Listy_120_397
PubMedSearch : Pohanka_2019_Bratisl.Lek.Listy_120_397
PubMedID: 31223019

Title : Inhibition of Acetylcholinesterase and Butyrylcholinesterase by a Plant Secondary Metabolite Boldine - Kostelnik_2018_Biomed.Res.Int_2018_9634349
Author(s) : Kostelnik A , Pohanka M
Ref : Biomed Res Int , 2018 :9634349 , 2018
Abstract : Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are two enzymes sensitive to various chemical compounds having ability to bind to crucial parts of these enzymes. Boldine is a natural alkaloid and it was mentioned in some older works that it can inhibit some kinds of AChE. We reinvestigated this effect on AChE and also on BChE using acetyl (butyryl) thiocholine and Ellman's reagents as standard substances for spectrophotometric assay. We found out IC50 of AChE equal to 372 mumol/l and a similar level to BChE, 321 mumol/l. We conclude our experiment by a finding that boldine is cholinesterase inhibitor; however we report significantly weaker inhibition than that suggested in literature. Likewise, we tried to investigate the mechanism of inhibition and completed it with in silico study. Potential toxic effect on cholinesterases in real conditions is also discussed.
ESTHER : Kostelnik_2018_Biomed.Res.Int_2018_9634349
PubMedSearch : Kostelnik_2018_Biomed.Res.Int_2018_9634349
PubMedID: 29850593

Title : [Cholinesterase activity assays and their use in the diagnosis of various pathological states including poisoning by neurotoxic agents] - Pohanka_2017_Ceska.Slov.Farm_66_147
Author(s) : Pohanka M
Ref : Ceska a Slovenska Farmacie , 66 :147 , 2017
Abstract : Cholinesterases are enzymes important for some nerve transmissions where the enzyme acetylcholinesterase plays a crucial role. The second enzyme, butyrylcholinesterase, is not necessary for the neurotransmission but it is involved in some detoxification reactions. A survey of literature, a discussion of diagnostic importance and the methods for an activity assay are presented in this review article. Liver failures, exposure to neurotoxic compounds, genetic dispositions are outlined here. In the field of assays, spectrophotometric, colorimetric and electrochemical tests are discussed.Key words: acetylcholinesterase butyrylcholinesterase poisoning liver function test pesticide nerve agent Alzheimer disease pathological state.
ESTHER : Pohanka_2017_Ceska.Slov.Farm_66_147
PubMedSearch : Pohanka_2017_Ceska.Slov.Farm_66_147
PubMedID: 29351376

Title : Galantamine has impact on immunity in mice exposed to keyhole limpet hemocyanin - Pohanka_2017_Bratisl.Lek.Listy_118_9
Author(s) : Pohanka M , Zakova J , Fusek J
Ref : Bratislavske Lekarske Listy , 118 :9 , 2017
Abstract : OBJECTIVES: In this work, we hypothesized whether galantamine could interact with the cholinergic anti-inflammatory pathway and modulate immunity this way. BACKGROUND: Galantamine is a drug used for the therapy of Alzheimer disease. The drug inhibits enzyme acetylcholinesterase in the central nervous system, which causes better availability of neurotransmitter acetylcholine.
METHODS: In the experiment, we immunized BALB/c laboratory mice by keyhole limpet hemocyanin (KLH) in combination with galantamine in a dose 0.02-0.5 mg/kg. The animals were sacrificed from 1 to 7 days after the substances applications and plasma was collected in order to examine immunochemical markers by enzyme-linked immunosorbent assay.
RESULTS: We found significant drop in production of immunoglobulins and interleukin (IL) 4 level while IL2, IL4 and tumour necrosis factor alpha remained unaltered for the whole experiment. We infer that galantamine causes better availability of acetylcholine also in blood system, where the neurotransmitter interacts with nicotinic acetylcholine receptors on macrophages and initiates cholinergic anti-inflammatory pathway.
CONCLUSIONS: In a conclusion, galantamine can cause lower efficacy of vaccination or immunity response to an infectious disease and the phenomenon should be taken into consideration in the current therapy (Tab. 1, Fig. 2, Ref. 24).
ESTHER : Pohanka_2017_Bratisl.Lek.Listy_118_9
PubMedSearch : Pohanka_2017_Bratisl.Lek.Listy_118_9
PubMedID: 28127976

Title : Construction of an Acetylcholinesterase Sensor Based on Synthesized Paramagnetic Nanoparticles, a Simple Tool for Neurotoxic Compounds Assay - Kostelnik_2017_Sensors.(Basel)_17_
Author(s) : Kostelnik A , Kopel P , Cegan A , Pohanka M
Ref : Sensors (Basel) , 17 : , 2017
Abstract : 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 microM 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.
ESTHER : Kostelnik_2017_Sensors.(Basel)_17_
PubMedSearch : Kostelnik_2017_Sensors.(Basel)_17_
PubMedID: 28338634

Title : Acetylcholinesterase Inhibitors Assay Using Colorimetric pH Sensitive Strips and Image Analysis by a Smartphone - Kostelnik_2017_Int.J.Anal.Chem_2017_3712384
Author(s) : Kostelnik A , Cegan A , Pohanka M
Ref : Int J Analytical Chemistry , 2017 :3712384 , 2017
Abstract : 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.
ESTHER : Kostelnik_2017_Int.J.Anal.Chem_2017_3712384
PubMedSearch : Kostelnik_2017_Int.J.Anal.Chem_2017_3712384
PubMedID: 28286520

Title : Anti-Parkinson Drug Biperiden Inhibits Enzyme Acetylcholinesterase - Kostelnik_2017_Biomed.Res.Int_2017_2532764
Author(s) : Kostelnik A , Cegan A , Pohanka M
Ref : Biomed Res Int , 2017 :2532764 , 2017
Abstract : Biperiden is a drug used in Parkinson disease treatment and it serves also as an antiseizures compound in organophosphates poisoning. It acts as antagonist of muscarinic receptor activated by acetylcholine while the enzyme acetylcholinesterase (AChE) cleaves acetylcholine in synaptic junction into choline and acetic acid. This enzyme is inhibited by various compounds; however there has not been proposed evidence about interaction with biperiden molecule. We investigated this interaction using standard Ellman's assay and experimental findings were critically completed with an in silico prediction by SwissDock docking software. Uncompetitive mechanism of action was revealed from Dixon plot and inhibition constant (Ki ) was calculated to be 1.11 mmol/l. The lowest predicted binding energy was -7.84 kcal/mol corresponding to H-bond between biperiden molecule and Tyr 341 residuum in protein structure of AChE. This interaction seems to be further stabilized by pi-pi interaction with Tyr 72, Trp 286, and Tyr 341. In conclusion, biperiden appears as a very weak inhibitor but it can serve as a lead structure in a pharmacological research.
ESTHER : Kostelnik_2017_Biomed.Res.Int_2017_2532764
PubMedSearch : Kostelnik_2017_Biomed.Res.Int_2017_2532764
PubMedID: 28785576

Title : Vaccination to Alzheimer Disease. Is it a Promising Tool or a Blind Way? - Pohanka_2016_Curr.Med.Chem_23_1432
Author(s) : Pohanka M
Ref : Curr Med Chem , 23 :1432 , 2016
Abstract : Alzheimer disease (AD) is an irreversible neurodegenerative disorder associated with cognitive dysfunction. The disease incidence has growing tendency worldwide with strong impact on healthcare funds. The fact that there is no effective therapy makes the disorder more serious. Currently, AD manifestation can be suppressed by having impact on enzyme acetylcholinesterase: donepezil, rivastigmine, and galantamine or ionotropic glutamate NMDA receptor ( memanitine). Contrary to the drugs effecting symptomatically, vaccination against amyloid plaques or neurofibrillary tangles and their precursors amyloid beta and hyperphosphorylated tau are expected to be more suitable. Huge numbers of works have been done on the issue. Unfortunately, the promising vaccines like the AN 1792 were halted during clinical trials because of adverse effects like meningoencephalitis. Monoclonal antibody specific to amyloid plaques, Bapineuzumab, was closest to the practical performance but the clinical trials were also stopped. The review summarizes facts about AD, opportunities in AD vaccination, and obstacles that limit the vaccination including reasons why the recent trials have fallen.
ESTHER : Pohanka_2016_Curr.Med.Chem_23_1432
PubMedSearch : Pohanka_2016_Curr.Med.Chem_23_1432
PubMedID: 27087245

Title : Color Change of Phenol Red by Integrated Smart Phone Camera as a Tool for the Determination of Neurotoxic Compounds - Kostelnik_2016_Sensors.(Basel)_16_
Author(s) : Kostelnik A , Cegan A , Pohanka M
Ref : Sensors (Basel) , 16 : , 2016
Abstract : The use of a cell phone as a detection system is easy, simple and does not require trained personnel, which is in contrast to standard laboratory instruments. This paper deals with immobilization of acetylcholinesterase (AChE) in a gelatin matrix, and phenol red, as an indicator of AChE activity, is used in order to establish a method that is easily compatible with a camera device. AChE splits acetylcholine into choline and acetic acid, which changes the pH of a medium, resulting in a phenol red color change. The coloration changed in presence of an AChE inhibitor. Measurements were performed on 3D-printed, tube-shaped holder, and digital photography, with subsequent analysis of red-green-blue (RGB), served for assay purposes. Calibration of AChE inhibitors, tacrine and galantamine, was performed, with limit of detection equal to 1.1 nM and 1.28 microM, respectively. Interferences were also measured, resulting in a proof-of-method stability. The method was further successfully validated for the standard Ellman's assay, and verified on murine plasma samples spiked with inhibitors.
ESTHER : Kostelnik_2016_Sensors.(Basel)_16_
PubMedSearch : Kostelnik_2016_Sensors.(Basel)_16_
PubMedID: 27618041

Title : Celecoxib is an inhibitor of enzyme acetylcholinesterase - Pohanka_2016_Neuro.Endocrinol.Lett_37_
Author(s) : Pohanka M
Ref : Neuro Endocrinol Lett , 37 : , 2016
Abstract : OBJECTIVES: Celecoxib is a nonsteroidal anti-inflammatory drug inhibiting enzyme cyclooxygenase-2 (COX-2). The drug was introduced in 1990s. In the work presented here, affinity of celecoxib to enzyme acetylcholinesterase (AChE) is inferred.
METHODS: Inhibition of human AChE by celecoxib was tested using standard spectrophotometric Ellman s method and extrapolation of experimental data by Dixon plot. Interaction between AChE and celecoxib was also predicted by molecular docking using Swiss dock software.
RESULTS: A non-competitive mechanism of inhibition was revealed and equilibrium inhibitory constant equal to 313+/-40 micromol/l was determined. Comparing to AChE, celecoxib was not proved as an inhibitor of enzyme butyrylcholinesterase (BChE). The lowest DeltaG was equal to -7.78 kcal/mol. In this case, celecoxib stacked sulfonamide moiety between TYR 337 and TYR 341 of alfa anionic subsite of active site. Cation-Pi interactions appears to be responsible for the inhibition.
CONCLUSIONS: Though the here revealed and characterized inhibition has lower effect in real conditions than inhibition of COX-2, the inhibitory effect would be utilized in the next research and development of new AChE inhibitors.
ESTHER : Pohanka_2016_Neuro.Endocrinol.Lett_37_
PubMedSearch : Pohanka_2016_Neuro.Endocrinol.Lett_37_
PubMedID: 28263539

Title : Evaluation of the benefit of the bispyridinium compound MB327 for the antidotal treatment of nerve agent-poisoned mice - Kassa_2016_Toxicol.Mech.Methods_26_334
Author(s) : Kassa J , Pohanka M , Timperley CM , Bird M , Green AC , Tattersall JE
Ref : Toxicol Mech Methods , 26 :334 , 2016
Abstract : The potency of the bispyridinium non-oxime compound MB327 [1,1'-(propane-1,3-diyl)bis(4-tert-butylpyridinium) diiodide] to increase the therapeutic efficacy of the standard antidotal treatment (atropine in combination with an oxime) of acute poisoning with organophosphorus nerve agents was studied in vivo. The therapeutic efficacy of atropine alone - or atropine in combination with an oxime, MB327, or both an oxime and MB237 - was evaluated by the determination of LD50 values of several nerve agents (tabun, sarin and soman) in mice with and without treatment. The addition of MB327 increased the therapeutic efficacy of atropine alone, and atropine in combination with an oxime, against all three nerve agents, although differences in the LD50 values only reached statistical significance for sarin. In conclusion, the addition of the compound MB327 to the standard antidotal treatment of acute poisonings with nerve agents was beneficial regardless of the chemical structure of the nerve agent, although at the dose employed, MB327 in combination with atropine, or atropine and an oxime, provided only a modest increase in protection ratio. These results from mice, and previous ones from guinea-pigs, provide consistent evidence for additional, albeit modest, efficacy resulting from the inclusion of the antinicotinic compound MB327 in standard antidotal therapy. Given the typically steep probit slope for the dose-lethality relationship for nerve agents, such modest increases in protection ratio could provide significant survival benefit.
ESTHER : Kassa_2016_Toxicol.Mech.Methods_26_334
PubMedSearch : Kassa_2016_Toxicol.Mech.Methods_26_334
PubMedID: 27097774

Title : Photography by Cameras Integrated in Smartphones as a Tool for Analytical Chemistry Represented by an Butyrylcholinesterase Activity Assay - Pohanka_2015_Sensors.(Basel)_15_13752
Author(s) : Pohanka M
Ref : Sensors (Basel) , 15 :13752 , 2015
Abstract : Smartphones are popular devices frequently equipped with sensitive sensors and great computational ability. Despite the widespread availability of smartphones, practical uses in analytical chemistry are limited, though some papers have proposed promising applications. In the present paper, a smartphone is used as a tool for the determination of cholinesterasemia i.e., the determination of a biochemical marker butyrylcholinesterase (BChE). The work should demonstrate suitability of a smartphone-integrated camera for analytical purposes. Paper strips soaked with indoxylacetate were used for the determination of BChE activity, while the standard Ellman's assay was used as a reference measurement. In the smartphone-based assay, BChE converted indoxylacetate to indigo blue and coloration was photographed using the phone's integrated camera. A RGB color model was analyzed and color values for the individual color channels were determined. The assay was verified using plasma samples and samples containing pure BChE, and validated using Ellmans's assay. The smartphone assay was proved to be reliable and applicable for routine diagnoses where BChE serves as a marker (liver function tests; some poisonings, etc.). It can be concluded that the assay is expected to be of practical applicability because of the results' relevance.
ESTHER : Pohanka_2015_Sensors.(Basel)_15_13752
PubMedSearch : Pohanka_2015_Sensors.(Basel)_15_13752
PubMedID: 26110404

Title : Possibility of Acetylcholinesterase Overexpression in Alzheimer Disease Patients after Therapy with Acetylcholinesterase Inhibitors - Kracmarova_2015_Acta.Medica.(Hradec.Kralove)_58_37
Author(s) : Kracmarova A , Drtinova L , Pohanka M
Ref : Acta Medica (Hradec Kralove) , 58 :37 , 2015
Abstract : Acetylcholinesterase is an enzyme responsible for termination of excitatory transmission at cholinergic synapses by the hydrolyzing of a neurotransmitter acetylcholine. Nowadays, other functions of acetylcholinesterase in the organism are considered, for example its role in regulation of apoptosis. Cholinergic nervous system as well as acetylcholinesterase activity is closely related to pathogenesis of Alzheimer disease. The mostly used therapy of Alzheimer disease is based on enhancing cholinergic function using inhibitors of acetylcholinesterase like rivastigmine, donepezil or galantamine. These drugs can influence not only the acetylcholinesterase activity but also other processes in treated organism. The paper is aimed mainly on possibility of increased expression and protein level of acetylcholinesterase caused by the therapy with acetylcholinesterase inhibitors.
ESTHER : Kracmarova_2015_Acta.Medica.(Hradec.Kralove)_58_37
PubMedSearch : Kracmarova_2015_Acta.Medica.(Hradec.Kralove)_58_37
PubMedID: 26455564

Title : More risk factors generate lower mortality - a useful advice for improved health in the world - Pohanka_2015_Bratisl.Lek.Listy_116_520
Author(s) : Pohanka M
Ref : Bratislavske Lekarske Listy , 116 :520 , 2015
Abstract : Caffeine (1,3,7-trimethylxanthine) is a plant secondary metabolite with a significant impact on multiple processes and regulatory pathways in the body. Though major part of the population meets caffeine via coffee, tea or chocolate, it has also an important role in pharmacology and it is used as a supplementary substance in medicaments. Currently, the ability of caffeine to ameliorate some neurodegenerative disorders is proved in some studies. This review describes basic data about caffeine including toxicity, pharmacokinetics, biological mechanism of the action, and metabolism. Beside this, promising applications of caffeine, new medicaments and derivatives are discussed. Relevant papers and inventions are depicted in the manuscript.Caffeine is a pharmacologically promising substance that deserves big consideration in the current research and development. The compound has several reasons to be an object of scientific interest and to be used for pharmacology purposes. Despite an extensive research for a long time, no significantly negative effects on human health were proved hence caffeine can be considered as a completely safe compound. The recent data about amelioration of neurodegenerative and other disorders are promising and deserving more work on the issue.ARTICLE HIGHLIGHTSCaffeine is a purine alkaloid from plants and it has a broad use in current pharmacology.Caffeine is a competitive antagonist of neurotransmitter adenosine on adenosine receptors.The substance is added as a supplementary to drugs and food.Besides interfering on adenosine receptors, caffeine interacts with acetylcholinesterase, monoamine oxidase, phosphodiesterase, ryanodine receptors and others.Current research is devoted to the role of caffeine in neurodegenerative diseases and immunity alteration.New chemical compounds based on caffeine moiety are prepared(Tab. 4, Fig. 6, Ref. 149).
ESTHER : Pohanka_2015_Bratisl.Lek.Listy_116_520
PubMedSearch : Pohanka_2015_Bratisl.Lek.Listy_116_520
PubMedID: 26435014

Title : Evaluation of 2,6-dichlorophenolindophenol acetate as a substrate for acetylcholinesterase activity assay - Pohanka_2015_J.Enzyme.Inhib.Med.Chem_30_796
Author(s) : Pohanka M , Holas O
Ref : J Enzyme Inhib Med Chem , 30 :796 , 2015
Abstract : Ellman's method is a standard protocol for the determination of cholinesterases activity. Though the method is ready for laboratory purposes, it has some drawbacks as well. In the current article, 2,6-dichloroindophenol acetate is performed as a chromogenic substrate suitable for acetylcholinesterase (AChE) activity examination. Michaelis constant and maximal velocity for 2,6-dichloroindophenol acetate were determined (38.0 microM and 244 pkat) and compared to the values for acetythiocholine (K(m) 0.18 mM; V(max) 5.1 nkat). Docking for 2,6-dichloroindophenol acetate and human AChE was done as well. In conclusion, 2,6-dichloroindophenol acetate seems to be suitable chromogenic substrate for AChE and spectrophotometry and based on this it can be easily performed whenever AChE activity should be tested.
ESTHER : Pohanka_2015_J.Enzyme.Inhib.Med.Chem_30_796
PubMedSearch : Pohanka_2015_J.Enzyme.Inhib.Med.Chem_30_796
PubMedID: 25672529

Title : Inhibitors of Acetylcholinesterase and Butyrylcholinesterase Meet Immunity - Pohanka_2014_Int.J.Mol.Sci_15_9809
Author(s) : Pohanka M
Ref : Int J Mol Sci , 15 :9809 , 2014
Abstract : Acetylcholinesterase (AChE) inhibitors are widely used for the symptomatic treatment of Alzheimer's disease and other dementias. More recent use is for myasthenia gravis. Many of these inhibitors interact with the second known cholinesterase, butyrylcholinesterase (BChE). Further, evidence shows that acetylcholine plays a role in suppression of cytokine release through a "cholinergic anti-inflammatory pathway" which raises questions about the role of these inhibitors in the immune system. This review covers research and discussion of the role of the inhibitors in modulating the immune response using as examples the commonly available drugs, donepezil, galantamine, huperzine, neostigmine and pyridostigmine. Major attention is given to the cholinergic anti-inflammatory pathway, a well-described link between the central nervous system and terminal effector cells in the immune system.
ESTHER : Pohanka_2014_Int.J.Mol.Sci_15_9809
PubMedSearch : Pohanka_2014_Int.J.Mol.Sci_15_9809
PubMedID: 24893223

Title : Postponed effect of neostigmine on oxidative homeostasis - Pohanka_2014_Interdiscip.Toxicol_7_134
Author(s) : Pohanka M
Ref : Interdiscip Toxicol , 7 :134 , 2014
Abstract : Cholinesterases are enzymes able to hydrolyze the neurotransmitter acetylcholine and thus to terminate transmission. Once the enzymes are inhibited, excitotoxicity can appear in the adjacent cells. It is well known that oxidative stress is involved in the toxicity of cholinesterase inhibitors. Commonly, stress follows inhibition of cholinesterases and disappears shortly afterwards. In the present experiment, it was decided to test the impact of an inhibitor, neostigmine, on oxidative stress in BALB/c mice after a longer interval. The animals were sacrificed three days after onset of the experiment and spleens and livers were collected. Reduced glutathione (GSH), glutathione reductase (GR), glutathione S-transferase (GST), thiobarbituric acid reactive substances (TBARS), ferric reducing antioxidant power (FRAP), caspase-3 and activity of acetylcholinesterase (AChE) were assayed. The tested markers were not altered with exceptions of FRAP. The FRAP values indicate accumulation of low molecular weight antioxidants in the examined organs. The role of low molecular weight antioxidants in the toxicity of AChE inhibitors is discussed.
ESTHER : Pohanka_2014_Interdiscip.Toxicol_7_134
PubMedSearch : Pohanka_2014_Interdiscip.Toxicol_7_134
PubMedID: 26109890

Title : Preparation and performance of a colorimetric biosensor using acetylcholinesterase and indoxylacetate for assay of nerve agents and drugs - Pohanka_2014_Interdiscip.Toxicol_7_215
Author(s) : Pohanka M , Vlcek V
Ref : Interdiscip Toxicol , 7 :215 , 2014
Abstract : Different toxic compounds can target the cholinergic nervous system. Acetylcholinesterase (AChE; EC is one of the most crucial components of the cholinergic nervous system and thus many of the toxins interact with this enzyme. As to inhibitors, nerve agents used as chemical warfare, some insecticides, and drugs influencing the cholinergic system are common examples of AChE inhibitors. Once inhibited by a neurotoxic compound, a serious cholinergic crisis can occur. On the other hand, sensitivity of AChE to the inhibition can be used for analytical purposes. In this study, a simple disposable biosensor with AChE as a recognition element was devised. AChE was immobilized onto a cellulose matrix and indoxylacetate was used as a chromogenic substrate. The enzyme reaction was assessed by the naked eye using arbitrary units and pyridostigmine, tacrine, paraoxon, carbofuran, soman and VX were assayed as selected inhibitors. A good stability of the biosensors was found, with no aging over a quarter of a year and minimal sensitivity to the interference of organic solvents. The limit of detection ranged from 10 to 100 nmol/L for the compounds tested with a sample volume of 40 microL.
ESTHER : Pohanka_2014_Interdiscip.Toxicol_7_215
PubMedSearch : Pohanka_2014_Interdiscip.Toxicol_7_215
PubMedID: 26109903

Title : Copper, aluminum, iron and calcium inhibit human acetylcholinesterase in vitro - Pohanka_2014_Environ.Toxicol.Pharmacol_37_455
Author(s) : Pohanka M
Ref : Environ Toxicol Pharmacol , 37 :455 , 2014
Abstract : Acetylcholinesterase (AChE) is an important part of cholinergic nerves where it participates in termination of neurotransmission. AChE can be inhibited by e.g. some Alzheimer disease drugs, nerve agents, and secondary metabolites. In this work, metal salts aluminum chloride, calcium chloride, cupric chloride, ferric chloride, potassium chloride, magnesium chloride and sodium chloride were tested for their ability to inhibit AChE. Standard Ellman assay based on human recombinant AChE was done and inhibition was measured using Dixon plot. No inhibition was proved for sodium, potassium and magnesium ions. However, aluminum, cupric, ferric and calcium ions were able to inhibit AChE via noncompetitive mechanism of inhibition. Though the inhibition is much weaker when compared to e.g. drugs with noncompetitive mechanism of action, biological relevance of the findings can be anticipated.
ESTHER : Pohanka_2014_Environ.Toxicol.Pharmacol_37_455
PubMedSearch : Pohanka_2014_Environ.Toxicol.Pharmacol_37_455
PubMedID: 24473150

Title : Voltammetric assay of butyrylcholinesterase in plasma samples and its comparison to the standard spectrophotometric test - Pohanka_2014_Talanta_119_412
Author(s) : Pohanka M
Ref : Talanta , 119 :412 , 2014
Abstract : Butyrylcholinesterase (BChE) is an enzyme abundantly constituted in the livers and released into blood where it is soluble. It may be found in the both plasma and serum. BChE can serve as a biochemical marker. BChE activity is typically measured by spectrophotometric Ellman's method. In the present work, voltammetric assay of cholinesterasemia is proposed as a simple and reliable method. In the experiments described here, limits of detections 4.57pkat for the spectrofotometric test and 1.14pkat for the voltammetric assay were determined. Interference caused by acetylcholinesterase (AChE) and organic solvents was characterized and counter measurement to the AChE caused interference was proposed. Finally, the both methods were correlated one to each other using mouse plasma spiked with carbofuran resulting in a promising coefficient of determination. In a conclusion, the voltammetric assay seems to be reliable and suitable for routine performance.
ESTHER : Pohanka_2014_Talanta_119_412
PubMedSearch : Pohanka_2014_Talanta_119_412
PubMedID: 24401433

Title : Novel tacrine\/acridine anticholinesterase inhibitors with piperazine and thiourea linkers - Hamulakova_2014_Int.J.Biol.Macromol_70C_435
Author(s) : Hamulakova S , Imrich J , Janovec L , Kristian P , Danihel I , Holas O , Pohanka M , Bohm S , Kozurkova M , Kuca K
Ref : Int J Biol Macromol , 70C :435 , 2014
Abstract : A new series of substituted tacrine/acridine and tacrine/tacrine dimers with aliphatic or alkylene-thiourea linkers was synthesized and the potential of these compounds as novel human acetylcholinesterase (hAChE) and human butyrylcholinesterase (hBChE) inhibitors with nanomolar inhibition activity was evaluated. The most potent AChE inhibitor was found to be homodimeric tacrine derivative 14a, which demonstrated an IC50 value of 2nM; this value indicates an activity rate which is 250-times higher than that of tacrine 1 and 7500-times higher than 7-MEOTA 15, the compounds which were used as standards in the study. IC50 values of derivatives 1, 9, 10, 14b and 15 were compared with the dissociation constants of the enzyme-inhibitor complex, Ki1, and the enzyme-substrate-inhibitor complex, Ki2, for. A dual binding site is presumed for the synthesized compounds which possess two tacrines or tacrine and acridine as terminal moieties show evidence of dual site binding. DFT calculations of theoretical desolvation free energies, DeltaDeltaGtheor, and docking studies elucidate these suggestions in more detail.
ESTHER : Hamulakova_2014_Int.J.Biol.Macromol_70C_435
PubMedSearch : Hamulakova_2014_Int.J.Biol.Macromol_70C_435
PubMedID: 25036600

Title : HI-6 modulates immunization efficacy in a BALB\/c mouse model - Pohanka_2013_Environ.Toxicol.Pharmacol_36_801
Author(s) : Pohanka M
Ref : Environ Toxicol Pharmacol , 36 :801 , 2013
Abstract : HI-6 is used as an antidote to nerve agents. It can also act as an antagonist to acetylcholine receptors (AChRs) including the nicotinic receptor, alpha 7 nAChR which is involved in regulating the immune response through macrophages. This experiment investigated the efficacy of HI-6 to regulate the immune response. Laboratory BALB/c mice received HI-6 and/or keyhole limpet hemocyanin (KLH) as an antigen. Antibody production was investigated after either 21 or 65 days when either single or repeated dose of antigen was applied. We confirmed that HI-6 significantly improved vaccination efficacy when KLH was given in a dose of 1mg/kg. The effect was dose dependent. A combination of HI-6 and KLH produced a vaccination of almost the same efficacy as that for Freund's complete adjuvant. The findings point at the suitability of HI-6 for improving vaccination efficacy at the level of immunity regulation by the nervous system.
ESTHER : Pohanka_2013_Environ.Toxicol.Pharmacol_36_801
PubMedSearch : Pohanka_2013_Environ.Toxicol.Pharmacol_36_801
PubMedID: 23958973

Title : Prophylaxis and Post-exposure Treatment of Intoxications Caused by Nerve Agents and Organophosphorus Pesticides - Kuca_2013_Mini.Rev.Med.Chem_13_2102
Author(s) : Kuca K , Jun D , Musilek K , Pohanka M , Karasova JZ , Soukup O
Ref : Mini Rev Med Chem , 13 :2102 , 2013
Abstract : Treatment of intoxications caused by nerve agents and organophosphorus pesticides consists of different approaches. The first approach is called prophylaxis or pre-exposure administration of antidotes such as cholinesterase reactivators or bioscavengers. The second, post-exposure treatment consists of anticholinergic drugs, acetylcholinesterase reactivators and anticonvulsants. This article is aimed at both mentioned approaches, especially focused on cholinesterase reactivators, which are a broad group of structurally different compounds that can be used in prophylaxis (separately or in combination with butyrylcholinesterase) and also as post-exposure treatment.
ESTHER : Kuca_2013_Mini.Rev.Med.Chem_13_2102
PubMedSearch : Kuca_2013_Mini.Rev.Med.Chem_13_2102
PubMedID: 24195661

Title : An acetylcholinesterase-based chronoamperometric biosensor for fast and reliable assay of nerve agents - Pohanka_2013_Sensors.(Basel)_13_11498
Author(s) : Pohanka M , Adam V , Kizek R
Ref : Sensors (Basel) , 13 :11498 , 2013
Abstract : The enzyme acetylcholinesterase (AChE) is an important part of cholinergic nervous system, where it stops neurotransmission by hydrolysis of the neurotransmitter acetylcholine. It is sensitive to inhibition by organophosphate and carbamate insecticides, some Alzheimer disease drugs, secondary metabolites such as aflatoxins and nerve agents used in chemical warfare. When immobilized on a sensor (physico-chemical transducer), it can be used for assay of these inhibitors. In the experiments described herein, an AChE- based electrochemical biosensor using screen printed electrode systems was prepared. The biosensor was used for assay of nerve agents such as sarin, soman, tabun and VX. The limits of detection achieved in a measuring protocol lasting ten minutes were 7.41 x 10-12 mol/L for sarin, 6.31 x 10-12 mol /L for soman, 6.17 x 10-11 mol/L for tabun, and 2.19 x 10-11 mol/L for VX, respectively. The assay was reliable, with minor interferences caused by the organic solvents ethanol, methanol, isopropanol and acetonitrile. Isopropanol was chosen as suitable medium for processing lipophilic samples.
ESTHER : Pohanka_2013_Sensors.(Basel)_13_11498
PubMedSearch : Pohanka_2013_Sensors.(Basel)_13_11498
PubMedID: 23999806

Title : A Resurrection of 7-MEOTA: A Comparison with Tacrine - Soukup_2013_Curr.Alzheimer.Res_10_893
Author(s) : Soukup O , Jun D , Karasova JZ , Patocka J , Musilek K , Korabecny J , Krusek J , Kaniakova M , Sepsova V , Mandikova J , Trejtnar F , Pohanka M , Drtinova L , Pavlik M , Tobin G , Kuca K
Ref : Curr Alzheimer Res , 10 :893 , 2013
Abstract : Alzheimer s disease (AD) is a progressive neurodegenerative dementia which currently represents one of the biggest threats for the human kind. The cure is still unknown and various hypotheses (cholinergic, amyloidal, oxidative, vascular etc.) are investigated in order to understand the pathophysiology of the disease and on this basis find an effective treatment. Tacrine, the first approved drug for the AD disease treatment, has been reported to be a multitargeted drug, however it was withdrawn from the market particularly due to its hepatotoxicity. Its derivative 7-methoxytacrine (7- MEOTA) probably due to the different metabolization does not exert this side effect. The aim of our study was to compare these two cholinesterase inhibitors from various, mainly cholinergic, points of view relevant for a potential AD drug. We found that 7-MEOTA does not fall behind its more well-known parent compound - tacrine. Furthermore, we found, that 7-MEOTA exerts better properties in most of the tests related to a possible AD treatment. Only the pharmacokinetics and a higher acetylcholinesterase and butyrylcholinesterase inhibitory potency would slightly give advantages to tacrine over 7-MEOTA, but concerning its lower toxicity, better antioxidant properties, interaction with muscarinic and nicotinic receptors and "safer" metabolization provide strong evidence for reconsider 7-MEOTA and its derivatives as candidate molecules for the treatment of AD.
ESTHER : Soukup_2013_Curr.Alzheimer.Res_10_893
PubMedSearch : Soukup_2013_Curr.Alzheimer.Res_10_893
PubMedID: 24093535

Title : Caffeine inhibits acetylcholinesterase, but not butyrylcholinesterase - Pohanka_2013_Int.J.Mol.Sci_14_9873
Author(s) : Pohanka M , Dobes P
Ref : Int J Mol Sci , 14 :9873 , 2013
Abstract : Caffeine is an alkaloid with a stimulant effect in the body. It can interfere in transmissions based on acetylcholine, epinephrine, norepinephrine, serotonin, dopamine and glutamate. Clinical studies indicate that it can be involved in the slowing of Alzheimer disease pathology and some other effects. The effects are not well understood. In the present work, we focused on the question whether caffeine can inhibit acetylcholinesterase (AChE) and/or, butyrylcholinesterase (BChE), the two enzymes participating in cholinergic neurotransmission. A standard Ellman test with human AChE and BChE was done for altering concentrations of caffeine. The test was supported by an in silico examination as well. Donepezil and tacrine were used as standards. In compliance with Dixon's plot, caffeine was proved to be a non-competitive inhibitor of AChE and BChE. However, inhibition of BChE was quite weak, as the inhibition constant, Ki, was 13.9 +/- 7.4 mol/L. Inhibition of AChE was more relevant, as Ki was found to be 175 +/- 9 micromol/L. The predicted free energy of binding was -6.7 kcal/mol. The proposed binding orientation of caffeine can interact with Trp86, and it can be stabilize by Tyr337 in comparison to the smaller Ala328 in the case of human BChE; thus, it can explain the lower binding affinity of caffeine for BChE with reference to AChE. The biological relevance of the findings is discussed.
ESTHER : Pohanka_2013_Int.J.Mol.Sci_14_9873
PubMedSearch : Pohanka_2013_Int.J.Mol.Sci_14_9873
PubMedID: 23698772

Title : Sulfur mustard induced oxidative stress and its alteration using asoxime (HI-6) - Pohanka_2013_Interdiscip.Toxicol_6_198
Author(s) : Pohanka M , Sobotka J , Svobodova H , Stetina R
Ref : Interdiscip Toxicol , 6 :198 , 2013
Abstract : Sulfur mustard (SM) is a blister agent with cytotoxic mechanism of action. There is no suitable treatment based on administration of an antidote. In this study, Wistar rats were exposed to SM in doses of 0-40 mg/kg body weight and treated with the compound HI-6. The treatment provided no significant effect on ferric reducing antioxidant power of blood and plasma. However, HI-6 caused an increase in the level of thiobarbituric acid reactive substances. This stressogenic response was presumably the cause of the significant elevation of the blood level of both glutathione reductase and reduced glutathione. HI-6 appears to be suitable for enhancing prophylactically oxidative stress protection from small oxidative insult.
ESTHER : Pohanka_2013_Interdiscip.Toxicol_6_198
PubMedSearch : Pohanka_2013_Interdiscip.Toxicol_6_198
PubMedID: 24678258

Title : Alzheimer s disease and oxidative stress: a review - Pohanka_2013_Curr.Med.Chem_21_356
Author(s) : Pohanka M
Ref : Curr Med Chem , 21 :356 , 2013
Abstract : Alzheimer s disease (AD) is a neurodegenerative disorder with no known cure and rapid rise in incidence. The predominant cognitive impairment is currently treated using cognitive enhancers like cholinesterase inhibitors. The two molecular hallmarks of AD are amyloid plaques created from an amyloid precursor protein and hyperphosphorylated tau protein that is deposited as neurofibrillary tangles inside neurons. A number of pathological mechanisms follow or precede these formations. Alteration in mitochondrial function and deposition of heavy metals are reported. The disease progression is enhanced by oxidative stress. However, the role of oxidative stress is not universally accepted. The current review covers and discusses the basic evidence and role of oxidative stress in AD development.
ESTHER : Pohanka_2013_Curr.Med.Chem_21_356
PubMedSearch : Pohanka_2013_Curr.Med.Chem_21_356
PubMedID: 24059239

Title : Butyrylcholinesterase as a biochemical marker - Pohanka_2013_Bratisl.Lek.Listy_114_726
Author(s) : Pohanka M
Ref : Bratislavske Lekarske Listy , 114 :726 , 2013
Abstract : Butyrylcholinesterase (BChE) is an enzyme expressed in multiple organs and abundant in plasma. BChE can fluctuate in course of several reasons while both hypercholiensterasemia and hypocholinesterasemia are known. Considering evidence of BChE activity alterations, hepatocellular carcinoma, chronic liver diseases and poisoning with carbamates or organophosphates can be diagnosed by activity assay. BChE is responsible for detoxification reactions, and the compounds such as cocaine, succinylcholine, and acetylsalicylic acid are degraded in the body. The detoxification can be slowed in patients carrying the K variant of the enzyme. Summarization of literature, discussion on the meaning of BChE in the body, and the principles of BChE assay in samples are described in the review (Tab. 2, Fig. 8, Ref. 86). Keywords: butyrylcholinesterase; acetylcholinesterase; liver function test; organophosphate; carbamate; poisoning; dibucaine number; Alzheimer s disease; BChE; AChE; K variant.
ESTHER : Pohanka_2013_Bratisl.Lek.Listy_114_726
PubMedSearch : Pohanka_2013_Bratisl.Lek.Listy_114_726
PubMedID: 24329513

Title : Spectrophotometric methods based on 2,6-dichloroindophenol acetate and indoxylacetate for butyrylcholinesterase activity assay in plasma - Pohanka_2013_Talanta_106_281
Author(s) : Pohanka M , Drtinova L
Ref : Talanta , 106 :281 , 2013
Abstract : Butyrylcholinesterase (BChE) is an enzyme presented in quite high level in blood plasma where it participates in detoxification reactions. Due to fact that the enzyme is constituted in livers, it is a marker of liver parenchyma function. It can be used for diagnosis of poisoning for e.g., nerve agents or carbofuran and intoxication by some drugs such as rivastigmine. The present experiment is devoted for the creation of new spectrophotometric tests for assay of BChE activity in biological samples. Standard Ellman's method was compared with use of 2,6-dichloroindophenol acetate and indoxylacetate as chromogenic substrates. Maximal velocities and Michaelis constants were calculated for the substrates. Considering calibration, 2,6-dichloroindophenol acetate provided the lowest limit of detection: 1.20 x 10(-9)kat and a long linear range. All methods were verified using pooled human plasma samples and tested for potential interferents. 2,6-dichloroindophenol acetate is recommended as suitable substrate for BChE assay in clinical diagnostics.
ESTHER : Pohanka_2013_Talanta_106_281
PubMedSearch : Pohanka_2013_Talanta_106_281
PubMedID: 23598128

Title : The progress in the cholinesterase quantification methods - Holas_2012_Expert.Opin.Drug.Discov_7_1207
Author(s) : Holas O , Musilek K , Pohanka M , Kuca K
Ref : Expert Opin Drug Discov , 7 :1207 , 2012
Abstract : Introduction: Determination of acetylcholinesterase and butyrylcholinesterase activity has become an important tool in drug design and discovery as well as in medicine and toxicology. There are a large number of compounds that are able to modulate cholinesterase activity. These compounds can be used for pharmacological management of various disorders (e.g., Alzheimer's disease, myasthenia Gravis). Moreover, organophosphate poisoning is frequently diagnosed via a cholinesterase activity assay. This broad variety of methods has been developed over the past decades for cholinesterase activity quantification. Areas covered: This review provides a summary of the methods that are based on specific properties of cholinesterases and their interactions with native or artificial substrates. The authors also aim to provide an overview of different techniques used for the determination of quantitative cholinesterase activity. Specifically, the authors describe and discuss the manometric, potentiometric, titrimetric, photometric, fluorometric, and radioisotopic methods. Expert opinion: Existing methods are able to cover most of the problems that arise during cholinesterase activity determination. Colorimetry according to Ellman has proved to be the most useful and versatile approach. It may be used in various protocols for the determination of pesticide or nerve agent exposure or for the development of new drugs. Its possible improvement lies in optimization of hemoglobin-rich samples. The progress of the most common methods (including Ellman) depends on miniaturization and modern physical platforms (e.g., optical fibers, chip methods, or nanotechnologies).
ESTHER : Holas_2012_Expert.Opin.Drug.Discov_7_1207
PubMedSearch : Holas_2012_Expert.Opin.Drug.Discov_7_1207
PubMedID: 23013366

Title : Carbamate insecticides in the czech republic: health and environmental impacts - Vlcek_2012_Mil.Med.Sci.Lett_81_2
Author(s) : Vlcek V , Pohanka M
Ref : Military Medical Science Letters , 81 :2 , 2012
Abstract : Carbamates neurotoxins are a group of compounds acting as pseudo-irreversible inhibitors of an enzyme acetylcholinesterase and butyrylcholinesterase. The compounds are well known as pesticides. Some of them, such as rivastigmine, pyridostigmine, and neostigmine, can be used as drugs for Myasthenia gravis or Alzheimer disease as well. The present review is aimed to summarize the basic facts about carbamate pesticides. Legislative aspects, e.g. law No. 326/2004 Coll., in the Czech Republic and examples of accidental exposures to carbamates in the country are provided, too. Other general provisions are determined by the EU, in particular by the Directive 91/414/EHS. The European legislative is discussed, too. Finally, examples of accidental exposures are introduced in the study.
ESTHER : Vlcek_2012_Mil.Med.Sci.Lett_81_2
PubMedSearch : Vlcek_2012_Mil.Med.Sci.Lett_81_2

Title : Tacrine is implicated in oxidative stress in the laboratory guinea pig model - Kracmarova_2012_Neuro.Endocrinol.Lett_33_136
Author(s) : Kracmarova A , Bandouchova H , Pikula J , Pohanka M
Ref : Neuro Endocrinol Lett , 33 :136 , 2012
Abstract : OBJECTIVES: Tacrine was the first acetylcholinesterase inhibitor approved for the treatment of Alzheimer disease. The compound is not available for therapeutic purposes as it was withdrawn due to hepatotoxicity of its metabolites. The hepatotoxicity can be decreased by alternative ways of drug administration avoiding thus the first pass effect. The present study is aimed to investigate the influence of intramuscularly administrated tacrine on oxidative stress.
METHODS: Laboratory guinea pigs were exposed to tacrine at doses of 0-800 mug/kg. The animals were euthanized 1 and 24 hours after the exposure. Parameters such as ferric reducing antioxidant power (FRAP), thiobarbituric acid reactive substances (TBARS), carbonylated proteins, caspase 3 activity, superoxide dismutase activity and glutathione reductase activity were assessed in the frontal, temporal and occipital lobe, cerebellum, liver, spleen, heart, and kidney. Moreover, levels of glucose, total and HDL cholesterol forms, triglycerides, blood urea nitrogen, creatinine, total bilirubin, total protein, albumin and activities of aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase and lactate dehydrogenase were assessed in plasma samples.
RESULTS: Activities of the enzymatic markers, level of carbonylated proteins in organs and levels of biochemical markers in plasma were only slightly influenced by tacrine. Dose-dependent elevation of the FRAP value was recognized in the brain tissues and the liver. The TBARS value was increased in the kidney and heart 1 and 24 hours, respectively, after exposure. CONCLUSION: In the study, the effect of tacrine on markers of oxidative stress was proved. Possible positive effects of tacrine on the antioxidant defence in the brain tissue were discussed.
ESTHER : Kracmarova_2012_Neuro.Endocrinol.Lett_33_136
PubMedSearch : Kracmarova_2012_Neuro.Endocrinol.Lett_33_136
PubMedID: 23353857

Title : Risk of combined exposure of birds to cyanobacterial biomass containing microcystins, acetylcholinesterase inhibitor and anticoagulant - Ondracek_2012_Neuro.Endocrinol.Lett_33_155
Author(s) : Ondracek K , Bandouchova H , Damkova V , Hilscherova K , Kral J , Osickova J , Mlcakova V , Pohanka M , Skochova H , Vitula F , Treml F , Pikula J
Ref : Neuro Endocrinol Lett , 33 :155 , 2012
Abstract : OBJECTIVES: The objective of this study was to examine the hypothesis that a combination of cyanobacterial biomass containing microcystins, acetylcholinesterase inhibitor and anticoagulant can enhance avian toxic effects produced by single exposures only.
METHODS: A total of 48 two-month-old Japanese quails (Coturnix coturnix japonica) with average body weight of 160 g were randomly divided into 8 experimental groups of six birds and sex ratio of 1:1. Experimental groups of control Japanese quails (C) and birds exposed to single and combined sub-lethal doses of paraoxon (P), bromadiolone (B), and microcystins in cyanobacterial biomass (M) included: C, P, P+B, B, B+M, P+M, M, and P+B+M. During the 10-day exposure birds in the respective groups received biomass containing 61.62 microg microcystins daily (i.e. 26.54 microg MC-RR, 7.62 microg MC-YR and 27.39 microg MC-LR), two 250 mug/kg doses of paraoxon, and two 500 mg/kg doses of bromadiolone. Group responses were compared using standard plasma biochemistry and antioxidant/oxidative stress parameters in tissues.
RESULTS: While single and double combinations of toxicants induced responses in individual biochemical parameters measured and evaluated using univariate statistical analysis, those in the triple exposure were most extensive. The principal component analysis of antioxidant/oxidative stress parameters (glutathione reductase, lipid peroxidation, and ferric reducing antioxidant power) in tissues (liver, kidney, heart, brain, lungs, gonads, and pectoralis major muscle) clearly separated the triple group (P+B+M) from all single and double exposure groups and the control and indicated thus marked joint effects in the overall pattern of antioxidant/oxidative stress responses of this group. The separation was driven by the modification of the ferric reducing antioxidant power levels in heart and brain and the cardiac lipid peroxidation level, in particular.
CONCLUSIONS: This experiment contributes to the understanding of the pathogenic mechanisms of combined sub-lethal exposure to natural toxins and agrochemicals and may be used for risk assessment of environmental pollution in birds.
ESTHER : Ondracek_2012_Neuro.Endocrinol.Lett_33_155
PubMedSearch : Ondracek_2012_Neuro.Endocrinol.Lett_33_155
PubMedID: 23353860

Title : Acetylcholinesterase inhibitors: a patent review (2008 - present) - Pohanka_2012_Expert.Opin.Ther.Pat_22_871
Author(s) : Pohanka M
Ref : Expert Opin Ther Pat , 22 :871 , 2012
Abstract : INTRODUCTION: Both acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are present in the body in large amounts. AChE is an important part of the cholinergic nervous system taking place in the central and peripheral nervous system. AChE is a target of several toxins such as insecticide carbofuran, nerve agents, sarin, soman, tabun and VX. Beside toxins, drugs for treatment of Alzheimer's disease and myasthenia gravis, such as galantamine, donepezil, rivastigmine, tacrine, huperzine, pyridostigmine and neostigmine, are known. AREAS COVERED: The review gives an overview of the importance of the cholinergic nervous system, the biochemistry of AChE and the role of AChE inhibitors. Current efforts to introduce potent drugs for Alzheimer's disease therapy and reduce toxicity, while keeping the maximal pharmacological effect, are also discussed. EXPERT OPINION: The current research effort into AChE inhibitors can be divided into two categories. First, new toxins useful for agricultural purposes and second, novel drugs that need to be prepared, although there is less interest in the new toxins. The research for drugs for Alzheimer's disease needs to focus on inhibitors that reduce the deposition of amyloid plaques, but do not initiate AChE expression.
ESTHER : Pohanka_2012_Expert.Opin.Ther.Pat_22_871
PubMedSearch : Pohanka_2012_Expert.Opin.Ther.Pat_22_871
PubMedID: 22768972

Title : Galantamine effect on tularemia pathogenesis in a BALB\/c mouse model - Pohanka_2012_Iran.Biomed.J_16_156
Author(s) : Pohanka M , Pavlis O , Pikula J
Ref : Iran Biomed J , 16 :156 , 2012
Abstract : BACKGROUND: Galantamine is a drug used for the treatment of Alzheimer's disease and some other cognitive disorders. It is an inhibitor of acetylcholinesterase; however, interaction with nicotinic acetylcholine receptors has also been reported. Owing to the significant role of cholinergic anti-inflammatory pathways in neuro-immunomodulation, we decided to examine the effect of galantamine on tularemia-infected BALB/c mice. METHODS: Animals were infected with Francisella tularensis LVS and treated with galantamine (0.1 mg/kg of body weight). Total mortality over the course of tularemia infection was assessed and interleukin 6 (IL-6) and interferon gamma (IFN-gamma) in plasma samples were measured by enzyme-linked immunosorbent assays. Apart from the cytokine assays, biochemical markers such as inorganic phosphate, uric acid, lactate dehydrogenase, gamma glutamyltransferase, creatinine phosphokinase and amylase were assayed. RESULTS: The modulation of immunity by galantamine depended on two opposing processes: up-regulation of IFN-gamma and down-regulation of IL-6. Tularemia infection resulted in significant nephropathy, as hyperphosphataemia and hyperuricaemia occurred in infected animals. In addition, galantamine resulted in the mitigation of nephropathy, and markers of kidney dysfunction were modulated. Alterations in mortality were also found in this study. CONCLUSIONS: Galantamine can significantly influence the immune response via the cholinergic anti-inflammatory pathway. Despite the decrease in IL-6 levels, galantamine treatment enhanced protection against the intracellular pathogen F. tularensis, resulting in the remission of some pathology and reduced mortality.
ESTHER : Pohanka_2012_Iran.Biomed.J_16_156
PubMedSearch : Pohanka_2012_Iran.Biomed.J_16_156
PubMedID: 23023217

Title : Toxicological scoring of Alzheimer's disease drug huperzine in a guinea pig model - Pohanka_2012_Toxicol.Mech.Methods_22_231
Author(s) : Pohanka M , Zemek F , Bandouchova H , Pikula J
Ref : Toxicol Mech Methods , 22 :231 , 2012
Abstract : Huperzine is a secondary metabolite in lycopods Huprzia and an inhibitor of acetylcholinesterase and antagonist of N-methyl-D-apartate receptor. Huperine is a suitable drug for the treatment of Alzheimer's disease as it is a part of traditional Chinese medicine. Currently, it undergoes clinical trials in the European Union and United States. The toxicological data about huperzine are missing and link between huperzine and oxidative stress has not been extensively investigated. For the above mentioned reasons, we organized experiment on a guinea pig model aimed at the investigation of adverse effects caused by huperzine. Guinea pigs were exposed to (-)-huperzine A in doses 5-625 microg/kg. Animals were sacrificed one day after exposure. Ferric reducing antioxidant power, thiobarbituric acid reactive substances, glutathione reductase, caspase 3 activity and selected biochemical markers (e.g. transaminases, blood urea nitrogen and glucose) were assayed. In frontal, parietal, temporal lobes and cerebellum, we found increase of antioxidants, glutathione reductase and oxidative stress markers in a dose dependent manner. Effects on liver, kidney and spleen were milder. We discuss ambivalent action of huperzine in the body and judge the huperzine action owing to recently reported experiments.
ESTHER : Pohanka_2012_Toxicol.Mech.Methods_22_231
PubMedSearch : Pohanka_2012_Toxicol.Mech.Methods_22_231
PubMedID: 22112162

Title : Biochemical insight into soman intoxication and treatment with atropine, HI-6, trimedoxime, and K203 in a rat model - Pohanka_2011_Bratisl.Lek.Listy_112_539
Author(s) : Pohanka M , Pikula J , Kuca K , Kassa J
Ref : Bratislavske Lekarske Listy , 112 :539 , 2011
Abstract : OBJECTIVE: The present experiment is based on biochemical assessment of nerve agent soman intoxication and atropine, respectively atropine and HI-6, trimedoxime or K203 treatment in rats. BACKGROUND: Nerve agents are toxic substances irreversibly inhibiting enzyme acetylcholinesterase (AChE). Treatment is typically based on application of atropine and oxime reactivator. Atropine is able to protect overstimulation of muscarinic acetylcholine receptors. Application of oxime reactivator enable return of AChE activity and full suppression of intoxication. METHODS: In a total, fifteen biochemical markers were assayed in plasma or blood of intoxicated animals. 42 rats were divided into 7 groups each 6 individuals. The first group was exposed to atropine; the second group was exposed to one LD50 of soman and atropine. The groups 3-5 were exposed in a same way as the second group and were treated with oxime reactivators: HI-6 (group 3), trimedoxime (4) and K203 (5). The sixth group was control treated with saline solution only. The last (seventh) group was intoxicated with soman only. RESULTS: The most striking shifts were found for blood acetylcholinesterase and plasma creatinine, glucose, inorganic phosphate as well as uric acid. Lactate dehydrogenase and aspartate aminotransferase assays were useless due to soman interference. CONCLUSION: It was demonstrated that treatment was able to protect poisoned animals from metabolic disorder represented by hyperglycemia and nephropathy represented by hyperuricemia and elevated creatinine. Soman exposure and treatment with the oxime reactivators and/or atropine contains quite complex and still not well understood side mechanisms (Tab. 2, Fig. 1, Ref. 25).
ESTHER : Pohanka_2011_Bratisl.Lek.Listy_112_539
PubMedSearch : Pohanka_2011_Bratisl.Lek.Listy_112_539
PubMedID: 21954536

Title : Preparation, in vitro screening and molecular modelling of symmetrical 4-tert-butylpyridinium cholinesterase inhibitors--analogues of SAD-128 - Musilek_2011_Bioorg.Med.Chem.Lett_21_150
Author(s) : Musilek K , Roder J , Komloova M , Holas O , Hrabinova M , Pohanka M , Dohnal V , Opletalova V , Kuca K , Jung YS
Ref : Bioorganic & Medicinal Chemistry Lett , 21 :150 , 2011
Abstract : Carbamate inhibitors (e.g., pyridostimine bromide) are used as a pre-exposure treatment for the prevention of organophosphorus poisoning. They work by blocking acetylcholinesterase's (AChE) native function and thus protect AChE against irreversible inhibition by organophosphorus compounds. However, carbamate inhibitors are known for many undesirable side-effects related to the carbamylation of AChE. In this Letter, 19 analogues of SAD-128 were prepared and evaluated as cholinesterase inhibitors. The screening results showed promising inhibitory ability of four compounds better to used standards (pralidoxime, obidoxime, BW284c51, ethopropazine, SAD-128). Four most promising compounds were selected for further molecular docking studies. The SAR was stated from obtained data. The former receptor studies were reported and discussed. The further in vivo studies were recommended in the view of OP pre-exposure treatment.
ESTHER : Musilek_2011_Bioorg.Med.Chem.Lett_21_150
PubMedSearch : Musilek_2011_Bioorg.Med.Chem.Lett_21_150
PubMedID: 21144749

Title : Cholinesterases, a target of pharmacology and toxicology - Pohanka_2011_Biomed.Pap.Med.Fac.Univ.Palacky.Olomouc.Czech.Repub_155_219
Author(s) : Pohanka M
Ref : Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub , 155 :219 , 2011
Abstract : BACKGROUND: Cholinesterases are a group of serine hydrolases that split the neurotransmitter acetylcholine (ACh) and terminate its action. Of the two types, butyrylcholinesterase and acetylcholinesterase (AChE), AChE plays the key role in ending cholinergic neurotransmission. Cholinesterase inhibitors are substances, either natural or man-made that interfere with the break-down of ACh and prolong its action. Hence their relevance to toxicology and pharmacology. METHODS AND RESULTS: The present review summarizes current knowledge of the cholinesterases and their inhibition. Particular attention is paid to the toxicology and pharmacology of cholinesterase-related inhibitors such as nerve agents (e.g. sarin, soman, tabun, VX), pesticides (e.g. paraoxon, parathion, malathion, malaoxon, carbofuran), selected plants and fungal secondary metabolites (e.g. aflatoxins), drugs for Alzheimer's disease (e.g. huperzine, metrifonate, tacrine, donepezil) and Myasthenia gravis (e.g. pyridostigmine) treatment and other compounds (propidium, ethidium, decamethonium). CONCLUSIONS: The crucial role of the cholinesterases in neural transmission makes them a primary target of a large number of cholinesterase-inhibiting drugs and toxins. In pharmacology, this has relevance to the treatment of neurodegenerative disorders.
ESTHER : Pohanka_2011_Biomed.Pap.Med.Fac.Univ.Palacky.Olomouc.Czech.Repub_155_219
PubMedSearch : Pohanka_2011_Biomed.Pap.Med.Fac.Univ.Palacky.Olomouc.Czech.Repub_155_219
PubMedID: 22286807

Title : On the universality of oxime Hl-7 - antidote for case of the nerve agent poisoning - Kuca_2011_Mil.Med.Sci.Lett_80_80
Author(s) : Kuca K , Musilek K , Karasova J , Jun D , Soukup O , Pohanka M , Ghosh KK , Hrabinova M
Ref : Military Medical Science Letters , 80 :80 , 2011
Abstract : Searching for the universal oxime, which could be able to reactivate acetylcholinesterase inhibited by various nerve agents is still topic of high interest. In this contribution, oxime HL-7, that was thoroughly discussed in the last decade, is evaluated . Its universality was tested in vitro using the rat brain homogenate as a source of the cholinesterases. The main members of the nerve agent family (tabun, sarin, soman, cyclosarin and VX) were used for this purpose. As shown, oxime HL-7 was able to reactivate cholinesterases inhibited by all tested nerve agents with the exception of tabun. Hence, it could not be designated as the broad-spectrum reactivator.
ESTHER : Kuca_2011_Mil.Med.Sci.Lett_80_80
PubMedSearch : Kuca_2011_Mil.Med.Sci.Lett_80_80

Title : Mono-oxime bisquaternary acetylcholinesterase reactivators with prop-1,3-diyl linkage-Preparation, in vitro screening and molecular docking - Musilek_2011_Bioorg.Med.Chem_19_754
Author(s) : Musilek K , Komloova M , Holas O , Horova A , Pohanka M , Gunn-Moore F , Dohnal V , Dolezal M , Kuca K
Ref : Bioorganic & Medicinal Chemistry , 19 :754 , 2011
Abstract : The treatment of organophosphorus (OP) poisoning consists of the administration of a parasympatholytic agent (e.g., atropine), an anticonvulsant (e.g., diazepam) and an acetylcholinesterase (AChE) reactivator (e.g., obidoxime). The AChE reactivator is the causal treatment of OP exposure, because it cleaves the OP moiety covalently bound to the AChE active site. In this paper, fourteen novel AChE reactivators are described. Their design originated from a former promising compound K027. These compounds were synthesized, evaluated in vitro on human AChE (hAChE) inhibited by tabun, paraoxon, methylparaoxon and DFP and then compared to commercial hAChE reactivators (pralidoxime, HI-6, trimedoxime, obidoxime, methoxime) or previously prepared compounds (K027, K203). Three of these novel compounds showed a promising ability to reactivate hAChE comparable or better than the used standards. Consequently, a molecular docking study was performed for three of these promising novel compounds. The docking results confirmed the apparent influence of pi-pi or cation-pi interactions and hydrogen bonding for reactivator binding within the hAChE active site cleft. The SAR features concerning the non-oxime part of the reactivator molecule are also discussed.
ESTHER : Musilek_2011_Bioorg.Med.Chem_19_754
PubMedSearch : Musilek_2011_Bioorg.Med.Chem_19_754
PubMedID: 21215642

Title : Assessment of acetylcholinesterase activity using indoxylacetate and comparison with the standard Ellman's method - Pohanka_2011_Int.J.Mol.Sci_12_2631
Author(s) : Pohanka M , Hrabinova M , Kuca K , Simonato JP
Ref : Int J Mol Sci , 12 :2631 , 2011
Abstract : Assay of acetylcholinesterase (AChE) activity plays an important role in diagnostic, detection of pesticides and nerve agents, in vitro characterization of toxins and drugs including potential treatments for Alzheimer's disease. These experiments were done in order to determine whether indoxylacetate could be an adequate chromogenic reactant for AChE assay evaluation. Moreover, the results were compared to the standard Ellman's method. We calculated Michaelis constant Km (2.06 x 10(-4) mol/L for acetylthiocholine and 3.21 x 10(-3) mol/L for indoxylacetate) maximum reaction velocity V(max) (4.97 x 10(-7) kat for acetylcholine and 7.71 x 10(-8) kat for indoxylacetate) for electric eel AChE. In a second part, inhibition values were plotted for paraoxon, and reactivation efficacy was measured for some standard oxime reactivators: obidoxime, pralidoxime (2-PAM) and HI-6. Though indoxylacetate is split with lower turnover rate, this compound appears as a very attractive reactant since it does not show any chemical reactivity with oxime antidots and thiol used for the Ellman's method. Thus it can be advantageously used for accurate measurement of AChE activity. Suitability of assay for butyrylcholinesterase activity assessment is also discussed.
ESTHER : Pohanka_2011_Int.J.Mol.Sci_12_2631
PubMedSearch : Pohanka_2011_Int.J.Mol.Sci_12_2631
PubMedID: 21731462

Title : Metrifonate alters antioxidant levels and caspase activity in cerebral cortex of Wistar rats - Pohanka_2011_Toxicol.Mech.Methods_21_585
Author(s) : Pohanka M , Novotny L , Pikula J
Ref : Toxicol Mech Methods , 21 :585 , 2011
Abstract : Metrifonate (trichlorfon) is an inhibitor of acetylcholinesterase (AChE). It was used as an Alzheimer's disease (AD) drug; however, the application was withdrawn due to adverse effects. Implication of metrifonate for the antioxidant status and regulation of apoptotic processes was evaluated in the present study. Wistar rats (six per group) were exposed subcutaneously to either 60 or 120 mg/kg of body weight of metrifonate and compared with the controls treated with saline only. Cerebral cortex and liver tissues were collected from animals 40 min after exposure. Activities of AChE, glutathione reductase, glutathione-S-transferase, caspase 3, total protein level, thiobarbituric acid reactive substances, reduced glutathione level and ferric reducing antioxidant power (FRAP) were assayed in the tissue samples. Metrifonate had only lower impact on oxidative stress in the liver. Cerebral cortex tissues had decreased AChE and increased caspase 3 activities as well as the FRAP level. Owing to the novel findings, suitability of metrifonate for AD therapy is discussed.
ESTHER : Pohanka_2011_Toxicol.Mech.Methods_21_585
PubMedSearch : Pohanka_2011_Toxicol.Mech.Methods_21_585
PubMedID: 21943232

Title : Preparation and in vitro screening of symmetrical bis-isoquinolinium cholinesterase inhibitors bearing various connecting linkage--implications for early Myasthenia gravis treatment - Musilek_2011_Eur.J.Med.Chem_46_811
Author(s) : Musilek K , Komloova M , Holas O , Hrabinova M , Pohanka M , Dohnal V , Nachon F , Dolezal M , Kuca K
Ref : Eur Journal of Medicinal Chemistry , 46 :811 , 2011
Abstract : Inhibitors of acetylcholinesterase are compounds widely used in the treatment of various diseases, such as Alzheimer's disease, glaucoma and Myasthenia gravis (MG). Compounds used in the therapy of MG posses a positive charge in the molecule to ensure peripheral effect of action and minimal blood-brain barrier penetration. The most prescribed carbamate inhibitors are however known for many severe side effects related to the carbamylation of AChE. This paper describes preparation and in vitro evaluation of 20 newly prepared bis-isoquinolinium inhibitors of potential concern for MG. The newly prepared compounds were evaluated in vitro on human recombinant AChE and human plasmatic butyrylcholinesterase (BChE). Their inhibitory ability was expressed as IC50 and compared to chosen standards ambenonium dichloride, edrophonium chloride, BW284c51 and ethopropazine hydrochloride. Three novel compounds presented promising inhibition (in nM range) of both enzymes in vitro better or similar to edrophonium and BW284c51, but worse to ambenonium. The novel inhibitors did not present higher selectivity toward AChE or BChE. The kinetic assay confirmed non-competitive inhibition of hAChE by two selected promising novel compounds. Two newly prepared compounds were also chosen for docking studies that confirmed apparent pi-pi or pi-cationic interactions aside the cholinesterases catalytic sites. The SAR findings were discussed.
ESTHER : Musilek_2011_Eur.J.Med.Chem_46_811
PubMedSearch : Musilek_2011_Eur.J.Med.Chem_46_811
PubMedID: 21236521

Title : The preparation, in vitro screening and molecular docking of symmetrical bisquaternary cholinesterase inhibitors containing a but-(2E)-en-1,4-diyl connecting linkage - Musilek_2011_J.Enzyme.Inhib.Med.Chem_26_245
Author(s) : Musilek K , Pavlikova R , Marek J , Komloova M , Holas O , Hrabinova M , Pohanka M , Dohnal V , Dolezal M , Gunn-Moore F , Kuca K
Ref : J Enzyme Inhib Med Chem , 26 :245 , 2011
Abstract : Carbamate inhibitors (e.g. pyridostigmine bromide) are used as a pre-treatment for the prevention of organophosphorus poisoning. They work by blocking the native function of acetylcholinesterases (AChE) and thus protect AChE against irreversible inhibition by organophosphorus compounds. However, carbamate inhibitors are known for their many undesirable side effects related to the carbamylation of AChE. In this paper, we describe 17 novel bisquaternary compounds and have analysed their effect on AChE inhibition. The newly prepared compounds were evaluated in vitro using both human erythrocyte AChE and human plasmatic butyrylcholinesterase. Their inhibitory ability was expressed as the half maximal inhibitory concentration (IC(5)(0)) and then compared to the standard carbamate drugs and two AChE reactivators. One of these novel compounds showed promising AChE inhibition in vitro (nM range) and was better than the currently used standards. Additionally, a kinetic assay confirmed the non-competitive inhibition of hAChE by this novel compound. Consequently, the docking results confirmed the apparent pi-pi or pi-cationic interactions with the key amino acid residues of hAChE and the binding of the chosen compound at the enzyme catalytic site.
ESTHER : Musilek_2011_J.Enzyme.Inhib.Med.Chem_26_245
PubMedSearch : Musilek_2011_J.Enzyme.Inhib.Med.Chem_26_245
PubMedID: 21406034

Title : Synthesis and in vitro evaluation of N-alkyl-7-methoxytacrine hydrochlorides as potential cholinesterase inhibitors in Alzheimer disease - Korabecny_2010_Bioorg.Med.Chem.Lett_20_6093
Author(s) : Korabecny J , Musilek K , Holas O , Binder J , Zemek F , Marek J , Pohanka M , Opletalova V , Dohnal V , Kuca K
Ref : Bioorganic & Medicinal Chemistry Lett , 20 :6093 , 2010
Abstract : All approved drugs for Alzheimer disease (AD) in clinical practice ameliorate the symptoms of the disease. Among them, acetylcholinesterase inhibitors (AChEIs) are used to increase the cholinergic activity. Among new AChEI, tacrine compounds were found to be more toxic compared to 7-MEOTA (9-amino-7-methoxy-1,2,3,4-tetrahydroacridine). In this Letter, series of 7-MEOTA analogues (N-alkyl-7-methoxytacrine) were synthesized. Their inhibitory ability was evaluated on recombinant human acetylcholinesterase (AChE) and plasmatic human butyrylcholinesterase (BChE). Three novel compounds showed promising results towards hAChE better to THA or 7-MEOTA. Three compounds resulted as potent inhibitors of hBChE. The SAR findings highlighted the C(6)-C(7)N-alkyl chains for cholinesterase inhibition.
ESTHER : Korabecny_2010_Bioorg.Med.Chem.Lett_20_6093
PubMedSearch : Korabecny_2010_Bioorg.Med.Chem.Lett_20_6093
PubMedID: 20817518

Title : Oxime K027: novel low-toxic candidate for the universal reactivator of nerve agent- and pesticide-inhibited acetylcholinesterase - Kuca_2010_J.Enzyme.Inhib.Med.Chem_25_509
Author(s) : Kuca K , Musilek K , Jun D , Pohanka M , Ghosh KK , Hrabinova M
Ref : J Enzyme Inhib Med Chem , 25 :509 , 2010
Abstract : Oxime K027 is a low-toxic bisquaternary compound originally developed as a reactivator of acetylcholinesterase (AChE) inhibited by nerve agents. The reactivation potency of K027 has been tested as a potential reactivator of AChE inhibited by tabun, sarin, cyclosarin, soman, VX, Russian VX, paraoxon, methylchlorpyrifos, and DDVP. The results show that oxime K027 reactivated AChE inhibited by almost all tested inhibitors to more than 10%, which is believed to be enough for saving the lives of intoxicated organisms. In the case of cyclosarin- and soman-inhibited AChE, oxime K027 did not reach sufficient reactivation potency.
ESTHER : Kuca_2010_J.Enzyme.Inhib.Med.Chem_25_509
PubMedSearch : Kuca_2010_J.Enzyme.Inhib.Med.Chem_25_509
PubMedID: 20192902

Title : Evaluation of aflatoxin B1--acetylcholinesterase dissociation kinetic using the amperometric biosensor technology: prospect for toxicity mechanism - Pohanka_2010_Protein.Pept.Lett_17_340
Author(s) : Pohanka M , Musilek K , Kuca K
Ref : Protein Pept Lett , 17 :340 , 2010
Abstract : Aflatoxins are group of secondary metabolites from moulds. The main toxic effect of aflatoxins on body is based on metabolic activation on cytochrome P450 system. Recently, some studies appointed at anticholinergic properties of aflatoxins and inhibition of acetylcholinesterases (AChE) was described. Inhibition is reversible; however, some questions arose. Is the interaction firmly enough to prevent distribution of aflatoxins in body? Could be AChE considered as a scavenger of aflatoxins? Amperometric biosensor with immobilized acetylcholinesterase was used for evaluation of aflatoxin B1 (AFB1) - AChE complex spontaneous dissociation, where AFB1 acts as an inhibitor. Displacement of solution with substrate and AFB1 by the intact one enabled estimation of dissociation kinetics. The dissociation rate constant k(dis) was found 0.0047 +/- 0.0005 s(-1). The half time (t(1/2)) of complex dissociation was 146 s. The achieved data appoint at fact that AChE could allow to distribute aflatoxins in body instead acting as a scavenger. Analytical impact of study is discussed, too.
ESTHER : Pohanka_2010_Protein.Pept.Lett_17_340
PubMedSearch : Pohanka_2010_Protein.Pept.Lett_17_340
PubMedID: 19508188

Title : Pralidoxime--the gold standard of acetylcholinesterase reactivators--reactivation in vitro efficacy - Kuca_2010_Bratisl.Lek.Listy_111_502
Author(s) : Kuca K , Hrabinova M , Soukup O , Tobin G , Karasova J , Pohanka M
Ref : Bratislavske Lekarske Listy , 111 :502 , 2010
Abstract : OBJECTIVE: In this work, we aim to summarize the universality of this compound, its reactivation potential when different cholinesterase inhibitors are used. BACKGROUND: Pralidoxime is considered as a gold standard of acetylcholinesterase reactivators--antidotes used in case of nerve agent poisonings. It has been commercially available for many years. However, several studies deem this oxime an old-fashion antidote. METHODS: Pralidoxime was synthesized at our department. The reactivating efficacy was tested on 10% (w/v) rat brain homogenate that had been incubated with appropriate inhibitor for 30 minutes to reach 96% inhibition of AChE. Then, pralidoxime was added for 10 minutes. Measurements were performed at 25 degrees C, pH 8, and 10(-3) and 10(-5) M concentrations of AChE reactivators. The activities of brain AChE were measured by a potentiostatic method. RESULTS: No sufficient reactivation was achieved at the concentration of 10(-5) M, which is a concentration that can be reached after administration of therapeutic doses. At a higher dose (10(-3) M), pralidoxime reactivated AChE inhibited by paraoxon, chlorpyrifos, Russian VX, VX and sarin. CONCLUSION: From the obtained results, it is clear that pralidoxime seems to be a poor reactivator of AChE inhibited by organophosphorous AChE inhibitors and thus cannot be labeled as a universal reactivator (Tab. 1, Fig. 3, Ref. 31).
ESTHER : Kuca_2010_Bratisl.Lek.Listy_111_502
PubMedSearch : Kuca_2010_Bratisl.Lek.Listy_111_502
PubMedID: 21180265

Title : Modulation of ionising radiation generated oxidative stress by HI-6 (asoxime) in a laboratory rat model - Pohanka_2010_Neuro.Endocrinol.Lett_31 Suppl 2_62
Author(s) : Pohanka M , Pejchal J , Horackova S , Kuca K , Bandouchova H , Damkova V , Pikula J
Ref : Neuro Endocrinol Lett , 31 Suppl 2 :62 , 2010
Abstract : OBJECTIVES: HI-6 is an antidotum suitable for treatment of intoxication by nerve agents. The recent investigation appointed its modulation of inflammatory response as well as vegetative nervous system activity. However, the present experiments were carried out in order to assess the antioxidant effect of HI-6 in irradiated animals. METHODS: male Wistar rats were irradiated by ionizing radiation (7.5 Gy, LD50/30). Animals were divided into four groups: i.e. controls (A), irradiated (B), treated with HI-6 (C), and both irradiated and treated with HI-6 (D). Ferric reducing antioxidant power (FRAP), thiobarbituric acid reactive substances (TBARS) and glutathione reductase activity were assayed in liver, spleen, plasma, and whole blood. Clinical biochemistry markers were determined in plasma samples. RESULTS: We found significantly increased FRAP levels in liver, while its levels decreased in the spleen of B group animals. Ionising radiation (B group) also significantly elevated TBARS values in spleen. HI-6 reversed FRAP and TBARS values to control levels. Glutathione reductase activity was significantly elevated in spleen and liver of animals exposed to HI-6 (C and D groups). Clinical biochemistry markers were shifted only slightly. The in vitro test confirmed the inhibitory effect of HI-6 towards acetylcholinesterase. CONCLUSIONS: In conclusion, HI-6 is potent in suppressing oxidative stress and might be a promising drug in the field of radiation protection.
ESTHER : Pohanka_2010_Neuro.Endocrinol.Lett_31 Suppl 2_62
PubMedSearch : Pohanka_2010_Neuro.Endocrinol.Lett_31 Suppl 2_62
PubMedID: 21187825

Title : Colorimetric Detectors Based on Acetylcholinesterase and Its Construction - Pohanka_2010_Mil.Med.Sci.Lett_79_9
Author(s) : Pohanka M , Vlcek V , Karasova JZ , Kuca K , Cabal J
Ref : Military Medical Science Letters , 79 :9 , 2010
Abstract : Colorimetric detectors based on recognition capability of acetylcholinesterase (AChE) are a suitable tool for fast but sensitive detection of nerve agents and some pesticides based on AChE inhibition. In this review article, basic biochemical methods appropriate for colorimetric estimation of AChE enzymatic activity are presented. Moreover, the available matrices suitable for construction of dipsticks and chemical as well as physical immobilization protocols for AChE binding on matrice surface are described, too. Detehit and ChP-71 are indicated as typical colorimetric detectors for assay of nerve agents.
ESTHER : Pohanka_2010_Mil.Med.Sci.Lett_79_9
PubMedSearch : Pohanka_2010_Mil.Med.Sci.Lett_79_9

Title : Immobilization of Acetylcholinesterase and a Construction of Biochemical Biosensor for Identification of Toxic Organophosphates. - Pohanka_2010_Mil.Med.Sci.Lett_79_105
Author(s) : Pohanka M , Drobik O , Krenkova Z , Karasova JZ , Cabal J , Kuca K
Ref : Military Medical Science Letters , 79 :105 , 2010
Abstract : Biosensors are analytical devices based on tight junction of so-called biorecognition element and a physicochemical transducer. In this study, a screen printed sensor with platinum working electrode as the transducer and acetylcholinesterase as the biorecognition element were used. Acetylcholinesterase was immobilized in a different way including sol-gel technologies, immobilization on graphite nanofibers, precipitation with glutaraldehyde and interception into spherical graphite microparticles. Efficacy for given immobilization protocols was estimated, and practical impact of the study was discussed.
ESTHER : Pohanka_2010_Mil.Med.Sci.Lett_79_105
PubMedSearch : Pohanka_2010_Mil.Med.Sci.Lett_79_105

Title : Testing of a Referential Method Based on Indoxylacetate as a Acetylcholinesterase Substrate for Identification of Reactivation Efficacy. - Pohanka_2010_Mil.Med.Sci.Lett_79_111
Author(s) : Pohanka M , Hrabinova M , Karasova JZ , Holas O , Kuca K , Cabal J
Ref : Military Medical Science Letters , 79 :111 , 2010
Abstract : Biochemical evaluation of acetylcholinesterase activity is an important factor for selection and evaluation of drugs, such as oxime reactivators, which modulate acetylcholinesterase activity. A spontaneous reaction between an Ellman reagent and an oxime reactivator is the main disadvantage of currently available Ellman's method used for identification of cholinesterase activity. In this study, an alternative chromogen, substrate indoxylacetate is used. It is suitable for evaluation of acetylcholinesterase reactivation efficacy without oxime interference in vitro. The method was successfully examined for five standard oxime reactivators (HI-6, methoxime, trimedoxime, obidoxime, and 2-PAM) and three organophosphate inhibitors: paraoxon-methyl, sarin and tabun.
ESTHER : Pohanka_2010_Mil.Med.Sci.Lett_79_111
PubMedSearch : Pohanka_2010_Mil.Med.Sci.Lett_79_111

Title : Monooxime-monocarbamoyl Bispyridinium Xylene-Linked Reactivators of Acetylcholinesterase-Synthesis, In vitro and Toxicity Evaluation, and Docking Studies - Musilek_2010_ChemMedChem_5_247
Author(s) : Musilek K , Holas O , Misik J , Pohanka M , Novotny L , Dohnal V , Opletalova V , Kuca K
Ref : ChemMedChem , 5 :247 , 2010
Abstract : Acetylcholinesterase (AChE) reactivators are crucial antidotes to organophosphate intoxication. A new series of 26 monooxime-monocarbamoyl xylene-linked bispyridinium compounds was prepared and tested in vitro, along with known reactivators (pralidoxime, HI-6, obidoxime, trimedoxime, methoxime, K107, K108 and K203), on a model of tabun- and paraoxon-, methylparaoxon- and DFP-inhibited human erythrocyte AChE. Although their ability to reactivate tabun-inhibited AChE did not exceed that of the previously known compounds, some newly prepared compounds showed promising reactivation of pesticide-inhibited AChE. The acute toxicity of the novel compounds was also determined. Docking studies using tabun-inhibited AChE were performed for three compounds of interest. The structure-activity relationship (SAR) study confirmed the apparent influence of the xylene linkage and carbamoyl moiety on the reactivation ability and toxicity of the agents.
ESTHER : Musilek_2010_ChemMedChem_5_247
PubMedSearch : Musilek_2010_ChemMedChem_5_247
PubMedID: 20058292

Title : Colorimetric dipstick for assay of organophosphate pesticides and nerve agents represented by paraoxon, sarin and VX - Pohanka_2010_Talanta_81_621
Author(s) : Pohanka M , Karasova JZ , Kuca K , Pikula J , Holas O , Korabecny J , Cabal J
Ref : Talanta , 81 :621 , 2010
Abstract : A dipstick for fast assay of nerve agents and organophosphate pesticides was developed. Indicator pH papers were used as detectors. The principle of the assay is based on enzymatic hydrolysis of acetylcholine into acetic acid and choline by acetylcholinesterase. Acidification of the reaction medium due to accumulation of acetic acid was visible. The colour changed from dark red to yellow as the pH indicator recognized pH shift. Presence of an organophosphate pesticide or a nerve agent results in irreversible inhibition of acetylcholinesterase intercepted on the dipstick. The inhibition stops the enzymatic reaction. The inhibition appears as no change of the medium pH. Three compounds were assayed: paraoxon-ethyl as representative organophosphate pesticides and nerve agents sarin and VX. The achieved limit of detection was 5 x 10(-8)M for paraoxon-ethyl and 5 x 10(-9)M for sarin and VX. Dipsticks were found stable for at least one month. Suitability of these dipsticks for routine assay is discussed.
ESTHER : Pohanka_2010_Talanta_81_621
PubMedSearch : Pohanka_2010_Talanta_81_621
PubMedID: 20188972

Title : Preparation and in vitro screening of symmetrical bispyridinium cholinesterase inhibitors bearing different connecting linkage-initial study for Myasthenia gravis implications - Musilek_2010_Bioorg.Med.Chem.Lett_20_1763
Author(s) : Musilek K , Komloova M , Zavadova V , Holas O , Hrabinova M , Pohanka M , Dohnal V , Nachon F , Dolezal M , Kuca K , Jung YS
Ref : Bioorganic & Medicinal Chemistry Lett , 20 :1763 , 2010
Abstract : Reversible inhibitors (e.g., pyridostigmine bromide, neostigmine bromide) of carbamate origin are used in the early treatment of Myasthenia gravis (MG) to block acetylcholinesterase (AChE) native function and conserve efficient amount of acetylcholine for decreasing number of nicotinic receptors. Carbamate inhibitors are known for many undesirable side effects related to the reversible inhibition of AChE. In contrast, this paper describes 20 newly prepared bispyridinium inhibitors of potential concern for MG. Although some compounds from this series have been known before, they were not assayed for cholinesterase inhibition yet. The newly prepared compounds were evaluated in vitro on human erythrocyte AChE and human plasmatic butyrylcholinesterase (BChE). Their inhibitory ability was expressed as IC(50) and compared to standard carbamate drugs. Three compounds presented promising inhibition (in muM range) of both enzymes in vitro similar to the used standards. The novel inhibitors did not present selectivity between AChE and BChE. Two newly prepared compounds were chosen for docking studies and confirmed apparent pi-pi or pi-cationic interactions aside enzyme's catalytic sites. The kinetics assay confirmed non-competitive inhibition of AChE by two best newly prepared compounds.
ESTHER : Musilek_2010_Bioorg.Med.Chem.Lett_20_1763
PubMedSearch : Musilek_2010_Bioorg.Med.Chem.Lett_20_1763
PubMedID: 20138518

Title : Passive diffusion of acetylcholinesterase oxime reactivators through the blood-brain barrier: influence of molecular structure - Karasova_2010_Toxicol.In.Vitro_24_1838
Author(s) : Karasova JZ , Pohanka M , Musilek K , Zemek F , Kuca K
Ref : Toxicol In Vitro , 24 :1838 , 2010
Abstract : In this in vitro study, high-performance liquid chromatography (HPLC) was used to determinate the penetration of 30 acetylcholinesterase (AChE) reactivators through the blood-brain barrier (BBB). According to our method, monoquaternary AChE reactivators were found to be able to penetrate the BBB. In addition to molecular structure, molecular weight appears to be an important factor for passive transport of oximes through the BBB. For bisquaternary reactivators, the connecting linker plays a key role in the ability to penetrate into the central nervous system (CNS): simple, short linkers tend to facilitate permeation. The location of groups on the pyridine ring also influences passive transport into the brain; the optimum position of the oxime group was found to be position four (para) and substitution of the oxime group on the pyridine ring by carbamoyl or amidoxime group markedly decreased penetration of AChE reactivators into the CNS.
ESTHER : Karasova_2010_Toxicol.In.Vitro_24_1838
PubMedSearch : Karasova_2010_Toxicol.In.Vitro_24_1838
PubMedID: 20546883

Title : Novel bisquaternary oximes--reactivation of acetylcholinesterase and butyrylcholinesterase inhibited by paraoxon - Kuca_2009_Molecules_14_4915
Author(s) : Kuca K , Musilova L , Palecek J , Cirkva V , Paar M , Musilek K , Hrabinova M , Pohanka M , Karasova JZ , Jun D
Ref : Molecules , 14 :4915 , 2009
Abstract : Four novel bisquaternary aldoxime cholinesterase reactivators differing in their chemical structure were prepared. Afterwards, their biological activity was evaluated for their ability to reactivate acetylcholinesterase (AChE; EC and butyrylcholinesterase (BuChE; EC inhibited by paraoxon. Their reactivation activity was compared with standard reactivators--pralidoxime, obidoxime and HI-6--which are clinically used at present. As it resulted, none of the prepared compounds surpassed obidoxime, which is considered to be the most potent compound if used for reactivation of AChE inhibited by paraoxon. In case of BuChE reactivation, two compounds (K053 and K068) achieved similar results as obidoxime.
ESTHER : Kuca_2009_Molecules_14_4915
PubMedSearch : Kuca_2009_Molecules_14_4915
PubMedID: 20032868

Title : Biochemical responses of juvenile and adult Japanese quails to cyanobacterial biomass - Peckova_2009_Neuro.Endocrinol.Lett_30 Suppl 1_199
Author(s) : Peckova L , Bandouchova H , Hilscherova K , Damkova V , Sedlackova J , Vitula F , Paskova V , Pohanka M , Kohoutek J , Pikula J
Ref : Neuro Endocrinol Lett , 30 Suppl 1 :199 , 2009
Abstract : OBJECTIVES: The aim of the present study was to evaluate differences between juvenile and adult Japanese quails in responses to the exposure to cyanobacterial biomass in the diet. DESIGN: The OECD 205 Guideline on Avian Dietary Toxicity Test (1984) was employed in the experiment. A total of 75 freshly hatched chicks and 30 adults were exposed to cyanobacterial biomass for 15 days and blood sampled daily and on days 5, 10 and 15, respectively. Japanese quail chicks and adults received the same daily dose of approximately 224.4 ng microcystins per gram of body weight. Biochemical responses were compared against controls. RESULTS: No Japanese quail chicks and adults died during the acute 15-day-cyanobacterial-biomass exposure. Biochemical responses to the biomass in diet were first observed from day 5 post exposure to cyanobacterial biomass both in chicks and adults and there were age-related differences in the parameters changed. The responses of adult birds included an increase in lactate dehydrogenase, a drop in glucose and the total antioxidant capacity as well as a 15 to 20 % inhibition of acetylcholinesterase activity. Japanese quail chicks exposed to cyanobacterial biomass for the first 15 days after hatching reacted by having hypoproteinaemia, increased concentrations of triglycerides, uric acid and the total antioxidant capacity and a drop in high-density lipoprotein cholesterol in the blood. CONCLUSIONS: Chicks were not found to be more susceptible to the effects of biomass exposure. It seems that, due to their physiological preparation for the oxidative stress associated with hatching, Japanese quail chicks were even better able to cope with the cyanobacterial-biomass-induced oxidative stress than adults.
ESTHER : Peckova_2009_Neuro.Endocrinol.Lett_30 Suppl 1_199
PubMedSearch : Peckova_2009_Neuro.Endocrinol.Lett_30 Suppl 1_199
PubMedID: 20027171

Title : In vitro reactivation of trichlorfon-inhibited butyrylcholinesterase using HI-6, obidoxime, pralidoxime and K048 - Pohanka_2009_J.Enzyme.Inhib.Med.Chem_24_680
Author(s) : Pohanka M , Jun D , Kuca K
Ref : J Enzyme Inhib Med Chem , 24 :680 , 2009
Abstract : Trichlorfon is a specific inhibitor of cholinesterases. It was typically used as an insecticide; however, trichlorfon was described as useful for symptomatic treatment of Alzheimer's disease some years ago. The presented study is aimed at reactivation of trichlorfon-inhibited butyrylcholinesterase since this enzyme play an important role in Alzheimer's disease as deputy for acetylcholinesterase and furthermore it could be applied as a scavenger in case of overdosing. We used in vitro reactivation test for considering only reactivation efficacy of butyrylcholinesterase that is inhibited by trichlorfon and not reactivation of butyrylcholinesterase inhibited by trichlorfon metabolic products. Four reactivators were used: HI-6, pralidoxime, obidoxime, and K048. Although all of the reactivators seem to be effective at 1 mM concentration, a lower concentration was not able ensure sufficient reactivation. There was also an observed lowering of reactivation efficacy when butyrylcholinesterase was exposed to trichlorfon for a longer time interval.
ESTHER : Pohanka_2009_J.Enzyme.Inhib.Med.Chem_24_680
PubMedSearch : Pohanka_2009_J.Enzyme.Inhib.Med.Chem_24_680
PubMedID: 18825528

Title : Effect of five acetylcholinesterase reactivators on tabun-intoxicated rats: induction of oxidative stress versus reactivation efficacy - Pohanka_2009_J.Appl.Toxicol_29_483
Author(s) : Pohanka M , Karasova JZ , Musilek K , Kuca K , Kassa J
Ref : J Appl Toxicol , 29 :483 , 2009
Abstract : Oxime reactivators HI-6, obidoxime, trimedoxime, K347 and K628 were investigated as drugs designed for treatment of tabun intoxication. The experiments were performed on rats in order to simulate real conditions. Rats were intoxicated with one LD(50 )of tabun and treated with atropine and mentioned reactivators. Activities of erythrocyte acetylcholinesterase (AChE), plasma butyrylcholinesterase (BChE) and brain AChE were measured as markers of reactivation efficacy. An estimation of low molecular weight antioxidant levels using cyclic voltammetry was the second examination parameter. The evaluation of cholinesterases activity showed good reactivation potency of blood AChE and plasma BChE by commercially available obidoxime and newly synthesized K347. The potency of oximes to reactivate brain AChE was lower due to the poor blood-brain barrier penetration of used compounds. Commercially available reactivator HI-6 and newly synthesized K628 caused oxidative stress measured by cyclic voltammetry as antioxidant level. The oxidative stress provoked by HI-6 and K628 was found to be significant on probability level P = 0.05. The others reactivators did not affect antioxidant levels.
ESTHER : Pohanka_2009_J.Appl.Toxicol_29_483
PubMedSearch : Pohanka_2009_J.Appl.Toxicol_29_483
PubMedID: 19338015

Title : Progress of biosensors based on cholinesterase inhibition - Pohanka_2009_Curr.Med.Chem_16_1790
Author(s) : Pohanka M , Musilek K , Kuca K
Ref : Curr Med Chem , 16 :1790 , 2009
Abstract : Biosensors are available and applicable for detection and characterization of specific inhibitors of many enzymes. In this review, biosensors based on fixed acetylcholinesterase (AChE) or butyrylcholinesterase (BChE) are presented. Inhibition of selected enzymes by various compounds, such as organophosphorus and carbamate pesticides, nerve agents (e.g. sarin or VX), and other natural toxins (e.g. aflatoxins), was employed to develop specific assays using biosensors only. Biosensor technology brings potential miniaturization and portability, when it is compared to standard methods. Construction of biosensors based on cholinesterases became a more important issue within the last decades. Novel approach with recombinant proteins, microelectrodes and immobilization protocol related to nanotechnologies opened new insight to the cholinesterase based biosensor construction and its perspective via routine assays. This review is focused on novel trends within such biosensors as a result of the known platform.
ESTHER : Pohanka_2009_Curr.Med.Chem_16_1790
PubMedSearch : Pohanka_2009_Curr.Med.Chem_16_1790
PubMedID: 19442145

Title : Effect of seven newly synthesized and currently available oxime cholinesterase reactivators on cyclosarin-intoxicated rats - Karasova_2009_Int.J.Mol.Sci_10_3065
Author(s) : Karasova JZ , Kassa J , Musilek K , Pohanka M , Novotny L , Kuca K
Ref : Int J Mol Sci , 10 :3065 , 2009
Abstract : Seven new oxime-based acetylcholinesterase reactivators were compared with three currently available ones (obidoxime, trimedoxime, HI-6) for their ability to lessen cholinesterase inhibition in blood and brain of cyclosarin-treated rats. Oximes were given at doses of 5% their LD(50) along with 21 mg/kg atropine five min before the LD(50) of cyclosarin (120 ug/kg) was administered. Blood and brain samples were collected 30 minutes later. The greatest difference between acetylcholinesterase inhibition in blood of cyclosarin-treated rats was found after administration of HI-6 (40%), compared to 22% for trimedoxime and 6% for obidoxime. Only two of the seven newly synthesized oximes had any effect (K203 at 7%, K156 at 5%). Effective oximes against cyclosarin-inhibited plasma butyrylcholinesterase were HI-6 (42%), trimedoxime (11%), and K156 (4%). The oximes were less effective in brain than in blood, with reactivation values for HI-6 30% against acetylcholinesterase and 10% against butyrylcholinesterase. Values for newly synthesized oximes were less than 10% for K206, K269 and K203.
ESTHER : Karasova_2009_Int.J.Mol.Sci_10_3065
PubMedSearch : Karasova_2009_Int.J.Mol.Sci_10_3065
PubMedID: 19742125

Title : Reactivation of human brain homogenate cholinesterases inhibited by Tabun using newly developed oximes K117 and K127 - Kuca_2009_Basic.Clin.Pharmacol.Toxicol_105_207
Author(s) : Kuca K , Cabal J , Jung YS , Musilek K , Soukup O , Jun D , Pohanka M , Musilova L , Karasova J , Novotny L , Hrabinova M
Ref : Basic Clin Pharmacol Toxicol , 105 :207 , 2009
Abstract : Newly developed acetylcholinesterase reactivators K117 [1,5-bis(4-hydroxyiminomethylpyridinium)-3-oxapentane dichloride] and K127 [(1-(4-hydroxyiminomethylpyridinium)-5-(4-carbamoylpyridinium)-3-oxapentane dibromide)] were tested for their potency to reactivate tabun-inhibited human brain cholinesterases. Pralidoxime and trimedoxime were chosen as standard reference reactivators. Human tissue was used, as that was closer on the real treatment of human beings. As a result, oxime K127 was found as the best tested reactivator according to the constant k(r), characterizing the overall reactivation process. On the contrary, the maximal reactivation ability expressed as percentage of reactivation was the best for trimedoxime. This differences were caused as a result of using the enzyme from different species. Due to this, experiments on human tissue should be conducted after in vitro and in vivo tests on animals to eliminate such important failures of promising oximes.
ESTHER : Kuca_2009_Basic.Clin.Pharmacol.Toxicol_105_207
PubMedSearch : Kuca_2009_Basic.Clin.Pharmacol.Toxicol_105_207
PubMedID: 19473310

Title : Effect of several new and currently available oxime cholinesterase reactivators on tabun-intoxicated rats - Karasova_2008_Int.J.Mol.Sci_9_2243
Author(s) : Karasova JZ , Kassa J , Jung YS , Musilek K , Pohanka M , Kuca K
Ref : Int J Mol Sci , 9 :2243 , 2008
Abstract : The therapeutical efficacies of eleven oxime-based acetylcholinesterase reactivators were compared in an in vivo (rat model) study of treatment of intoxication caused by tabun. In this group there were some currently available oximes (obidoxime, trimedoxime and HI-6) and the rest were newly synthesized compounds. The best reactivation efficacy for acetylcholinesterase in blood (expressed as percent of reactivation) among the currently available oximes was observed after administration of trimedoxime (16%) and of the newly synthesized K127 (22432) (25%). The reactivation of butyrylcholinesterase in plasma was also studied; the best reactivators were trimedoxime, K117 (22435), and K127 (22432), with overall reactivation efficacies of approximately 30%. Partial protection of brain ChE against tabun inhibition was observed after administration of trimedoxime (acetylcholinesterase 20%; butyrylcholinesterase 30%) and obidoxime (acetylcholinesterase 12%; butyrylcholinesterase 16%).
ESTHER : Karasova_2008_Int.J.Mol.Sci_9_2243
PubMedSearch : Karasova_2008_Int.J.Mol.Sci_9_2243
PubMedID: 19330072

Title : The effect of HI-6 on cholinesterases and on the cholinergic system of the rat bladder - Soukup_2008_Neuro.Endocrinol.Lett_29_759
Author(s) : Soukup O , Pohanka M , Tobin G , Jun D , Fusek J , Musilek K , Marek J , Kassa J , Kuca K
Ref : Neuro Endocrinol Lett , 29 :759 , 2008
Abstract : OBJECTIVES: The current standard treatment of organophosphate poisoning consists of an administration of anticholinergic drugs and cholinesterase reactivators (oximes). Oximes can react - except their reactivating effect on cholinesterases - directly with cholinoreceptors. HI-6 is an oxime that may have an inhibitory effect on the muscarinic receptors, too. METHODS: In our work, we have investigated an influence of HI-6 on the acetylcholinesterase (AChE), butyrylcholinesterase (BChE) and on the muscarinic receptors in vitro. The study was conducted using biosensor technique and on the rat bladder using in vitro test (tissue bath; methacholine as muscarinic agonist). IC50 for BChE from human serum was determined to be 1.01x10-6 M and for human erythrocytes AChE 3.31x10-6 M, respectively. CONCLUSION: We assume that the demonstrated contractile response can be attributed to the inhibition of the AChE at the lower concentration and to a predominant inhibition of muscarinic receptor at higher concentration of compound tested.
ESTHER : Soukup_2008_Neuro.Endocrinol.Lett_29_759
PubMedSearch : Soukup_2008_Neuro.Endocrinol.Lett_29_759
PubMedID: 18987577

Title : Cholinesterase biosensor construction - a review - Pohanka_2008_Protein.Pept.Lett_15_795
Author(s) : Pohanka M , Jun D , Kalasz H , Kuca K
Ref : Protein Pept Lett , 15 :795 , 2008
Abstract : Biosensors using cholinesterases as the biorecognition component have been used to assay organophosphates and carbamates for a long time. In this review, some strategies convenient for biosensor construction are presented. Solutions for cholinesterase immobilization and output signal monitoring are presented as the basic presumptions for successful biosensor construction.
ESTHER : Pohanka_2008_Protein.Pept.Lett_15_795
PubMedSearch : Pohanka_2008_Protein.Pept.Lett_15_795
PubMedID: 18855751

Title : New performance of biosensor technology for Alzheimer's disease drugs: in vitro comparison of tacrine and 7-methoxytacrine - Pohanka_2008_Neuro.Endocrinol.Lett_29_755
Author(s) : Pohanka M , Kuca K , Kassa J
Ref : Neuro Endocrinol Lett , 29 :755 , 2008
Abstract : Two drugs were tested using electrochemical biosensor with immobilized acetylcholinesterase (AChE). The first was commercialized drug tacrine (known also as Cognex) used for treatment of cognitive manifestation of Alzheimer\'s disease (AD). The second one was its 7-methoxy derivate (7-MEOTA) that has not been marketed. We determined the IC50 (6.67+/-0.92)x10-7 M for tacrine and (1.66+/-1.43)x10-9 M for 7-MEOTA. In this in vitro study, 7-MEOTA acts as stronger inhibitor of AChE and in this way could be more favorable for treatment of cognitive manifestation of AD. Our study shows that biosensor technology could be used as a quick and cheap tool for testing of promising AChE inhibitors (AD drug candidates).
ESTHER : Pohanka_2008_Neuro.Endocrinol.Lett_29_755
PubMedSearch : Pohanka_2008_Neuro.Endocrinol.Lett_29_755
PubMedID: 18987590

Title : Diagnosis of Intoxication by the Organophosphate VX: Comparison Between an Electrochemical Sensor and Ellman s Photometric Method - Pohanka_2008_Sensors.(Basel)_8_5229
Author(s) : Pohanka M , Hrabinova M , Kuca K
Ref : Sensors (Basel) , 8 :5229 , 2008
Abstract : An electrochemical sensor is introduced as a tool applicable for diagnosis of intoxication by cholinesterase inhibitors caused by the well-known nerve agent VX. The traditional Ellman method was chosen for comparison with the sensor's analytical parameters. Both methods are based on estimation of blood cholinesterase inhibition as a marker of intoxication. While Ellman s method provided a limit of detection of 5.2 10-7 M for blood containing VX, the electrochemical sensor was able to detect 4.0 10-7 M. Good correlation between both methods was observed (R = 0.92). The electrochemical sensor could be considered a convenient tool for a fast yet accurate method, easily available for field as well as laboratory use. Time and cost savings are key features of the sensor-based assay.
ESTHER : Pohanka_2008_Sensors.(Basel)_8_5229
PubMedSearch : Pohanka_2008_Sensors.(Basel)_8_5229
PubMedID: 27873811

Title : Amperometric Biosensors for Real Time Assays of Organophosphates - Pohanka_2008_Sensors.(Basel)_8_5303
Author(s) : Pohanka M , Jun D , Kuca K
Ref : Sensors (Basel) , 8 :5303 , 2008
Abstract : An amperometric biosensor based on acetylcholinesterase (AChE) immobilized in gelatin was used to develop an assay for the organophosphate paraoxon. The more traditional manner employing preincubation was used for comparison between measurement procedures, although the aim of the study was to examine the performance of the biosensor for real time monitoring of organophosphates. The biosensor was immersed in a reaction chamber and paraoxon was injected inside. We were able to detect 200 pg of paraoxon within one minute or 2.5 ppb when the biosensor was preincubed in the sample solution for 15 minutes. The practical impact and expectations are discussed.
ESTHER : Pohanka_2008_Sensors.(Basel)_8_5303
PubMedSearch : Pohanka_2008_Sensors.(Basel)_8_5303
PubMedID: 27873815

Title : Photometric microplate assay for estimation of the efficacy of paraoxon-inhibited acetylcholinesterase reactivation - Pohanka_2008_J.Enzyme.Inhib.Med.Chem_23_781
Author(s) : Pohanka M , Jun D , Kuca K
Ref : J Enzyme Inhib Med Chem , 23 :781 , 2008
Abstract : Photometric microplate assay was performed for testing of paraoxon-inhibited acetylcholinesterase (AChE) using three reactivators for reactivation purposes: obidoxime, pralidoxime, and HI-6. 3-D graphs (percent of reactivation vs. concentration of reactivator and vs. time of reactivator effecting) were constructed for each reactivator to compare their efficacy. The best results were obtained using obidoxime where reactivation was near to 80%. Suitability of photometric microplates for following of reactivation procedures is discussed.
ESTHER : Pohanka_2008_J.Enzyme.Inhib.Med.Chem_23_781
PubMedSearch : Pohanka_2008_J.Enzyme.Inhib.Med.Chem_23_781
PubMedID: 18615283

Title : Amperometric biosensor for pesticide methamidophos assay - Pohanka_2007_Acta.Medica.(Hradec.Kralove)_50_239
Author(s) : Pohanka M , Kuca K , Jun D
Ref : Acta Medica (Hradec Kralove) , 50 :239 , 2007
Abstract : Amperometric biosensor based on enzyme acetylcholinesterase (AChE; EC was tested for pesticide methamidophos assay. Biosensor consists from four screen printed platinum electrodes on ceramic strip. AChE was physically adsorbed onto the electrode surface. The measuring principle was based on the inhibition of AChE activity in the presence of methamidophos. The proposed method limit of detection was 2.45 nM, responding to 3.46 pg of methamidophos detected absolutely when we consider the sample volume.
ESTHER : Pohanka_2007_Acta.Medica.(Hradec.Kralove)_50_239
PubMedSearch : Pohanka_2007_Acta.Medica.(Hradec.Kralove)_50_239
PubMedID: 18290547