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International Journal of Molecular... Nov 2022Seven pyridoxal dioxime quaternary salts (-) were synthesized with the aim of studying their interactions with human acetylcholinesterase (AChE) and...
Seven pyridoxal dioxime quaternary salts (-) were synthesized with the aim of studying their interactions with human acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The synthesis was achieved by the quaternization of pyridoxal monooxime with substituted 2-bromoacetophenone oximes (phenacyl bromide oximes). All compounds, prepared in good yields (43-76%) and characterized by 1D and 2D NMR spectroscopy, were evaluated as reversible inhibitors of cholinesterase and/or reactivators of enzymes inhibited by toxic organophosphorus compounds. Their potency was compared with that of their monooxime analogues and medically approved oxime HI-6. The obtained pyridoxal dioximes were relatively weak inhibitors for both enzymes ( = 100-400 µM). The second oxime group in the structure did not improve the binding compared to the monooxime analogues. The same was observed for reactivation of VX-, tabun-, and paraoxon-inhibited AChE and BChE, where no significant efficiency burst was noted. In silico analysis and molecular docking studies connected the kinetic data to the structural features of the tested compound, showing that the low binding affinity and reactivation efficacy may be a consequence of a bulk structure hindering important reactive groups. The tested dioximes were non-toxic to human neuroblastoma cells (SH-SY5Y) and human embryonal kidney cells (HEK293).
Topics: Humans; Butyrylcholinesterase; Acetylcholinesterase; Cholinesterase Reactivators; Molecular Docking Simulation; Cholinesterase Inhibitors; HEK293 Cells; Neuroblastoma; Oximes; Pyridoxal; Ligands
PubMed: 36362178
DOI: 10.3390/ijms232113388 -
Toxicology in Vitro : An International... Feb 2023Neuropathy target esterase (NTE) has been proven to act as a lysophospholipase (LysoPLA) and phospholipase B (PLB) in mammalian cells. In this study, we took human...
Neuropathy target esterase (NTE) has been proven to act as a lysophospholipase (LysoPLA) and phospholipase B (PLB) in mammalian cells. In this study, we took human neuroblastoma SK-N-SH cells as the research object and explored the effect of NTE on phospholipid homeostasis. The results showed that phosphatidylcholine (PC) and lysophosphatidylcholine (LPC) levels significantly increased (> 40%), while glycerophosphocholine (GPC) decreased (below 60%) after NTE gene was knockdown in the cells (NTE < 30% of control), which were prepared by gene silencing with dsRNA-NTE. However, in the NTE-overexpressed cells (NTE > 50% of control), which were prepared by expressing recombinant catalytic domain of NTE, LPC remarkably decreased (below 80%) and GPC enhanced (> 40%). Mipafox, a neuropathic organophosphorus compound (OP), significantly inhibited NTE-LysoPLA and NTE-PLB activities (> 95-99% inhibition at 50 μM), which was accompanied with a decreased GPC level (below 40%) although no change of the PC and LPC levels was observed; while paraoxon, a non-neuropathic OP, suppresses neither the activities of NTE-phospholipases nor the levels of PC, LPC, and GPC. Thus, we concluded that both the stable up- or down-regulated expression of NTE gene and the loss of NTE-LysoPLA/PLB activities disrupts phospholipid homeostasis in the cells although the inhibition of NTE activity only decreased GPC content without altering PC and LPC levels.
Topics: Humans; Carboxylic Ester Hydrolases; Homeostasis; Lysophosphatidylcholines; Lysophospholipase; Mammals; Neuroblastoma; Organophosphorus Compounds; Phosphatidylcholines; Phospholipids
PubMed: 36336212
DOI: 10.1016/j.tiv.2022.105509 -
Analytical Chemistry Nov 2022Mn(II)-based electron spin resonance (ESR) spectroscopy was used for detecting butyrylcholinesterase (BChE) and organophosphorus pesticides (OPs). MnO nanosheets were...
Label-Free and Ultrasensitive Detection of Butyrylcholinesterase and Organophosphorus Pesticides by Mn(II)-Based Electron Spin Resonance Spectroscopy with a Zero Background Signal.
Mn(II)-based electron spin resonance (ESR) spectroscopy was used for detecting butyrylcholinesterase (BChE) and organophosphorus pesticides (OPs). MnO nanosheets were synthesized with manganese chloride and hydrogen peroxide. With the catalysis of BChE, S-butyrylthiocholine iodide (BTCh) was hydrolyzed into thiocholine which has a reducing -SH group. In the presence of thiocholine, MnO nanosheets were broken down and Mn(IV) in MnO nanosheets was reduced into Mn(II). Mn is a paramagnetic ion and gives a good ESR signal. In contrast, MnO nanosheets have no ESR signal and need not be separated from Mn. Mn can be determined directly by ESR spectroscopy, and no further sensing probe is needed. ESR spectroscopy based on directly detecting Mn is much simpler than those using other probes besides MnO. The ESR signal of Mn is proportional to the catalytic activity of BChE. OPs which inhibit the activity of BChE can also be detected by probing the ESR signal of Mn. Since there is no ESR signal of MnO nanosheets, the background signal in the absence of BChE was close to zero. The limit of detection (LOD) of BChE was as low as 0.042 U L. The standard curve for determining the OP paraoxon was established by measuring the inhibition of BChE by paraoxon, and the LOD of paraoxon was found to be 0.076 ng mL. The spiked Chinese cabbage extract samples were analyzed, and the experimental results indicated that the recoveries were from 96.5 to 102.8%. The planted Chinese cabbage was sprayed with the paraoxon solution, and the residue amount of paraoxon in the extract was estimated by the method. The result obtained by the present method was consistent with that obtained by HPLC, which proved the practicability of this new method.
Topics: Butyrylcholinesterase; Manganese Compounds; Pesticides; Paraoxon; Organophosphorus Compounds; Electron Spin Resonance Spectroscopy; Oxides; Thiocholine
PubMed: 36332200
DOI: 10.1021/acs.analchem.2c03708 -
Biosensors & Bioelectronics Jan 2023Enzyme-mediated dephosphorylation reaction is the important approach to realize the inactivation and detection of hazardous phosphate chemicals. To date, many...
Enzyme-mediated dephosphorylation reaction is the important approach to realize the inactivation and detection of hazardous phosphate chemicals. To date, many phosphatases-like nanozymes (e.g., CeO) have demonstrated the catalytic hydrolysis ability of the phosphomonoesters, rather than phosphotriester, and the CeO nanozyme only work under relatively harsh conditions of high temperature, and large dosage. Thus, exploration of efficient nanozymes for the rapid dephosphorylation of phosphotriester under mild conditions remains a challenge. Here, a novel CeO@NC nanozyme is developed with excellent phosphatases-like activity based on substrate synergistic effect, in which, CeO nanoparticles embedded in N-doped carbon (NC) material. Taking paraoxon as the model substrate, such CeO@NC nanozyme can drive rapid dephosphorylation of phosphotriester over a broad temperature range, which not only significantly outperforms natural phosphatases and neat CeO, but also can preserve >80% of the optimal activity after exposure of harsh conditions, such as strong acidic/basic medium, high temperature of up to 80 °C. The excellent catalytic performance could be due to that Ce(IV)/Ce(III) species act as the active sites to realize the polarization and hydrolysis of P-O bond while NC template works as the synergistic group to adsorb the substrate. Furthermore, a simple colorimetric assay is developed for the rapid and selective detection of paraoxon. Overall, this work not only develops a highly efficient phosphatases-like nanozyme via substrate synergetic strategy, but also opens an interesting avenue for the rapid detection of organophosphorus pesticides.
Topics: Paraoxon; Colorimetry; Cerium; Organophosphorus Compounds; Pesticides; Biosensing Techniques; Phosphoric Monoester Hydrolases
PubMed: 36323162
DOI: 10.1016/j.bios.2022.114841 -
Chemosphere Jan 2023Cytosolic phospholipase A2 (cPLA2) belongs to a large family of proteins and plays a crucial role in the regulation of arachidonic acid metabolism and inflammation...
Cytosolic phospholipase A2 (cPLA2) belongs to a large family of proteins and plays a crucial role in the regulation of arachidonic acid metabolism and inflammation cascade in zebrafish (Danio rerio). This enzyme with a molecular weight of 85 kDa, has two distinct domains. One is the regulatory and calcium-dependent (Ca) domain called C2, the other is the catalytic α/β hydrolase Ca-independent domain, where serine and aspartic acid catalytic dyad residues are present. We investigated the interaction of malathion and their organophosphate metabolites in the cPLA2 using in silico tools. Molecular docking results showed hydrophobic interactions with the paraoxon and catalytic site residue (Ser 223). Malathion increases intracellular Ca due to endoplasmic reticulum influx which in turn activities phospholipase A2 and arachidonic acid release. Molecular docking and homology modelling of proteins and ligands could be a complementary tool for ecotoxicology and environment pollution assessment.
Topics: Animals; Zebrafish; Cytosol; Malathion; Arachidonic Acid; Molecular Docking Simulation; Phospholipases A2; Calcium; Phospholipases A2, Cytosolic
PubMed: 36306964
DOI: 10.1016/j.chemosphere.2022.136984 -
ACS Applied Materials & Interfaces Oct 2022Defense against chemical warfare agents (CWAs) is regarded as a top priority for the protection of humanity, but it still depends on physical protection with severe...
Feasible Detoxification Coating Material for Chemical Warfare Agents Using Poly(methyl methacrylate)-Branched Poly(ethyleneimine) Copolymer and Metal-Organic Framework Composites.
Defense against chemical warfare agents (CWAs) is regarded as a top priority for the protection of humanity, but it still depends on physical protection with severe limitations such as residual toxicity and post-treatment requirement. In this study, a strategically designed functional polymeric substrate was composited with a metal-organic framework catalyst to remove toxicity immediately. A series of PMMA-BPEI copolymers exhibited high processability as a coating and accelerated the catalytic activity of Zr(IV)-based metal-organic framework catalysts (UiO-66). Among them, PMB_40 composite coating on a cotton fabric, containing a PMMA-BPEI copolymer (PMMA/BPEI = 1/2) and 40% of UiO-66 catalyst, can efficiently decompose nerve agent simulants (methyl-paraoxon) under both liquid phase ( = 0.14 h) and humidified ( = 4.8 h) conditions. Moreover, a real agent, GD, was decomposed 100% by PMB_40 in 4 h at 25 °C and 65% relative humidity. On the basis of superior catalytic activity, the PMB composites are anticipated to be a potential material for active chemical protection coating.
PubMed: 36288400
DOI: 10.1021/acsami.2c15961 -
Structure (London, England : 1993) Nov 2022Organophosphorus (OP) compounds, including nerve agents and some pesticides, covalently bind to the catalytic serine of human acetylcholinesterase (hAChE), thereby...
Organophosphorus (OP) compounds, including nerve agents and some pesticides, covalently bind to the catalytic serine of human acetylcholinesterase (hAChE), thereby inhibiting acetylcholine hydrolysis necessary for efficient neurotransmission. Oxime antidotes can reactivate the OP-conjugated hAChE, but reactivation efficiency can be low for pesticides, such as paraoxon (POX). Understanding structural and dynamic determinants of OP inhibition and reactivation can provide insights to design improved reactivators. Here, X-ray structures of hAChE with unaged POX, with POX and oximes MMB4 and RS170B, and with MMB4 are reported. A significant conformational distortion of the acyl loop was observed upon POX binding, being partially restored to the native conformation by oximes. Neutron vibrational spectroscopy combined with molecular dynamics simulations showed that picosecond vibrational dynamics of the acyl loop soften in the ∼20-50 cm frequency range. The acyl loop structural perturbations may be correlated with its picosecond vibrational dynamics to yield more comprehensive template for structure-based reactivator design.
Topics: Humans; Acetylcholinesterase; Paraoxon; Crystallography, X-Ray; Cholinesterase Inhibitors; Oximes; Organophosphorus Compounds; Neutrons; Pesticides
PubMed: 36265484
DOI: 10.1016/j.str.2022.09.006 -
Global Challenges (Hoboken, NJ) Sep 2022Glyphosate is a globally applied herbicide yet it has been relatively undetectable in-field samples outside of gold-standard techniques. Its presumed nontoxicity toward...
Glyphosate is a globally applied herbicide yet it has been relatively undetectable in-field samples outside of gold-standard techniques. Its presumed nontoxicity toward humans has been contested by the International Agency for Research on Cancer, while it has been detected in farmers' urine, surface waters and crop residues. Rapid, on-site detection of glyphosate is hindered by lack of field-deployable and easy-to-use sensors that circumvent sample transportation to limited laboratories that possess the equipment needed for detection. Herein, the flavoenzyme, glycine oxidase, immobilized on platinum-decorated laser-induced graphene (LIG) is used for selective detection of glyphosate as it is a substrate for GlyOx. The LIG platform provides a scaffold for enzyme attachment while maintaining the electronic and surface properties of graphene. The sensor exhibits a linear range of 10-260 m, detection limit of 3.03 m, and sensitivity of 0.991 nA m . The sensor shows minimal interference from the commonly used herbicides and insecticides: atrazine, 2,4-dichlorophenoxyacetic acid, dicamba, parathion-methyl, paraoxon-methyl, malathion, chlorpyrifos, thiamethoxam, clothianidin, and imidacloprid. Sensor function is further tested in complex river water and crop residue fluids, which validate this platform as a scalable, direct-write, and selective method of glyphosate detection for herbicide mapping and food analysis.
PubMed: 36176938
DOI: 10.1002/gch2.202200057 -
Environmental Toxicology and Chemistry Dec 2022The acetylcholinesterase (AChE) inhibition assay has been frequently applied for environmental monitoring to capture insecticides such as organothiophosphates (OTPs) and...
The acetylcholinesterase (AChE) inhibition assay has been frequently applied for environmental monitoring to capture insecticides such as organothiophosphates (OTPs) and carbamates. However, natural organic matter such as dissolved organic carbon (DOC) co-extracted with solid-phase extraction from environmental samples can produce false-negative AChE inhibition in free enzyme-based AChE assays. We evaluated whether disturbance by DOC can be alleviated in a cell-based AChE assay using differentiated human neuroblastoma SH-SY5Y cells. The exposure duration was set at an optimum of 3 h considering the effects of OTPs and carbamates. Because loss to the airspace was expected for the more volatile OTPs (chlorpyrifos, diazinon, and parathion), the chemical loss in this bioassay setup was investigated using solid-phase microextraction followed by chemical analysis. The three OTPs were relatively well retained (loss <34%) during 3 h of exposure in the 384-well plate, but higher losses occurred on prolonged exposure, accompanied by slight cross-contamination of adjacent wells. Inhibition of AChE by paraoxon-ethyl was not altered in the presence of up to 68 mg /L Aldrich humic acid used as surrogate for DOC. Binary mixtures of paraoxon-ethyl and water extracts showed concentration-additive effects. These experiments confirmed that the matrix in water extracts does not disturb the assay, unlike purified enzyme-based AChE assays. The cell-based AChE assay proved to be suitable for testing water samples with effect concentrations causing 50% inhibition of AChE at relative enrichments of 0.5-10 in river water samples, which were distinctly lower than corresponding cytotoxicity, confirming the high sensitivity of the cell-based AChE inhibition assay and its relevance for water quality monitoring. Environ Toxicol Chem 2022;41:3046-3057. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.
Topics: Humans; Acetylcholinesterase; Paraoxon; Water Quality; Neuroblastoma; Insecticides; Organothiophosphates; Carbamates; Cholinesterase Inhibitors
PubMed: 36165561
DOI: 10.1002/etc.5490 -
Analytica Chimica Acta Oct 2022Sensors based on colorimetry, fluorescence, and electrochemistry have been widely employed to detect acetylcholinesterase and its inhibitors, however, there are only a...
Sensors based on colorimetry, fluorescence, and electrochemistry have been widely employed to detect acetylcholinesterase and its inhibitors, however, there are only a minority of strategies for AChE detection based on photothermal method. This work reports a versatile dual-mode colorimetric and photothermal biosensing platform for acetylcholinesterase (AChE) detection and its inhibitor (paraoxon-ethyl, a model of AChE inhibitors) monitor based on Fe-N-C/HO/3,3',5,5'-tetramethylbenzidine (TMB) system. The Fe-N-C with abundant active Fe-Nx sites shows outstanding peroxidase-mimicking activity and can be used to promote the generation of •OH by HO to oxidize TMB. However, the introduction of mercapto molecules tending to coordinate with metal atoms result in the block of action site in Fe-N-C, thereby decrease its peroxidase-mimetic activity. The designed biosensor principle is based on the block of active sites of Fe-N-C by thiocholine (TCh, one kind of mercapto molecules) that can be produced by acetylthiocholine (ATCh) in the presence of AChE. Under optimum conditions, the limit of detection (LOD) for AChE activity is 1.9 mU mL (colorimetric) and 2.2 mU mL (photothermal), while for paraoxon-ethyl is 0.012 μg mL (colorimetric) and 0.013 μg mL (photothermal), respectively. The assay we proposed not only can be designed to monitor AChE detection and its inhibitors, but also can be easily extended for the detection of other biomolecules relate to the generation or consumption of HO.
Topics: Acetylcholinesterase; Acetylthiocholine; Benzidines; Biosensing Techniques; Colorimetry; Hydrogen Peroxide; Paraoxon; Peroxidases; Thiocholine
PubMed: 36156227
DOI: 10.1016/j.aca.2022.340383