Presence Of Acetylcholinesterase Research Articles

A molecular container providing supramolecular protection against acetylcholine hydrolysis.

Alzheimer's disease (AD) is characterized by cognitive decline, often attributed to the deficiency of acetylcholine, which can undergo hydrolysis by acetylcholinesterase (AChE) within the biological milieu. Here, we report a supramolecular strategy that takes advantage of confinement effects to inhibit such a hydrolysis process, shedding some light on AD therapy. A water-soluble and bowl-shaped molecule, hexacarboxylated tribenzotriquinacene (TBTQ-C6), was employed to shield acetylcholine (G1) from enzymatic degradation through host-guest binding interactions. Our study revealed highly efficient host-guest interactions with a binding ratio of 1 : 3, resulting in a significant reduction in acetylcholine hydrolysis from 91.1% to 7.4% in the presence of AChE under otherwise identical conditions. Furthermore, TBTQ-C6 showed potential for attenuating the degradation of butyrylcholine (G2) by butyrylcholinesterase (BChE). The broader implications of this study extend to the potential use of molecular containers in various biochemical and pharmacological applications, opening new avenues for research in the field of neurodegenerative diseases.

Low levels of endogenous cholinesterases support the choice of cows, sheep and goats for the transgenic expression of human butyrylcholinesterase in milk

Butyrylcholinesterase purified from human plasma (Hu BChE) as well as recombinant (r) Hu BChE are candidate enzymes that can protect humans from toxicity of organophosphorus compounds (OPs). Domestic animals such as cows, pigs, sheep, and goats have been used for the transgenic expression of a variety of valuable therapeutic proteins. Indeed, rHu BChE was successfully expressed in the milk of transgenic goats, but the presence of any endogenous cholinesterases (ChE) in milk would interfere with the isolation of expressed rHu BChE. The aim of this study was to determine the presence of endogenous ChEs in bovine, ovine, caprine, and porcine milk to determine the suitability of these species for the production of rHu BChE. Using acetyl- and butyryl- thiocholine as substrates, ChE activity (2–4 U/mL) was detected in pig milk only. ChE activities in milk from other animals were <0.01 U/mL and could only be detected following enrichment on procainamide-Sepharose gel. Two different methods based on measuring activity in the presence of acetylcholinesterase (AChE)- or BChE- specific inhibitors were used to estimate the proportions of AChE and BChE activities in enriched milk. Monoclonal antibodies (MAbs), against fetal bovine serum AChE that recognize AChEs from ruminants only, were also used to confirm the identity of AChEs. While bovine and ovine milk contain both AChE and BChE activities, caprine and porcine milk contain predominantly BChE activity. The presence of very low ChE activity supports the choice of cows, sheep, and goats for the transgenic expression of rHu BChE in milk.

A biochemistry‐oriented drug design: synthesis, anticancer activity, enzymes inhibition, molecular docking studies of novel 1,2,4-triazole derivatives

In this study, we researched the reactions of 5-(5-bromofuran-2-yl)-4-methyl-1,2,4-triazole-3-thiol and 5-thiophene-(3-ylmethyl)-4R-1,2,4-triazole-3-thiols with some halogen-containing compounds, a number of new compounds were synthesized (1.1–1.5 and 2.1–2.8). These compounds showed excellent to good inhibitory activities on acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes. For obtaining the effects of these compounds on AChE and BChE enzymes were determined spectrophotometrically according to Ellman. IC50 values of these enzymes were ranging between 1.63 and 17.68 nM for AChE and 8.71 and 84.02 nM for BChE. After, prostate cancer is the second leading cause of cancer-related mortality for men over the age of 65 in developed countries. Current treatment options remain limited in the treatment of advanced-stage prostate cancer leading to biochemical recurrence in almost 40% of the patients. Therefore, there is an urgent need for development of novel therapeutic tools for treatment of prostate cancer patients. In this study, we aimed at analyzing the potential of all compounds against prostate cancer cells. We found that, of the tested compounds, 2.1, 2.2 and 2.3 showed significant cytotoxic activities against PC3 prostate cancer cells, although their effect on the viability of normal prostate cells was limited. These findings suggest their selective targeting potential for prostate cancer cells and offer them as candidate therapeutic agents against prostate cancer. The inhibitory activities of some chemical compounds, such as (1.1–1.5 and 2.1–2.8) were assessed by performing the molecular docking study in the presence of AChE, BChE and prostate cancer protein. MM/GBSA methods are calculated binding free energy. Finally, ADME/T analysis was performed to examine the drug properties of the 13 studied molecules. Communicated by Ramaswamy H. Sarma

Development of a carbon quantum dot-based sensor for the detection of acetylcholinesterase and the organophosphate pesticide

In this study, a proper and reliable fluorometric method is introduced for screening acetylcholinesterase (AChE) and its inhibitors, using carbon quantum dots (CQDs) as the signal reporter. Pure, S-doped, and P-doped CQDs, were synthesized and their recoverable fluorescence quenching properties were observed, when exposed to Hg2+, Cu2+, and Fe3+ quenching ions, respectively. The study on the recovery of their emission showed that after the introduction of another guest substance with a stronger affinity to the quenching ions, their fluorescence is restored. The Design Expert software was employed to compare the performance of the three CQDs, as fluorescent probes, based on their quenching efficiency and the percentage of their emission recovery in the presence of AChE and acetylthiocholine (ATCh). Based on the statistical analysis, among the studied CQDs, S-doped CQD was the most suitable candidate for sensor designing. The detection mechanism for the proposed S-doped CQD-based sensor is as follows: The strong binding of Cu2+ ions to carboxyl groups of S-doped CQD quenches the fluorescence signal. Then, hydrolysis of ATCh into thiocholine (TCh) in the presence of AChE causes fluorescence recovery, due to the stronger affinity of Cu2+ to the TCh, rather than the CQD. Finally, in the presence of malathion and chlorpyrifos inhibitors, AChE loses its ability to hydrolyze ATCh to TCh, so the fluorescence emission remains quenched. Based on the proposed detection technique, the designed sensor showed detection limits of 1.70 ppb and 1.50 ppb for malathion and chlorpyrifos, respectively.

Chemical composition, cholinesterase, and α-glucosidase inhibitory activity of the essential oils of some Iranian native Salvia species

BackgroundThe plants from Salvia genus contain widely distributed species which have been used in folk medicine as well as pharmaceutical and food industries.MethodsThe chemical composition of 12 native Iranian Salvia species (14 plants) was identified using gas chromatography-mass spectrometry (GC–MS). Also, the inhibitory activity of all essential oils (EOs) was evaluated toward α-glucosidase and two types of cholinesterase (ChE) using spectrophotometric methods. The in vitro α-glucosidase inhibition assay was performed by the determination of p-nitrophenol (pNP) obtained from the enzymatic dissociation of p-nitrophenol-α-D-glucopyranoside (pNPG) as the substrate. In vitro ChE inhibitory assay was conducted based on the modified Ellman’s procedure using the measurement of 5-thio-2-nitrobenzoic acid produced from the hydrolysis of thiocholine derivatives as the substrate, in the presence of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE).ResultsTotally, 139 compounds were detected and caryophyllene oxide and trans-β-caryophyllene were the most abundant compounds in all EOs. The yield of EOs extracted from the plants were also calculated in the range of 0.06 to 0.96% w/w. Herein, α-glucosidase inhibitory activity of 8 EOs was reported for the first time and among all, S. spinosa L. was found to be the most potent inhibitor (90.5 inhibition at 500 μg/mL). Also, the ChE inhibitory activity of 8 species was reported for the first time and our results showed that the BChE inhibitory effect of all EOs was more potent than that of AChE. The ChE inhibition assay indicated that S. mirzayanii Rech.f. & Esfand. collected from Shiraz was the most potent inhibitor (72.68% and 40.6% at the concentration of 500 μg/mL, toward AChE and BChE, respectively).ConclusionsIt seems that native Salvia species of Iran could be considered in the development of anti-diabetic and anti-Alzheimer's disease supplements.

Construction of a ratiometric fluorescent sensing platform based on near-infrared carbon dots for organophosphorus pesticides detection.

In this work, a convenient ratiometric fluorescent platform was designed to measure organophosphorus pesticides (OPs) based on acetylcholinesterase (AChE), acetylthiocholine (ATCh), manganese dioxide nanosheets (MnO2), near-infrared carbon dots (RCDs) and o-phenylenediamine (OPD). In this platform, a direct oxidation of OPD by MnO2 generated the luminescent product 2,3-diaminophenolazine (DAP) through intrinsic oxidase activity, while RCDs served as a fluorescent reference indicator. In the presence of AChE and ATCh, the enzymatic hydrolysate thiocholine (TCh) would reduce MnO2 nanosheets to Mn2+, leading to the quenching of DAP fluorescence. On the other hand, OPs can inhibit the catabolism of ATCh by AChE thus acting as a recognizer of OPs. According to these reactions, OPs were quantitatively analyzed by the intensity ratio of fluorescence emitted from RCDs and DAP (F560/F676). The constructed platform can detect OPs with the range of 0.2-0.6μM with a detection limit of 4.3nM. Figure A ratiometric fluorescent probe based on carbon dots was obtained and using it to determine the concentration of organophosphorus pesticides.

A visualized sensor based on layered double hydroxides with peroxidase-like activity for sensitive acetylcholinesterase assay.

Acetylcholinesterase (AChE) plays a crucial role in biological neurotransmission. The aberrant expression of AChE is associated with various neurodegenerative diseases. Therefore, it is of great significance to develop a simple and highly sensitive AChE analysis platform. Herein, a simple colorimetric sensor was constructed for sensitive detection of AChE based on the peroxidase-like catalytic activity of Ni/Co layered double hydroxides (Ni/Co LDHs). In this sensor, the fabricated Ni/Co LDHs possess high peroxidase-like activity, enabling rapid catalysis of o-phenylenediamine (OPD) to produce yellow oxOPD in the presence of H2O2. This peroxidase-like activity of Ni/Co LDHs was found to be effectively inhibited by the presence of AChE. It is speculated that the combination of AChE on the outer surface of Ni/Co LDHs through non-covalent interaction may cover the active sites and hinder their adsorption to the substrates, leading to the failure of OPD oxidation. As a result, the yellow color from oxOPD is related to the AChE concentration, enabling the direct AChE assay in an equipment-free manner. In addition, the fabricated Ni/Co LDHs could be modified on a paper surface to obtain a paper-based analytical device for visualized colorimetric detection of AChE. The as-proposed sensor shows high sensitivity to AChE with a detection limit down to 6.6 μU mL-1. Therefore, this naked-eye paper-based sensor is capable of on-site and real-time detection of AChE, and has outstanding application prospects in clinical diagnosis and biomedical fields.

CRISPR/Cas12a-based fluorescence assay for the detection of acetylcholinesterase activity

Here we report an ultrasensitive and simple acetylcholinesterase (AChE) detection method by utilizing CRISPR/Cas12a collateral cleavage activity. In this design, the activator ssDNA of CRISPR/Cas12a was absorbed and sealed in the synthesized MnO2 nanosheets. The hydrolysate substrate, acetylthiocholine (ATCh), in the presence of AChE could reduce MnO2 nanosheets to Mn2+, thereby releasing the activator ssDNA, which subsequently activated the collateral cleavage ability of CRISPR/Cas12a to generate a fluorescent signal. As expected, AChE was successfully determined with high sensitivity based on the signal amplification ability of CRISPR/Cas12a collateral cleavage. Under the optimized conditions, the detection limit of AChE could reach as low as 0.0087 U/mL. In addition, for the detection of spiked AChE in human serum samples, our assay demonstrated excellent specificity and good recovery, demonstrating its practical applicability. This work would provide new insight into the construction of novel CRISPR/Cas12a strategies for the detection of enzymes beyond molecular diagnostics.

Dual-mode colorimetric-photothermal sensing platform of acetylcholinesterase activity based on the peroxidase-like activity of Fe–N–C nanozyme

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/H2O2/3,3′,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 H2O2 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−1 (colorimetric) and 2.2 mU mL−1 (photothermal), while for paraoxon-ethyl is 0.012 μg mL−1 (colorimetric) and 0.013 μg mL−1 (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 H2O2.

Synthesis and Biological Activity of Functionally Substituted Pyrimidine and Pyran Derivatives on the Basis of Isatylidene Malononitriles

AbstractIn this study, it was shown that pyrimidine and imidazopyridine derivatives were obtained from the multicomponent reaction of isatylidene malononitriles with malononitrile and diamines at room temperature. Also, the interaction of 2‐(5‐bromo‐2‐oxoindoline‐3‐ylidene)with malononitrile and benzoyl acetone (or ethyl‐4‐chlorine‐acetoacetate) was carried out and the formation of the corresponding pyran derivatives was observed.The structure of the synthesized compounds was confirmed by 1H, 13CNMR spectroscopy and X‐ray analysis. The acetylcholinesterase (AChE) inhibitor compounds recorded as important therapeutic drugs for the therapy of Alzheimer's disease. Also, novel complexes effectively inhibited AChE enzyme, with Ki values in the range of 4.56 to 8.21 μM. For this enzyme, it was obtained with IC50 values in the range of 4.04 to 9.85 μM. For α‐glycosidase enzyme the most effective Ki values were for 4 a and 10 compounds with Ki values of 31.48 and 32.63 μM, respectively. The molecular docking study was employedto investigate of chemical activities and interaction of synthesized compounds with low Ki in the presence of AChE, butyrylcholinesterase, and α‐Glycosidase. The results revealed that some of the compounds, like compound 9 have a good binding affinity to AChE with a docking score of −6.243 (kcal/mol). This compound can affect the enzyme activity by attachment to essential residues of the catalytic domain of the enzyme. The ADME/T analysis was also performed and revealed that compounds have the potential to be utilized as medicine.

Bioactivity of hamamelitannin, flavokawain A, and triacetyl resveratrol as natural compounds: Molecular docking study, anticolon cancer, and anti-Alzheimer potentials.

In this study, we worked on anticolon cancer effects and anti-Alzheimer's disease with molecular docking studies. Hamamelitannin, flavokawain A, and triacetyl resveratrol compounds showed good inhibitory activities on acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) enzymes. The inhibition effects of flavokawain A, hamamelitannin, and triacetyl resveratrol on AChE and BuChE enzymes were determined spectrophotometrically conforming to Ellman. IC50 values of these enzymes were ranging between 0.95±0.12 and 93.27±8.14nM for AChE and 5.71±0.77 and 52.10±8.41nM for BuChE. The inhibitory activities of some chemical compounds such as flavokawain A, hamamelitannin, and triacetyl resveratrol were assessed by performing the molecular docking study in the presence of AChE and BuChE. Also, the features of the ligand-enzyme complex had value of -7.722kcal/mol for flavokawain A against AChE and -5.530kcal/mol against BuChE. The molecular docking calculations indicated the probable interactions and their characteristics at an atomic level. Due to the outcomes gained from docking, the affinity of the chemical compounds to the enzymes was considerable. In vitro cell viabilities of flavokawain A, hamamelitannin, and triacetyl resveratrol with various concentrations on SW620, DLD-1, HT29, HCT8, and HCT116 were investigated by MTT assay with Doxorubicin as the control compound.

MnO2 Nanosheet-Based colorimetric sensor Array: Toward identification of organophosphorus pesticides

In recent years, two-dimensional (2D) nanomaterials have attracted more and more attention because of their high specific surface area and excellent physical and chemical properties. We herein demonstrate two kinds of 2D nanomaterials, i.e., MnO2 nanosheets and UFO-shaped Au@MnO2 nanosheets (denoted as MnO2 NSs and AM NSs), for qualitative identification and quantitative detection of organophosphorus pesticides (OPs). In the presence of acetylcholinesterase (AChE), acetylthiocholine iodide (ATCh) can be hydrolyzed to thiocholine (TCh), which is used as a reducing agent to etch ultrathin MnO2 nanosheets. During the etching process, the color of AM NSs solution changes from green to pink, and the MnO2 NSs solution changes from dark brown to light brown. OPs can inhibit the activity of AChE and thus inhibit the etching of MnO2. Based on this principle, five OPs (i.e., ethion, bromophos-ethyl, chlorpyrifos-methyl, trichlorfon, and omethoate) at 0.5 μg·L−1 were successfully identified by this sensor array. Besides, the distinction for diverse concentrations of each OP and different ratios of OP mixtures was demonstrated. The sensor array also reveals excellent anti-interference ability toward OPs in the presence of different potentially interfering substances. The feasibilities of the sensor array were further validated by differentiating the OPs in grape samples.

A sensitive and selective fluorescent probe for acetylcholinesterase: Synthesis, performance, mechanism and application

The detection of acetylcholinesterase (AChE) activity is of great significance for studying the physiological functions of AChE and clinical diagnosis of pesticide poisoning. Herein, a small-molecule fluorescent probe BDFA was rationally designed and readily synthesized via a one-step reaction, which enables qualitative and quantitative detection of AChE. BDFA emits a slight fluorescence in an aqueous medium, while the fluorescence is significantly enhanced under the catalysis of AChE. Mechanism studies reveal that BDFA eliminates the N, N-dimethyl carbamate protective group in the presence of AChE and then spontaneously undergoes intramolecular cyclization conversion to generate an intense fluorescent product. Based on the above mechanism, BDFA exhibits a sensitive, selective, rapid and stable “turn-on” fluorescence response to AChE, without interference from pH, ions, thiols, amino acids and other enzymes. The fluorescence intensity of BDFA at 525 nm has a linear relationship with the AChE concentration in the range of 0.0045–1.0 U/mL, and the detection limit is 4.5 mU/mL. Moreover, BDFA is suitable for rapidly diagnosing AChE activity in blood samples, thus providing an efficient and convenient tool for diagnosing organophosphorus and carbamate pesticide poisoning. Compared with the reported AChE fluorescent probes, BDFA exhibits apparent advantages including simple synthesis, low detection limit and fast response speed.

Graphitic-phase C3N4 nanosheets combined with MnO2 nanosheets for sensitive fluorescence quenching detection of organophosphorus pesticides

In this study, we have developed a sensitive approach to measure organophosphorus pesticides (OPs) using graphitic-phase C3N4 nanosheets (g-C3N4) combined with a nanomaterial-based quencher, MnO2 nanosheets (MnO2 NS). Since MnO2 NS can quench the fluorescence of g-C3N4 via the inner-filter effect (IFE), enzymatic hydrolysate (thiocholine, TCh) can efficiently trigger the decomposition of MnO2 nanosheets in the presence of acetylcholinesterase (AChE) and acetylthiocholine (ATCh), resulting in the fluorescence recovery of g-C3N4. OPs, as inhibitors to AChE activity, can prevent the generation of TCh and decomposition of MnO2 nanosheets while exhibiting fluorescence quenching. Therefore, the AChE-ATCh-MnO2-g-C3N4 system can be utilized to quantitatively detect OPs based on g-C3N4 fluorescence. Under optimal conditions, the linear ranges for the determination of parathion-methyl (PM) and 2,2-dichlorovinyl dimethyl phosphate (DDVP) were found to be 0.1–2.1 ng/mL and 0.5–16 ng/mL, respectively, with limits of detection of 0.069 ng/mL and 0.20 ng/mL, respectively. The advantages of this assay are user-friendliness, ease of use, and cost effectiveness compared to other more sophisticated analytical instruments.

Novel inhibitors of AChE and Aβ aggregation with neuroprotective properties as lead compounds for the treatment of Alzheimer's disease

A series of sulfone analogs of donepezil were designed and synthesized as novel acetylcholinesterase (AChE) inhibitors with the potent inhibiting Aβ aggregation and providing neuroprotective effects as potential modalities for Alzheimer's disease (AD). Most of the target compounds displayed effective inhibition of AChE, especially compound 24r which displayed powerful inhibitory activity (IC50 = 2.4 nM). Kinetic and docking studies indicated that compound 24r was a mixed-type inhibitor. Furthermore, in glyceraldehyde (GA)-exposed SH-SY5Y differentiated neuronal cells, compound 24r could potently inhibit AChE, reduce tau phosphorylation at S396 residue, provide neuroprotection by rescuing neuronal morphology and increasing cell viability. It was also found to reduce amyloid aggregation in the presence of AChE. In addition, compound 24r showed evident protections from mitochondrial membrane dysfunction and oxidative stress in okadaic acid-induced pharmacological models. Moreover, compound 24r exhibited more effective treatment prospects in vivo than donepezil, including a moderate blood-brain barrier permeability, a more potent AChE inhibitory activity and behavioral improvement in scopolamine-induced cognition-impaired mice model at a much lower dose. Collectively, compound 24r is a promising lead compound for further investigation to discovery and development of new anti-AD agents.

Lipid Membrane State Change by Catalytic Protonation and the Implications for Synaptic Transmission.

In cholinergic synapses, the neurotransmitter acetylcholine (ACh) is rapidly hydrolyzed by esterases to choline and acetic acid (AH). It is believed that this reaction serves the purpose of deactivating ACh once it has exerted its effect on a receptor protein (AChR). The protons liberated in this reaction, however, may by themselves excite the postsynaptic membrane. Herein, we investigated the response of cell membrane models made from phosphatidylcholine (PC), phosphatidylserine (PS) and phosphatidic acid (PA) to ACh in the presence and absence of acetylcholinesterase (AChE). Without a catalyst, there were no significant effects of ACh on the membrane state (lateral pressure change mN/m). In contrast, strong responses were observed in membranes made from PS and PA when ACh was applied in presence of AChE (>5 mN/m). Control experiments demonstrated that this effect was due to the protonation of lipid headgroups, which is maximal at the pK (for PS: ; for PA: ). These findings are physiologically relevant, because both of these lipids are present in postsynaptic membranes. Furthermore, we discussed evidence which suggests that AChR assembles a lipid-protein interface that is proton-sensitive in the vicinity of pH . Such a membrane could be excited by hydrolysis of micromolar amounts of ACh. Based on these results, we proposed that cholinergic transmission is due to postsynaptic membrane protonation. Our model will be falsified if cholinergic membranes do not respond to acidification.

Sensitivity Dependence on the Crystal Forms of a Fluorescence Quencher for Silicon Quantum Dots and Its Use in Acetylcholinesterase Assay.

Acetylcholinesterase (AChE) plays crucial roles in the nervous system, and thus the reliable assay of its activity is of great significance for the diagnosis of nervous diseases. In this work, we report a fluorescent sensing platform with silicon quantum dots (Si-QDs) as a fluorescence oscillator and nano iron oxyhydroxide (α-, β-, and γ-FeOOH) as a quencher for the assay of AChE. FeOOH with α-, β-, and γ-crystal forms quenches the fluorescence of Si-QDs at λex/λem = 350/438 nm, which is retrieved in the presence of AChE and its substrate acetylthiocholine (ATCh) to provide an off-on strategy with a high signal/noise ratio. It is interesting that the sensitivity of AChE sensing is closely related to the crystal forms of FeOOH, with the highest sensitivity by adopting α-FeOOH as the quencher. A linear calibration is achieved within 0.02-1.4 U/L along with a limit of detection of 0.016 U/L. The sensing strategy was demonstrated by the AChE assay in human blood, plasma, and hemocytes.

Simple and Label‐Free Liquid‐Crystal‐Based Detection of Acetylcholinesterase through Interactions between Liquid Crystals and Oil‐in‐Water Emulsion Droplets

AbstractHere, we report a liquid crystal (LC)‐based sensing method for the detection of acetylcholinesterase (AChE) and its inhibitor. The LC and glyceryl trioleate (GT)‐in‐water emulsion droplets were decorated with myristoylcholine chloride (Myr). The decoration helps to prevent the coalescence between the LC and GT emulsion droplets and therefore keeps the LCs in the nematic state, resulting in a bright optical image. In the presence of AChE, Myr was hydrolyzed, causing coalescence of LC and GT emulsion droplets, inducing the phase transition of LCs from the nematic to the isotropic state, resulting in a dark optical image. The biosensor exhibited a limit of detection (LOD) of 100 ng/mL. Moreover, the biosensor was highly specific for AChE and showed a high sensitivity for the detection of AChE inhibitor, with a LOD of 10 μg/mL. The biosensor is a simple and convenient sensing platform for the detection of AChE and its inhibitors.

Enhanced oxidase-like activity of g-C3N4 nanosheets supported Pd nanosheets for ratiometric fluorescence detection of acetylcholinesterase activity and its inhibitor

The undesirable enzymatic activity of nanozymes under near neutral pH condition and the traditional single signal output always restrict the analytical application of nanozyme-based biosensors. Herein, graphitic carbon nitride nanosheets supported palladium nanosheets composite (Pd/g-C3N4) with both oxidase-like activity and fluorescent property is synthesized. Notably, Pd/g-C3N4 exhibits enhanced oxidase-like activity compared to Pd NSs under pH 7.4. By combining Pd/g-C3N4 with o-phenylenediamine (OPD), a ratiometric fluorescence assay for acetylcholinesterase (AChE) activity detection is developed. Pd/g-C3N4 can catalyze oxidation of nonfluorescent OPD to fluorescent oxidized OPD (oxOPD, Em = 565 nm), which can quench fluorescence of g-C3N4 supporter (Em = 441 nm) through fluorescence resonance energy transfer (FRET). However, in presence of AChE, acetylthiocholine can be hydrolyzed into thiocholine, which will block the oxidase-like activity of Pd/g-C3N4 and then hamper the FRET process. This ratiometric fluorescence assay is also viable to screen AChE inhibitor. This work will guide design of ratiometric fluorescence assay based on nanozymes with improved enzymatic activity.

Colorimetric detection of acetylcholinesterase and its inhibitor based on thiol-regulated oxidase-like activity of 2D palladium square nanoplates on reduced graphene oxide.

Aconvenient and sensitive colorimetricassay for acetylcholinesterase (AChE) and its inhibitor has been designedbased on the oxidase-like activity of {100}-faceted Pd square nanoplates which are grown in situ on reduced graphene oxide (PdSP@rGO). PdSP@rGO can effectively catalyze the oxidation of colorless 3,3',5,5'-tetramethylbenzidine (TMB) without the assistance of H2O2 to generate blue oxidized TMB (oxTMB) with a characteristic absorption peak at 652nm. In the presence of AChE, acetylthiocholine (ATCh), a typical AChE substrate, is hydrolyzed to thiocholine (TCh). The generated TCh can effectively inhibit the PdSP@rGO-triggered chromogenic reaction of TMB via cheating with Pd, resulting in color fading and decrease in absorbance. Thus, a sensitive probe for AChE activity is constructed with a working range of 0.25-5mUmL-1 and a limit of detection (LOD) of 0.0625mUmL-1. Furthermore, because of the inhibition effect of tacrine on AChE, tacrine is also detected through the colorimetric AChE assay system within the concentrations range 0.025-0.4μM with a LOD of 0.00229μM. Hence, a rapid and facile colorimetric procedure to sensitively detect AChE and its inhibitor can be anticipated through modulating the oxidase-like activity of PdSP@rGO. Colorimetric method for detection of AChE and its inhibitor is established by modulating the oxidase mimetic activity of {100}-faceted Pd square nanoplates on reduced graphene oxide (PdSP@rGO).