Acetylcholinesterase Enzyme Research Articles

Exploring the Antioxidant and Neuroprotective Potential of Muscari armeniacum: Phenolic Profiling and Enzyme Inhibition

This study aimed to investigate the antioxidant and enzyme inhibitory properties of Muscari armeniacum, a plant native to Türkiye, which is known for its bioactive compound content. Ethanol extracts of Muscari armeniacum were prepared and analyzed using a variety of bioanalytical methods to assess its potential health-promoting properties. The antioxidant capacity was evaluated through Fe³⁺-Fe²⁺ reduction capacity, CUPRAC, DPPH, and ABTS radical scavenging activities. Additionally, the total phenolic and flavonoid contents of the extracts were determined, as these compounds are critical indicators of antioxidant potential. The antioxidant activity of Muscari armeniacum was compared with five standard antioxidants commonly used for benchmarking: BHA, α-tocopherol, BHT, trolox, and ascorbic acid. The extract demonstrated a high flavonoid content (23.24 µg QE/mg extract) and a significant amount of phenolics (14.38 µg GAE/mg extract), with the flavonoid content being particularly noteworthy. In addition to its antioxidant properties, the enzymatic activity assays revealed that the extract significantly inhibited acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes, which are associated with neurodegenerative diseases such as Alzheimer’s. The IC₅₀ values were determined to be 54.14 mg/mL (R² = 0.9695) for AChE and 58.73 mg/mL (R² = 0.9609) for BChE, indicating moderate enzyme inhibition. These results suggest that Muscari armeniacum could serve as a valuable source of bioactive compounds with significant antioxidant and enzyme inhibitory activities. Its potential applications in the treatment of oxidative stress-related diseases and neurodegenerative disorders make it a promising candidate for further pharmacological studies. This research underscores the importance of exploring native plants for their medicinal and therapeutic potential, contributing to the development of alternative treatments for challenging health conditions.

Investigation of the Effects of Light, Darkness, and Dim Light on Rat Brain Tissue: A Biochemical and Histological Study.

This study evaluates acetylcholinesterase (AChE) enzyme activity levels, oxidative stress parameters, histopathological findings, and serum melatonin levels in rat brain tissue. 32 male Wistar Albino rats were randomly divided into four groups: Control, Light, Dark, Dim light (n = 8 each group). After a 30 day experiment, brain tissues were collected to measure AChE, glutathione S-transferase (GST), glutathione (GSH), and malondialdehyde (MDA) levels and conduct histopathological analyses. Serum melatonin levels were also measured. In this study, we observed a significant increase in MDA levels in dim light, dark, and light groups. AChE and α-GST enzyme activity levels were significantly decreased in the dark group compared with the other groups. Additionally, there was a statistical difference in melatonin levels between the light and dark groups. In the light microscope examination of the sections stained with hematoxylin-eosin from the dark group brain tissue, mild perineuronal edema was observed in all areas. Our study is the first to compare the effects of three groups on the brain: continuous light, continuous darkness, and dim light at night. Additionally, it is the only study to examine the effects of light exposure differences on the brain AChE levels.

Mitochondrial Gene Expression of Three Different Dragonflies Under the Stress of Chlorpyrifos

Chlorpyrifos (CPF) is an organophosphate insecticide that is extensively utilized globally due to its effectiveness against over 200 pest species. CPF exhibits its toxicity primarily through the inhibition of the acetylcholinesterase (AChE) enzyme, while mitochondrial damage and dysfunction have also been observed. The present study quantified the transcript levels of mitochondria protein-coding genes (mtPCGs) using quantitative real-time polymerase chain reaction (RT-qPCR) in samples of larvae of three dragonfly species (A. parthenope, E. elegans, and G. confluens) under different levels of CPF stress. By exposing larvae from uncontaminated populations to 0.05 μg/L CPF for 24 h, the transcript levels of seven mtPCGs in A. parthenope were significantly increased (p < 0.05) by 1.89 ± 0.42-fold for COI, 4.30 ± 0.24-fold for COIII, 5.94 ± 0.17-fold for ND1, 4.69 ± 0.56-fold for ND2, 3.44 ± 0.48-fold for ND4, 2.19 ± 0.53-fold for ND4L, and 5.05 ± 0.36-fold for Cytb, respectively. In E. elegans, the transcript levels of ND1, ND2, and ND4 increased by 1.23 ± 0.15, 1.48 ± 0.31, and 1.98 ± 0.25-fold, respectively (p < 0.05). In G. confluens, the transcript levels of COI, COIII, and ND4 increased by 1.56 ± 0.13, 1.50 ± 0.26, and 3.74 ± 0.40-fold, respectively (p < 0.01). It was demonstrated that the transcript levels of different mtPCGs showed significant up-regulation in the three different dragonfly larvae under CPF stress in the absence of mortality. ND4 was significantly increased in all three species, indicating that it is an important target gene. The present study underscores the response of mitochondrial gene expression in larvae of three different species in response to CPF pollutants, indicating that pesticide influences can potentially alter mitochondrial gene expression and potentially act as a method for assessing aquatic ecosystem health.

Resveratrol-Based Carbamates as Selective Butyrylcholinesterase Inhibitors: Design, Synthesis, Computational Study and Biometal Complexation Capability

Considering our previous experience in the design of new cholinesterase inhibitors, especially resveratrol analogs, in this research, the basic stilbene skeleton was used as a structural unit for new carbamates designed as potentially highly selective butyrylcholinesterase (BChE) inhibitors with excellent absorption, distribution, metabolism, excretion and toxicity ADMET properties. The inhibitory activity of newly prepared carbamates 1–13 was tested toward the enzymes acetylcholinesterase (AChE) and BChE. In the tested group of compounds, the leading inhibitors were 1 and 7, which achieved excellent selective inhibitory activity for BChE with IC50 values of 0.12 ± 0.09 μM and 0.38 ± 0.01 μM, respectively. Both were much more active than the standard inhibitor galantamine against BChE. Molecular docking of the most promising inhibitor candidates, compounds 1 and 7, revealed that stabilizing interactions between the active site residues of BChE and the ligands involve π-stacking, alkyl-π interactions, and, when the carbamate orientation allows, H-bond formation. MD analysis confirmed the stability of the obtained complexes. Some bioactive resveratrol-based carbamates displayed complex-forming capabilities with Fe3+ ions as metal centers. Spectrophotometric investigation indicated that they coordinate one or two metal ions, which is in accordance with their chemical structure, offering two binding sites: an amine and a carboxylic group in the carbamate moiety. Based on the obtained in silico, experimental and computational results on biological activity in the present work, new carbamates 1 and 7 represent potential selective BChE inhibitors as new therapeutics for neurological disorders.

Production of neuroprotective compounds from barley (Hordeum vulgare L.) germinated under different conditions during the malting process

Abstract The effect of different barley germination conditions on generation of neuroprotective compounds was studied. Malts were obtained by germination at 20 °C for 35 hr (M1), 20 °C for 72 hr (M2), 25 °C for 35 hr (M3), and 25 °C for 72 hr (M4). M4 showed higher phenylalanine ammonia lyase (PAL) activity and total phenolic acid content than the others. Moreover, a direct relationship (r2: 0.9802) between protein hydrolysis degree and malt protease activity was obtained. Regarding neuroprotective properties, all malts inhibited the acetylcholinesterase, tyrosinase, and prolyl oligopeptidase enzymes. However, M4 showed the highest inhibitory activity. This effect was due to the higher proportion of 595 and 392 Da peptides with high hydrophobicity and the higher phenolic acid content in this malt. Thus, the temperature and time of barley germination influenced the enzymatic activity of proteases and PAL, which modified the activity and profile of neuroprotective compounds in malts.

A Computational Study of Phenothiazine Derivatives as Acetylcholinesterase Inhibitors Targeting Alzheimer's Disease.

Alzheimer's disease is a neurodegenerative disorder that affects learning, memory and behavioral turbulence in elderly patients. Acetylcholinesterase (AChE) inhibitors act as anti-Alzheimer's agents. Phenothiazine derivatives are considered momentous anti-Alzheimer's agents because of their AChE inhibitory activity. The elevated levels and increased expression of this protein have been associated with Alzheimer's disease. Coumarin-fused phenothiazines have emerged as significant anti-Alzheimer's agents due to their notable receptor inhibitory activity. Some unique phenothiazine analogs were designed, and computational studies were conducted to explore their inhibitory activity against the AChE enzyme (PDB id: 4EY7) by using the Schrodinger suite-2019-4. Docking studies were conducted by using the Glide module; binding free energies were calculated by means of the Prime MM-GBSA module, and Molecular dynamics (MD) simulation was performed by using the Desmond module of the Schrodinger suite. Glide scores were used to find out the binding affinity of the ligands with the target 4EY7. The compounds exhibited enhanced hydrophobic interactions and formed hydrogen bonds, effectively impeding Acetylcholinesterase. The Glide scores for the compounds ranged from -13.4237 to -8.43439, surpassing the standard (Donepezil) with a score of -16.9898. Interestingly, a positive value was obtained for the MM-GBSA binding of the potent inhibitor. To gain insights into the dynamic behavior of the protein A8, molecular dynamics (MD) simulations were employed. Based on the results, the study concludes that phenothiazine derivatives show promise as acetylcholinesterase inhibitors. Compounds with notable Glide scores are poised to exhibit significant anti-Alzheimer's activity, suggesting their potential therapeutic efficacy. Further in vitro and in vivo investigations are warranted to validate and explore the therapeutic potentials of these compounds.

Bioactive fraction of Musa balbisiana seed mitigates D-galactose-induced brain aging via SIRT1/PGC-1α/FoxO3a activation and intestinal barrier dysfunction by modulating gut microbiota and core metabolites

Aging is an inevitable biological process, and emerging research has highlighted the potential of dietary and pharmacological interventions to decelerate the trajectory of age-related diseases and prolong the health span. This study evaluates the protective effects of Musa balbisiana seed on healthy aging using D-galactose-induced accelerated aging rats. The results suggested that the bioactive ethyl acetate fraction of Musa balbisiana seed extract (BF) exhibited protective effects against aging-induced oxidative stress by reducing oxidative DNA damage, advanced glycation end-product formation, and malondialdehyde levels while restoring antioxidant and glyoxalase enzyme activities. BF also ameliorated neurodegeneration by decreasing acetylcholinesterase enzyme activity and amyloid beta plaque formation. Histopathological analysis demonstrated the protective effects of BF against brain aging, liver disruption, renal damage, and intestinal barrier dysfunction. BF further restored intestinal permeability by upregulating the tight junctions (zonula occludens 1 and 2, claudin 1,2,3 and 4, and occludin) and mucin (mucin 2 and mucin 5ac) gene expression while downregulating the expression of inflammatory cytokines (IL-1β, IL-6, and TNF-α). BF significantly induced the phosphorylation of FoxO3a proteins and upregulated the gene expression of SIRT1, PGC-1α, and TFAM in the hippocampus. Next-generation sequencing (NGS) of 16s rRNA amplicons of fecal metagenomics DNA and metabolites profiling showed that BF intervention restructured the gut microbiota and altered core metabolites related to cholesterol metabolism. Overall, our findings demonstrated the multifaceted protective effects of Musa balbisiana seed against D-galactose-induced aging.

Assessment of Lethal Dose and Lethal Time of Diazinon in Swiss Albino Mice

Approximately two thirds of deaths and millions of nonfatal cases are attributed to organophosphorus pesticides (OPP) annually. Diazinon, a prominent member of the OP pesticides, was introduced for both agriculture and public health usages during. For several decades, the compound was among the top pesticides in use in many countries, including Libya. The toxicity of this compound is based on its ability to inhibiting the acetylcholinesterase enzyme (AchE), which is vitally important for normal nerve impulse transmission. This study evaluated the acute toxicity of diazinon through acute exposure testing on female Swiss albino mice. In the acute toxicity test, 72 mice were subcutaneously injected with doses of diazinon ranging from 320 to 480 μL/ kg. Mortality and symptoms were recorded over four days, with the median lethal dose found to be 403.5 μL/kg. Based on the findings, the present study suggests that stringent controls and surveillance must be placed on diazinon and similar chemicals.

Conjugation of Triterpenic Acids with 3-Aminoquinuclidine Moiety: An Approach to Acetylcholinesterase Mixed or Uncompetitive Type Inhibitors.

Alzheimer's disease (AD) poses a significant public health issue. Despite the fact that today there are several methods of maintenance therapy, one of the most widely used methods is designed to correct the deficiency of acetylcholine. In the search for new potential inhibitors of cholinesterase enzymes, eight new derivatives of 3-oxo- or 2,3-indolo-triterpenic acid conjugated with amino-quinuclidine bicyclic cores were designed and synthesized. Then, the obtained compounds were screened in Ellman's assays for their ability to inhibit acetylcholinesterase enzyme, and for each of the active compounds, the type of inhibition was determined. The obtained results demonstrate the dependence of the activity on the triterpenoid structure and the type of substituents. The best activity for ursolic acid derivatives was observed for the 3-oxoamide 8, with an IC50 value of 0.43 µM, acting as a mixed-type inhibitor. In turn, for the oleanane type, the amide with an indole unit in the A ring 11 exhibited the best activity with an IC50 value of 0.47 µM (while the ursane-type analog was weakly active) and led to an uncompetitive type of inhibition. Thus, 3-amidoquinuclidine-triterpenoids conjugates could be considered novel inhibitors of acetylcholinesterase with a different mechanism of action.

Multi-Target Quinoxaline Derivatives for Alzheimer’s Disease: Inhibitory Activities Against AChE and BACE-1 Enzymes

New choline esterase inhibitors and B-secretase inhibitors present promising treatment options for the treatment of Alzheimer's disease (AD). In this study, molecular docking was performed using our chemistry library to discover lead compounds. Molecular docking was employed to predict binding affinities, while molecular dynamics (MD) simulations provided insights into the stability of the ligand–enzyme interactions. To improve the activity, 12 new derivatives were designed and synthesized based on the lead compound obtained. The structures of the synthesized compounds were identified by 1H-NMR,13C-NMR, and HRMS techniques. Their activities on choline esterase enzymes and Beta-secretase 1 enzyme were elucidated through in vitro studies. Compound 4f had an IC50 = 0.026 ± 0.001 µ. value against the acetylcholinesterase (AChE) enzyme and an IC50 = 0.125 ± 0.005 µ. value against the BACE-1 enzyme. The excellent activity of compound 4f was supported by molecular docking and MD simulation studies.

LC-MS/MS Analysis and Biological Activities of Different Parts of Ziziphora capitate L.

The Ziziphora species, classified under the Lamiaceae family, have a strong aromatic property. Ziziphora species have been used in folk medicine as sedative, gastric, aphrodisiac, bloating, and degassing. In the current study, the phenolic and flavanoid content of ethanol extracts of Ziziphora capitata L. species of flower, leaf, branch, mixed, and root parts was determined by the LC-MS/MS device. In addition, the antioxidant and cytotoxic activities of the extracts, as well as their inhibitory effects on enzymes (antihypertensive, AchE (acetylcholinesterase), BchE (butyrylcholinesterase), elastase, tyrosinase, collagenase and urease), were determined. The LC-MS/MS results showed that quinic acid (25578, 5842, 25171, 14055, 10597 µg g-1, respectively) was found in higher amounts in flower, leaf, branch, mixed, and root extracts of Z. capitata species compared to other components. Additionally, rosmarinic acid (17097 µg g-1), cynaroside (8432), and hesperidin (8067) were found to be major components. It was observed that the flower extract of the species exhibited strong antioxidant activity (IC50: 37.18±1.36 µg mL-1, 9.89±0.45, A0.5:16.27±0.02, respectively) in DPPH, ABTS and CUPRAC methods. It was concluded that the leaf extract of Z. capitata species had a strong cytotoxic effect on HT-29 (colon cancer cell line) (viability %: 9.26±0.69). It was observed that the root part of the species exhibited higher activity in butyrylcholinesterase (BChE) enzyme inhibition activity (inhibition %: 40.56±0.88) than other parts. It was determined that Z. capitata extracts did not show acetylcholinesterase, urease, tyrosinase, elastase, collagenase, and antihypertensive enzyme activity or showed low activity. As a result, it is thought that the flower extract of the Z. capitata species has better results in terms of the examined parameters, whereas the leaf extract needs to be subjected to more detailed in vitro and in vivo research conducted to be used in the pharmaceutical industry as a result of its cytotoxic effect against colon cancer cell lines.

Determination of in vitro Antioxidant, Anticholinergic, and Antiepileptic Activities of some Medicinal and Aromatic Plant Extracts

Medicinal and aromatic plants such as Crocus cancellatus, Scilla Siberica subsp. armena, Juniperus oxycedrus subsp. oxycedrus and Anthriscus nemorosa have many different biological activities. While antioxidants are significant in preventing many diseases, inhibition of metabolic enzymes is also effective in preventing many diseases. In this study, antioxidant activities of water, ethanol, and dichloromethane extracts of four different medicinal and aromatic plant species were determined by 1,1-diphenyl-2-picrylhydrazyl (DPPH•) and 2,20-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS•+) radical scavenging and Cu2+, Fe+3, and Fe3+-2,4,6-tris(2-pyridyl)-S-triazine (TPTZ) reducing assays. Enzyme inhibition studies were performed with metabolic enzymes acetylcholinesterase, butyrylcholinesterase, carbonic anhydrase I and II isoenzymes. The ethanol extract of A. nemorosa showed the highest activity in DPPH and ABTS assays (IC50: 17.36 µg mL-1, IC50: 7.02 µg mL-1). In the Fe3+ reducing assay, the dichloromethane extract of A. nemorosa showed the highest activity (1.96±0.060 µg mL-1). In the Cu2+ reducing assay, the dichloromethane extract of J. oxycedrus showed the highest activity (1.773±0.066 µg mL-1). In the Fe3+-TPTZ reducing assay, the ethanol extract of S. siberica showed the highest activity (1.256±0.011 µg mL-1). In the enzyme inhibition results, it was determined that all plants and all extracts inhibited the enzymes studied. As a result of this study, it was determined that these four medicinal and aromatic plants have high biological activities.

Acetylcholinesterase (AChE) Insensitivity of Aedes aegypti Exposured by Citronella Oil (Cymbopogon nardus)

Aedes mosquitoes are vectors of viruses, namely the dengue virus. Treatment with insecticides is no longer effective if the target mosquito is resistant. Therefore, efforts are needed to overcome the resistance of these mosquitoes. Inhibition of the enzyme acetylcholinesterase (AChE) is one way the synthetic insecticide Profenofos works; if AChE is inhibited, acetylcholine accumulation will disrupt the nervous system. Insensitivity to AChE is one mechanism of mosquito resistance to insecticides. Based on this, the problem is formulated as follows: is the increase in susceptibility of Aedes aegypti mosquitoes to alphacymethrin, citronella oil, and a combination of both caused by a decrease in the insensitivity of mosquito AChE? This study aims to determine the AChE insensitivity of Aedes aegypti mosquitoes to citronella oil, a combination of citronella oil and alphacymethrin 0.03%), compared to the positive control (insecticide alphacymethrin 0.03%). Percentage of mosquito mortality after being held for 24 hours: The results showed no difference between the two groups of mosquitoes without treatment (A) and 100 µL/L citronella oil (B), F count 0.000, Sig. 1.000, significance value <0.05. Exposure to 100 µL/L citronella oil on instar III-IV mosquito larvae did not make mosquitoes more susceptible or more resistant to 0.03% alphacypermethrin insecticide. Based on the absorption value analysis results using the T-test between group A/ without treatment and group B with 100µL/L citronella oil treatment showed a Sig. (2-tailed) value of 0.547>0.05 means there is no significant difference between the two groups.

Biological Implications of the Intrinsic Deformability of Human Acetylcholinesterase Induced by Diverse Compounds: A Computational Study.

The enzyme acetylcholinesterase (AChE) plays a crucial role in the termination of nerve impulses by hydrolyzing the neurotransmitter acetylcholine (ACh). The inhibition of AChE has emerged as a promising therapeutic approach for the management of neurological disorders such as Lewy body dementia and Alzheimer's disease. The potential of various compounds as AChE inhibitors was investigated. In this study, we evaluated the impact of natural compounds of interest on the intrinsic deformability of human AChE using computational biophysical analysis. Our approach incorporates classical dynamics, elastic networks (ENM and NMA), statistical potentials (CUPSAT and SWOTein), energy frustration (Frustratometer), and volumetric cavity analyses (MOLE and PockDrug). The results revealed that cyanidin induced significant changes in the flexibility and rigidity of AChE, especially in the distribution and volume of internal cavities, compared to model inhibitors such as TZ2PA6, and through a distinct biophysical-molecular mechanism from the other inhibitors considered. These findings suggest that cyanidin could offer potential mechanistic pathways for future research and applications in the development of new treatments for neurodegenerative diseases.

The Effects of Lamium garganicum L. Subsp. lasioclades (Stapf.) R. Mill Plant Against Fibroblast (U2OS Cell), Acetylcholinesterase, Glutathione S-Transferase: An In Vitro, In Silico Biological Activity Screening Study.

In this study, some biological activities of extracts of Lamium garganicum subsp. lasioclades (Lgl) have been evaluated as well as identified the phenolic composition. Concentration ranges of 31.25, 62.5, 125, 250, and 500µg/mL were applied to determine the extract's anticancer properties. Significant results were obtained against the osteosarcoma cell line (U2OS cell) compared to normal human umbilical vein endothelial cells (HUVEC). To determine the antioxidant activities, ABTS, DPPH, FRAP, and CUPRAC methods were studied in vitro. Enzyme inhibition effects of methanol extract against the glutathione S-transferase (GST) and acetylcholinesterase (AChE) enzymes were investigated. IC50 values were calculated as 12.96µL/mL for AChE and 13.02µg/mL for GST, respectively. The phenolic contents of the plant extract were analyzed by HPLC. The interaction mechanisms of protein-ligand complexes formed by AChE and GST receptors with gallic acid and rutin were investigated by molecular docking studies. The stability of the complexes formed between receptors and ligands was confirmed by root mean square deviation (RMSD), root mean square fluctuation (RMSF), number of average hydrogen bonding interactions (Hb), and radius of gyration (Rg) analyses obtained from 100ns molecular dynamics simulations.

Novel Medical Countermeasures for Nerve Agent and Pharmaceutical Based Agent Poisoning

Nerve agents are organophosphorus compounds which inhibit acetylcholinesterase (AChE) enzyme. Existing AChE reactivators (Oximes) have several limitations in mean of reactivation potential, broad action spectrum, and penetration rate through blood-brain barrier. Ongoing studies focus on design and synthesis of novel oximes. Pharmaceutical based agents like fentanyl abuse becomes an important public health threat. Naloxone and naltrexone that are centrally acting opioid receptor antagonists, are used for reversing the effects of the opioid overdose.

Neuroprotective Assessment of Nutraceutical (Betanin) in Neuroblastoma Cell Line SHSY-5Y: An in-Vitro and in-Silico Approach.

The cognitive dysfunction in the brain cause severe pathological consequences such as Alzheimer's disease (AD), Parkinson's disease. The current treatments are cost expensive and also cause negative side effects. Therefore it is inevitable to develop natural phyto-compounds as a drug like molecules to treat neurodegenerative diseases. In this context, we have assayed the neuroprotective effects of betanin, an indole derivative, in the neuroblastoma cell line SHSY-5Y cells. The neuroprotective effect was investigated in the β-amyloid (Aβ) - induced SHSY-5Y cells; betanin (25µg) protected the SHSY-5Y cells from the toxic effects and maintained the cell viability. Moreover, the acridine orange and ethidum Bromide staining decipher that treatment of betanin in the Aβ-induced SHSY-5Y cells maintain the cell viablity sustainably. The Reactive Oxygen Species (ROS) assay infers that betanin quenches the generation of free radicals progressively in the Aβ-induced SHSY-5Y cells. In addition, the autophagy determination by flow cytometry revealed that betanin induces autophagy to remove the neurodegenerated cells. Further, we examined the docking and simulation patterns with the angiotensin-converting enzyme (ACE), TNF-α converting enzyme (TACE), glycogen synthase kinase 3 (GK3), and acetylcholinesterase enzymes (AChE) and amyloid precursor protein (APP). The insilico docking analysis denotes that betanin had a significant docking score with the target molecules. Thus, from the invitro and insilico studies, betanin strongly inhibit the toxic effects of Aβand protect the cells from degeneration.

Haloperidol-Induced Catalepsy and Its Correlations with Acetylcholinesterase Activity in Different Brain Structures of Mice.

Antipsychotic medicines are used to treat several psychological disorders and some symptoms caused by dementia and schizophrenia. Haloperidol (Hal) is a typical antipsychotic usually used to treat psychosis; however, its use causes motor or extrapyramidal symptoms (EPS) such as catalepsy. Hal blocks the function of presynaptic D2 receptors on cholinergic interneurons, leading to the release of acetylcholine (ACh), which is hydrolyzed by the enzyme acetylcholinesterase (AChE). This study was designed to investigate the Hal-inhibitory effects on AChE activity in regions representative of the cholinergic system of mice and potential associations between cataleptic effects generated by Hal using therapeutic doses and their inhibitory effects on AChE. The distribution of the AChE activity in the different regions of the brain followed the order striatum > hippocampus > (prefrontal cortex/hypothalamus/ cerebellum) > brainstem > septo-hippocampal system. In ex vivo assays, Hal inhibited AChE activity obtained from homogenate tissue of the striatum, hippocampus, and septo-hippocampal system in a concentration-dependent manner. The inhibitory concentration of 50% of enzyme activity (IC50) indicated that the septo-hippocampal system required a higher concentration of Hal (IC50 = 202.5 µmol·L-1) to inhibit AChE activity compared to the striatum (IC50 = 162.5 µmol·L-1) and hippocampus (IC50 = 145 µmol·L-1). In in vivo assays, male Swiss mice treated with concentrations of Hal higher than 0.1 mg·kg-1 induced cataleptic effects. Positive correlations with Spearman's correlation were observed only between the lack of cataleptic effect and the decreased AChE activity of the hippocampus in the mice treated with 0.01 mg·kg-1 of Hal but not in the striatum and septo-hippocampal system. Our results suggest that Hal could increase cholinergic effects via AChE inhibition, in addition to its dopamine antagonist effect, as an alternative approach to the treatment of behavioral disturbances associated with dementia.

Enhanced Bioactivity of Quercetin-Tetrahydroisoquinoline Derivatives: Effect on Lipophilicity, Enzymes Inhibition, Antioxidant Potential, and Cytotoxicity.

Quercetin, a well-known flavonoid with significant medicinal potential, was derivatized at the C8 position with a tetrahydroisoquinoline (THIQ) moiety, and physicochemical and pharmacological properties, inhibition potential, antioxidant activity, and cytotoxicity of new compounds were evaluated. Physicochemical and pharmacological properties, including lipophilicity, membrane permeability, and P-glycoprotein substrate affinity, were assessed theoretically using the SwissADME software. The metal-chelating ability of the new compounds was evaluated on metal ions Fe2+, Zn2+, and Cu2+, whose homeostasis disruption is linked to the development of Alzheimer's disease. Inhibition potential was tested on the cholinergic enzymes acetylcholinesterase and butyrylcholinesterase, as well as Na+, K+-ATPase, an enzyme commonly overexpressed in tumours. Antioxidant potential was assessed using the DPPH assay. Cytotoxicity studies were conducted on healthy MRC-5 cells and three cancer cell lines: HeLa, MDA-231, and MDA-468. The results indicated that derivatization of quercetin with THIQ yielded compounds with lower toxicity, preserved chelating ability, improved antioxidant potential, increased selectivity toward the cholinergic enzyme butyrylcholinesterase, and enhanced inhibition potential toward Na+, K+-ATPase and butyrylcholinesterase compared to quercetin alone. Therefore, the synthesized derivatives represent compounds with an improved profile and could be promising candidates for further optimization in developing drugs for neurodegenerative and cancer diseases.

Identification of anticholinesterase active compounds from ethylacetate fraction of hydroalcoholic extract of Itrifal Sana using TLC-bioautography-MS and its validation using in silico molecular approach.

Traditional formulations are used extensively throughout the world due to their holistic approach to health and wellness with the fewest possible adverse effects. Itrifal Sana is a traditional unani polyherbal formulation, a unique combination that makes it synergistically potent, capable of providing dual benefits for health and well-being. Even though the formulation is frequently utilised, there is no scientific evidence to support its therapeutic efficiency. The present study was designed to detect and identify bioactives responsible for acetylcholinesterase inhibitory activity by TLC-bioautography-MS and its validation using an in silico molecular approach. Authentication of the formulation was done using macroscopy and powder microscopy. Quality control was done using UPLC-MS fingerprint analysis. TLC-bioautography-MS was done to detect the bioactives responsible for acetylcholinesterase inhibitory activity and the findings were validated using in-silico approach. The TLC-MS bioautography revealed the presence of rosmarinic acid, kaempferol, and apigenin as potential bioactive anticholinesterase metabolites. UPLC-MS analysis demonstrated the separation of 48 phytocompounds in the best active fraction of formulation. In silico analysis of identified metabolites showed acetylcholinesterase inhibitory activity of ten identified metabolites, moreover, rosmarinic acid and lobeline showed the best potential activity. Our findings indicated that Itrifal Sana, which was investigated for the first time, has an enormous potential for managing AD caused by acetylcholinesterase enzyme inhibition. It was derived through successfully attempted TLC-bioautography-MS and in silico approach; however, further research on their full efficacy using in vitro cell line studies, in vivo studies, pharmacokinetics studies, and toxicity studies is still needed. TLC-bioautography-MS and in silico molecular approach offer much more effective, accurate, and reliable results than the conventional methods in the identification and validation of bioactive components from IS, a polyherbal formulation helps to advance the development of natural product-based therapeutics for cholinesterase dysfunctional diseases.