Rapid screening of acetylcholinesterase active contaminants in water: A solid phase microextraction-based ligand fishing approach.

Effect-directed analysis (EDA) has been increasingly used for screening toxic contaminants in the environment, but conventional EDA procedures are often time-consuming and labor-extensive. This challenges the use of EDA for toxicant identification in the scenarios when quick answers are demanded. Herein, a solid phase microextraction ligand fishing (SPME-LF) strategy has been proposed as a rapid EDA approach for identifying acetylcholinesterase (AChE) active compounds in water. The feasibility of ligand fishing techniques for screening AChE active chemicals from environmental mixtures was first verified by a membrane separation method. Then, SPME fibers were prepared through self-assembly of boronic acid groups with AChE via co-bonding and applied for SPME-LF. As AChE coated SPME fibers selectively enriched AChE-active compounds from water, comparing sorbing compounds by the SPME fibers with and without AChE coating can quickly distinguish AChE toxicants in mixtures. Compared with conventional EDA, SPME-LF does not require repeating sample separations and bioassays, endowing SPME-LF with the merits of low-cost, labor-saving, and user-friendly. It is believed that cost-efficient and easy-to-use SPME-LF strategy can potentially be a rapid EDA method for screening receptor-specific toxicants in aquatic environment, especially applicable in time-sensitive screening.

Nanoplasmonic biosensors for multicolor visual analysis of acetylcholinesterase activity and drug inhibitor screening in point-of-care testing.

The monitoring of acetylcholinesterase (AChE) activity and the screening of its inhibitors are significance of the diagnosis and drug therapy of nervous diseases. A metal ions-mediated signal amplification strategy was developed for the highly sensitive and multicolor assay of AChE activity and visually screening its drug inhibitors. After the specific reaction between AChE and acetylthiocholine (ATCh), the hydrolysis product thiocholine (TCh) can directly and decompose the α-FeOOH nanorods (NRs) to release amounts of Fe2+, which was regarded as Fenton reagent to efficiently catalyze H2O2 to produce ·OH. Then, the as-formed ·OH can further largely shorten the gold nanobipyramids (Au NBPs), generating a series of palpable color variations. The linear range for AChE activity was 0.01-500.0 U/L with the limit of detection as low as 0.0074 U/L. The vivid visual effects could be easily distinguished for the multicolor assay of AChE activity by naked eye in visible light. To achieve the point-of-care testing, Au NBPs were further assembled on polymeric electrospun nanofibrous films (ENFs) surface as test strips for the easy-to-use test of AChE activity by RGB values with a smartphone. Fascinatingly, this proposed strategy can be used for the visual screening AChE inhibitors or non-inhibitors. Comparing with the clinical drugs (rivastigmine tartrate, and donepezil), some natural alkaloids such as evodiamine, caffeine, camptothecin, and berberine hydrochloride were selected as inhibitor modes to confirm the drug screening capability of this method. This proposed strategy may have great potential in the other disease-related enzymatic biomarkers assay and the rapid screening of drug therapy.

In Vitro Antibacterial, Antioxidant, Anticholinesterase, and Antidiabetic Activities and Chemical Composition of Salvia balansae.

CONTEXT: Salvia balansae de Noé (S. balansae) (Lamiaceae) is known to be an important plant used in folk medicine as an herbal remedy in Algeria. OBJECTIVE: The purpose of the present study was to demonstrate the phytochemical composition, antioxidant activities, enzyme inhibitory activities, and antimicrobial activities of S. balansae extracts. MATERIALS AND METHODS: A methanolic extract and a petroleum ether extract from the aerial parts of the plant were assessed for their chemical composition. HPLC-MS and HPLC-DAD assessed the content of phenols, GC-MS the fatty acid composition, and ICP-MS the mineral profiles of the plant. Additionally, we evaluated the bioactivities of S. balansae extracts by the DPPH, ABTS, and CUPRAC assays, including the antioxidant potential against AChE, BChE, α-amylase, and α-glucosidase for enzyme inhibition. The antibacterial and antifungal activities of the methanolic extract were determined by the disc diffusion test against several strains of bacteria and yeasts. RESULTS: Our findings revealed that the aerial parts of S. balansae were rich in phytochemical components and contained large amounts of minerals. Quantitative analysis of phenolic compounds by HPLC-DAD revealed the presence of 12 compounds in three major classes, flavonoids, hydroxycinnamic acid, and phenolic acid derivatives, with 0.61, 0.45, and 0.29 mg/g of extract, respectively. Nine phenolic constituents were quantified by HPLC-MS analysis; catechin (72.5%) was the main compound, followed by myricetin (21.7%). The fatty acid composition of the S. balansae petroleum ether extract by GC-MS analysis was quantified. Seventeen compounds, including palmitic acid, were identified as the major fatty acids. The antioxidant activity of the S. balansae extracts was measured by three different methods: the methanol extract provided better results than the petroleum ether extract, and interesting values were noted for the DPPH, ABTS, and CUPRAC assays of 242.7 ± 7.44, 124.1 ± 9.70, and 222.9 ± 6.05 µg/mL, respectively. The enzyme inhibition activity of the plant could not be determined. The antimicrobial results of the methanolic extract obtained from the disc diffusion method, followed by measurements of MIC, MBC, and MFC against several bacteria and yeasts, indicated that S. balansae exhibited noticeable antimicrobial and antifungal activities. CONCLUSIONS: These results provided new data about the main phenolic compounds and biological activities of extracts of the aerial parts of S. balansae, which might be an alternative source for synthetic bioactive compounds.

Parkia from Cerrado: phytochemical bioprospection, toxicity and in vitro bioactivities of bark and flower extracts.

Parkia platycephala is the only species of the genus Parkia that is endemic to the brazilian Cerrado and the tree symbol of the state of Tocantins, but there are still few studies regarding its bioprospecting. In this study, we aimed to investigate the phytochemical composition, toxicity and bioactivities of the bark and flower of Parkia platycephala. Hot sequential extractions (Soxhlet) were performed using methanol and hydroethanolic solution (70%), after degreasing the sample (hexane). The presence of flavonoids, tannins, steroids and alkaloids was detected in the preliminary screening. Trilinolein, (Z)-9-octadecenamide, 3-O-methyl-d-glucose were detected by Gas Chromatography coupled to Mass Spectrometry (GC-MS). In the Liquid Chromatography with Diode Array Detector (LC-PDA) analysis, it was detected exclusively ferulic acid (bark) and ellagic acid (flower). The ethanolic extract of the bark (IC50=10.69 ± 0.35 µgmL-1) has an antioxidant potential (DPPH• radical) higher than that of the rutin standard (IC50=15.85 ± 0.08 µgmL-1). All extracts showed excellent anticholinesterase potential (Ellman), with emphasis on the ethanol extract of the flower (IC50 =5.34 ± 0.12 µgmL-1). Regarding toxicity (Artemia salina), the methanolic extract of the bark and the ethanolic extract of the flower presented high and moderate levels, respectively. Such results limit the concentrations of biological activities in this study, however, the antioxidant and anticholinesterase indices fall short of toxicity. The results demonstrated promising antioxidant and anticholinesterase activities of both the bark and the flower of Parkia platycephala.

Composition and Anti-Acetylcholinesterase Properties of the Essential Oil of the Ecuadorian Endemic Species Eugenia valvata McVaugh.

Alzheimer's disease is a global health problem due to the scarcity of acetylcholinesterase inhibitors, the basis for symptomatic treatment of this disease; this requires new approaches to drug discovery. In this study, we investigated the chemical composition and anticholinesterase activity of Eugenia valvata McVaugt (Myrtaceae) collected in southern Ecuador, which was obtained as an essential oil (EO) with a yield of 0.124 ± 0.03% (w/w); as a result of the chemical composition analysis, a total of 58 organic compounds were identified-representing 95.91% of the total volatile compounds-using a stationary phase based on 5% phenyl-methylpolysiloxane, as analyzed via gas chromatography coupled to mass spectrometry (GC-MS) and flame ionization detection (GC-FID). The main groups were hydrocarbon sesquiterpenes (37.43%), oxygenated sesquiterpenes (31.08%), hydrocarbon monoterpenes (24.14%), oxygenated monoterpenes (0.20%), and other compounds (3.058%). Samples were characterized by the following compounds: α-pinene (22.70%), α-humulene (17.20%), (E)-caryophyllene (6.02%), citronellyl pentanoate (5.76%), 7-epi-α-eudesmol (4.34%) and 5-iso-cedranol (3.64%); this research was complemented with an enantioselective analysis carried out using 2,3-diethyl-6-tert-butyldimethylsilyl-ß-cyclodextrin as a stationary phase chiral selector. As a result, α-pinene, limonene, and α-cadinene enantiomers were identified; finally, in the search for new active principles, the EO reported strong anticholinesterase activity with an IC50 of 53.08 ± 1.13 µg/mL, making it a promising candidate for future studies of Alzheimer's disease.

Tsaokoic Acid: A New Bicyclic Nonene from the Fruits of Amomum tsao-ko with Acetylcholinesterase Inhibitory Activity.

A new bicyclic nonene, tsaokoic acid (1), was isolated from the fruits of Amomum tsao-ko, together with three known compounds (2-4). The structure of 1 was elucidated by analyzing spectroscopic data including 1D and 2D NMR spectra and compounds 2-4 were identified as tsaokoin, vanillin, and tsaokoarylone, respectively, by comparing their NMR spectra with previously reported data. Compounds 1-4 showed possible inhibitory activity against acetylcholinesterase (AChE) in silico molecular docking simulations. They were submitted to in vitro assay system and exhibited moderate inhibitory activity with IC50 values of 32.78, 41.70, 39.25, and 31.13 µM, respectively.

Efficient Combination of Complex Chromatography, Molecular Docking and Enzyme Kinetics for Exploration of Acetylcholinesterase Inhibitors from Poria cocos.

Poria cocos (P. cocos) is a traditional Chinese medicinal product with the same origin as medicine and food. It has diuretic, anti-inflammatory and liver protection properties, and has been widely used in a Chinese medicine in the treatment of Alzheimer's disease (AD). This study was conducted to explore the activity screening, isolation of acetylcholinesterase inhibitors (AChEIs), and in vitro inhibiting effect of P. cocos. The aim was to develop a new extraction process optimization method based on the Matlab genetic algorithm combined with a traditional orthogonal experiment. Moreover, bio-affinity ultrafiltration combined with molecular docking was used to screen and evaluate the activity of the AChEIs, which were subsequently isolated and purified using high-speed counter-current chromatography (HSCCC) and semi-preparative high-performance liquid chromatography (semi-preparative HPLC). The change in acetylcholinesterase (AChE) activity was tested using an enzymatic reaction kinetics experiment to reflect the inhibitory effect of active compounds on AChE and explore its mechanism of action. Five potential AChEIs were screened via bio-affinity ultrafiltration. Molecular docking results showed that they had good binding affinity for the active site of AChE. Meanwhile, the five active compounds had reversible inhibitory effects on AChE: Polyporenic acid C and Tumulosic acid were non-competitive inhibitors; 3-Epidehydrotumulosic acid was a mixed inhibitor; and Pachymic acid and Dehydrotrametenolic acid were competitive inhibitors. This study provided a basis for the comprehensive utilization of P. cocos and drug development for the treatment of AD.

Integrating Effect-Directed Analysis and Chemically Indicative Mass Spectral Fragmentation to Screen for Toxic Organophosphorus Compounds.

Analytical chemists are often challenged to screen for bioactive compounds in complex matrices, sometimes without a priori knowledge of the exact compound of interest. Therefore, "flagging" techniques, highlighting common characteristics of bioactive compounds, are highly sought after. In this work, we demonstrate a double flagging method, where unknown organophosphorus acetylcholinesterase inhibitors are "flagged" out of a complex matrix by the presence of organophosphorus-indicative ions as well as their acetylcholinesterase inhibition. This is accomplished by flagging the LC chromatographic retention time of phosphorus-indicative ions using accurate mass high-energy in-source CID products, and the retention time of acetylcholinesterase inhibiting compounds using a parallel microfractionation-based bioassay. We successfully apply this method to screen VX, VM, and RVX nerve agents as well as methomyl, a carbamate pesticide, out of soil and whole blood samples at low µM to sub-µM concentrations. This methodology can be easily extended to diverse chemical families and biological activities of interest.

Phytochemical investigation and biological activity of Doronicum pardalianches L. roots against Alzheimer's disease.

The methanol extract of Doronicum pardalianches L. was fractionated using n-hexane, chloroform, and ethyl acetate to evaluate their cholinesterase (ChE) inhibitory activity via modified Ellman's method. It was perceived that only the ethyl acetate fraction was active toward acetylcholinesterase (AChE) with IC50 value of 172.21 µg/mL. Also, all fractions showed no butyrylcholinesterase (BChE) inhibitory activity. The ethyl acetate fraction was also investigated for its neuroprotectivity and metal chelating ability (Zn2+, Fe2+, and Cu2+) which demonstrated desired activity. Phytochemical analysis of the ethyl acetate fraction led to isolation and identification of formononetin 7-O-ß-D-glucopyranoside which has not been previously reported for this plant.

Natural Antioxidants, Tyrosinase and Acetylcholinesterase Inhibitors from Cercis glabra Leaves.

Cercis glabra is a plant belonging to the legume family, whose flowers and barks are commonly used as food and traditional Chinese medicines. However, its leaves are usually disposed of as wastes. This research comprehensively investigated the bioactive constituents of C. glabra leaves, and two new phenolic, ceroffesters A-B (1-2) and thirteen known compounds (3-15) were isolated. Their structures were elucidated by spectroscopic methods such as nuclear magnetic resonance (1D NMR and 2D NMR), high-resolution electrospray ionization mass spectra (HR-ESI-MS), optical rotatory dispersion (ORD) and electronic circular dichroism (ECD). All of them were assessed for their antioxidant activities through ABTS, DPPH and PTIO methodologies, and evaluated for inhibitory activities against two enzymes (mushroom tyrosinase and acetylcholinesterase). As a result, compounds 3-6, 10 and 13 exhibited evident antioxidant activities. Meanwhile, compounds 5, 10 and 13 showed the most potent tyrosinase inhibitory activities, with IC50 of 0.64, 0.65 and 0.59 mM, and compared with the positive control of 0.63 mM (kojic acid). In the initial concentration of 1 mg/mL, compounds 3, 5 and 6 demonstrated moderate inhibitory activities against acetylcholinesterase with 85.27 ± 0.06%, 83.65 ± 0.48% and 82.21 ± 0.09%, respectively, compared with the positive control of 91.17 ± 0.23% (donepezil). These bioactive components could be promising antioxidants, tyrosinase and acetylcholinesterase inhibitors.

Divulging the anti-acetylcholinesterase activity of Colletotrichum lentis strain KU1 extract as sustainable AChE active site inhibitors.

Alzheimer's disease (AD), also called senile dementia is a neurodegenerative disease seen commonly in the elderly and is characterised by the formation of ß-amyloid plaques and neurofibrillary tangles (NFT). Though a complete understanding of the disease is lacking, recent studies showed the role of the enzyme acetylcholinesterase (AChE) in pathogenesis. Finding new lead compounds from natural sources has always been a quest for researchers. Endophytic fungi are a set of microbes that reside within plants without causing any harm. This study focuses on screening endophytes for the production of active acetylcholinesterase inhibitors. Five endophytic fungi were isolated from Catharanthus roseus and screened for AChE inhibitory activity. Three isolates were found to inhibit AChE inhibitory activity and were distinguished based on molecular and microscopic methods. The mycelial extract was taken for the bioassay-guided column chromatography and TLC was performed on the active fraction. The GC-MS and NMR analysis identified the active compounds in the extract as 9-hexadecen-1-ol and erucamide. Molecular docking studies revealed that the compounds are thermodynamically feasible and have significant glide scores. Computational studies revealed that the hydroxyl group of 9-hexadecen-1-ol forms a hydrogen bond with Ser 293 in the active site of AChE, whereas the active site interactions were predominantly hydrophobic in the case of erucamide and are reflected in AChE inhibition assays.

Polyphenols composition and the biological effects of six selected small dark fruits.

The health-promoting activities of fruits are in the limelight in view of the growing risks posed by civilisational diseases and are connected with polyphenols. The present study examined bilberry, blueberry, blackcurrant, redcurrant, cherry and plum for their polyphenolic content and biological activities. The contents of total polyphenolic compounds and their subclasses were determined. Liquid chromatography high-resolution mass spectrometry was used to characterise the polyphenolic profiles. Small dark fruits' antioxidant, anti-inflammatory, and anti-cholinesterase activities were also extensively determined. Significant qualitative and quantitative differences in the analysed fruits' polyphenols composition and biological activities were demonstrated. The highest polyphenolic contents and antioxidant activities were established in blackcurrant fruit, but bilberry also had our attention due to an additional mild influence on antioxidant enzymes. The condensed tannin content in small dark fruits is developed. All tested fruits exhibit anti-inflammatory and anti-cholinesterase activities.

Phospholipids modifications, genotoxic and anticholinesterase effects of pepper fruit (Dennettia tripetala G. Baker) extract in Swiss mice.

The toxicity of D. tripetala fruit extract to mice was investigated using data obtained from lipidomic analyses, comet and Acetylcholinesterase (AChE) assays. Mice (n = 8) were exposed for 30 days via oral gavage to vehicle (5% Tween 80) (negative control), D. tripetala extract (100, 200 and 400 mg/kg) and 40 mg/kg methyl methanesulfonate (MMS) (positive control). The profile of compounds in the fruit extract was analyzed using gas chromatography-mass spectrometry. Out of the total of 32 compounds identified, considerable amount of established insecticidal compounds such as 2-phenylnitroethane, cis-vaccenic acid, linalool and linoleic acid were detected. Fruit extract did not induce DNA damage relative to negative control. Percentage gain in body weights differed significantly across the four weeks. Significantly highest and lowest brain AChE activity was observed in animals exposed to 200 and 400 mg/kg D. tripetala, respectively. Fruit extract modulated the brain phospholipid profile due to significant fold changes of 48 lipid species out of the total of 280 lipid species. High number of differentially expressed phosphatidylcholine (PC) species and significant levels of phosphatidylethanolamine (PE) at 400 mg/kg suggests that activation of inflammation and methylation pathways are the most plausible mechanisms of D. tripetala toxicity to mouse brain tissue.

A green photocatalytic-biosensor for colorimetric detection of pesticide (carbaryl) based on inhibition of acetylcholinesterase.

Carbaryl is a widely-used carbamate pesticide and the detection of its residues in environmental, food and clinical samples is of great importance. In this sturdy, we developed a green photocatalytic-biosensor based on double strand DNA-SYBR green I complex for sensitively colorimetric detection of carbaryl. This green photocatalytic-biosensor can oxidize 3,3',5,5'-tetramethylbenzidine (TMB) into blue ox-TMB. Meanwhile thiocholine is catalytically produced by acetylcholinesterase (AChE) to directly reduce blue ox-TMB into colorless TMB. But the activity of AChE will be suppressed by carbaryl, thus generating less thiocholine and resulting in more ox-TMB for colorimetric analysis. After the careful optimization of sensing conditions (2 µM for DNA concentration, 50 × concentration for SYBR Green I, 10 min for illumination time), the lowest detectable concentration for carbaryl is 0.008 ng/mL with a linear response in the range of 0.01-0.25 ng/mL. In addition, this photocatalytic-biosensor has good selectivity over non-target chemicals (acetamiprid, atrazine, carbendazim, melamine, bisphenol A, estradiol). It also allows detection of pesticides in real samples verified by a standard HPLC method.

High-performance thin-layer chromatography in combination with an acetylcholinesterase-inhibition bioassay with pre-oxidation of organothiophosphates to determine neurotoxic effects in storm, waste, and surface water.

Pesticides such as organothiophosphates (OTPs) are neurotoxically active and enter the aquatic environment. Bioassays, using acetylcholinesterase (AChE), a suitable substrate and reactant, can be applied for the photometric detection of AChE-inhibiton (AChE-I) effects. The oxidized forms of OTPs, so-called oxons, have higher inhibition potentials for AChE. Therefore, a higher sensitivity is achieved for application of oxidized samples to the AChE assay. In this study, the oxidation of malathion, parathion, and chlorpyrifos by n-bromosuccinimide (NBS) was investigated in an approach combining high-performance thin-layer chromatography (HPTLC) with an AChE-I assay. Two AChE application approaches, immersion and spraying, were compared regarding sensitivity, precision, and general feasibility of the OTP effect detection. The oxidation by NBS led to an activation of the OTPs and a strong increase in sensitivity similar to the oxons tested. The sensitivity and precision of the two application techniques were similar, although the spray method was slightly more sensitive to the oxidized OTPs. The 10% inhibition concentrations (IC10) for the spray approach were 0.26, 0.75, and 0.35 ng/spot for activated malathion, parathion, and chlorpyrifos, respectively. AChE-I effect recoveries in samples from a stormwater retention basin and receiving stream were between 69 and 92% for malathion, parathion, and chlorpyrifos. The overall workflow, including sample enrichment by solid-phase extraction, HPTLC, oxidation of OTPs, and AChE-I assay, was demonstrated to be suitable for the detection of AChE-I effects in native water samples. An effect of unknown origin was found in a sample from a stormwater retention basin.

A critical comparison between an ultra-high-performance liquid chromatography triple quadrupole mass spectrometry (UHPLC-QqQ-MS) method and an enzyme assay for anti-cholinesterase pesticide residue detection in cereal matrices.

Analytical method development for the control of pesticide residues occurring in significant dietary foodstuffs is of utmost importance considering their potential impact on consumer health and food market sustainability. Depending on the purpose, either instrumental analysis, mainly chromatographic methods, or screening assays, mostly using biorecognition affinity, are commonly used, featuring different advantages and drawbacks. To practically compare these two different types of analytical strategies, we applied them for the detection of (i) 97 organophosphate (OP) and carbamate (CM) pesticide residues in wheat flour and (ii) carbofuran (a carbamate insecticide) in wheat, rye and maize flour samples. Regarding high-end analysis, an ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC-QqQ-MS) method was developed and validated achieving low limits of quantification (LOQs, from 0.002 to 0.040 mg kg-1) and a short chromatographic run (12 min). In terms of bioanalytical methods, a fast (17 min) and cost-efficient (∼0.01€ per sample) acetylcholinesterase (AChE) microplate assay for carbofuran screening was utilized. Importantly, carbofuran was the strongest of the 11 OP and CM tested pesticides achieving a half maximal inhibitory concentration (IC50) of 0.021 µM whilst the assay detectability was at the parts per billion level in all three cereal matrices. Based on the attained results, a critical discussion is presented providing the analytical merits and bottlenecks for each case and a wider outlook related to the application of analytical methods in the food safety control analytical scheme.

Phytochemicals of Avocado Residues as Potential Acetylcholinesterase Inhibitors, Antioxidants, and Neuroprotective Agents.

Avocado (Persea americana) is a widely consumed fruit and a rich source of nutrients and phytochemicals. Its industrial processing generates peels and seeds which represent 30% of the fruit. Environmental issues related to these wastes are rapidly increasing and likely to double, according to expected avocado production. Therefore, this work aimed to evaluate the potential of hexane and ethanolic peel (PEL-H, PEL-ET) and seed (SED-H, SED-ET) extracts from avocado as sources of neuroprotective compounds. Minerals, total phenol (TPC), total flavonoid (TF), and lipid contents were determined by absorption spectroscopy and gas chromatography. In addition, phytochemicals were putatively identified by paper spray mass spectrometry (PSMS). The extracts were good sources of Ca, Mg, Fe, Zn, ω-6 linoleic acid, and flavonoids. Moreover, fifty-five metabolites were detected in the extracts, consisting mainly of phenolic acids, flavonoids, and alkaloids. The in vitro antioxidant capacity (FRAP and DPPH), acetylcholinesterase inhibition, and in vivo neuroprotective capacity were evaluated. PEL-ET was the best acetylcholinesterase inhibitor, with no significant difference (p > 0.05) compared to the control eserine, and it showed neither preventive nor regenerative effect in the neuroprotection assay. SED-ET demonstrated a significant protective effect compared to the control, suggesting neuroprotection against rotenone-induced neurological damage.

In-vitro antioxidant potential and acetylcholinesterase inhibitory effect of Ficus benghalensis aerial root extract.

Aim and objective: The aim of the current study was to evaluate the antioxidant effect, acetylcholinesterase (AChE) inhibitory effect and phytochemical screening of different extracts of aerial root extract of Ficus benghalensis using in-vitro methods. Methods: The aerial root extract was prepared by successive extraction method using different organic solvents having increasing order of polarity. FB aerial root extract was screened for preliminary phytochemical analysis. FB aerial root extracts were evaluated for in-vitro acetylcholinesterase inhibitory effect by the Ellman's method and anti-oxidant potential by DPPH assay and hydroxyl radical neutralizing assay. Results: Preliminary phytochemical screening of FB extracts indicate the existence of the phytochemicals such as phenols, alkaloids, flavonoids, glycosides, anthraquinones, tannins and steroids. The results of the DPPH assay, hydroxyl radical scavenging assay and AChE inhibitory assay show that chloroform and ethyl acetate extracts are having significant antioxidant activity and acetylcholinesterase inhibitory effect as compared to the other extracts, respectively. Conclusion: The results of the current study suggest that the aerial root extract of FB might be a potential drug source for treatment of neurodegenerative disorders like Alzheimer disease.

Electrochemical Biosensor for Markers of Neurological Esterase Inhibition.

A novel, integrated experimental and modeling framework was applied to an inhibition-based bi-enzyme (IBE) electrochemical biosensor to detect acetylcholinesterase (AChE) inhibitors that may trigger neurological diseases. The biosensor was fabricated by co-immobilizing AChE and tyrosinase (Tyr) on the gold working electrode of a screen-printed electrode (SPE) array. The reaction chemistry included a redox-recycle amplification mechanism to improve the biosensor's current output and sensitivity. A mechanistic mathematical model of the biosensor was used to simulate key diffusion and reaction steps, including diffusion of AChE's reactant (phenylacetate) and inhibitor, the reaction kinetics of the two enzymes, and electrochemical reaction kinetics at the SPE's working electrode. The model was validated by showing that it could reproduce a steady-state biosensor current as a function of the inhibitor (PMSF) concentration and unsteady-state dynamics of the biosensor current following the addition of a reactant (phenylacetate) and inhibitor phenylmethylsulfonylfluoride). The model's utility for characterizing and optimizing biosensor performance was then demonstrated. It was used to calculate the sensitivity of the biosensor's current output and the redox-recycle amplification factor as a function of experimental variables. It was used to calculate dimensionless Damkohler numbers and current-control coefficients that indicated the degree to which individual diffusion and reaction steps limited the biosensor's output current. Finally, the model's utility in designing IBE biosensors and operating conditions that achieve specific performance criteria was discussed.

High-throughput enzyme inhibition screening of 44 Iranian medicinal plants via piezoelectric spraying of planar cholinesterase assays.

A rapid and straightforward approach was developed for screening the acetyl- and butyrylcholinesterase (ChE) inhibitory activity of 44 Iranian medicinal plant extracts at laboratory scale. After a fast ChE inhibitory pre-testing of samples applied as band pattern, 40 out of the 44 Iranian medicinal plant extracts were selected. These were adjusted in the application volume depending on their inhibition activity, applied on both plate sides and simultaneously developed in a horizontal developing chamber. Different mobile phases were studied to achieve maximum separation of ChE inhibitors and minimum co-elution with matrix. Contrary to immersion, the piezoelectric spraying reduced the consumption of assay solutions, prevented zone tailing, zone shift and cross-contamination, and homogeneously covered the entire plate surface with the assay solutions. The ChE inhibitors of the six most bioactive plant extracts were tentatively assigned by high-resolution mass spectrometry in combination with the spectral and chromatographic information obtained.