Toxicity Evaluation of Anti-cholinesterasic Amphibian Extracts by MTT and an Optimized Artemia salina Test.

The amphibian skin is an important source of bioactive compounds. Recently, our workgroup reported the bioactivity of new extracts from the Hylidae, Microhylidae and Leptodactylidae families against several pathways involved in Alzheimer's disease. However, since cytotoxicity can be a limiting factor for their applicability, we evaluated the toxicity of nine amphibian skin extracts with reported anticholinesterase activity, using the traditional MTT assay and an optimized Artemia salina test. The proposed improvement, guided by experimental design, aims to reduce the amount of biological sample needed. Overall, we proved that the active extracts were non-toxic at effective concentration against cholinesterases (AChE/BChE), positioning the amphibian skin as a promising and preliminary safe source of bioactive compounds in the anti-Alzheimer's treatment. Interestingly, we demonstrated that both toxicity assays can discriminate between toxic and non-toxic samples. We propose the A. salina bioassay as a reliable and cost-effective alternative for early toxicity screening.

B-esterase measurements and other blood related biomarkers in loggerhead sea turtles (Caretta caretta) as indicators of health status.

The loggerhead sea turtle (Caretta caretta) has been selected as sentinel species by the Marine Strategy Framework Directive (MSFD) descriptor 10 in relation to marine litter. In this, and other protected species, there is a need to develop conservative pollution biomarkers equally informative of chemical exposures to those traditionally carried out in metabolic organs, such as the liver. With this aim, plasma from turtles undergoing rehabilitation at the Fundació Oceanogràfic rescue centre (Arca del Mar) were selected and tested for B-esterase measurements. Hydrolysis rates of acetylcholinesterase (AChE), butyrylcholinesterase (BChE) and carboxylesterases (CEs) using four commercial substrates were undertaken on 191 plasma samples. Results indicated that acetylthiocholine was the most adequate substrate of cholinesterases and butyrate esters for CE measures. The correlation of these parameters with well-established blood biochemistry measurements was analysed. B-esterase measures in wild specimens were discussed in relation to age group, pathology on admission to the rescue centre and season; moreover, contrasts with long-term resident turtles were also made. Although this study provides baseline data on B-esterase measures in a large sample size for this species, more complementary information is still needed in terms of population genetics, chemical exposures, and in relation to other biochemical parameters before they can be confidently applied in wild specimens within the regulatory MSFD.

In-Vitro, In-Vivo, Molecular Docking and ADMET Studies of 2-Substituted 3,7-Dihydroxy-4H-chromen-4-one for Oxidative Stress, Inflammation and Alzheimer's Disease.

Plants' bioactives are well-known safe drugs for vital diseases. Flavones and Flavonoid-rich dietary supplements are known to exhibit neuroprotective potential. In this study, we isolated a flavone 2-(3,4-dimethoxyphenyl)-3,7-dihydroxy-4H-chromen-4-one from Notholirion thomsonianum and it was evaluated against various targets of the oxidative stress-related neurological disorders. The compound showed excellent acetyl and butyrylcholinesterase inhibitions in its profile, giving IC50 values of 1.37 and 0.95 µM, respectively. Similarly, in in-vitro MAO-B assay, our flavone exhibited an IC50 value of 0.14 µM in comparison to the standard safinamide (IC50 0.025 µM). In in-vitro anti-inflammatory assay, our isolated compound exhibited IC50 values of 7.09, 0.38 and 0.84 µM against COX-1, COX-2 and 5-LOX, respectively. The COX-2 selectivity (SI) of the compound was 18.70. The compound was found safe in animals and was very effective in carrageenan-induced inflammation. Due to the polar groups in the structure, a very excellent antioxidant profile was observed in both in-vitro and in-vivo models. The compound was docked into the target proteins of the respective activities and the binding energies confirmed the potency of our compound. Furthermore, absorption, distribution, metabolism, excretion, and toxicity (ADMET) results showed that the isolated flavone has a good GIT absorption ability and comes with no hepatic and cardiotoxicity. In addition, the skin sensitization test, in-vitro human cell line activation test (h-CLAT) and KeratinoSens have revealed that isolated flavone is not skin sensitive with a confidence score of 59.6% and 91.6%. Herein, we have isolated a natural flavone with an effective profile against Alzheimer's, inflammation and oxidative stress. The exploration of this natural flavone will provide a baseline for future research in the field of drug development.

Synthesis, in Vitro Cholinesterase Inhibition, Molecular Docking, DFT, and ADME Studies of Novel 1,3,4-Oxadiazole-2-Thiol Derivatives.

A series of 1,3,4-oxadiazole-2-thiol derivatives bearing various alkyl or aryl moieties were designed, synthesized, and characterized using modern spectroscopic methods to yield 17 compounds (6a-6q) that were screened for acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzymes in the search for 'lead' compounds for Alzheimer's disease treatment (AD). The compounds 6q, 6p, 6k, 6o, and 6l showed inhibitory capability against AChE and BChE, with IC50 values ranging from 11.73±0.49 to 27.36±0.29 µM for AChE and 21.83±0.39 to 39.43±0.44 µM for BChE, inhibiting both enzymes within a limited range. The SAR ascertained that the substitution of the aromatic moiety had a profound effect on the AChE and BChE inhibitory potential as compared to the aliphatic substitutions which were supported by the molecular docking studies. The drug-likeness of the most synthesized compounds was confirmed by in silico ADME investigations. These results were additionally supplemented by the molecular orbital analysis (HOMO-LUMO) and electrostatic potential maps got from DFT calculations. ESP maps expose that on all structures, there are two potential binding sites conquered by the most positive and most negative districts.

Multi-target neuroprotective effects of herbal medicines for Alzheimer's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Alzheimer's disease is the most common form of dementia, but its treatment options remain few and ineffective. To find new therapeutic strategies, natural products have gained interest due to their neuroprotective potential, being able to target different pathological hallmarks associated with this disorder. Several plant species are traditionally used due to their empirical neuroprotective effects and it is worth to explore their mechanism of action. AIM OF THE STUDY: This study intended to explore the neuroprotective potential of seven traditional medicinal plants, namely Scutellaria baicalensis, Ginkgo biloba, Hypericum perforatum, Curcuma longa, Lavandula angustifolia, Trigonella foenum-graecum and Rosmarinus officinalis. The safety assessment with reference to pesticides residues was also aimed. MATERIALS AND METHODS: Decoctions prepared from these species were chemically characterized by HPLC-DAD and screened for their ability to scavenge four different free radicals (DPPH•, ABTS•+, O2•‒ and •NO) and to inhibit enzymes related to neurodegeneration (cholinesterases and glycogen synthase kinase-3ß). Cell viability through MTT assay was also evaluated in two different brain cell lines, namely non-tumorigenic D3 human brain endothelial cells (hCMEC/D3) and NSC-34 motor neurons. Furthermore, and using GC, 21 pesticides residues were screened. RESULTS: Regarding chemical composition, chromatographic analysis revealed the presence of several flavonoids, phenolic acids, curcuminoids, phenolic diterpenoids, one alkaloid and one naphthodianthrone in the seven decoctions. All extracts were able to scavenge free radicals and were moderate glycogen synthase kinase-3ß inhibitors; however, they displayed weak to moderate acetylcholinesterase and butyrylcholinesterase inhibition. G. biloba and L. angustifolia decoctions were the less cytotoxic to hCMEC/D3 and NSC-34 cell lines. No pesticides residues were detected. CONCLUSIONS: The results extend the knowledge on the potential use of plant extracts to combat multifactorial disorders, giving new insights into therapeutic avenues for Alzheimer's disease.

Design and synthesis of novel tacrine-indole hybrids as potential multitarget-directed ligands for the treatment of Alzheimer's disease.

The authors report on the synthesis and biological evaluation of new compounds whose structure combines tacrine and indole moieties. Tacrine-indole heterodimers were designed to inhibit cholinesterases and ß-amyloid formation, and to cross the blood-brain barrier. The most potent new acetylcholinesterase inhibitors were compounds 3c and 4d (IC50 = 25 and 39 nM, respectively). Compound 3c displayed considerably higher selectivity for acetylcholinesterase relative to human plasma butyrylcholinesterase in comparison to compound 4d (selectivity index: IC50 [butyrylcholinesterase]/IC50 [acetylcholinesterase] = 3 and 0.6, respectively). Furthermore, compound 3c inhibited ß-amyloid-dependent amyloid nucleation in the yeast-based prion nucleation assay and displayed no dsDNA destabilizing interactions with DNA. Compounds 3c and 4d displayed a high probability of crossing the blood-brain barrier. The results support the potential of 3c for future development as a dual-acting therapeutic agent in the prevention and/or treatment of Alzheimer's disease.

Phytochemical Analysis, In Vitro Anticholinesterase, Antioxidant Activity and In Vivo Nootropic Effect of Ferula ammoniacum (Dorema ammoniacum) D. Don. in Scopolamine-Induced Memory Impairment in Mice.

BACKGROUND: Ferula ammoniacum (D. Don) is one of the endemic medicinal plants that is traditionally used to treat a number of diseases. Although the plant has been used to enhance memory, the investigational evidence supporting the nootropic effect was unsubstantial. Hence, the rationale for this study was to assess the potential beneficial effect of F. ammoniacum seed extracts on learning and memory in mice. METHODS: The powdered plant samples (aerial parts) were subjected to extraction ad fractionation. Among the extracts, crude and ethyl acetate extracts were screened for major phytochemicals through HPLC analysis. All the extracts were evaluated for the in vitro anticholinesterase (AChE and BChE) and antioxidant potentials. Among the extracts the active fraction was further assessed for improving learning and memory in mice using behavioural tests like Y-maze and novel object recognition test (NORT) using standard protocols. After behavioural tests, all the animals were sacrificed and brains tissues were assessed for the ex vivo anticholinesterase and antioxidant potentials. RESULTS: Phytochemicals like chlorogenic acid, quercetin, mandelic acid, phloroglucinol, hydroxy benzoic acid, malic acid, epigallocatechin gallate, ellagic acid, rutin, and pyrogallol were identified in crude methanolic extract (Fa.Met) and ethyl acetate fraction (Fa.EtAc) through HPLC. Fa.EtAc and Fa.Chf extracts more potently inhibited AChE and BChE with IC50 values of 40 and 43 µg/mL, and 41 and 42 µg/mL, respectively. Similarly highest free radical scavenging potential was exhibited by Fa.EtAc fraction against DPPH (IC50 = 100 µg/mL) and ABTS (IC50 = 120 µg/mL). The extract doses, 100 and 200 mg/kg body weight significantly (p < 0.01) improved the short-term memory by increasing the percent spontaneous alternation in the Y-maze test along with increasing discrimination index in the NORT that clearly indicated the enhancement in the recognition memory of mice. CONCLUSION: The extracts more potently scavenged the tested free radicals, exhibited anticholinesterase activities, improved the learning abilities and reduced the memory impairment induced by scopolamine in mice model thus suggesting that these extracts could be effectively used for the management of oxidative stress, neurodegenerative diseases and memory loss.

Interactions between Ginkgo biloba L. and Scutellaria baicalensis Georgi in multicomponent mixtures towards cholinesterase inhibition and ROS scavenging.

This study gives new insights to understand the type of interactions between Ginkgo biloba L. and Scutellaria baicalensis Georgi, two Chinese medicinal plants with well documented neuroprotective effects, on three targets in Alzheimer's disease (AD): acetylcholinesterase (AChE) and butyrylcholnesterase (BuChE) inhibition and hydrogen peroxide scavenging. Individual samples, binary mixtures with different proportions of both plant species, and also a commercial multicomponent combination containing both plants together with unroasted Coffea arabica L. and quercetin-3-O-rutinoside were used to perform this in vitro evaluation. Sample phenolic profiles were also determined by HPLC-DAD, showing the presence of several flavonoid glycosides, phenolic acids and a methylxanthine. In order to investigate the possible synergism/antagonism interaction, data obtained were analyzed by CompuSyn software. The results showed that G. biloba and S. baicalensis alone display better activities than in mixtures, most of the interactions exhibiting different degrees of antagonism. A slight synergism interaction was only observed for the commercial multicomponent mixture tested against H2O2. Further analysis was carried out to understand which compounds could be responsible for the antagonistic interaction. Seventeen single pure compounds present in all extracts were tested against AChE inhibition, most of them displaying weak or no activity. Only caffeine had a remarkable activity. Five different binary and quaternary mixture compositions were design to deepen the interaction between these compounds, revealing mainly phenolic acid-flavonoid, flavonoid-flavonoid and methylxanthine-flavonoid-phenolic acid antagonistic interactions. These results clearly show that, for the targets evaluated, there is no potentiation of the neuroprotective effect by combining S. baicalensis and G. biloba extracts.

Effect of Andrographis paniculata leaves extract on neurobehavioral and biochemical indices in scopolamine-induced amnesic rats.

Andrographis paniculata is a medicinal herb that is used to treat various disease conditions due to its pharmacological properties. Thus, this study sought to assess the effect of A. paniculata extract on neurobehavioral and some biochemical parameters in scopolamine-induced amnesic rats. Thirty-five male rats were divided into seven groups and treated with aqueous extract of A. paniculata (50 and 500 mg/kg) and donepezil (5 mg/kg) for 14 days before administration of scopolamine. Behavioral studies (Morris water maze and Y-maze) were carried out to evaluate cognitive dysfunction in scopolamine-induced rats. Biochemical assays such as cholinesterases (AChE and BChE), monoamine oxidase (MAO), and purinergic activities were determined. Results revealed the presence of orientin, quercetin, caffeic acid, apigenin, and gallic acid in A. paniculata. Also, findings from this study showed that aqueous extract of A. paniculata had a modulatory effect on scopolamine-induced cognitive impairment and could be used in the management of memory loss. PRACTICAL APPLICATIONS: Aqueous extract of A. paniculata characterized revealed the presence of polyphenols which are antioxidants. The inhibitory activity possessed by A. paniculata on some enzymes linked to neurodegeneration could be due to the antioxidant activity. Given this, we recommend that results gotten from this study could be used to develop treatment therapy for neurodegeneration. However, in-depth studies should be carried out on the toxic effect of A. paniculata to ascertain a safe dose for treatment.

Electrochemical screening of genoprotective and antioxidative effectiveness of Origanum vulgare L. and its functionality in the prevention of neurodegenerative disorders.

A new way of electrochemical DNA sensor using as a screening tool for the determination of phytochemicals with high genoprotective functionality is proposed. The biosensor's detection layer was prepared with double stranded deoxyribonucleic acid (ds DNA) that were subjected to oxidative stress induced by •OH radicals generated by Fenton reaction. The oxidized guanine derivative, 8-oxo-7,8-dihydro-2'-deoxyguanosine, was treated as an indicator of DNA oxidative damage. This derivative may cause mutation through its ability to pair with adenine. The abnormalities of DNA structure and DNA repair system are known to be directly related to progressive neurodegeneration. The present study showed that during oxidative stress, the 2.5% oregano extract protected guanine from undergoing oxidation to 8-oxoguanine. The results revealed that this genoprotective effectiveness can make oregano a very efficient protective barrier against oxidative stress. Due to these unique properties of oregano we propose the recipe of a functional bread with its addition. It was found that the functionality of the prepared bread was not limited to antioxidative activity but also is expressed in the inhibition of cholinesterases. These findings indicate that oregano can act as an important component in the therapeutic diet recommended in neurodegenerative disorders.

Kinetics and computational analysis of cholinesterase inhibition by REVERC3, a bisdemethoxycurcumin-rich Curcuma longa extract: Relevance to the treatment of Alzheimer's disease.

Objectives: Cholinesterase inhibition is a common strategy to treat Alzheimer's disease. In this study, we have investigated the cholinesterase inhibitory effects of a first-of-its-kind turmeric extract (REVERC3) having enriched content of bisdemethoxycurcumin as major active curcuminoid. Methods: The inhibition studies were performed using Ellman's colorimetric assay. The kinetics of acetylcholinesterase and butyrylcholinesterase was determined in the presence of REVERC3 using the Lineweaver-Burk double reciprocal plots. Furthermore, we used AutoDock tools to predict the binding of bisdemethoxycurcumin with the active sites of cholinesterases. Results: REVERC3 showed 4.8- and 5.39-fold higher inhibitory potential of acetylcholinesterase and butyrylcholinesterase with IC50 values of 29.08 and 33.59 µg/mL, respectively, compared to the regular turmeric extract. The mode of binding of REVERC3 was competitive in the case of acetylcholinesterase while it was uncompetitive for the inhibition of butyrylcholinesterase. Docking analysis revealed that bisdemethoxycurcumin, the major constituent of REVERC3, has different preferences of binding in the active sites of acetylcholinesterase and butyrylcholinesterase. However, the best binding pose predictions are in line with the experimental binding mode of the cholinesterases. Conclusion: These results indicate that bisdemethoxycurcumin-enriched turmeric extract could improve the cholinergic functions via dual inhibition of cholinesterases. However, the predominant role of bisdemethoxycurcumin in REVERC3 must be further validated using preclinical studies and clinical trials.

Effect of Solanum macrocarpon Linn leaf aqueous extract on the brain of an alloxan-induced rat model of diabetes.

OBJECTIVE: This study was designed to evaluate the protective effect of aqueous extract of Solanum macrocarpon Linn leaf in the brain of an alloxan-induced rat model of diabetes. METHODS: The experimental model of diabetes was induced by a single intraperitoneal injection of freshly prepared alloxan. Rats were then divided into six groups: normal control, diabetes control, diabetes group treated with metformin, and three diabetes groups treated with different concentrations of S. macrocarpon. Rats were sacrificed on day 14 of the experiment and different brain biochemical parameters were assessed and compared between groups. RESULTS: Administration of different doses of S. macrocarpon leaf aqueous extract was associated with significantly reduced levels of fasting blood glucose, lipid peroxidation, neurotransmitters, cholinesterases, cyclooxygenase-2 and nitric oxide compared with diabetes control rats. In addition, antioxidant enzyme activities were significantly increased in diabetes rats administered 12.45, 24.9 and 49.8 mg/kg body weight of S. macrocarpon versus diabetes control rats. CONCLUSION: Aqueous extract of S. macrocarpon Linn leaf may be useful in the management of diabetic neuropathy.

Norditerpenoids with Selective Anti-Cholinesterase Activity from the Roots of Perovskia atriplicifolia Benth.

Inhibition of cholinesterases remains one of a few available treatment strategies for neurodegenerative dementias such as Alzheimer's disease and related conditions. The current study was inspired by previous data on anticholinesterase properties of diterpenoids from Perovskia atriplicifolia and other Lamiaceae species. The acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibition by the three new natural compounds-(1R,15R)-1-acetoxycryptotanshinone (1), (1R)-1-acetoxytanshinone IIA (2), and (15R)-1-oxoaegyptinone A (3)-as well as, new for this genus, isograndifoliol (4) were assessed. Three of these compounds exhibited profound inhibition of butyrylcholinesterase (BChE) and much weaker inhibition of acetylcholinesterase (AChE). All compounds (1-4) selectively inhibited BChE (IC50 = 2.4, 7.9, 50.8, and 0.9 µM, respectively), whereas only compounds 3 and 4 moderately inhibited AChE (IC50 329.8 µM and 342.9 µM). Molecular docking and in silico toxicology prediction studies were also performed on the active compounds. Natural oxygenated norditerpenoids from the traditional Central Asian medicinal plant P. atriplicifolia are selective BChE inhibitors. Their high potential makes them useful candidate molecules for further investigation as lead compounds in the development of a natural drug against dementia caused by neurodegenerative diseases.

Chemical Profiling and Cholinesterase Inhibitory Activity of Five Phaedranassa Herb. (Amaryllidaceae) Species from Ecuador.

It is estimated that 50 million people in the world live with dementia, 60-70% of whom suffer from Alzheimer's disease (AD). Different factors are involved in the development of AD, including a reduction in the cholinergic neurotransmission level. The Amaryllidaceae plant family contains an exclusive, large, and still understudied alkaloid group characterized by a singular skeleton arrangement and a broad spectrum of biological activities. The chemistry and biodiversity of Ecuadorian representatives of the Phaedranassa genus (Amaryllidaceae) have not been widely studied. In this work, five Ecuadorian Phaedranassa species were examined in vitro for their activity towards the enzymes acetyl- (AChE) and butyrylcholinesterase (BuChE), and the alkaloid profile of bulb extracts was analyzed by GC-MS. The species Phaedranassa cuencana and Phaedranassa dubia showed the most AChE and BuChE inhibitory activity, respectively. To obtain insight into the potential role of the identified alkaloids in these inhibitory effects, docking experiments were carried out, and cantabricine showed in silico inhibitory activity against both cholinesterase structures. Our results show that Amaryllidaceae species from Ecuador are a potential source of new drugs for the palliative treatment of AD.

Evaluation of neuroprotective and anti-amnesic effects of Elaeagnus umbellata Thunb. On scopolamine-induced memory impairment in mice.

BACKGROUND: Elaeagnus umbellata is abundantly found in Himalayan regions of Pakistan which is traditionally used to treat various health disorders. However, the experimental evidence supporting the anti-amnesic effect is limited. Therefore the study was aimed to evaluate the prospective beneficial effect of E. umbellata on learning and memory in mice. OBJECTIVES: To assess neuroprotective and anti-amnesic effects of E. umbellata fruit extracts and isolated compounds on the central nervous system. METHODS: Major phytochemical groups present in methanolic extract of E. umbellata were qualitatively determined. The total phenolic and flavonoid contents were also determined in extract/fractions of E. umbellata. On the basis of in vitro promising anticholinesterases (AChE & BChE) and antioxidant activities observed for CHF. Ext and isolated compound-I (Chlorogenic acid = CGA), they were further evaluated for learning and memory in normal and scopolamine-induced cognitive impairment in mice using memory behavioral tests such as the Y maze and Novel object recognition using standard procedures. The test sample were further assessed for in vivo anticholinesterases (AChE & BChE) and DPPH free radical scavenging activities in mice brain sample and finally validated by molecular docking study using GOLD software. RESULTS: The extract/fractions and isolated compounds were tested for their anticholinesterase and antioxidant potentials. The CHF. Ext and CGA showed maximum % inhibition of tested cholinesterases and free radicals. The CHF. Ext and CGA reversed the effects of scopolamine in mice. The CHF. Ext and CGA significantly increased the alternate arm returns and % spontaneous alteration performance while escape latency times (second) significantly decreased in Y maze test. The CHF. Ext and CGA significantly increased the time spent with novel object and also increased the discrimination index in the Novel object recognition test. Furthermore, molecular docking was used to validate the mechanism of cholinesterases inhibition of isolated compounds. CONCLUSION: The data obtained from behavioral and biochemical studies (AChE/BChE and DPPH/ABTS inhibition) have shown that E. umbellata possessed significant memory enhancing potency. These results suggest that E. umbellata extract possess potential antiamnesic effects and amongst the isolated compounds, compound I could be more effective anti-amnesic therapeutics. However, further studies are needed to identify the exact mechanism of action.

Exploring Structure-Activity Relationship in Tacrine-Squaramide Derivatives as Potent Cholinesterase Inhibitors.

Tacrine was the first drug to be approved for Alzheimer's disease (AD) treatment, acting as a cholinesterase inhibitor. The neuropathological hallmarks of AD are amyloid-rich senile plaques, neurofibrillary tangles, and neuronal degeneration. The portfolio of currently approved drugs for AD includes acetylcholinesterase inhibitors (AChEIs) and N-methyl-d-aspartate (NMDA) receptor antagonist. Squaric acid is a versatile structural scaffold capable to be easily transformed into amide-bearing compounds that feature both hydrogen bond donor and acceptor groups with the possibility to create multiple interactions with complementary sites. Considering the relatively simple synthesis approach and other interesting properties (rigidity, aromatic character, H-bond formation) of squaramide motif, we combined this scaffold with different tacrine-based derivatives. In this study, we developed 21 novel dimers amalgamating squaric acid with either tacrine, 6-chlorotacrine or 7-methoxytacrine representing various AChEIs. All new derivatives were evaluated for their anti-cholinesterase activities, cytotoxicity using HepG2 cell line and screened to predict their ability to cross the blood-brain barrier. In this contribution, we also report in silico studies of the most potent AChE and BChE inhibitors in the active site of these enzymes.

Flavonoids as Prospective Neuroprotectants and Their Therapeutic Propensity in Aging Associated Neurological Disorders.

Modern research has revealed that dietary consumption of flavonoids and flavonoids-rich foods significantly improve cognitive capabilities, inhibit or delay the senescence process and related neurodegenerative disorders including Alzheimer's disease (AD). The flavonoids rich foods such as green tea, cocoa, blue berry and other foods improve the various states of cognitive dysfunction, AD and dementia-like pathological alterations in different animal models. The mechanisms of flavonoids have been shown to be mediated through the inhibition of cholinesterases including acetylcholinesterase (AChE), and butyrylcholinesterase (BChE), ß-secretase (BACE1), free radicals and modulation of signaling pathways, that are implicated in cognitive and neuroprotective functions. Flavonoids interact with various signaling protein pathways like ERK and PI3-kinase/Akt and modulate their actions, thereby leading to beneficial neuroprotective effects. Moreover, they enhance vascular blood flow and instigate neurogenesis particularly in the hippocampus. Flavonoids also hamper the progression of pathological symptoms of neurodegenerative diseases by inhibiting neuronal apoptosis induced by neurotoxic substances including free radicals and ß-amyloid proteins (Aß). All these protective mechanisms contribute to the maintenance of number, quality of neurons and their synaptic connectivity in the brain. Thus flavonoids can thwart the progression of age-related disorders and can be a potential source for the design and development of new drugs effective in cognitive disorders.

Aqueous-ethanol extracts of some South African seaweeds inhibit beta-amyloid aggregation, cholinesterases, and beta-secretase activities in vitro.

In this study, we evaluated the anti-amyloidogenic, anticholinesterase, and antioxidant potentials of hydroethanolic extracts of Ecklonia maxima (ECK), Gelidium pristoides (GLD), Gracilaria gracilis (GCL), and Ulva lactuca (ULT). The effect of the extracts on ß-amyloid (Aß1-42 ) peptide were determined using electron microscope. The effects of the extracts on ß-secretase and cholinesterase activities, as well as their radical scavenging and metal chelating activities were also assessed. Electron micrographs revealed that ECK, GLD, GCL, and ULT incubated with Aß1-42 at different intervals (0-96 hr) showed very low levels of fibrils compared to the control. The extracts also inhibited ß-secretase, acetylcholinesterase, and butyrylcholinesterase activities in a dose-dependent manner. Furthermore, the extracts scavenged hydroxyl radicals and were able to chelate Fe2+ in a dose-dependent manner. Our findings suggest that the seaweed extracts are potential sources of lead compounds and novel inhibitors of ß-amyloid aggregation, ß-secretase, and cholinesterases for the management of Alzheimer's diseases. PRACTICAL APPLICATIONS: Seaweeds have been identified as good sources of naturally occurring bioactive compounds with several medicinal properties. They are commonly used as functional foods and development of nutraceuticals, dietary supplements, and cosmeceuticals. However, the neuroprotective effects of many species of seaweeds have not been fully explored. The findings of this study suggests that Gracilaria gracilis, Ulva lactuca, Ecklonia maxima, and Gelidium pristoides are potential sources of cholinesterase, beta-secretase, and amyloid protein aggregation inhibitors. Hence, this support the use of these seaweeds as alternative sources of antioxidants and natural compounds with neuroprotective potentials for the management of Alzheimer's disease.

Phenolic composition, antioxidant activity, anticholinesterase potential and modulatory effects of aqueous extracts of some seaweeds on ß-amyloid aggregation and disaggregation.

Context: Seaweeds contain bioactive compounds with different biological activities. They are used as functional ingredients for the development of therapeutic agents to combat degenerative diseases. Objective: This study investigated the phenolic composition, antioxidant activity, cholinesterase inhibitory and anti-amyloidogenic activities of aqueous extracts of Gracilaria beckeri (J.Agardh) Papenfuss (Gracilariaceae) (RED-AQ), Ecklonia maxima (Osbeck) Papenfuss (Lessoniaceae) (ECK-AQ), Ulva rigida (C.Agardh) Linnaeus (Ulvaceae) (URL-AQ) and Gelidium pristoides (Turner) Kützing (Gelidiaceae) (GEL-AQ). Materials and methods: Phenolic composition of the seaweed extracts was determined using liquid chromatography mass spectrometry. Radical scavenging and metal chelating activities were assessed in vitro. The effect of the extracts (21-84 µg/mL) on acetylcholinesterase and butyrylcholinesterase activities were also investigated using an in vitro colorimetric assay. Transmission electron microscope and thioflavin-T fluorescence assay were used to examine the anti-amyloidogenic activities of the extracts. Results: Phloroglucinol, catechin, epicatechin 3-glucoside were identified in the extracts. ECK-AQ (IC50=30.42 and 280.47 µg/mL) exhibited the highest OH• scavenging and metal chelating activities, while RED-AQ (41.23 and 334.45 µg/mL) exhibited the lowest. Similarly, ECK-AQ (IC50 = 49.41 and 52.11 µg/mL) exhibited higher inhibitory effects on acetylcholinesterase and butyrylcholinesterase activities, while RED-AQ (64.56 and 63.03 µg/mL) showed the least activities. Rapid formation of ß-amyloid (Aß1-42) fibrils and aggregates was observed in electron micrographs of the control after 72 and 96 h. The reduction of Aß1-42 aggregates occurred after co-treatment with the seaweed extracts. Discussion and conclusion: ECK-AQ, GEL-AQ, URL-AQ and RED-AQ may possess neuroprotective potential and could be explored for the management of Alzheimer's disease.

Chlorella sorokiniana and Chlorella minutissima exhibit antioxidant potentials, inhibit cholinesterases and modulate disaggregation of ß-amyloid fibrils

BACKGROUND: Microalgae are aquatic chlorophyll-containing organisms comprising unicellular microscopic forms, and their biomasses are potential sources of bioactive compounds, biofuels and food-based products. However, the neuroprotective effects of microalgal biomass have not been fully explored. In this study, biomass from two Chlorella species was characterized, and their antioxidant, anticholinesterase and anti-amyloidogenic activities were investigated. RESULTS: GC­MS analysis of the extracts revealed the presence of some phenols, sterols, steroids, fatty acids and terpenes. Ethanol extract of Chlorella sorokiniana (14.21 mg GAE/g) and dichloromethane extract of Chlorella minutissima (20.65 mg QE/g) had the highest total phenol and flavonoid contents, respectively. All the extracts scavenged 2,2-diphenyl-1-picrylhydrazyl, 2,2-azinobis (3-ethylbenzothiazoline-6-sulfonate) and hydroxyl radicals. The highest metal chelating activity of the extracts was observed in the ethanol extracts of C. minutissima (102.60 µg/mL) and C. sorokiniana (107.84 µg/mL). Furthermore, the cholinesterase inhibitory activities of the extracts showed that ethanol extract of C. sorokiniana (13.34 µg/mL) exhibited the highest acetylcholinesterase inhibitory activity, while dichloromethane extract of C. minutissima (11.78 µg/mL) showed the highest butyrylcholinesterase inhibitory activity. Incubation of the ß-amyloid protein increased the aggregation of amyloid fibrils after 96 h. However, ethanol extract of C. sorokiniana and C. minutissima inhibited further aggregation of Aß1­42 and caused disaggregation of matured protein fibrils compared to the control. This study reveals the modulatory effects of C. sorokiniana and C. minutissima extracts on some mediators of Alzheimer's disease and provides insights into their potential benefits as functional food, nutraceutics or therapeutic agent for the management of this neurodegenerative disease.