The exploration of novel Alzheimer's therapeutic agents from the pool of FDA approved medicines using drug repositioning, enzyme inhibition and kinetic mechanism approaches.

Novel drug development is onerous, time consuming and overpriced process with particularly low success and relatively high enfeebling rates. To overcome this burden, drug repositioning approach is being used to predict the possible therapeutic effects of FDA approved drugs in different diseases. Herein, we designed a computational and enzyme inhibitory mechanistic approach to fetch the promising drugs from the pool of FDA approved drugs against AD. The binding interaction patterns and conformations of screened drugs within active region of AChE were confirmed through molecular docking profiles. The possible associations of selected drugs with AD genes were predicted by pharmacogenomics analysis and confirmed through data mining. The stability behaviour of docked complexes (Drugs-AChE) were checked by MD simulations. The possible therapeutic potential of repositioned drugs against AChE were checked by in vitro analysis. Taken together, Cinitapride displayed a comparable results with standard and can be used as possible therapeutic agent in the treatment of AD.

Green Synthesis of Silver Nanoparticles using Bidens Frondosa Extract and their Tyrosinase Activity.

Herbal nanoparticles gain lot of attention because of their pharmaceutical importance. The present study reports the eco-friendly synthesis, characterization and their tyrosinase activity of silver nanoparticles (AgNPs) using aqueous extract of Bidens frondosa. The appearance of brown color indicated the formation of B. frondosa AgNPs. The Formation of AgNPs was confirmed by UV-Vis spectroscopy, FTIR, FESEM and EDS analysis. The formation of herbal AgNPs of size ranging 20-70 nm further was assured by energy dispersive X-ray spectroscopy (EDS) and field emission scanning electron microscopy (FESEM). The mushroom tyrosinase inhibitory activity of synthesized AgNPs was evaluated. Nanoparticles were found to have significant higher tyrosinase inhibitory activity compared to control. The IC50 values of crude extract, AgNP and Kojic acid were found to be 9, 15, and 2.37 µg/mL, respectively. AgNPs of B. frondosa may be considered as potential candidate for the production of medical and cosmetic products.

Pharmacoinformatics exploration of polyphenol oxidases leading to novel inhibitors by virtual screening and molecular dynamic simulation study.

Polyphenol oxidases (PPOs)/tyrosinases are metal-dependent enzymes and known as important targets for melanogenesis. Although considerable attempts have been conducted to control the melanin-associated diseases by using various inhibitors. However, the exploration of the best anti-melanin inhibitor without side effect still remains a challenge in drug discovery. In present study, protein structure prediction, ligand-based pharmacophore modeling, virtual screening, molecular docking and dynamic simulation study were used to screen the strong novel inhibitor to cure melanogenesis. The 3D structures of PPO1 and PPO2 were built through homology modeling, while the 3D crystal structures of PPO3 and PPO4 were retrieved from PDB. Pharmacophore modeling was performed using LigandScout 3.1 software and top five models were selected to screen the libraries (2601 of Aurora and 727, 842 of ZINC). Top 10 hit compounds (C1-10) were short-listed having strong binding affinities for PPO1-4. Drug and synthetic accessibility (SA) scores along with absorption, distribution, metabolism, excretion and toxicity (ADMET) assessment were employed to scrutinize the best lead hit. C4 (name) hit showed the best predicted SA score (5.75), ADMET properties and drug-likeness behavior among the short-listed compounds. Furthermore, docking simulations were performed to check the binding affinity of C1-C10 compounds against target proteins (PPOs). The binding affinity values of complex between C4 and PPOs were higher than those of other complexes (-11.70, -12.1, -9.90 and -11.20kcal/mol with PPO1, PPO2, PPO3, or PPO4, respectively). From comparative docking energy and binding analyses, PPO2 may be considered as better target for melanogenesis than others. The potential binding modes of C4, C8 and C10 against PPO2 were explored using molecular dynamics simulations. The root mean square deviation and fluctuation (RMSD/RMSF) graphs results depict the significance of C4 over the other compounds. Overall, bioactivity and ligand efficiency profiles suggested that the proposed hit may be more effective inhibitors for melanogenesis.

Isolation, characterization, and in silico, in vitro and in vivo antiulcer studies of isoimperatorin crystallized from Ostericum koreanum.

CONTEXT: Ostericum koreanum (Maxim.) Kitagawa (Apiaceae) roots are traditionally used as an analgesic and antiulcer agent. However, the antiulcer potential of isoimperatorin isolated from O. koreanum has not yet been explored. AIM: To evaluate the antiulcer activity of isoimperatorin isolated from the roots of O. koreanum. MATERIALS AND METHODS: Isoimperatorin was isolated as cubic crystals by repeated column chromatography of the ethyl acetate fraction and structure was verified with 1H NMR, 13C NMR and high-resolution mass spectrometry (HRMS-FAB). The crystals obtained were analyzed with the single crystal X-ray method. The MTT assay was used to determine its cytotoxicity against chondrocytes at different concentrations (0.0-737.74 µM, 24 h). The in vivo antiulcer activity of isoimperatorin (40 mg/kg) was determined against ethanol-, indomethacin- and pyloric ligation-induced ulcers in Sprague-Dawley rats. Furthermore, the effect of isoimperatorin (0.0-737.74 µM, 24 h) on the expression of type II collagen in chondrocytes was determined using western blot method. The in vitro urease inhibitory activity of isoimperatorin (0-80 µM) and molecular docking was also performed against urease. RESULTS AND DISCUSSION: Isoimperatorin demonstrated significant inhibitory activity (IC50 36.43 µM) against urease as compared to the standard drug thiourea (IC50 33.57 µM) without cytotoxic effects. It provided 70.9%, 67.65% and 54.25% protection in ulcer models induced by ethanol, indomethacin and pyloric ligation, respectively. Isoimperatorin showed the highest expression level of type II collagen at 368.87 µM. The docking results confirmed strong binding affinity with the target protein. CONCLUSION: Isoimperatorin may be used to develop antiulcer drugs with decreased side effects.

Acetylcholinesterase immobilization and characterization, and comparison of the activity of the porous silicon-immobilized enzyme with its free counterpart.

A successful prescription is presented for acetylcholinesterase physically adsorbed on to a mesoporous silicon surface, with a promising hydrolytic response towards acetylthiocholine iodide. The catalytic behaviour of the immobilized enzyme was assessed by spectrophotometric bioassay using neostigmine methyl sulfate as a standard acetycholinesterase inhibitor. The surface modification was studied through field emission SEM, Fourier transform IR spectroscopy, energy-dispersive X-ray spectroscopy, cathode luminescence and X-ray photoelectron spectroscopy analysis, photoluminescence measurement and spectrophotometric bioassay. The porous silicon-immobilized enzyme not only yielded greater enzyme stability, but also significantly improved the native photoluminescence at room temperature of the bare porous silicon architecture. The results indicated the promising catalytic behaviour of immobilized enzyme compared with that of its free counterpart, with a greater stability, and that it aided reusability and easy separation from the reaction mixture. The porous silicon-immobilized enzyme was found to retain 50% of its activity, promising thermal stability up to 90°C, reusability for up to three cycles, pH stability over a broad pH of 4-9 and a shelf-life of 44 days, with an optimal hydrolytic response towards acetylthiocholine iodide at variable drug concentrations. On the basis of these findings, it was believed that the porous silicon-immobilized enzyme could be exploited as a reusable biocatalyst and for screening of acetylcholinesterase inhibitors from crude plant extracts and synthesized organic compounds. Moreover, the immobilized enzyme could offer a great deal as a viable biocatalyst in bioprocessing for the chemical and pharmaceutical industries, and bioremediation to enhance productivity and robustness.

Berberine induces dedifferentiation by actin cytoskeleton reorganization via phosphoinositide 3-kinase/Akt and p38 kinase pathways in rabbit articular chondrocytes.

Osteoarthritis is a nonrheumatologic joint disease characterized by progressive degeneration of the cartilage extracellular matrix. Berberine (BBR) is an isoquinoline alkaloid used in traditional Chinese medicine, the majority of which is extracted from Huang Lian (Coptis chinensis). Although numerous studies have revealed the anticancer activity of BBR, its effects on normal cells, such as chondrocytes, and the molecular mechanisms underlying its actions remain elusive. Therefore, we examined the effects of BBR on rabbit articular chondrocytes, and the underlying molecular mechanisms, focusing on actin cytoskeletal reorganization. BBR induced dedifferentiation by inhibiting activation of phosphoinositide-3(PI3)-kinase/Akt and p38 kinase. Furthermore, inhibition of p38 kinase and PI3-kinase/Akt with SB203580 and LY294002, respectively, accelerated the BBR-induced dedifferentiation. BBR also caused actin cytoskeletal architecture reorganization and, therefore, we investigated if these effects were involved in the dedifferentiation. Disruption of the actin cytoskeleton by cytochalasin D reversed the BBR-induced dedifferentiation by activating PI3-kinase/Akt and p38 kinase. In contrast, the induction of actin filament aggregation by jasplakinolide accelerated the BBR-induced dedifferentiation via PI3-kinase/Akt inhibition and p38 kinase activation. Taken together, these data suggest that BBR strongly induces dedifferentiation, and actin cytoskeletal reorganization is a crucial requirement for this effect. Furthermore, the dedifferentiation activity of BBR appears to be mediated via PI3-kinase/Akt and p38 kinase pathways in rabbit articular chondrocytes.

Antidiabetic and antiacetylcholinesterase effects of ethyl acetate fraction of Chaenomeles sinensis (Thouin) Koehne fruits in streptozotocin-induced diabetic rats.

The present study was intended to examine the effects of the supplementation of active α-glucosidase, α-amylase and lipase inhibitory ethyl acetate (CSE) fraction from the fruits of Chaenomeles sinensis (Thouin) Koehne on blood glucose (BG), triglyceride (TG), total cholesterol (TC), high density lipoprotein cholesterol (HDL-C), alanine aminotransferase (ALT), aspartate aminotransaminase (AST), acetylcholinesterase (AChE) and antioxidant levels. The diabetic rats were treated orally with CSE at the doses of 50 and 100 mg/kg bw for 14 days. BG, TC, TG, HDL-C, ALT, AST and AChE levels were significantly reduced; on the other hand antioxidant levels were significantly increased in the treated groups. These observations suggest protective effects of CSE against STZ-induced diabetic dementia model.

Screening of Korean Medicinal Plant Extracts for α-Glucosidase Inhibitory Activities.

Glycosidases are the enzymes involved in various biochemical processes related to metabolic disorders and diseases. Therefore, much effort has been focused on searching novel pharmacotherapy for the treatment of these ailments from medicinal plants due to higher safety margins. To pursue these efforts, the present study was performed to evaluate the α-glucosidase inhibitory activities of thirty Korean medicinal plant extracts. Among the plants studied, Euonymus sachalinensis, Rhododendron schlippenbachii, Astilbe chinensis and Juglans regia showed the strongest α-glucosidase inhibitory activity with IC50 values of 10, 20, 30 and 80 µg/mL, respectively. In addition, Meliosma oldhamii and Symplocos chinensis showed moderate α-glucosidase inhibition with IC50 values of 150 and 220 µg/mL, respectively. Therefore, they might prove to be a potential natural source for the treatment of metabolic ailments such as, diabetes, and need further investigations.

Evaluation of antiglycosidase and anticholinesterase activities of Boehmeria nivea.

In this era, major community worldwide is suffering from diabetes type II, cancer and neurodegenerative disorders. To overcome these diseases, in the screening of Korean medicinal plants, we studied the whole plant of Boehmeria nivea (B. nivea). The methanolic leaf, stem and root extracts of B. nivea and their respective n-hexane, methylene chloride (CH(2)Cl(2)), ethyl acetate (EtOAc), n-butanol (BuOH) and aqueous fractions were investigated for their total phenolic content (TPC), 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging activity, alpha-glucosidase, beta-glucosidase, alpha-galactosidase, beta-galactosidase, acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) enzyme inhibition activities. Profound TPC and DPPH free radical scavenging activities were observed in the EtOAc and BuOH fractions of root, where the BuOH fraction showed high-pitched alpha-glucosidase inhibition and the EtOAc layer showed the maximum beta-glucosidase inhibition. Furthermore, the leaf extract demonstrated the highest beta-galactosidase inhibitory activity, but no alpha-galactosidase inhibition was seen in any of the plant parts. Notable BChE and moderate AChE inhibitory activity was found in whole plant. It can be suggested that whole plant of B. nivea provides a strong biochemical rationale as one of the good choices for the treatment of diabetes type II, cancer and neurodegenerative diseases (AD, etc).

Effect of 1,2,3,4,6-penta-O-galloyl-beta-D-glucose on elastase and hyaluronidase activities and its type II collagen expression.

In the current era, natural products are gaining prime attention in the fields of cosmeceuticals and pharmaceuticals due to higher safety margins and biological functions, as they have a considerable amount of potential in treating different ailments. Thus, to find effective elastase and hyaluronidase inhibitors from natural resources, fifty Korean plants were screened, and the fruit of Terminalia chebula RETZIUS (Combretaceae) was selected for further structural isolation due to its potent efficacy. The methanol crude extract of the fruits showed 80% elastase and 87% hyaluronidase enzyme inhibition activities at 1 mg/mL. The crude extract, upon bioassay-directed fractionation, led to the isolation of compound 1, whose structure was found by spectral analysis to be 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (PGG). PGG displayed significant elastase and hyaluronidase inhibitory activities with IC50, values of 57 microg/mL and 0.86 mg/mL, respectively; also, treatment of PGG on rabbit articular chondrocytes significantly induced the type II collagen expression. Based on elastase and hyaluronidase inhibitions, and type II collagen expression, it could be suggested that PGG might have an influence on skin conditions when used cosmetically as an active anti-aging ingredient with no cytotoxicity; also, it might be beneficial in relieving painful joint conditions, and thus have relevance for treating arthritis. Therefore, it can be concluded that PGG may prove to be an active ingredient in cosmeceutical and pharmaceutical formulations, and that it definitely merits further in vivo investigations.

Observations of Forsythia koreana methanol extract on mast cell-mediated allergic reactions in experimental models.

To explore effects of Forsythia koreana methanol extract (FKME) on mast cell-mediated allergic and inflammatory properties, the effect of FKME was evaluated on compound 48/80-induced systemic anaphylaxis, ear swelling, and anti-dinitrophenyl (DNP) immunoglobulin E (IgE)-induced passive cutaneous anaphylaxis (PCA). In addition, the effect of FKME was investigated on the histamine release from rat peritoneal mast cells (RPMCs) stimulated by compound 48/80, which promotes histamine release. The human mast cell line HMC-1 was stimulated by phorbol 12-myristate 13-acetate plus calcium ionophore A23187. Activated HMC-1 can produce several proinflammatory and chemotactic cytokines such as tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-6, and IL-8. Cytokine levels in the culture supernatant were measured by an enzyme-linked immunosorbent assay. Cytotoxicity by FKME was determined by a 3-(4,5-dimethylthiazol-2-yl)-diphenyl-tetrazolium bromide (MTT) assay. FKME inhibited compound 48/80-induced systemic anaphylactic shock and ear swelling in mice. When 1 g/kg FKME was pretreated or posttreated with mice, compound 48/80-induced mice morality was 50 and 66.7%, respectively. One gram per kilogram of FKME pretreatment inhibited ear-swelling responses derived from compound 48/80 by 29.75%. A PCA reaction was inhibited by 17.9%. In an in vitro model, FKME (1 mg/ml) inhibited histamine release from the RPMCs by 13.8% and TNF-alpha, IL-6, and IL-8 production from HMC-1 cells by 71.16% (P < 0.001), 86.72% (P < 0.001), and 44.6%, respectively. However, FKME had no cytotoxic effects on cell viability. In conclusion, FKME inhibited not only systemic anaphylaxis and ear swelling induced by compound 48/80 but also inhibited a PCA reaction induced by anti-DNP IgE in vivo. Treatment with FKME showed significant inhibitory effects on histamine, TNF-alpha, IL-6, and IL-8 release from mast cells.

Cytokine reporter mouse system for screening novel IL12/23 p40-inducing compounds.

Cytokines interleukin (IL) 12 and 23 play critical roles in linking innate and adaptive immune responses. They are members of heterodimeric cytokines, sharing a subunit p40. Although IL12/23 p40 is mainly induced in macrophages and dendritic cells (DCs) after stimulation with microbial Toll-like receptor ligands, methods to monitor the cells that produce IL12/23 p40 in vivo are limited. Recently, the mouse model to track p40-expressing cells with fluorescent reporter, yellow fluorescent protein, has been developed. Macrophages and DCs from these mice faithfully reported p40 induction using the fluorescent marker. Here we took advantage of these reporter mice to screen bio-compounds for p40-inducing activity. After screening hundreds of compounds, we found several extracts inducing IL12/23 p40 gene expression. Treatment of DCs with these extracts induced the expression of MHC class II and co-stimulatory molecules, which implies that these might be useful as adjuvants. Next, the in vivo target immune cells of candidate compounds were examined. The reporter system can be useful to identify cells producing IL12 or IL23 in vivo as well as in vitro. Thus, our cytokine reporter system proved to be a valuable reagent for screening for immunostimulatory molecules and identification of target cells in vivo.

Screening of some medicinal plants for anti-lipase activity.

In order to find new pancreatic lipase (triacylglycerol lipase, EC 3.1.1.3) inhibitors from natural sources, 75 medicinal plants belonging to different families were screened for their anti-lipase activity, using a radioactive method. On evaluating the results, methanolic extracts of three plants namely, Eriochloa villosa (Thunb.) Kunth, Orixa japonica Thunb. and Setaria italica (L.) Palib., exhibited strong in vitro anti-lipase activity (above 80%). These plants will be studied further to elucidate the structure and chemical properties of the active compound responsible for anti-lipase action.

In vitro anti-tyrosinase activity of 5-(hydroxymethyl)-2-furfural isolated from Dictyophora indusiata.

The inhibition of mushroom tyrosinase by methanolic extract of Dictyophora indusiata was evaluated and the bioactive component was characterized and identified as 5-(hydroxymethyl)-2-furfural (HMF) by chromatographic and spectroscopic means. Kinetic studies revealed it to be a noncompetitive inhibitor for the oxidation of L-DOPA. On the basis of these findings some related analogues were also tested for their anti-tyrosinase activity, in order to gain more insight into structure and activity relationship among these heterocyclic compounds.