In silico evaluation of the compounds of the ayurvedic drug, AYUSH-64, for the action against the SARS-CoV-2 main protease.

BACKGROUND: Outbreak of Corona Virus Disease in late 2019 (COVID-19) has become a pandemic global Public health emergency. Since there is no approved anti-viral drug or vaccine declared for the disease and investigating existing drugs against the COVID-19. OBJECTIVE: AYUSH-64 is an Ayurvedic formulation, developed and patented by Central Council of Research in Ayurvedic Sciences, India, has been in clinical use as anti-malarial, anti-inflammatory, anti-pyretic drug for few decades. Thus, the present study was undertaken to evaluate AYUSH-64 compounds available in this drug against Severe Acute Respiratory Syndrome-Corona Virus (SARS-CoV-2) Main Protease (Mpro; PDB ID: 6LU7) via in silico techniques. MATERIALS AND METHODS: Different molecular docking software's of Discovery studio and Auto Dock Vina were used for drugs from selected AYUSH-64 compounds against SARS-CoV-2. We also conducted 100 ns period of molecular dynamics simulations with Desmond and further MM/GBSA for the best complex of AYUSH-64 with Mpro of SARS-CoV-2. RESULTS: Among 36 compounds of four ingredients of AYUSH-64 screened, 35 observed to exhibits good binding energies than the published positive co-crystal compound of N3 pepetide. The best affinity and interactions of Akuammicine N-Oxide (from Alstonia scholaris) towards the Mpro with binding energy (AutoDock Vina) of -8.4 kcal/mol and Discovery studio of Libdock score of 147.92 kcal/mol. Further, molecular dynamics simulations with MM-GBSA were also performed for Mpro- Akuammicine N-Oxide docked complex to identify the stability, specific interaction between the enzyme and the ligand. Akuammicine N-Oxide is strongly formed h-bonds with crucial Mpro residues, Cys145, and His164. CONCLUSION: The results provide lead that, the presence of Mpro- Akuammicine N-Oxide with highest Mpro binding energy along with other 34 chemical compounds having similar activity as part of AYUSH-64 make it a suitable candidate for repurposing to management of COVID-19 by further validating through experimental, clinical studies.

Anti-depressant activity of Erythrina variegata bark extract and regulation of monoamine oxidase activities in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Erythrina variegata, commonly referred to as 'Indian coral tree' belongs to the Fabaceae family. It is a plant native to the coast of India, China and is distributed in tropical and subtropical regions worldwide. In traditional medicine, E. variegata is known to exhibit anxiolytic and anti-convulsant activities and has been used as a nervine sedative. As per the Indian Materia Medica, E. variegata barks have been traditionally known to act on the central nervous system. However, there is a lack of data demonstrating this. AIM OF THE STUDY: Our study focuses on previously unreported anti-depressant activity of E. variegata bark ethanolic extract (EBE) and determination of its mechanism of action possibly through regulation of monoamine oxidase activity in mouse brain homogenates. MATERIALS AND METHODS: EBE was characterized using standard protocols for phytochemical analysis, followed by liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS) analysis. The compounds in EBE (previously reported for anti-depressant activity), were further studied by LC-MS/MS and GC-MS/MS analysis. Anti-depressant activity of EBE (50, 100, 200 and 500 mg/kg) was evaluated in Swiss albino mice using acute and chronic tail suspension test (TST) and forced swim test (FST) models. Furthermore, the possible mechanism of action of EBE was evaluated using the chronic unpredictable mild stress (CUMS) model, wherein inhibitory effects on monoamine oxidase (MAO) A and B were assessed by spectrophotometric-chemical analysis in mouse whole brain homogenates. RESULTS: EBE showed significant reduction in immobility time periods in both TST (acute: 50, 100 and 200 mg/kg and chronic: 100 and 200 mg/kg) and FST (acute: 200 mg/kg and chronic: 100, 200 and 500 mg/kg) models. Moreover, the locomotor activity test confirmed that acute and chronic administration of EBE did not significantly affect the motor activity of mice. In the CUMS model, EBE when administered alone (100 and 200 mg/kg) and in combination (50, 100 and 200 mg/kg) with escitalopram (15 mg/kg), showed significant reductions in immobility time periods compared to the control group, in the acute FST performed on 22nd day of CUMS. Furthermore, when administered alone (50, 100 and 200 mg/kg), EBE showed significant inhibition in MAO-A and B activities compared to the control group. When used in combination, EBE (50, 100 and 200 mg/kg) showed synergistic action with escitalopram (15 mg/kg), resulting in significantly greater inhibition of MAO-A and B activities, compared to both EBE alone and escitalopram alone. Phytochemical analysis of EBE revealed presence of sugars, steroids, glycosides, alkaloids and tannins. LC-MS, LC-MS/MS, GC-MS and GC-MS/MS analysis identified components in EBE, previously reported for their anti-depressant activity. CONCLUSIONS: The study thus concluded the anti-depressant like activity of EBE. The study identified components present in EBE that may be responsible for its anti-depressant activity. The possible mechanism of action of EBE was also investigated in the CUMS model, wherein inhibitory effects of EBE on MAO-A and B activities in the mouse brain were demonstrated. Furthermore, the study confirms the traditional use of E. variegata barks in CNS related activities through its anti-depressant like activity.

Anti-inflammatory evaluation and characterization of leaf extract of Ananas comosus.

Ananas comosus (L.) Merr (Pineapple) is a tropical plant with an edible fruit. In the present study, the potential anti-inflammatory activity of A. comosus leaf extract (ALE) was studied. ALE prepared using soxhlet apparatus was subjected to preliminary qualitative phytochemical analysis and quantitative estimations of flavonoids and tannins. The components present in ALE were identified using liquid chromatography-mass spectrometry (LC-MS) and gas chromatography-mass spectrometry (GC-MS). Inhibitory effects of ALE on protein denaturation, and proteinase activity were assessed. Its effect on secretion of pro-inflammatory cytokines and inflammatory mediators by lipopolysaccharide-stimulated macrophages was also analyzed. Further, its anti-inflammatory activity in carrageenan-induced inflammatory rat model was examined. The preliminary qualitative phytochemical analysis revealed presence of flavonoids, phenols, tannins, carbohydrates, glycosides, and proteins in the extract. Total flavonoids and total tannins were 0.17 ± 0.006 mg equivalent of quercetin/g of ALE and 4.04 ± 0.56 mg equivalent of gallic acid/g of ALE. LC-MS analysis identified the presence of 4-hydroxy pelargonic acid, 3,4,5-trimethoxycinnamic and 4-methoxycinnamic acid, whereas GC-MS analysis identified the presence of campesterol and ethyl isoallocholate that have been previously reported for anti-inflammatory activity. ALE showed significant inhibition of protein denaturation and proteinase activity and also controlled secretion of tumour necrosis factor-α, interleukin-1ß and prostaglandins, as well as the generation of reactive oxygen species by activated macrophages. ALE also significantly decreased carrageenan-induced acute paw edema. The study, therefore, identified the components present in ALE that may be responsible for its anti-inflammatory activity and thus demonstrated its potential use against acute inflammatory diseases.

Anti-rheumatic activity of Ananas comosus fruit peel extract in a complete Freund's adjuvant rat model.

CONTEXT: Rheumatoid arthritis is a chronic, autoimmune and systemic inflammatory disease, which targets synovial joints leading to joint destruction mediated in part by migration of inflammatory cells into the synovial tissue. OBJECTIVE: The present study evaluates the anti-rheumatic effect of a methanol extract of Ananas comosus (L.) Merr. (Bromeliaceae) peel in rats. MATERIALS AND METHODS: Anti-rheumatic activity of crude extract of peels of A. comosus in complete Freund's induced arthritis model in rats was studied at doses of 50, 100, 250 and 500 mg/kg b.w. for 21 days. Parameters such as paw size, levels of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), C-reactive proteins (CRP) and prostaglandins (PGE2) were analysed. RESULTS: Oral administration of the extract significantly reduced the swelling in the paw of rats (EC50 65.1 ± 2.95 mg/kg b.w.) with a maximal inhibition of 77.01 ± 10.53% on 21st day at 500 mg/kg b.w. The extract also significantly reduced the levels of SOD, CAT and GPx in liver (EC50 26.84 ± 16.37, 68.37 ± 19.22, 106.54 ± 34.81 mg/kg b.w., respectively), kidney (EC50 261.75 ± 81.5, 176.38 ± 8.08, 14.32 ± 6.64, mg/kg b.w., respectively) and spleen (EC50 152.14 ± 39.57, 83.97 ± 14.6, 47.1 ± 10.45 mg/kg b.w., respectively); and CRP (EC50 36.37 ± 12.4 mg/kg b.w.) and PGE2 (EC50 191.06 ± 71.54 mg/kg b.w.) in tissue homogenate and serum, respectively, at 500 mg/kg b.w. as compared to arthritic control group. DISCUSSION AND CONCLUSION: These results suggest that A. comosus fruit peel extract exerts anti-rheumatic activity.

Synthesis, characterization and in vitro study of biocompatible cinnamaldehyde functionalized magnetite nanoparticles (CPGF Nps) for hyperthermia and drug delivery applications in breast cancer.

Cinnamaldehyde, the bioactive component of the spice cinnamon, and its derivatives have been shown to possess anti-cancer activity against various cancer cell lines. However, its hydrophobic nature invites attention for efficient drug delivery systems that would enhance the bioavailability of cinnamaldehyde without affecting its bioactivity. Here, we report the synthesis of stable aqueous suspension of cinnamaldehyde tagged Fe3O4 nanoparticles capped with glycine and pluronic polymer (CPGF NPs) for their potential application in drug delivery and hyperthermia in breast cancer. The monodispersed superparamagnetic NPs had an average particulate size of ∼ 20 nm. TGA data revealed the drug payload of ∼ 18%. Compared to the free cinnamaldehyde, CPGF NPs reduced the viability of breast cancer cell lines, MCF7 and MDAMB231, at lower doses of cinnamaldehyde suggesting its increased bioavailability and in turn its therapeutic efficacy in the cells. Interestingly, the NPs were non-toxic to the non-cancerous HEK293 and MCF10A cell lines compared to the free cinnamaldehyde. The novelty of CPGF nanoparticulate system was that it could induce cytotoxicity in both ER/PR positive/Her2 negative (MCF7) and ER/PR negative/Her2 negative (MDAMB231) breast cancer cells, the latter being insensitive to most of the chemotherapeutic drugs. The NPs decreased the growth of the breast cancer cells in a dose-dependent manner and altered their migration through reduction in MMP-2 expression. CPGF NPs also decreased the expression of VEGF, an important oncomarker of tumor angiogenesis. They induced apoptosis in breast cancer cells through loss of mitochondrial membrane potential and activation of caspase-3. Interestingly, upon exposure to the radiofrequency waves, the NPs heated up to 41.6 °C within 1 min, suggesting their promise as a magnetic hyperthermia agent. All these findings indicate that CPGF NPs prove to be potential nano-chemotherapeutic agents in breast cancer.

Bactericidal Effect of Selected Antidiarrhoeal Medicinal Plants on Intracellular Heat-Stable Enterotoxin-Producing Escherichia coli.

Diarrhoeal diseases due to enterotoxigenic Escherichia coli continue to be a cause of global concern. Medicinal plants have been gaining popularity as promising antidiarrhoeal agents. In the present study, four antidiarrhoeal plants, viz. Aegle marmelos, Cyperus rotundus, Psidium guajava and Zingiber officinale were screened against a heat-stable toxin-producing enterotoxigenic E. coli strain. Decoctions of these plants were studied for their effect on intracellular killing of the bacterial strain using murine monocytic cell line, J774. [(3)H] thymidine release assay was used to evaluate the apoptotic/necrotic effect. All plants at concentrations <1% enhanced intracellular killing of the bacteria by J774 cells. However, at higher concentrations, the decoctions induced apoptosis in J774 cells. The study demonstrates that these plants could control diarrhoea caused by heat-stable toxin-producing enterotoxigenic E. coli through their immunomodulatory effect.

Antiplasmodial potential of extracts from two species of genus Blumea.

CONTEXT: A number of Blumea (Asteraceae) species are being used in traditional Chinese and Indian folklore medicines to cure various diseases including cancer, fungal and bacterial infections. OBJECTIVE: To evaluate the in vitro antiplasmodial potential and cytotoxicity of various extracts and fractions of B. membranacea DC and B. eriantha DC and high performance liquid chromatography (HPLC) chemical fingerprinting of their crude extracts. MATERIALS AND METHODS: The aerial parts and roots of B. membranacea and B. eriantha were extracted with ethanol and the extracts were successively partitioned with n-hexane, ethyl acetate and n-butanol, which were later evaluated for their in vitro antiplasmodial activity against Plasmodium falciparum NF-54 and in vitro cytotoxicities against non-cancerous Vero cell line. HPLC chemical fingerprinting was performed on extracts of B. membranacea and B. eriantha. RESULTS: The n-hexane (MA1), ethyl acetate (MA2) fractions of aerial parts and n-butanol (MR3) fraction of roots of B. membranacea showed IC50 values of 17.4, 19.0 and 3.3 µg/mL respectively, while the n-hexane (EA1), ethyl acetate (EA2) fractions of aerial parts and ethyl acetate (ER2) fraction of roots of B. eriantha showed IC50 values of 25.0, 26.5 and 15.6 µg/mL, respectively, against P. falciparum NF-54. All these fractions were non-toxic to Vero cells. DISCUSSION AND CONCLUSION: Both B. membranacea and B. eriantha possesses a high degree of selective antiplasmodial activity (selectivity index up to >60) and hence, may find their use in antimalarial phytopharmaceuticals as well as in discovery of a safer and novel antimalarial lead.

Novel antiplasmodial agents from Christia vespertilionis.

The roots, leaves and stems of Christia vespertilionis were separately and successively extracted with methanol and aqueous-methanol (1:4, v/v) and were evaluated in vitro for their antiplasmodial potential against Plasmodium falciparum NF-54. The aqueous-methanolic stem (AS) extract was the most active (IC50 7.5 microg/mL) followed by the methanolic leaf (ML) extract (IC50 32.0 microg/mL). The in vivo antimalarial activity of the combined plant extract of C. vespertilionis was also assessed in P. berghei infected mice, which showed 87.8% suppression of parasitaemia as compared with complete suppression by chloroquine on day 8. Finally, detailed chemical investigation of C. vespertilionis resulted in the isolation and characterization of fifteen compounds (1-15), of which two (1 and 4) are being reported for the first time from nature. The novel compound 1 possesses potent antiplasmodial activity (IC50 = 9.0 microg/mL).

Newer insights into the mechanism of action of Psidium guajava L. leaves in infectious diarrhoea.

BACKGROUND: Psidium guajava L., Myrtaceae, is used widely in traditional medicine for the treatment of diarrhoea, dysentery, gastroenteritis, stomachaches, and indigestion. However, the effect of the leaf extract of P. guajava on the pathogenesis of infectious diarrhoea has not been studied. The present study evaluates the effect of a hot aqueous extract (decoction) of dried leaves of P. guajava on parameters associated with pathogenicity of infectious diarrhoea. The aim was to understand its possible mechanism(s) of action in controlling infectious diarrhoea and compare it with quercetin, one of the most reported active constituents of P. guajava with antidiarrhoeal activity. METHODS: The crude decoction and quercetin were studied for their antibacterial activity and effect on virulence features of common diarrhoeal pathogens viz. colonization of epithelial cells and production and action of enterotoxins. Colonization as measured by adherence of enteropathogenic Escherichia coli (EPEC) and invasion of enteroinvasive E. coli (EIEC) and Shigella flexneri was assessed using HEp-2 cell line. The production of E. coli heat labile toxin (LT) and cholera toxin (CT) and their binding to ganglioside monosialic acid (GM1) were studied by GM1-ELISA whereas the production and action of E. coli heat stable toxin (ST) was assessed by suckling mouse assay. RESULTS: The decoction of P. guajava showed antibacterial activity towards S. flexneri and Vibrio cholerae. It decreased production of both LT and CT and their binding to GM1. However, it had no effect on production and action of ST. The decoction also inhibited the adherence of EPEC and invasion by both EIEC and S. flexneri to HEp-2 cells. Quercetin, on the other hand, had no antibacterial activity at the concentrations used nor did it affect any of the enterotoxins. Although it did not affect adherence of EPEC, it inhibited the invasion of both EIEC and S. flexneri to HEp-2 cells. CONCLUSION: Collectively, the results indicate that the decoction of P. guajava leaves is an effective antidiarrhoeal agent and that the entire spectrum of its antidiarrhoeal activity is not due to quercetin alone.

Studies on the antidiarrhoeal activity of Aegle marmelos unripe fruit: validating its traditional usage.

BACKGROUND: Aegle marmelos (L.) Correa has been widely used in indigenous systems of Indian medicine due to its various medicinal properties. However, despite its traditional usage as an anti-diarrhoeal there is limited information regarding its mode of action in infectious forms of diarrhoea. Hence, we evaluated the hot aqueous extract (decoction) of dried unripe fruit pulp of A. marmelos for its antimicrobial activity and effect on various aspects of pathogenicity of infectious diarrhoea. METHODS: The decoction was assessed for its antibacterial, antigiardial and antirotaviral activities. The effect of the decoction on adherence of enteropathogenic Escherichia coli and invasion of enteroinvasive E. coli and Shigella flexneri to HEp-2 cells were assessed as a measure of its effect on colonization. The effect of the decoction on production of E. coli heat labile toxin (LT) and cholera toxin (CT) and their binding to ganglioside monosialic acid receptor (GM1) were assessed by GM1-enzyme linked immuno sorbent assay whereas its effect on production and action of E. coli heat stable toxin (ST) was assessed by suckling mouse assay. RESULTS: The decoction showed cidal activity against Giardia and rotavirus whereas viability of none of the six bacterial strains tested was affected. It significantly reduced bacterial adherence to and invasion of HEp-2 cells. The extract also affected production of CT and binding of both LT and CT to GM1. However, it had no effect on ST. CONCLUSION: The decoction of the unripe fruit pulp of A. marmelos, despite having limited antimicrobial activity, affected the bacterial colonization to gut epithelium and production and action of certain enterotoxins. These observations suggest the varied possible modes of action of A. marmelos in infectious forms of diarrhoea thereby validating its mention in the ancient Indian texts and continued use by local communities for the treatment of diarrhoeal diseases.

Antimicrobial potential of Glycyrrhiza glabra roots.

The present study was aimed to investigate antimicrobial potential of Glycyrrhiza glabra roots. Antimycobacterial activity of Glycyrrhiza glabra was found at 500 microg/mL concentration. Bioactivity guided phytochemical analysis identified glabridin as potentially active against both Mycobacterium tuberculosis H(37)Ra and H(37)Rv strains at 29.16 microg/mL concentration. It exhibited antimicrobial activity against both Gram-positive and Gram-negative bacteria. Our results indicate potential use of licorice as antitubercular agent through systemic experiments and sophisticated anti-TB assay.

Studies on Pongamia pinnata (L.) Pierre leaves: understanding the mechanism(s) of action in infectious diarrhea.

While data are available on the effect of medicinal plants on intestinal motility and their antibacterial action, there is a paucity of information on their mode of action on various aspects of diarrheal pathogenicity, namely colonization to intestinal epithelial cells and production/action of enterotoxins. Crude decoction of dried leaves of Pongamia pinnata was evaluated for its antimicrobial (antibacterial, antigiardial and antirotaviral) effect; and its effect on production and action of enterotoxins (cholera toxin, CT; Escherichia coli labile toxin, LT; and E. coli stable toxin, ST); and adherence of enteropathogenic E. coli and invasion of enteroinvasive E. coli and Shigella flexneri to epithelial cells. The decoction had no antibacterial, antigiardial and antirotaviral activity, but reduced production of CT and bacterial invasion to epithelial cells. The observed results indicated that the crude decoction of P. pinnata has selective antidiarrheal action with efficacy against cholera and enteroinvasive bacterial strains causing bloody diarrheal episodes.