Phyto-therapeutic potential of Withania somnifera: Molecular mechanism and health implications.

Withania somnifera, the plant named Indian ginseng, Ashwagandha, or winter cherry, has been used since ancient times to cure various health ailments. Withania somnifera is rich in constituents belonging to chemical classes like alkaloids, saponins, flavonoids, phenolic acids, and withanolides. Several chemotypes were identified based on their phytochemical composition and credited for their multiple bioactivities. Besides, exhibiting neuroprotective, immunomodulatory, adaptogenic, anti-stress, bone health, plant has shown promising anti-cancer properties. Several withanolides have been reported to play a crucial role in cancer; they target cancer cells by different mechanisms such as modulating the expression of tumor suppressor genes, apoptosis, telomerase expression, and regulating cell signaling pathway. Though, many treatments are available for cancer; however, to date, no assured reliable cure for cancer is made available. Additionally, synthetic drugs may lead to development of resistance in time; therefore, focus on new and natural drugs for cancer therapeutics may prove a longtime effective alternative. This current report is a comprehensive combined analysis upto 2023 with articles focused on bio-activities of plant Withania somnifera from various sources, including national and international government sources. This review focuses on understanding of various mechanisms and pathways to inhibit uncontrolled cell growth by W. somnifera bioactives, as reported in literature. This review provides a recent updated status of the W. somnifera on pharmacological properties in general and anti-cancer in particular and may provide a guiding resource for researchers associated with natural product-based cancer research and healthcare management.

Identification of starvation-mimetic bioactive phytocomponent from Withania somnifera using in-silico molecular modelling and flow cytometry-based analysis for the management of malaria.

Multidrug resistance episodes in malaria increased from 3.9% to 20% from 2015 to 2019. Synchronizing the clinical manifestation in chronological sequence led to a unique impression on glucose demand (increased up to 100-fold) by the parasite-infected RBCs. Hence, restriction in the glucose uptake to parasite-infected RBCs could be an alternative approach to conquer the global burden of malaria to a greater extent. A C28 steroidal lactone Withaferin A (WS-3) isolated from Withania somnifera leave extract shows better thermodynamically stable interactions with the glucose transporters (GLUT-1 and PfHT) to standard drugs metformin and lopinavir. MD simulations for a trajectory period of 100 ns reflect stable interactions with the interactive amino acid residues such as Pro141, Gln161, Gln282, Gln283, Trp388, Phe389, and Phe40, Asn48, Phe85, His168, Gln169, Asn311 which potentiating inhibitory activity of WS-3 against GLUT-1 and PfHT respectively. WS-3 was non-hemotoxic (%hemolysis <5%) for a high concentration of up to 1 mg/ml in the physiological milieu. However, the %hemolysis significantly increased up to 30.55 ± 0.929% in a parasitophorous simulated environment (pH 5.0). Increased hemolysis of WS-3 could be due to the production of ROS in an acidic environment. Further, the inhibitory activity of WS-3 against both glucose transporters was supported with flow cytometry-based analysis of parasite-infected RBCs. Results show that WS-3 has low mean fluorescence intensities for both target proteins compared to conventional drugs, suggesting a potential sugar transporter inhibitor against GLUT-1 and PfHT for managing malaria. Communicated by Ramaswamy H. Sarma.

Revisiting the Multifaceted Therapeutic Potential of Withaferin A (WA), a Novel Steroidal Lactone, W-ferinAmax Ashwagandha, from Withania Somnifera (L) Dunal.

Withania somnifera (L.) Dunal, abundant in the Indian subcontinent as Ashwagandha or winter cherry, is a herb of unprecedented therapeutic value. The number of ailments for which crude Ashwagandha extract can be used as a preventive or curative is practically limitless; and this explains why its use has been in vogue in ancient Ayurveda since at-least about four thousand years. The therapeutic potential of Ashwagandha mainly owes from its reservoir of alkaloids (isopelletierine, anaferine), steroidal lactones (withanolides) and saponins with an extra acyl group (sitoindoside VII and VIII). Withaferin A is an exceptionally potent withanolide which is found in high concentrations in W. somnifera plant extracts. The high reactivity of Withaferin A owes to the presence of a C-28 ergostane network with multiple sites of unsaturation and differential oxygenation. It interacts with the effectors of multiple signaling pathways involved in inflammatory response, oxidative stress response, cell cycle regulation and synaptic transmission and has been found to be significantly effective in inducing programmed cell death in cancer cells, restoring cognitive health, managing diabetes, alleviating metabolic disorders, and rejuvenating the overall body homeostasis. Additionally, recent studies suggest that Withaferin A (WA) has the potential to prevent viral endocytosis by sequestering TMPRSS2, the host transmembrane protease, without altering ACE-2 expression. The scope of performing subtle structural modifications in this multi-ring compound is believed to further expand its pharmacotherapeutic horizon. Very recently, a novel, heavy metal and pesticide free formulation of Ashwagandha whole herb extract, with a significant amount of WA, termed W-ferinAmax Ashwagandha, has been developed. The present review attempts to fathom the present and future of this wonder molecule with comprehensive discussion on its therapeutic potential, safety and toxicity.Key teaching pointsWithania somnifera (L.) Dunal is a medicinal plant with versatile therapeutic values.The therapeutic potential of the plant owes to the presence of withanolides such as Withaferin A.Withaferin A is a C-28 ergostane based triterpenoid with multiple reactive sites of therapeutic potential.It is effective against a broad spectrum of ailments including neurodegenerative disorders, cancer, inflammatory and oxidative stress disorders and it also promotes cardiovascular and sexual health.W-ferinAmax Ashwagandha, is a heavy metal and pesticide free Ashwagandha whole herb extract based formulation with significant amount of Withaferin A.

Clinical pharmacokinetic evaluation of Withania somnifera (L.) Dunal root extract in healthy human volunteers: A non-randomized, single dose study utilizing UHPLC-MS/MS analysis.

ETHNOPHARMACOLOGICAL RELEVANCE: Withania somnifera (L.) Dunal; (Solanaceae), commonly known as Ashwagandha, is one of the most significant medicinal herbs in 'Ayurveda', a traditional Indian medicine used for centuries with evidence in scriptures. Ashwagandha was mentioned in old Ayurvedic medical literature such as Charaka Samhita and Sushruta Samhita for improving weight and strength, with multiple citations for internal and exterior usage in emaciation and nourishing the body. Ethnopharmacological evidence revealed that it was used to relieve inflammation, reduce abdominal swelling, as a mild purgative, and treat swollen glands. The root was regarded as a tonic, aphrodisiac, and emmenagogue in the Unani tradition of the Indian medicinal system. Further, Ashwagandha has been also described as an Ayurvedic medicinal plant in the Ayurvedic Pharmacopoeia of India extending informed therapeutic usage and formulations. Despite the widespread ethnopharmacological usage of Ashwagandha, clinical pharmacokinetic parameters are lacking in the literature; hence, the findings of this study will be relevant for calculating doses for future clinical evaluations of Ashwagandha root extract. AIM: This study aimed to develop a validated and highly sensitive bioanalytical method for quantifying withanosides and withanolides of the Ashwagandha root extract in human plasma to explore its bioaccessibility. Further to apply a developed method to perform pharmacokinetics of standardized Withania somnifera (L.) Dunal root extract (WSE; AgeVel®/Witholytin®) capsules in healthy human volunteers. METHODS: A sensitive, reliable, and specific ultra-high pressure liquid chromatography-mass spectrometry (UHPLC-MS/MS) method was developed and validated for the simultaneous quantification of five major withanosides and withanolides (withanoside IV, withanoside V, withanolide A, withaferin A, and 12-deoxy-withastramonolide) in human plasma. Further for the study, eighteen healthy male volunteers (18-45 years) were enrolled in a non-randomized, open-label, single period, single treatment, clinical pharmacokinetic study and given a single dose (500 mg) of WSE (AgeVel®/Witholytin®) capsules containing not less than 7.5 mg of total withanolides under fasting condition. Later, pharmacokinetic profiles were assessed using the plasma concentration of each bioactive constituent Vs. time data. RESULTS: For all five constituents, the bioanalytical method demonstrated high selectivity, specificity, and linearity. There was no carryover, and no matrix effect was observed. Furthermore, the inter-day and intra-day precision and accuracy results fulfilled the acceptance criteria. Upon oral administration of WSE capsules, Cmax was found to be 0.639 ± 0.211, 2.926 ± 1.317, 2.833 ± 0.981, and 5.498 ± 1.986 ng/mL for withanoside IV, withanolide A, withaferin A, and 12-deoxy-withastramonolide with Tmax of 1.639 ± 0.993, 1.361 ± 0.850, 0.903 ± 0.273, and 1.375 ± 0.510 h respectively. Further, withanoside V was also detected in plasma; but its concentration was found below LLOQ. CONCLUSION: The novel and first-time developed bioanalytical method was successfully applied for the quantification of five bio-active constituents in human volunteers following administration of WSE capsules, indicating that withanosides and withanolides were rapidly absorbed from the stomach, have high oral bioavailability, and an optimum half-life to produce significant pharmacological activity. Further, AgeVel®/Witholytin® was found safe and well tolerated after oral administration, with no adverse reaction observed at a 500 mg dose.

Withania somnifera extracts induced attenuation of HIV-1: a mechanistic approach to restrict viral infection.

BACKGROUND: Several anti-retroviral drugs are available against Human immunodeficiency virus type-1, but have multiple adverse side effects. Hence, there is an incessant compulsion for effectual anti-retroviral agents with minimal or no intricacy. Traditionally, natural products have been the most successful source for the development of new medications. Withania somnifera, also known as Ashwagandha, is the utmost treasured medicinal plant used in Ayurveda, which holds the potential to give adaptogenic, immunomodulatory, and antiviral effects. However, its effect on HIV-1 replication at the cellular level has never been explored. Herein, we focused on the anti-HIV-1 activity and the probable mechanism of action of hydroalcoholic and aqueous extracts of Withania somnifera roots and its phytomolecules. METHODS: The cytotoxicity of the extracts was determined through MTT assay, while the in vitro anti-HIV-1 activity was assessed in TZM-bl cells against the HIV-1 strains of X4 and R5 subtypes. Results were confirmed in peripheral blood mononuclear cells, using the HIV-1 p24 antigen assay. Additionally, the mechanism of action was determined through the Time of Addition assay, which was further validated through the series of enzymatic assays, i.e. HIV-1 Integrase, Reverse transcriptase, and Protease assays. To explore the role of the identified active metabolites of Withania somnifera in antiretroviral activity, molecular docking analyses were performed against these key HIV-1 replication enzymes. RESULTS: The hydroalcoholic and aqueous extracts of Withania somnifera roots were found to be safer at the sub-cytotoxic concentrations and exhibited their ability to inhibit replication of two primary isolates of HIV-1 through cell-associated and cell-free assays, in dose-dependent kinetics. Several active phytomolecules found in Withania somnifera successfully established hydrogens bonds in the active binding pocket site residues responsible for the catalytic activity of HIV replication and therefore, signifying their role in the attenuation of HIV-1 infection as implied through the in silico molecular docking studies. CONCLUSIONS: Our research identified both the hydroalcoholic and aqueous extracts of Withania somnifera roots as potent inhibitors of HIV-1 infection. The in silico analyses also indicated the key components of Withania somnifera with the highest binding affinity against the HIV-1 Integrase by 12-Deoxywithastramonolide and 27-Hydroxywithanone, HIV-1 Protease by Ashwagandhanolide and Withacoagin, and HIV-1 Reverse transcriptase by Ashwagandhanolide and Withanolide B, thereby showing possible mechanisms of HIV-1 extenuation. Overall, this study classified the role of Withania somnifera extracts and their active compounds as potential agents against HIV-1 infection.

A critical assessment of the whole plant-based phytotherapeutics from Withania somnifera (L.) Dunal with respect to safety and efficacy vis-a-vis leaf or root extract-based formulation.

Withania somnifera (L.) Dunal, popularly known as Ashwagandha or Indian ginseng, is well acclaimed for its health-enhancing effects, including its potent immunomodulatory, anti-inflammatory, neuroprotective, and anti-tumorigenic properties. The prime biological effectors of these attributes are a diverse group of ergostane-based steroidal lactones termed withanolides. Withanones and withanosides are distributed differentially across the plant body, whereas withanolides and withanones are known to be more abundant in leaves, while withanosides are found exclusively in the roots of the plants. Standardized W. somnifera extract is Generally Recognized as Safe (GRAS)-affirmed, however, moderate to severe toxic manifestations may occur at high dosages. Withaferin A, which also happens to be the primary bioactive ingredient for the effectiveness of this plant. There have been contrasting reports regarding the distribution of withaferin A in W. somnifera. While most reports state that the roots of the plant have the highest concentrations of this phytochemical, several others have indicated that leaves can accumulate withaferin A in proportionately higher amounts. A comprehensive survey of the available reports suggests that the biological effects of Ashwagandha are grossly synergistic in nature, with many withanolides together mediating the desired physiological effect. In addition, an assorted formulation of withanolides can also neutralize the toxic effects (if any) associated with withaferin A. This mini-review presents a fresh take on the recent developments regarding the safety and toxicity of the plant, along with a critical assessment of the use of roots against leaves as well as whole plants to develop therapeutic formulations. Going by the currently available scientific evidence, it is safe to infer that the use of whole plant formulations instead of exclusively root or leaf recipes may present the best possible option for further exploration of therapeutic benefits from this novel medicinal plant.HighlightsTherapeutic potential of withanolides owes to the presence of α,ß unsaturated ketone which binds to amines, alcohols, and esters and 5ß, 6ß epoxy group which react with side chain thiols of proteins.At concentrations above NOAEL (no observed adverse effect level), the same mechanisms contribute towards toxicity of the molecule.Although withanosides are found exclusively in roots, whole plants have higher contents of withanones and withanolides.Whole plant-based formulations have other metabolites which can nullify the toxicity associated with roots.Extracts made from whole plants, therefore can holistically impart all therapeutic benefits as well as mitigate toxicity.

Simultaneous estimation of five biomarkers of neuroprotective herb Ashwagandha NMITLI-118R AF1 in rat plasma and brain using LC-ESI-MS/MS: Application to its pharmacokinetic and stability studies.

Withania Somnifera (WS) is a popular nutritional supplement in the USA, Europe, and Asia, known for its pharmacological effects on neurological disorders. However, the bioanalytical method development, validation, and pharmacokinetics of WS NMITLI-118R AF1 biomarkers Withanolide A (WLD A), Withanone (WNONE), Withanolide B (WLD B), Withaferin A (WF A), and 12 Deoxywithastramonolide (12 DEOXY) in rats have not been comprehensively explored. This study aimed to develop and validate a sensitive and selective LC-ESI-MS/MS method for these biomarkers in male Sprague Dawley rats plasma and brain matrix. Rats were divided into eight groups, each containing five rats. A plant extract of NMITLI-118R AF1 at 50 mg/kg was orally administered to the rats for in-vivo pharmacokinetic investigation. All the analytes had a linear calibration curve (r2 > 0.999), and intra-day and inter-day precision (%) were found in the range of 2.46 - 13.71% and accuracy were within the acceptable range (±15%). The biomarkers of NMITLI-118R AF1 were found stable in in-vitro plasma and simulated gastro-intestinal fluids. The observed (Cmax) and (Tmax) values for the biomarkers in the systemic circulation were WLD A (5.59 ± 0.34 ng/mL, Tmax 1.00 ± 0.00 h), WNONE (6.28 ± 0.41 ng/mL, Tmax 0.95 ± 0.11 h), WLD B (6.45 ± 2.87 ng/mL, Tmax 0.95 ± 0.11 h), WF A (6.50 ± 0.27 ng/mL, Tmax 1.00 ± 0.00 h), and 12 DEOXY (5.68 ± 0.39 ng/mL, Tmax 1.00 ± 0.00 h). In contrast to the old method, our approach exhibits a lower limit of quantification (LLOQ), shorter run time (less than10 min), and enables the detection of WF A and WNONE in fresh rat plasma by other quantitative analysis of mass spectrometry (m/z) [M]+. Shows high sample volumes for both, larger plasma volumes, costlier sample collection techniques dried blood spot (DBS), more expensive solid phase extraction techniques (SPE) and longer analysis time 14 min. Moreover, our method requires a smaller sample volume 10 µL, offers faster analysis time 4 min, and achieves a higher sensitivity 1 ng/mL. This is the first report of a comprehensive study on in-vitro and in-vivo pharmacokinetics of NMITLI-118R AF1 biomarkers, which may aid in further pre-clinical and clinical trial investigations.

Sleep-promoting activity of amylase-treated Ashwagandha (Withania somnifera L. Dunal) root extract via GABA receptors.

Ashwagandha (Withania somnifera L. Dunal), an Indian medicinal plant that has been used for centuries to treat insomnia, exhibits a variety of biological activities, such as improving cognitive function, immunity and anxiety. In this study, the effect of enzyme-treated Ashwagandha root extract (EA) and on sleep was evaluated using rodent models. Starch contained in the Ashwagandha root extract was removed by amylase treatment to prepare EA. To evaluate the sleep-promoting activity of EA, a pentobarbital-induced sleep test and electroencephalogram analysis were performed. In addition, the sleep-promoting mechanism of EA was elucidated by analyzing the expression of sleep-related receptors. In the pentobarbital-induced sleep test, EA dose-dependently increased sleep duration. Additionally, electroencephalogram analysis revealed that EA significantly increased δ-wave and non-rapid eye movement sleep times, which are involved in deep sleep, thereby improving sleep quality and quantity. EA also effectively relieved caffeine-induced insomnia symptoms. Furthermore, the γ-aminobutyric acid (GABA) content in the brain and mRNA and protein expression of GABAA, GABAB1, and serotonin receptors were significantly increased by EA compared to the normal group. In particular, EA showed sleep-promoting activity by binding to various GABAA receptor sites. Collectively, EA exhibited sleep-promoting activity through the GABAergic system and may be used as a functional material to improve sleep deprivation.

Molecular Mechanism behind the Safe Immunostimulatory Effect of Withania somnifera.

Withania somnifera (L.) Dunal (family Solanaceae) is a medicinal plant known for, among many pharmacological properties, an immune boosting effect. Our recent study revealed that its key immunostimulatory factor is lipopolysaccharide of plant-associated bacteria. This is peculiar, because, although LPS can elicit protective immunity, it is an extremely potent pro-inflammatory toxin (endotoxin). However, W. somnifera is not associated with such toxicity. In fact, despite the presence of LPS, it does not trigger massive inflammatory responses in macrophages. To gain insights into the safe immunostimulatory effect of W. somnifera, we conducted a mechanistic study on its major phytochemical constituent, withaferin A, which is known for anti-inflammatory activity. Endotoxin-triggered immunological responses in the presence and absence of withaferin A were characterized by both in vitro macrophage-based assay and in vivo cytokine profiling in mice. Collectively, our results demonstrate that withaferin A selectively attenuates the pro-inflammatory signaling triggered by endotoxin without impairing other immunological pathways. This finding provides a new conceptual framework to understand the safe immune-boosting effect of W. somnifera and possibly other medicinal plants. Furthermore, the finding opens a new opportunity to facilitate the development of safe immunotherapeutic agents, such as vaccine adjuvants.

Three-Way Cell-Based Screening of Antistress Compounds: Identification, Validation, and Relevance to Old-Age-Related Pathologies.

A variety of environmental stress stimuli have been linked to poor quality of life, tissue dysfunctions, and ailments including metabolic disorders, cognitive impairment, and accelerated aging. Oxidative, metal, and hypoxia stresses are largely associated with these phenotypes. Whereas drug development and disease therapeutics have advanced remarkably in last 3 decades, there are still limited options for stress management. Because the latter can effectively decrease the disease burden, we performed cell-based screening of antistress compounds by recruiting 3 chemical models of oxidative (paraquat), metal (cadmium nitrate), or hypoxia (cobalt chloride) stresses. The screening of 70 compounds for their ability to offer protection against oxidative, metal, and hypoxia stresses resulted in the selection of 5 compounds: Withaferin-A (Wi-A), methoxy Withaferin-A (mWi-A), Withanone (Wi-N), triethylene glycol (TEG), and Ashwagandha (Withania somnifera) leaf M2-DMSO extract (M2DM). Molecular assays revealed that whereas stress caused increase in (a) apoptosis, (b) reactive oxygen species accumulation coupled with mitochondrial depolarization, (c) DNA double-strand breaks, and (d) protein aggregation, low nontoxic doses of the selected compounds caused considerable protection. Furthermore, Wi-N, TEG, and their mixture-treated normal human fibroblasts (at young, mature, and senescent stages representing progressively increasing accumulation of stress) showed increase in proliferation. Taken together, these results suggested 3-way (oxidative, metal, and hypoxia) antistress potential of Wi-N and TEG that may be useful for management of environmental and old-age-related pathologies.

In Silico Study on the Interactions, Molecular Docking, Dynamics and Simulation of Potential Compounds from Withania somnifera (L.) Dunal Root against Cancer by Targeting KAT6A.

Cancer is characterized by the abnormal development of cells that divide in an uncontrolled manner and further take over the body and destroy the normal cells of the body. Although several therapies are practiced, the demand and need for new therapeutic agents are ever-increasing because of issues with the safety, efficacy and efficiency of old drugs. Several plant-based therapeutics are being used for treatment, either as conjugates with existing drugs or as standalone formulations. Withania somnifera (L.) Dunal is a highly studied medicinal plant which is known to possess immunomodulatory activity as well as anticancer properties. The pivotal role of KAT6A in major cellular pathways and its oncogenic nature make it an important target in cancer treatment. Based on the literature and curated datasets, twenty-six compounds from the root of W. somnifera and a standard inhibitor were docked with the target KAT6A using Autodock vina. The compounds and the inhibitor complexes were subjected to molecular dynamics simulation (50 ns) using Desmond to understand the stability and interactions. The top compounds (based on the docking score of less than -8.5 kcal/mol) were evaluated in comparison to the inhibitor. Based on interactions at ARG655, LEU686, GLN760, ARG660, LEU689 and LYS763 amino acids with the inhibitor WM-8014, the compounds from W. somnifera were evaluated. Withanolide D, Withasomniferol C, Withanolide E, 27-Hydroxywithanone, Withanolide G, Withasomniferol B and Sitoindoside IX showed high stability with the residues of interest. The cell viability of human breast cancer MCF-7 cells was evaluated by treating them with W. Somnifera root extract using an MTT assay, which showed inhibitory activity with an IC50 value of 45 µg/mL. The data from the study support the traditional practice of W. somnifera as an anticancer herb.

Biodiversity, Biochemical Profiling, and Pharmaco-Commercial Applications of Withania somnifera: A Review.

Withania somnifera L. Dunal (Ashwagandha), a key medicinal plant native to India, is used globally to manage various ailments. This review focuses on the traditional uses, botany, phytochemistry, and pharmacological advances of its plant-derived constituents. It has been reported that at least 62 crucial and 48 inferior primary and secondary metabolites are present in the W. somnifera leaves, and 29 among these found in its roots and leaves are chiefly steroidal compounds, steroidal lactones, alkaloids, amino acids, etc. In addition, the whole shrub parts possess various medicinal activities such as anti-leukotriene, antineoplastic, analgesic, anti-oxidant, immunostimulatory, and rejuvenating properties, mainly observed by in vitro demonstration. However, the course of its medical use remains unknown. This review provides a comprehensive understanding of W. somnifera, which will be useful for mechanism studies and potential medical applications of W. somnifera, as well as for the development of a rational quality control system for W. somnifera as a therapeutic material in the future.

Insights into the potential of withanolides as Phosphodiesterase-4 (PDE4D) inhibitors.

Medicinal herbs have been used as traditional medicines for centuries. The molecular mechanism of action of their bioactive molecules against various diseases or therapeutic targets is still being explored. Here, the active compounds (withanolides) of a well-known Indian medicinal herb, Ashwagandha (Withania somnifera), have been studied for their most potential therapeutic targets and their mechanism of action using ligand-based screening and receptor-based approaches. Ligand-based screening predicted the six top therapeutic targets, namely, Protein kinase C alpha (PRKCA), Protein kinase C delta (PRKCD), Protein kinase C epsilon (PRKCE), Androgenic Receptor (AR), Cycloxygenase-2 (PTGS-2) and Phosphodiesterase-4D (PDE4D). Further, when these predictions were validated using receptor-based studies, i.e. molecular docking, molecular dynamics simulation and free energy calculations, it was found that PDE4D was the most potent target for four withanolides, namely, Withaferin-A, 17-Hydroxywithaferin-A, 27-Hydroxywithanone and Withanolide-R. These compounds had a better binding affinity and similar interactions as that of an already known inhibitor (Zardaverine) of PDE4D. These results warrant further in-vitro and in-vivo investigations to examine their therapeutic potential as an inhibitor of PDE4D.Communicated by Ramaswamy H. Sarma.

DPP-4 inhibition mediated antidiabetic potential of phytoconstituents of an aqueous fruit extract of Withania coagulans (Stocks) Dunal: in-silico, in-vitro and in-vivo assessments.

The DPP-4 inhibition is an interesting target for the development of antidiabetic agents which promotes the longevity of GPL-1(Glucagon-like peptide 1). The current study was intended to assess DPP-4(Dipeptidyl Peptidase-4) inhibition mediated antidiabetic effect of phytocompounds of an aqueous fruit extract of Withania coagulans (Stocks) Dunal by in-vitro, in-silico and in-vivo approaches. The phytoconstituents screening was executed by LCMS (Liquid Chromatography with tandem mass spectrometry). The in-vitro and in-vivo, DPP-4 assays were performed by using available kits. The in-vitro DPP-4 activity was inhibited up to 68.3% by the test extract. Accordingly, in-silico determinations of molecular docking, molecular dynamics and pharmacokinetics were performed between the target enzyme DPP-4 and leading phytocompounds. The molecular dynamics authenticated the molecular docking data by crucial parameters of cytosolic milieu by the potential energy, RSMD (Root Mean Square Deviation), RSMF (Root Mean Square Fluctuation), system density, NVT (Number of particles at fixed volume, ensemble) and NPT (Number of particles at fixed pressure, ensemble). Accordingly, ADMET predictions assessed the druggability profile. Subsequently, the course of the test extract and the sitagliptin (positive control), instigated significant (p ≤ 0.001) ameliorations in HOMA indices and the equal of antioxidants in nicotinamide-streptozotocin induced type 2 diabetic animal model. Compassionately, the histopathology represented increased pancreatic cellular mass which caused in restoration of histoarchitectures. It has been concluded that phytoconstituents in W. coagulans aqueous fruit extract can regulate DPP-4, resulting in improved glucose homeostasis and enhanced endocrinal pancreatic cellular mass.Communicated by Ramaswamy H. Sarma.

Phytochemistry, allelopathy and anticancer potentiality of Withania somnifera (L.) Dunal (Solanaceae).

Withania somnifera is a wild plant that shows great activity and safety against several human diseases. The current research explored the plant's chemical composition and allelopathic effects on Rumex dentatus (recipient plant). Moreover, anticancer activity is also tested against four types of human cancer cell lines. Chemical analysis of W. somnifera showed a high percentage of saponins and tannins, while glycosides, alkaloids, and flavonoids occurred in the second order. Results of the allelopathic experiments revealed significant inhibition of the R. dentatus plumule and radicle lengths as well as their relative dry weights. In addition, significant reductions in some primary metabolites of R. dentatus, like non-reducing and total sugar as well as soluble proteins, were determined. Cytotoxic potentiality of W. somnifera was also proved against four different cancer lines, namely; human hepatocellular carcinoma cell line (HepG2), human non-small cell lung cancer cell line (A549), human breast cancer cell line (MCF7), and colon cancer cell line (CaCo2) with IC50 value of about 38, 19, 27, and 24 𝜇g/ml, respectively.

Characterization and anticancer potential of Withania somnifera fruit bioactives (a native species to Pakistan) using gas chromatography-mass spectrometer, nuclear magnetic resonance and liquid chromatography-mass spectrometry-electrospray ionization.

INTRODUCTION: Withania somnifera (W. somnifera) is a plant with remarkable pharmacological properties. The plant has an impressive profile of medicinal uses in the folk medicine system of several civilizations. AIM: This comprehensive study is aimed to characterize phytochemicals in fruit of W. somnifera and tested for anticancer potential to find out active candidate in disease prevention and treatment. METHODS: The bioactive components from W. somn-ifera fruit were extracted with polar and non-polar solvents. Anticancer potential of the isolated bioactive was assessed against different cancer cell lines through MTT assay and Incucytes imaging analysis. The extracts were characterized for secondary metabolites using GC-MS (gas chromatography-mass spectrometer), LCMS (liquid chromatography-mass spectrometry)-ESI (electrospray Ionization) and 1H-NMR (electrospray Ionization) techniques. RESULTS: Both freeze-dried and rotary evaporator con-densed extracts exhibited anticancer potential against MDA-MB-231, MCF7- SKOV3 and SKBR3 cell lines. The tested extracts have cell growth inhibition potential ag-ainst mammalian cancer cell line. Hexacosanedioic acid purified from -hexane extract through HPLC was inves-tigated for its cytotoxicity against breast cancer cell line SKBR3 by using Incucytes imaging analysis. CONCLUSION: We found that a variety of bioactive compounds existed in this plant. One identified compound that was not investigated for cytotoxicity in previous studies was purified and its application showed cytotoxicity on breast cancer cell lines. A number of bioactive identified from fruit may have an effective potential for development into chemotherapy drugs.

Promising Antioxidant and Antimicrobial Potencies of Chemically-Profiled Extract from Withania aristata (Aiton) Pauquy against Clinically-Pathogenic Microbial Strains.

Withania aristata (Aiton) Pauquy, a medicinal plant endemic to North African Sahara, is widely employed in traditional herbal pharmacotherapy. In the present study, the chemical composition, antioxidant, antibacterial, and antifungal potencies of extract from the roots of Withania aristata (Aiton) Pauquy (RWA) against drug-resistant microbes were investigated. Briefly, RWA was obtained by maceration with hydro-ethanol and its compounds were identified by use of high-performance liquid chromatography (HPLC). The antioxidant activity of RWA was determined by use of ferric-reducing antioxidant power (FRAP), 1,1-diphenyl-2-picrylhydrazyl (DPPH), and total antioxidant capacity (TAC). The evaluation of the antimicrobial potential of RWA was performed against drug-resistant pathogenic microbial strains of clinical importance by use of the disc diffusion agar and microdilution assays. Seven compounds were identified in RWA according to HPLC analysis, including cichoric acid, caffeic acid, apigenin, epicatechin, luteolin, quercetin, and p-catechic acid. RWA had excellent antioxidant potency with calculated values of 14.0 ± 0.8 µg/mL (DPPH), 0.37 ± 0.08 mg/mL (FRAP), 760 ± 10 mg AAE/g (TAC), and 81.4% (ß-carotene). RWA demonstrated good antibacterial potential against both Gram-negative and Gram-positive bacteria, with inhibition zone diameters ranging from 15.24 ± 1.31 to 19.51 ± 0.74 mm, while all antibiotics used as drug references were infective, except for Oxacillin against S. aureus. Results of the minimum inhibitory concentration (MIC) assay against bacteria showed that RWA had MIC values ranging from 2.13 to 4.83 mg/mL compared to drug references, which had values ranging from 0.031 ± 0.003 to 0.064 ± 0.009 mg/mL. Similarly, respectable antifungal potency was recorded against the fungal strains with inhibition zone diameters ranging from 25.65 ± 1.14 to 29.00 ± 1.51 mm compared to Fluconazole, used as a drug reference, which had values ranging from 31.69 ± 1.92 to 37.74 ± 1.34 mg/mL. Results of MIC assays against fungi showed that RWA had MIC values ranging from 2.84 ± 0.61 to 5.71 ± 0.54 mg/mL compared to drug references, which had values ranging from 2.52 ± 0.03 to 3.21 ± 0.04 mg/mL. According to these outcomes, RWA is considered a promising source of chemical compounds with potent biological properties that can be beneficial as natural antioxidants and formulate a valuable weapon in the fight against a broad spectrum of pathogenic microbes.

Withania somnifera (L.) Dunal whole-plant extracts exhibited anti-sporotrichotic effects by destabilizing peripheral integrity of Sporothrix globosa yeast cells.

Chronic topical cases of Sporotrichosis, a chronic fungal infection caused by the ubiquitously present cryptic members of the Sporothrix species complex, are treated with oral administrations of itraconazole. However, severe pulmonary or disseminated cases require repeated intra-venous doses of amphotericin B or even surgical debridement of the infected tissue. The unavoidable adverse side-effects of the current treatments, besides the growing drug resistance among Sporothrix genus, demands exploration of alternative therapeutic options. Medicinal herbs, due to their multi-targeting capacity, are gaining popularity amidst the rising antimicrobial recalcitrance. Withania somnifera is a well-known medicinal herb with reported antifungal activities against several pathogenic fungal genera. In this study, the antifungal effect of the whole plant extract of W. somnifera (WSWE) has been explored for the first time, against an itraconazole resistant strain of S. globosa. WSWE treatment inhibited S. globosa yeast form growth in a dose-dependent manner, with IC50 of 1.40 mg/ml. Minimum fungicidal concentration (MFC) was found to be 50 mg/ml. Sorbitol protection and ergosterol binding assays, revealed that anti-sporotrichotic effects of WSWE correlated well with the destabilization of the fungal cell wall and cell membrane. This observation was validated through dose-dependent decrease in overall ergosterol contents in WSWE-treated S. globosa cells. Compositional analysis of WSWE through high performance liquid chromatography (HPLC) exhibited the presence of several anti-microbial phytochemicals like withanone, withaferin A, withanolides A and B, and withanoside IV and V. Withanone and withaferin A, purified from WSWE, were 10-20 folds more potent against S. globosa than WSWE, thus, suggesting to be the major phytocompounds responsible for the observed anti-sporotrichotic activity. In conclusion, this study has demonstrated the anti-sporotrichotic property of the whole plant extract of W. somnifera against S. globosa that could be further explored for the development of a natural antifungal agent against chronic Sporotrichosis.

Withania somnifera - a magic plant targeting multiple pathways in cancer related inflammation.

BACKGROUND: Deregulated inflammatory responses are known to play a pivotal role in cancer initiation and progression. Tumor microenvironment is associated with the presence of a diverse array of inflammatory reactions, which further help tumor growth, metastasis and drug resistance. Withania somnifera is known to curb proliferation of cancer cells and lower inflammatory responses. PURPOSE: In order to minimize the inflammation, cancer treatments often include immunomodulatory drugs. However, given the side effects of both of the cytotoxic cancer drugs and synthetic immunomodulatory agents, there is a need to develop novel anti-inflammatory agents for improved cancer therapy. A number of reports indicate that bioactive phytochemicals derived from W. somnifera exhibit anti-inflammatory capabilities in cancer. A deeper look into the underlying molecular mechanisms implicated in W. somnifera mediated anti inflammation is lacking, which is essential to fully understand the potential of this magical plant in cancer. Therefore, in the present review we are summarizing various reports, which describe mechanistic understanding of W. somnifera in cancer related inflammation. STUDY DESIGN AND METHODOLOGY: In order to gather information on the molecular pathways affected by W. somnifera in cancer related inflammation, 'PubMed' and 'Science Direct' databases were searched using keywords Withania, cancer inflammation, and Withaferin A. Selected literature was analyzed to cover the role of inflammation in cancer, usage and side effects of anti-inflammatory drugs, W. somnifera as an immunomodulatory agent in cancer, molecular pathways modulated by W. somnifera in various preclinical models, and clinical trials using W. somnifera as an anti-inflammatory agent. RESULTS: Upon literature survey we found that both W. somnifera extracts and Withaferin-A, exhibit anti inflammatory activities in various preclinical cancer models. W. somnifera modulates a number of signaling pathways such as NF-kB, JAK-STAT and AP1 to reduce cancer related inflammation. Anti inflammatory properties of W. somnifera might be effective in the treatment of drug resistance in cancers. Based on its promising effects against cancer associated inflammation in preclinical studies, W. somnifera derived products are being tested in clinical trials. CONCLUSION: Several preclinical studies demonstrated anti-inflammatory potential of W. somnifera in a variety of cancers. While a few clinical trials are investigating the role of W. somnifera in various diseases, focused studies on its role in cancer related inflammation are lacking. Additionally, its anti-inflammatory effects offer targeting of senescence associated secretory phenotype (SASP), which is speculated to play a critical role in chemoresistance. Apart from targeting cancer cell proliferation, anti-inflammatory effects of Withania provide double advantage in cancer management. Therefore, clinical trials to target cancer related inflammation using W. somnifera as a drug, should be performed to validate its advantages in cancer therapy.

Unveiling the effect of Withania somnifera on neuronal cytoarchitecture and synaptogenesis: A combined in vitro and network pharmacology approach.

Withania somnifera (WS), is known for its remarkable contribution in herbal medicine and Ayurveda, which is therapeutically applied to improve memory and anxiety in patients. However, the pharmacological details of this plant on memory boosting yet remained undefined. This study provides mechanistic insights on the effect of ethanol solution extract of the whole plant of WS (WSEE) on neuritogenesis by combining in vitro and in silico network pharmacology approaches. WSEE promoted significant neuronal growth through early differentiation, axodendritic arborization, and synaptogenesis on primary hippocampal neurons. The network pharmacological study confirmed that the neuritogenic activity is potentially mediated by modulating the neurotrophin signaling pathway, where NRTK1 (TrkA) was revealed as the primary target of WS secondary metabolites. This neurotrophic activity of WSEE was significantly stifled by the presence of TrkA inhibitor, which further confirms the TrkA-dependent activity of WSEE. In addition, a molecular docking study suggested steroidal lactones present in the WS might act as nerve growth factor (NGF)-mimetics, activating TrkA by binding to the NGF-binding domain. As a whole, the findings of the study suggest a significant role of WSEE on neuritogenesis and its potential to function as a therapeutic agent and in drug designing for the prevention and treatment of memory-related neurological disorders.