Chemical, microbial and safety profiling of a standardized Withania somnifera (Ashwagandha) extract and Withaferin A, a potent novel phytotherapeutic of the millennium

Background:Withania somnifera (L.) Dunal, popularly known as Ashwagandha, is an ethnomedicinal plant with multiple pharmacotherapeutic applications. The diverse medicinal properties of the plant are largely due to the presence of withanolides, a group of C28 ergostane based steroidal lactones, with several sites of unsaturation and oxygenation. Withaferin A, a major with anolide present in Ashwagandha plant accounts for its emerging new roles to treat cancer, arthritis, inflammatory responses, immunomodulatory properties, and neuronal disorders. The root and leaf extracts are specifically important constituent materials for the development of phytotherapeutics, mostly intended for oral consumption. Several studies have been carried out to delineate the toxic manifestations of the extract for human consumption. Objective:Establish the broad-spectrum safety of W-ferinAmax ashwagandha (WFA). Study: This investigation demonstrated a novel, standardized W-ferinAmax ashwagandha (WFA) extraction technology from the whole herb of Withania sominfera, conducted HPLC analysis to identify the constituents, detected the heavy metals, microbiological contaminants, pesticides contaminants, and safety profile. Results:A novel extraction technology was employed to obtain WFA from the whole plant of Withania sominfera. HPLC analysis revealed that WFA contains a total of 15.4% Withanolides. In particular, Withaferin A, Withanoside IV, and Withanolide A contents were 6.469%, 1.022%, and 0.073%, respectively. The extract contained only 0.403 ppm of heavy metals out of which traces of arsenic, mercury and lead were detected, and cadmium was absent. USP recommended 80 residue basic pesticide screen indicated that the extraction was safe for human consumption. It was also found to be free from pathogenic microbes as assessed by the absence of E. coli and other coliforms, Salmonella and Staphylococcus species. Conclusion: The data generated cumulatively indicated that WFA is safe for further downstream processing to and for human consumption

Molecular design of the γδT cell receptor ectodomain encodes biologically fit ligand recognition in the absence of mechanosensing

High-acuity αβT cell receptor (TCR) recognition of peptides bound to major histocompatibility complex molecules (pMHCs) requires mechanosensing, a process whereby piconewton (pN) bioforces exert physical load on αβTCR–pMHC bonds to dynamically alter their lifetimes and foster digital sensitivity cellular signaling. While mechanotransduction is operative for both αβTCRs and pre-TCRs within the αβT lineage, its role in γδT cells is unknown. Here, we show that the human DP10.7 γδTCR specific for the sulfoglycolipid sulfatide bound to CD1d only sustains a significant load and undergoes force-induced structural transitions when the binding interface-distal γδ constant domain (C) module is replaced with that of αβ. The chimeric γδ–αβTCR also signals more robustly than does the wild-type (WT) γδTCR, as revealed by RNA-sequencing (RNA-seq) analysis of TCR-transduced Rag2−/− thymocytes, consistent with structural, single-molecule, and molecular dynamics studies reflective of γδTCRs as mediating recognition via a more canonical immunoglobulin-like receptor interaction. Absence of robust, force-related catch bonds, as well as γδTCR structural transitions, implies that γδT cells do not use mechanosensing for ligand recognition. This distinction is consonant with the fact that their innate-type ligands, including markers of cellular stress, are expressed at a high copy number relative to the sparse pMHC ligands of αβT cells arrayed on activating target cells. We posit that mechanosensing emerged over ∼200 million years of vertebrate evolution to fulfill indispensable adaptive immune recognition requirements for pMHC in the αβT cell lineage that are unnecessary for the γδT cell lineage mechanism of non-pMHC ligand detection

Utilization of pineapple leaf agro-waste for extraction of fibre and the residual biomass for vermicomposting

172-177A special type of machine with metal knife scrapper roller and serrated roller has been developed and used to scrap out the waxy layer and at the same time macerating and breaking the leaf surface for ease of retting to extract the pineapple leaf fibres. Pineapple leaf contains 2.5-3.5% strong, white and silky textile grade fibre embedded by a top waxy layer within the leaf. After removing the top waxy layer, fibre has been extracted from the pineapple leaf by retting in water. The residual green sludge has been used for vermicomposting after appropriate treatment using earthworm species African night crawler (Eudrilus eugeniae) as inoculums. The vermicomposting process was complete within 45 days. This vermicompost from pineapple leaf agro-waste is found to be rich in plant nutrients. The combined technology package for the extraction of fibre from pineapple leaf and utilization of the residual biomass debris from the pineapple leaf scratching machine for vermicomposting is economically viable and remunerative for the pineapple cultivators

Unveiling the Mode of Interaction of Berberine Alkaloid in Different Supramolecular Confined Environments: Interplay of Surface Charge between Nano-Confined Charged Layer and DNA

In this Article, we demonstrate a detailed characterization of binding interaction of berberine chloride (BBCl) with calf-thymus DNA (CT-DNA) in buffer solution as well as in two differently charged reverse micelles (RMs). The photophyscial properties of this alkaloid have been modulated within these microheterogeneous bioassemblies. The mode of binding of this alkaloid with DNA is of debate to date. However, fluorescence spectroscopic measurements, circular dichroism (CD) measurement, and temperature-dependent study unambiguously establish that BBCl partially intercalates into the DNA base pairs. The nonplanarity imposed by partial saturation in their structure causes the nonclassical types of intercalation into DNA. Besides the intercalation, electrostatic interactions also play a significant role in the binding between BBCl and DNA. DNA structure turns into a condensed form after encapsulation into RMs, which is followed by the CD spectra and microscopy study. The probe location and dynamics in the nanopool of the RMs depended on the electrostatic interaction between the charged surfactants and cationic berberine. The structural alteration of CT-DNA from B form to condensed form and the interplay of surface charge between RMs and DNA determine the interaction between the alkaloid and DNA in RMs. Time-resolved study and fluorescence anisotropy measurements successfully provide the binding interaction of BBCl in the nanopool of the RMs in the absence and in the presence of DNA. This study motivates us to judge further the potential applicability of this alkaloid in other biological systems or other biomimicking organized assemblies

Comparative Fluorescence Resonance Energy-Transfer Study in Pluronic Triblock Copolymer Micelle and Niosome Composed of Biological Component Cholesterol: An Investigation of Effect of Cholesterol and Sucrose on the FRET Parameters

The formation of pluronic triblock copolymer (F127)–cholesterol-based niosome and its interaction with sugar (sucrose) molecules have been investigated. The morphology of F127–cholesterol -based niosome in the presence of sucrose has been successfully demonstrated using dynamic light scattering (DLS) and transmission electron microscopic (TEM) techniques. The DLS profiles and TEM images clearly suggest that the size of the niosome aggregates increases significantly in the presence of sucrose. In addition to structural characterization, a detailed comparative fluorescence resonance energy transfer (FRET) study has been carried out in these F127-containing aggregates, involving coumarin 153 (C153) as donor (D) and rhodamine 6G (R6G) as an acceptor (A) to monitor the dynamic heterogeneity of the systems. Besides, time-resolved anisotropy and fluorescence correlation spectroscopy measurements have been carried out to monitor the rotational and lateral diffusion motion in these F127–cholesterol-based aggregates using C153 and R6G, respectively. During the course of FRET study, we have observed multiple time constants of FRET inside the F127–cholesterol-based niosomes in contrast with the F127 micelle. This corresponds to the presence of more than one preferential donor–acceptor (D–A) distance in niosomes than in F127 micelle. FRET has also been successfully used to probe the effect of sucrose on the morphology of F127–cholesterol-based niosome. In the presence of sucrose, the time constant of FRET further increases as the D–A distances increase in sucrose-decorated niosome. Finally, the excitation-wavelength-dependent FRET studies have indicated that as the excitation of donor molecules varies from 408 to 440 nm the contribution of the faster rise component of the acceptor enhances considerably, which clearly establishes the dynamics heterogeneity of both systems. Our findings also indicate that FRET is completely intravesicular in nature in these block copolymer-cholesterol-based aggregates

Picosecond Solvation and Rotational Dynamics: An Attempt to Reinvestigate the Mystery of Alcohol–Water Binary Mixtures

In this article, we have investigated the anomalous behavior of two alcohol–water (<i>tert</i>-butyl alcohol (TBA)–water and ethanol–water) binary mixtures using femtosecond fluorescence upconversion technique. Recently, Gupta and Patey (Gupta, R.; Patey, G. N. <i>J. Chem. Phys</i>. <b>2012</b>, 137, 034509(1)–034509(12)) have used four force fields to simulate TBA–water binary mixtures. Surprisingly, two of them do not identify any aggregation of TBA molecules. We have calculated average solvation time of Coumarin 480 (C480) using two different methods. Our results indicate slowdown in solvation time in the mole fraction ranges <i>X</i>_T = 0.09–0.15, <i>X</i>_T = 0.40–0.46 and <i>X</i>_E = 0.06–0.08, <i>X</i>_E = 0.20–0.25 for TBA–water and ethanol–water binary mixtures, respectively. Additionally, we have detected another anomalous region at <i>X</i>_T ∼ 0.03. Slow solvation responses in the ranges <i>X</i>_T = 0.40–0.46 and <i>X</i>_E = 0.20–0.25 are probably due to the higher shear viscosity of the medium. However, <i>X</i>_T = 0.09–0.15 and <i>X</i>_E = 0.06–0.08 are the manifestation of aggregation induced structural transition of alcohol molecules. Hindered rotation of C480 in the ranges <i>X</i>_T = 0.04–0.09 and <i>X</i>_E = 0.03–0.07 corroborates our solvation dynamics results. From temperature dependent anisotropy measurements, we have shown that aggregation of alcohol molecules increases with increase in temperature

Modulation of the Excited-State Dynamics of 2,2′-Bipyridine-3,3′-diol in Crown Ethers: A Possible Way To Control the Morphology of a Glycine Fibril through Fluorescence Lifetime Imaging Microscopy

In this article, we have investigated the modulation of excited-state intramolecular double proton transfer (ESIDPT) dynamics of 2,2′-bipyridine-3,3′-diol (BP­(OH)₂) in two crown ethers (CEs), namely, 18-Crown-6 (18C6) and 15-Crown-5 (15C5). From steady-state UV–visible measurements, we have shown that there is no significant interaction between the dienol tautomeric form of BP­(OH)₂ and two CEs. However, in the presence of CEs, an additional emission band (∼415 nm) is generated along with the diketo tautomer band (∼465 nm). In time-resolved analysis, we have observed the generation of ∼260 ps rise component in the presence of 18C6. Therefore, by combining the results of steady-state and time-resolved emissions, we have proposed that the water-assisted ESIDPT route of BP­(OH)₂ generates a hydronium ion (H₃O⁺) in the excited state. 18C6 binds nicely to this H₃O⁺ ion. As a result, retarded ESIDPT dynamics is observed in 18C6. However, as 15C5 cannot bind H₃O⁺ properly, no rise component is found. With the addition of potassium chloride (KCl), the contribution of the rise component decreases due to unavailability of free 18C6 cavity to capture the H₃O⁺ ion generated in the excited state. Addition of calcium chloride (CaCl₂) leads to complete removal of the rise component due to the inhibition of the water-assisted ESIDPT route. From wavelength-dependent behavior, we have observed that the rise component is present only at 465 nm in 18C6. We have also shown that the fibrillar morphology of glycine can be successfully probed through fluorescence lifetime imaging microscopy using BP­(OH)₂ as an imaging agent. Modulation of fibrillar morphology has been found in the presence of two CEs. The interaction of glycine fiber with CEs can be explained by lifetime distribution analysis

Vesicles Formation by Zwitterionic Micelle and Poly‑l‑lysine: Solvation and Rotational Relaxation Study

The stable unilamellar vesicles formation, having large potential applications in biological as well as biomedical fields, has been investigated in aqueous solution composed of a zwitterionic surfactant, <i>N</i>-hexadecyl-<i>N</i>,<i>N</i>-dimethylammonio-1-propanesulfonate (SB-16), and water-soluble cationic poly­(amino acid), poly-l-lysine (PLL). Dynamic light scattering (DLS), transmission electron microscopy (TEM), and other optical spectroscopic techniques revealed the transformation of SB-16 micelles in aqueous solutions into stable unilamellar vesicles above a certain concentration (0.008 to 0.1% w/v) of PLL. Solvation and rotational dynamics of coumarin 480 (C-480) give the information on hydration behavior around the headgroup regions of SB-16 micelle and SB-16/PLL vesicle. It was observed that the hydration nature around the headgroup regions of SB-16/PLL vesicular system is higher than the head group regions of micellar system. Thus, PLL permits more water molecules in the headgroup regions of vesicular system