Green synthesized silver nanoparticles from Phoenix dactylifera synergistically interact with bioactive extract of Punica granatum against bacterial virulence and biofilm development.

The global rise of antibiotic resistance poses a substantial risk to mankind, underscoring the necessity for alternative antimicrobial options. Developing novel drugs has become challenging in matching the pace at which microbial resistance is evolving. Recently, nanotechnology, coupled with natural compounds, has emerged as a promising solution to combat multidrug-resistant bacteria. In the present study, silver nanoparticles were green-synthesized using aqueous extract of Phoenix dactylifera (variety Ajwa) fruits and characterized by UV-vis spectroscopy, X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Scanning electron microscopy (SEM) coupled with Energy dispersive X-ray analysis (EDX), Transmission electron microscopy (TEM) and Thermogravimetric-differential thermal analysis (TGA-DTA). The in-vitro synergy of green synthesized P. dactylifera silver nanoparticle (PD-AgNPs) with selected antibiotics and bioactive extract of Punica granatum, i.e., ethyl acetate fraction (PGEF), was investigated using checkerboard assays. The most effective synergistic combination was evaluated against the QS-regulated virulence factors production and biofilm of Pseudomonas aeruginosa PAO1 by spectroscopic assays and electron microscopy. In-vivo anti-infective efficacy was examined in Caenorhabditis elegans N2 worms. PD-AgNPs were characterized as spherical in shape with an average diameter of 28.9 nm. FTIR analysis revealed the presence of functional groups responsible for the decrease and stabilization of PD-AgNPs. The signals produced by TGA-DTA analysis indicated the generation of thermally stable and pure crystallite AgNPs. Key phytocompounds detected in bioactive fractions include gulonic acid, dihydrocaffeic acid 3-O-glucuronide, and various fatty acids. The MIC of PD-AgNPs and PGEF ranged from 32 to 128 µg/mL and 250-500 µg/mL, respectively, against test bacterial strains. In-vitro, PD-AgNPs showed additive interaction with selected antibiotics (FICI 0.625-0.75) and synergy with PGEF (FICI 0.25-0.375). This combination inhibited virulence factors by up to 75 % and biofilm formation by 84.87 % in P. aeruginosa PAO1. Infected C. elegans worms with P. aeruginosa PAO1 had a 92.55 % survival rate when treated with PD-AgNPs and PGEF. The combination also reduced the reactive oxygen species (ROS) level in C. elegans N2 compared to the untreated control. Overall, these findings highlight that biosynthesized PD-AgNPs and bioactive P. granatum extract may be used as a potential therapeutic formulation against MDR bacteria.

Human Retinal Ganglion Cells Respond to Evolutionarily Conserved Chemotropic Cues for Intra Retinal Guidance and Regeneration.

Retinal ganglion cells (RGCs) connect the retina with the higher centers in the brain for visual perception. Their degeneration leads to irreversible vision loss in patients with glaucoma. The mechanism underlying human RGCs (hRGCs) axon growth and guidance remains poorly understood because hRGCs are born during development and connections with the central targets are established before birth. Here, using RGCs directly generated from human embryonic stem cells, we demonstrate that hRGCs express a battery of guidance receptors. These receptors allow hRGCs to read the spatially arrayed chemotropic cues in the developing rat retina for the centripetal orientation of axons toward the optic disc, suggesting that the mechanism of intraretinal guidance is conserved in hRGCs. The centripetal orientation of hRGCs axons is not only in response to chemorepulsion but also involves chemoattraction, mediated by Netrin-1/DCC interaction. The spatially arrayed chemotropic cues differentially influence hRGCs physiological responses, suggesting that neural activity of hRGCs and axon growth may be coupled during inter-retinal guidance. In addition, we demonstrate that Netrin-1/DCC interaction, besides promoting axon growth, facilitates hRGCs axon regeneration by recruiting the mTOR signaling pathway. The diverse influence of Netrin-1/DCC interaction ranging from axon growth to regeneration may involve recruitment of multiple intracellular signaling pathways as revealed by transcriptome analysis of hRGCs. From the perspective of ex vivo stem cell approach to glaucomatous degeneration, our findings posit that ex vivo generated hRGCs can read the intraretinal cues for guidance toward the optic disc, the first step required for connecting with the central target to restore vision.

Efflux Pump Inhibition-Based Screening and Anti-Infective Evaluation of Punica granatum Against Bacterial Pathogens.

Drug efflux pumps contribute to bacterial multidrug resistance (MDR), reducing antibiotic effectiveness and causing treatment failures. Besides their role in MDR, efflux pumps also assist in the transportation of quorum sensing (QS) signal molecules and increased the tolerance of biofilms. Recently, the search for efflux pump inhibitors from natural sources, including anti-infective plants, has gained attention as a potential therapy against drug-resistant bacteria. In this study, 19 traditional Indian medicinal plants were screened for their efflux pump inhibitory activity against Escherichia coli TGI. The promising extract, i.e., Punica granatum was subsequently fractioned in the solvents of increasing polarity. Among them, at sub-MIC active EPI fraction was PGEF (P. granatum ethyl acetate fraction), further investigated for anti-infective potential against Chromobacterium violaceum 12,472, Pseudomonas aeruginosa PAO1, and Serratia marcescens MTCC 97. PGEF was also evaluated for in vivo efficacy in Caenorhabditis elegans model. Major phytocompounds were analyzed by mass spectroscopic techniques. At respective Sub-MIC, PGEF reduced violacein production by 71.14% in C. violaceum 12,472. Moreover, PGEF inhibited pyocyanin (64.72%), pyoverdine (48.17%), protease (51.35%), and swarming motility (44.82%) of P. aeruginosa PAO1. Furthermore, PGEF reduced the production of prodigiosin and exoprotease by 64.73% and 61.80%, respectively. Similarly, at sub-MIC, PGEF inhibited (≥ 50%) biofilm development in all test pathogens. The key phytocompounds detected in active fraction include 5-hydroxymethylfurfural, trans-p-coumaric acid 4- glucoside, (-)-Epicatechin 3'-O-glucuronide, and ellagic acid. Interestingly, PGEF also demonstrated anti-infective efficacy against the PAO1-infected C. elegans test model and highlighting its therapeutic potential as an anti-infective agent to combat drug-resistant problems.

Human retinal ganglion cell axon regeneration by recapitulating developmental mechanisms: effects of recruitment of the mTOR pathway.

The poor axon regeneration in the central nervous system (CNS) often leads to permanent functional deficit following disease or injury. For example, degeneration of retinal ganglion cell (RGC) axons in glaucoma leads to irreversible loss of vision. Here, we have tested the hypothesis that the mTOR pathway regulates the development of human RGCs and that its recruitment after injury facilitates axon regeneration. We observed that the mTOR pathway is active during RGC differentiation, and using the induced pluripotent stem cell model of neurogenesis show that it facilitates the differentiation, function and neuritogenesis of human RGCs. Using a microfluidic model, we demonstrate that recruitment of the mTOR pathway facilitates human RGC axon regeneration after axotomy, providing evidence that the recapitulation of developmental mechanism(s) might be a viable approach for facilitating axon regeneration in the diseased or injured human CNS, thus helping to reduce and/or recover loss of function.

Chemical induction of neurogenic properties in mammalian Müller glia.

Müller glia (MG), cells that maintain homeostasis in the retina, are dormant stem cells that can regenerate neurons upon injury. However, the regenerative property of MG, which is reproducibly displayed in the lower vertebrates, is not readily observed in the mammals even upon forced expression of regulatory genes or exposure to growth factors. Here, we demonstrate a reproducible unmasking of the neurogenic properties of enriched rodent MG by serial exposure to different combinations of small molecules. The enriched MG, in response to changing culture conditions, silenced glia-specific genes and acquired transcriptional signature of neurons, accompanied by upregulation of genes known to regulate neuronal potential of MG. The MG-derived neurons expressed immunoreactivities corresponding to neuronal proteins and displayed electrophysiological features of immature neurons. Our study presents a proof of principle of pharmacological activation of neurogenic properties of mammalian MG, which may be utilized for therapeutic regeneration.

Assessment of Foodborne Bacterial Pathogens in Buffalo Raw Milk Using Polymerase Chain Reaction Based Assay.

Milk is a putrescible commodity that is extremely prone to microbial contamination. Primarily, milk and dairy products are believed to be easily contaminated by pathogenic microorganisms, including Listeria monocytogenes, Salmonella spp., and Staphylococcus aureus. The microbiological quality of raw milk and dairy products regarding foodborne pathogens is of paramount importance due to concern of human health. In this study 400 buffalo raw milk samples were screened for assessing the prevalence of L. monocytogenes, Salmonella spp., and S. aureus. This study implemented uniplex-polymerase chain reaction (u-PCR) and multiplex-polymerase chain reaction (m-PCR) assays for the fast simultaneous detection of these pathogens comparing to the conventional culturing methods. Raw milk samples were found contaminated with the prevalence of 2.2%, 4.0%, and 14.2% for L. monocytogenes, Salmonella spp., and S. aureus, respectively. These pathogens were detected with the optimized polymerase chain reaction assays after 6 h of enrichment. u-PCR and m-PCR demonstrated the limit of detection as 104, 102, and 10 cells/mL after 6, 12, 18, and 24 h for each culture of the pathogens. A high sensitivity (10 colony-forming unit [CFU]/mL) of the m-PCR protocol was noted. The developed protocol is a cost-effective and rapid method for the simultaneous detection of pathogens associated with raw milk and dairy industries.

Mapping developmental trajectories and subtype diversity of normal and glaucomatous human retinal ganglion cells by single-cell transcriptome analysis.

Glaucoma is characterized by a progressive degeneration of retinal ganglion cells (RGCs), leading to irreversible vision loss. Currently, there is no effective treatment for RGC degeneration. We used a disease-in-a-dish stem cell model to examine the developmental susceptibility of RGCs to glaucomatous degeneration, which may inform on the formulation of therapeutic approaches. Here, we used single-cell transcriptome analysis of SIX6 risk allele (SIX6risk allele ) primary open angle glaucoma patient-specific and control hRGCs to compare developmental trajectories in terms of lineage- and stage-specific transcriptional signature to identify dysregulated stages/genes, and subtype composition to estimate the relative vulnerability of RGCs to degeneration because their ability to regenerate axons are subtype-specific. The developmental trajectories, beginning from neural stem cells to RGCs, were similar between SIX6risk allele and control RGCs. However, the differentiation of SIX6risk allele RGCs was relatively stalled at the retinal progenitor cell stage, compromising the acquisition of mature phenotype and subtype composition, compared with controls, which was likely due to dysregulated mTOR and Notch signaling pathways. Furthermore, SIX6risk allele RGCs, as compared with controls, expressed fewer genes corresponding to RGC subtypes that are preferentially resistant to degeneration. The immature phenotype of SIX6risk allele RGCs with underrepresented degeneration-resistant subtypes may make them vulnerable to glaucomatous degeneration.

Plumbagin inhibits quorum sensing-regulated virulence and biofilms of Gram-negative bacteria: in vitro and in silico investigations.

The global rise in antimicrobial resistance and lack of discovery of new antimicrobials have created serious concerns. Targeting quorum sensing (QS) and biofilms of pathogenic bacteria is considered a promising approach in antimicrobial drug discovery. This study explored the inhibitory effect of plumbagin against biofilms and QS of Chromobacterium violaceum, Serratia marcescens and Pseudomonas aeruginosa. Violacein production in C. violaceum 12472 was reduced by >80%. The virulent traits of P. aeruginosa PAO1 such as pyocyanin, rhamnolipid and proteases were also inhibited at sub-minimum inhibitory concentrations. Moreover, the biofilms of the test bacteria were reduced by 56-70%. Plumbagin reduced the bacterial adherence and colonization on solid surface. Computational studies gave closer insights regarding the possible modes of action. Molecular dynamics simulations revealed that the protein complexes were quite stable under physiological conditions. This study provides both experimental and computational evidence regarding the efficacy of plumbagin against biofilms and the QS-controlled virulence factors of Gram-negative bacteria.

Stabilizing a Lithium Metal Battery by an In Situ Li2S-modified Interfacial Layer via Amorphous-Sulfide Composite Solid Electrolyte.

A novel strategy has been proposed to produce in situ Li2S at the interfacial layer between lithium anode and the solid electrolyte, by using an amorphous-sulfide-LiTFSI-poly(vinylidene difluoride) (PVDF) composite solid electrolyte (SLCSE). Besides retarding the decomposition of PVDF in CSE, the Li2S-modified interfacial layer (SMIL) also improves the wettability between lithium metal and SLCSE which in turn optimizes the lithium deposition process. Our density functional theory calculation results reveal that the migration energy barrier of Li passing through SMIL is much lower than that of Li passing through LiF-modified interfacial layer (FMIL) formed from the decomposition of PVDF. The as-prepared SLCSE shows a Li ionic transference number of 0.44 and Li ion conductivity of 3.42 × 10-4 S/cm at room temperature, and the Li||SLCSE||LiFePO4 cell exhibits an outstanding rate performance with a capacity of 153, 144, 131, and 101 mAh/g at a current density of 0.05, 0.10, 0.25, and 0.50 mA/cm2, respectively.

Cumin Prevents 17ß-Estradiol-Associated Breast Cancer in ACI Rats.

Breast cancer (BC) is a leading cause of cancer deaths in women in less developed countries and the second leading cause of cancer death in women in the U.S. In this study, we report the inhibition of E2-mediated mammary tumorigenesis by Cuminum cyminum (cumin) administered via the diet as cumin powder, as well as dried ethanolic extract. Groups of female ACI rats were given either an AIN-93M diet or a diet supplemented with cumin powder (5% and 7.5%, w/w) or dried ethanolic cumin extract (1%, w/w), and then challenged with subcutaneous E2 silastic implants (1.2 cm; 9 mg). The first appearance of a palpable mammary tumor was significantly delayed by both the cumin powder and extract. At the end of the study, the tumor incidence was 96% in the control group, whereas only 55% and 45% animals had palpable tumors in the cumin powder and extract groups, respectively. Significant reductions in tumor volume (660 ± 122 vs. 138 ± 49 and 75 ± 46 mm3) and tumor multiplicity (4.21 ± 0.43 vs. 1.16 ± 0.26 and 0.9 ± 0.29 tumors/animal) were also observed by the cumin powder and cumin extract groups, respectively. The cumin powder diet intervention dose- and time-dependently offset E2-related pituitary growth, and reduced the levels of circulating prolactin and the levels of PCNA in the mammary tissues. Mechanistically, the cumin powder diet resulted in a significant reversal of E2-associated modulation in ERα, CYP1A1 and CYP1B1. Further, the cumin powder diet reversed the expression levels of miRNAs (miR-182, miR-375, miR-127 and miR-206) that were highly modulated by E2 treatment. We analyzed the composition of the extract by GC/MS and established cymene and cuminaldehyde as major components, and further detected no signs of gross or systemic toxicity. Thus, cumin bioactives can significantly delay and prevent E2-mediated mammary tumorigenesis in a safe and effective manner, and warrant continued efforts to develop these clinically translatable spice bioactives as chemopreventives and therapeutics against BC.

Faecal and nitrate contamination in the groundwater of Mardan district, Pakistan.

This study aimed to determine the status of groundwater contamination with faecal coliform and nitrate in the rural areas of Mardan district, Pakistan. Both analytes require regular monitoring according to the National Environmental Quality Standards (NEQS) of Pakistan. Groundwater samples (n = 100) were collected from 25 villages across four zones. Samples were analysed for physicochemical parameters including pH, electrical conductivity (EC), Escherichia coli (E. coli) contamination, nitrite, and nitrate ([Formula: see text] and [Formula: see text]). Whilst the average concentrations of [Formula: see text] in the water samples were within the permissible limits of 50 mg L-1 set by the World Health Organisation (WHO) and NEQS two villages exceeded the safety limits. Non-carcinogenic health risks of [Formula: see text] were estimated in terms of average daily dose (ADD) and hazard quotient (HQ). The HQ values for [Formula: see text] were > 1 for children signifying potential health risks; however, the adult population had HQ < 1 which indicates no risk. Groundwater samples tested positive for E. coli contamination in 13 villages, suggesting that residents may be living at risk of various microbial diseases due to drinking of contaminated water. The findings of this study provide valuable baseline data for groundwater researchers, policymakers, and the local public health department.

miR-29c regulates neurogliogenesis in the mammalian retina through REST.

In the developing central nervous system, including its simple and accessible model retina, neurogenesis is followed by gliogenesis. However, the mechanism underlying the neurogliogenic switch remains poorly understood despite the identification of several regulatory genes, associated with the lineage identity and transition. The mechanism may involve cross talks between regulatory genes, facilitated through microRNAs. Here, we posit miR-29c as one of the regulatory miRNAs that may influence neuronal versus glial differentiation. We observed that the temporal patterns of miR-29c expression corresponded with late retinal histogenesis, the stage in the developing retina when neurogliogenic decision predominantly occurs. Examination of the effects of miR-29c on neurogliogenesis by the perturbation of function approach revealed that miR-29c preferentially facilitated differentiation of late RPCs into rod photoreceptors and bipolar cells, the late-born neurons, at the expense of Müller glia, the sole glia generated by retinal progenitor cells. We further observed that miR-29c facilitated neurogenesis and inhibited gliogenesis by regulating the expression of RE-1 silencing transcription factor (REST), which encodes a transcriptional repressor of cell cycle regulators and neuronal genes. Thus, miR-29c may influence neurogliogenic decision in the developing retina by regulating the instructive out put of a molecular axis helmed by REST.

The retroviral accessory proteins S2, Nef, and glycoMA use similar mechanisms for antagonizing the host restriction factor SERINC5.

Serine incorporator 5 (SERINC5) is a recently identified restriction factor that blocks virus entry but is antagonized by three unrelated retroviral accessory proteins. The S2 protein from equine infectious anemia virus (EIAV) has been reported to reduce SERINC5 expression at steady-state levels likely via the endocytic pathway; however, the precise mechanism is still unclear. Here, we investigated how EIAV S2 protein down-regulates SERINC5 compared with down-regulation induced by Nef from HIV-1 and glycoMA proteins from murine leukemia virus (MLV). Using bimolecular fluorescence complementation (BiFC) assay and immunoprecipitation (IP), we detected an interaction between S2 and SERINC5. We found that this interaction relies on the S2 myristoylation site, indicating that it may occur on the plasma membrane. S2 internalized SERINC5 via receptor-mediated endocytosis and targeted it to endosomes and lysosomes, resulting in a ubiquitination-dependent decrease in SERINC5 expression at steady-state levels. Both BiFC and IP detected a glycoMA-SERINC5 interaction, but a Nef-SERINC5 interaction was detected only by BiFC. Moreover, S2 and glycoMA down-regulated SERINC5 more effectively than did Nef. We further show that unlike Nef, both S2 and glycoMA effectively down-regulate SERINC2 and also SERINC5 from Xenopus tropicalis (xSERINC5). Moreover, we detected expression of the equine SERINC5 (eSERINC5) protein and observed that its expression is much weaker than expression levels of SERINC5 from other species. Nonetheless, eSERINC5 had a strong antiviral activity that was effectively counteracted by S2. We conclude that HIV-1, EIAV, and MLV share a similar mechanism to antagonize viral restriction by host SERINC5.

Murine Leukemia Virus Glycosylated Gag Reduces Murine SERINC5 Protein Expression at Steady-State Levels via the Endosome/Lysosome Pathway to Counteract SERINC5 Antiretroviral Activity.

Glycosylated Gag (glycoGag) is an accessory protein expressed by most gammaretroviruses, including murine leukemia virus (MLV). MLV glycoGag not only enhances MLV replication and disease progression but also increases human immunodeficiency virus type 1 (HIV-1) infectivity as Nef does. Recently, SERINC5 (Ser5) was identified as the target for Nef, and the glycoGag Nef-like activity has been attributed to the Ser5 antagonism. Here, we investigated how glycoGag antagonizes Ser5 using MLV glycoMA and murine Ser5 proteins. We confirm previous observations that glycoMA relocalizes Ser5 from plasma membrane to perinuclear punctated compartments and the important role of its Y36XXL39 motif in this process. We find that glycoMA decreases Ser5 expression at steady-state levels and identify two other glycoGag crucial residues, P31 and R63, for the Ser5 downregulation. The glycoMA and Ser5 interaction is detected in live cells using a bimolecular fluorescence complementation assay. Ser5 is internalized via receptor-mediated endocytosis and relocalized to Rab5+ early, Rab7+ late, and Rab11+ recycling endosomes by glycoMA. Although glycoMA is not polyubiquitinated, the Ser5 downregulation requires Ser5 polyubiquitination via the K48- and K63-linkage, resulting in Ser5 destruction in lysosomes. Although P31, Y36, L39, and R63 are not required for glycoMA interaction with Ser5, they are required for Ser5 relocalization to lysosomes for destruction. In addition, although murine Ser1, Ser2, and Ser3 exhibit very poor antiviral activity, they are also targeted by glycoMA for lysosomal destruction. We conclude that glycoGag has a broad activity to downregulate SERINC proteins via the cellular endosome/lysosome pathway, which promotes viral replication.IMPORTANCE MLV glycoGag not only enhances MLV replication but also increases HIV-1 infectivity similarly as Nef. Recent studies have discovered that both glycoGag and Nef antagonize a novel host restriction factor Ser5 and promote viral replication. Compared to Nef, the glycoGag antagonism of Ser5 is still poorly understood. MLV glycoGag is a transmembrane version of the structural Gag protein with an extra 88-amino-acid leader region that determines its activity. We now show that glycoGag interacts with Ser5 in live cells and internalizes Ser5 via receptor-mediated endocytosis. Ser5 is polyubiquitinated and relocalized to endosomes and lysosomes for massive destruction. In addition to the previously identified tyrosine-based sorting signal, we find two more important residues for Ser5 relocalization and downregulation. We also find that the Ser5 sensitivity to glycoGag is conserved in the SERINC family. Together, our findings highlight the important role of endosome/lysosome pathway in the enhancement of viral replication by viral proteins.

Green synthesis of silver nanoparticles using Carum copticum: Assessment of its quorum sensing and biofilm inhibitory potential against gram negative bacterial pathogens.

Antimicrobial resistance among pathogenic bacteria has become a global threat to human health. Due to poor progress in development of new antimicrobial drugs, there is a need for the development of novel alternative strategies to combat the problem of multidrug resistance. Moreover, there is focus on ecofriendly approach for the synthesis nanoparticles having efficient medicinal properties including antivirulence properties to tackle the emergence of multi-drug resistance. Targeting quorum sensing controlled virulence factors and biofilms has come out to be a novel anti-infective drug target. The silver nanoparticles (Ag@CC-NPs) were synthesized from aqueous extract of Carum copticum and characterized using UV-vis absorption spectroscopy, fourier-transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Ag@CC-NPs were checked for its ability to inhibit quorum sensing-mediated virulence factors and biofilms against three test pathogens at sub-MIC values. There was ~75% inhibition of violacein production by Ag@CC-NPs against C. violaceum. The P. aeruginosa virulence factors such as pyocyanin production, pyoverdin production, exoprotease activity, elastase activity, swimming motility and rhamnolipid production were inhibited by 76.9, 49.0, 71.1, 53.3, 89.5, and 60.0% at sub-MIC. Moreover, virulence factors of S. marcescens viz. prodigiosin production, exoprotease activity, and swarming motility was reduced by 78.4, 67.8, and 90.7%. Ag@CC-NPs also exhibited broad-spectrum antibiofilm activity with 77.6, 86.3, and 75.1% inhibition of biofilms of P. aeruginosa, S. marcescens, and C. violaceum respectively. The biofilm formation on glass coverslip was reduced remarkably as evident from SEM and CLSM analysis. The findings revealed the in vitro efficacy of Ag@CC-NPs against bacterial pathogens and can be exploited in the development of alternative therapeutic agent in management of bacterial infections for topical application, mainly wound infection, or coating of surfaces to prevent bacterial adherence on medical devices.

Development and validation of a spectrofluorimetric method for the analysis of tolfenamic acid in pure and tablet dosage form.

Tolfenamic acid (TA) is commonly used in humans and animals because of its anti-inflammatory, antipyretic, and analgesic effects. So far, no study has been carried out to develop a validated spectrofluorimetric method for determination of TA. Therefore, the present study aimed to develop and validate a simple, accurate, rapid, economical, and precise spectrofluorimetric method to assay TA in its pure and dosage forms, and also in degraded solutions. The fluorimetric method had higher sensitivity compared with the spectrophotometric and high-performance liquid chromatography methods and could determine the drug at the microgram level. Optimum pH of TA for maximum fluorescence intensity was 3, and its two pKa values were calculated as 1.95 and 4.05. The proposed method was validated according to the guidelines of the International Council for Harmonisation, and parameters such as linearity, range, accuracy, precision, sensitivity, robustness, specificity, and solution stability were tested. Stress-induced degradation studies on TA did not affect the accuracy and precision of the proposed method. The results obtained indicated that the method was linear over the concentration range 0.2-0.9 × 10-3 M with good accuracy, precision, and robustness for assay of TA in its pure and its tablet dosage forms and was comparable statistically with the British Pharmacopoeia method.

Authentication of various commercially available crude drugs using different quality control testing parameters.

The object of this study is to investigate the quality of various plant materials used in the preparation of herbal formulations using different methods of standardization to confirm their purity, safety and efficacy. However, it is uncertain whether these raw materials comply with the standards prescribed in the pharmacopeias. In the present study six raw materials' i.e. Foeniculum vulgarae, Curcuma longa, Aloe barbadensis, Plantago ovata, Zingiber officinale and Glycyrrhiza glabra have been obtained from the market and various quality control tests including microscopic evaluation, physico-chemical characteristics, thin layer chromatography (TLC), spectrophotometric assay (British Pharmacopoeia) and Fourier transform infrared spectroscopy (FTIR) have been performed to determine their compliance with the standards. The TLC has been used for the identification of the active ingredients on comparison of their Rf values with the reference standard. FTIR Spectra of these materials have been obtained to assign the functional groups present in the components of a particular material. Although these findings provide a significant data to herbal drug manufacturers for authentication of commercially available plant materials used in various herbal formulation.

Light transmission properties of pharmaceutical liquid bottles and evaluation of their photoprotective efficacy.

The light sensitive pharmaceutical dosage forms are well protected from light by packing in light protective bottles especially the colored glass and plastic bottles. In the present study the transmission characteristics of transparent glass bottle, amber glass bottle, polyvinyl chloride amber plastic bottle (PVC) and low density polyethylene semi-opaque plastic bottles (LDPE) (empty and drug filled) have been evaluated and the data compared for compliance with Pharmacopoeial limits of percentage transmission. The variations in thickness affect the amount of light transmitted through the bottles. For an average thickness, the transmission of bottles was not uniform indicated the effect of manufacturing variables on the transmission of light. The drug filled bottles showed an increase in light transmission probably as a result of interaction between drug and bottle components. The leaching of any coloring agents from glass bottles or the pigments from plastic bottles into the solution during storage appeared to increase the transmission of light which could be detrimental to photosensitive drugs in a formulation. The light protective efficacy of bottles was in the order: Semi-opaque plastic (LDPE) > amber plastic (PVC) > amber glass. The photoprotection of aqueous solution of riboflavin as a model compound in these bottles has been studied and its shelf-lives and stability ratio were determined.

Lin28a regulates neurogliogenesis in mammalian retina through the Igf signaling.

In the developing central nervous system (CNS) the majority of neurons are born before the generation of glia. Emerging evidence implicates heterochronic gene, Lin28 in the temporal switch between two distinct lineages. However, the respective contributions of Lin28a and Lin28b in neurogliogenesis remain poorly understood. Here, we have examined the relative involvement of Lin28a and Lin28b in mammalian retina, a simple and accessible CNS model where neurogliogenic decision largely occurs postnatally. Examination of Lin28a/b involvement during late histogenesis by the perturbation of function approaches revealed that while Lin28b did not influence differentiation in general Lin28a facilitated and antagonized the generation of neurons and glia, respectively. Silencing of Lin28a expression in vitro and its conditional deletion in vivo during early histogenesis led to premature generation of glia. The instructive role of Lin28a on neuronal differentiation was revealed by its influence to suppress glial-specific genes and directly differentiate glia along the neuronal lineage. This function of Lin28a is likely mediated through the Igf signaling, as inhibition of the pathway abrogated Lin28a-mediated neurogliogenesis. Thus, our observations suggest that Lin28a is an important intrinsic factor that acts in concert with cell-extrinsic factors like Igfs, coordinating the developmental bias of the progenitors and niche, respectively, for the successive generation of neurons and glia.

Broad-spectrum quorum sensing and biofilm inhibition by green tea against gram-negative pathogenic bacteria: Deciphering the role of phytocompounds through molecular modelling.

The emerging prevalence of multidrug-resistance in Gram-negative pathogens, due to conventional antimicrobial therapeutics, has led the researchers to emphasize on development of alternative novel strategies to suppress the bacterial virulence and pathogenicity through inhibition of quorum sensing (QS) and biofilms. QS is a bacterial communication system to produce density-dependent response via chemical signalling that controls pathogenesis and biofilms formation. Leaves of green tea are used worldwide as beverage which is also known for its broad-spectrum therapeutic efficacy. In this work, we have identified and characterized the most bioactive faction of green tea extract and evaluated the anti-QS and antibiofilm activity of green tea ethyl acetate fraction (GTEF) i.e. most active fraction, on three different Gram-negative bacterial pathogens. GTEF inhibited the violacein production by >75% in C. violaceum 12472. Many virulence factors of P. aeruginosa PAO1 viz. pyocyanin, pyoverdin, exoprotease, elastase, rhamnolipid production, and swimming motility were remarkably reduced in presence of sub-MICs of GTEF. Moreover, prodigiosin, protease activity, cell surface hydrophobicity, and swimming of S. marcescens MTCC 97 were also decreased significantly by the supplementation of GTEF in culture media. GTEF exhibited broad-spectrum antibiofilm action with >80% reduction in biofilm formation of test pathogens. In silico studies gave a mechanistic insight of action of GTEF. Molecular modelling revealed that phytoconstituents detected by GC/MS exhibited affinity (in order of 104 M-1) towards AHL synthases (LasI and EsaI). The molecular binding between phytocompounds and receptor proteins (LasR, RhlR, and PqsR) of QS circuit was also energetically favourable (ΔG°≥ 5.0 kcal mol-1) and supported by hydrogen bonds and hydrophobic interactions. These compounds were found to be docked in ligand binding domain of CviR and occupied same cavity as that of its antagonist. Squalene and thunbergol interacted with LasA at tartaric acid binding pocket and the complex was strengthened with binding energy -5.9 kcal mol-1. Moreover, interaction of thunbergol with biofilm-associated proteins viz. PilT and PilY1, might be disabling the pilus assembly and consequently inhibiting biofilm formation. In vivo validation of results suggested the protective role GTEF against QS-mediated pathogenicity and it might become a novel non-antibiotic QS inhibitor to control bacterial infection.