Photocatalytic activity of biosynthesized silver nanoparticle fosters oxidative stress at nanoparticle interface resulting in antimicrobial and cytotoxic activities.

Inside the biological milieu, nanoparticles with photocatalytic activity have potential to trigger cell death non-specifically due to production of reactive oxygen species (ROS) upon reacting with biological entities. Silver nanoparticle (AgNP) possessing narrow band gap energy can exhibit high light absorption property and significant photocatalytic activity. This study intends to explore the effects of ROS generated due to photocatalytic activity of AgNP on antimicrobial and cytotoxic propensities. To this end, AgNP was synthesized using the principle of green chemistry from the peel extract of Punica granatum L., and was characterized using UV-Vis spectroscope, transmission electron microscope and x-ray diffraction, and so forth. The antimicrobial activity of AgNP against studied bacteria indicated that, ROS generated at AgNP interface develop stress on bacterial membrane leading to bacterial cell death, whereas Alamar Blue dye reduction assay indicated that increased cytotoxic activity with increasing concentrations of AgNP. The γH2AX activity assay revealed that increasing the concentrations of AgNP increased DNA damaging activity. The results altogether demonstrated that both antimicrobial and cytotoxic propensities are triggered primarily due interfacial ROS generation by photocatalytic AgNP, which caused membrane deformation in bacteria and DNA damage in HT1080 cells resulting in cell death.

Mesosphaerum suaveolens Essential Oil Attenuates Inflammatory Response and Oxidative Stress in LPS-Stimulated RAW 264.7 Macrophages by Regulating NF-κB Signaling Pathway.

Mesosphaerum suaveolens (L.) Kuntze (Syn. Hyptis suaveolens (L.) Poit.) is a wild essential-oil-bearing plant having multiple uses in traditional medicine, perfumery, food, agriculture, and pharmaceutical industries. The present paper is the first report on the in vitro anti-inflammatory effects of the leaf essential oil of M. suaveolens (MSLEO) and unravels its molecular mechanism in LPS-stimulated RAW 264.7 macrophage cells. GC-MS analysis of the essential oil (EO) isolated from the leaves by hydro-distillation led to the identification of 48 constituents, accounting for 90.55% of the total oil, and ß-caryophyllene (16.17%), phyllocladene (11.85%), abietatriene (11.46%), and spathulenol (7.89%) were found to be the major components. MSLEO treatment had no effect on the viability of RAW 264.7 cells up to a concentration of 100 µg/mL, and the EO was responsible for a reduction in proinflammatory cytokines like IL-6, IL-1ß, and TNF-α, a decrease in intracellular ROS production, and the restoration of oxidative damage by elevating the levels of endogenous antioxidative enzymes like CAT, SOD, GPx, and GSH. RT-qPCR analysis indicated that MSLEO reduced the mRNA expression levels of iNOS and COX-2 as compared to the LPS-induced group. In addition, a confocal microscopy analysis showed that MSLEO inhibited the translocation of NF-κB from the cytosol to the nucleus. The results of this experiment demonstrate that MSLEO possesses significant anti-inflammatory potential by preventing the activation of NF-κB, which, in turn, inhibits the downstream expression of other inflammatory mediators associated with the activation of the NF-κB pathway in LPS-induced RAW 264.7 cells. Thus, the leaf essential oil of M. suaveolens may prove to be a promising therapeutic agent for the treatment of inflammation, and targeting the NF-κB signaling pathway may be considered as an attractive approach for anti-inflammatory therapies.

Variation in Yield, Chemical Composition and Biological Activities of Essential Oil of Three Curcuma Species: A Comparative Evaluation of Hydrodistillation and Solvent-Free Microwave Extraction Methods.

The essential oils of three medicinally important Curcuma species (Curcuma alismatifolia, Curcuma aromatica and Curcuma xanthorrhiza) were extracted using conventional hydro-distillation (HD) and solvent free microwave extraction (SFME) methods. The volatile compounds from the rhizome essential oils were subsequently analysed by GC-MS. The isolation of essential oils of each species was carried out following the six principles of green extraction and comparison was made between their chemical composition, antioxidant, anti-tyrosinase and anticancer activities. SFME was found to be more efficient than HD in terms of energy savings, extraction time, oil yield, water consumption and waste production. Though the major compounds of essential oils of both the species were qualitatively similar, there was a significant difference in terms of quantity. The essential oils extracted through HD and SFME methods were dominated by hydrocarbon and oxygenated compounds, respectively. The essential oils of all Curcuma species exhibited strong antioxidant activity, where SFME was significantly better than HD with lower IC50 values. The anti-tyrosinase and anticancer properties of SFME-extracted oils were relatively better than that of HD. Further, among the three Curcuma species, C. alismatifolia essential oil showed the highest rates of inhibition in DPPH and ABTS assay, significantly reduced the tyrosinase activity and exhibited significant selective cytotoxicity against MCF7 and PC3 cells. The current results suggested that the SFME method, being advanced, green and fast, could be a better alternative for production of essential oils with better antioxidant, anti-tyrosinase and anticancer activities for application in food, health and cosmetic industries.

Derivatives of Cinnamic Acid Esters and Terpenic Diversity in Volatiles of Thirty-Six Sand Ginger (Kaempferia galanga L.) Accessions of Eastern India Revealing Quality Chemovars.

The essential oil of Kaempferia galanga L. commonly known as sand ginger has increased its demand in national and international market for decades. Cinnamic acid esters like ethyl-p-methoxy cinnamate (EPMC) and ethyl cinnamate (EC) are major constituents in its essential oil. In spite of the high demand for the plant as raw material, identification of quality chemovars having high essential oil (EO) yield and constituents is still at an infant stage. With this in mind, we have evaluated the EO yield of 36 accessions from three provinces of Eastern India, which varied within a range of 0.41 ± 0.01 to 2.63 ± 0.03 v/w. Further, a total of 65 compounds were detected by gas chromatography and mass spectrometry (GC-MS) with area percentages varying from 76.16 to 97.3%. EPMC was found to be the major component in 14 accessions with area percentages varying from 10.7% to 41.1%, whereas other 22 accessions showed EC as the major constituent, varying from 16% to 29.1%. Further, a diversity study among accessions was performed by agglomerative hierarchical clustering (AHC) and principal component analysis (PCA) analysis based on the abundance of identified constituents, which categorized all 36 accessions into three clusters. Thus, the present study helps to identify quality chemovar K.g16 and K.g14 with respect to oil yield and constituents, respectively, which could be used to guide commercial cultivation and further improvement of the taxa.

Neocinnamomum caudatum Essential Oil Ameliorates Lipopolysaccharide-Induced Inflammation and Oxidative Stress in RAW 264.7 Cells by Inhibiting NF-κB Activation and ROS Production.

Neocinnamomum caudatum (Lauraceae) plant is used in the traditional system of medicine and is considered a potential source of edible fruits, spices, flavoring agents and biodiesel. The leaves, bark and roots of the species are used by local communities for the treatment of inflammatory responses, such as allergies, sinusitis and urinary tract infections. However, there is no scientific evidence to support the molecular mechanism through which this plant exerts its anti-inflammatory effect. The aim of the current research was to characterize the chemical constituents of bark (NCB) and leaf (NCL) essential oil of N. caudatum and to elucidate its anti-inflammatory action in lipopolysaccharide (LPS)-treated RAW 264.7 cells. Essential oils extracted by hydrodistillation were further subjected to gas chromatography mass spectrometry (GC-MS) analysis. The major constituents in bark essential oil identified as ß-pinene (13.11%), α-cadinol (11.18%) and α-pinene (10.99%), whereas leaf essential oil was found to be rich in ß-pinene (45.21%), myrcene (9.97%) and α-pinene (9.27%). Treatment with NCB and NCL at a concentration of 25 µg/mL exerted significant anti-inflammatory activity by significantly reducing LPS-triggered nitric oxide (NO) production to 45.86% and 61.64%, respectively, compared to the LPS-treated group. In the LPS-treated group, the production of proinflammatory cytokines, such as tumor necrosis factor (TNF)-α, interleukin (IL)-6 and IL-1ß, decreased after treatment with essential oil, alleviating the mRNA levels of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX)-2. The essential oil also inhibited the production of intracellular ROS and attenuated the depletion of mitochondrial membrane potential in a concentration-dependent manner. Pretreatment with NCB also reduced nuclear factor kappa-B (NF-κB)/p65 translocation and elevated the levels of endogenous antioxidant enzymes in LPS-induced macrophages. The present findings, for the first time, demonstrate the anti-inflammatory potential of both bark and leaf essential oils of N. caudatum. The bark essential oil exhibited a significantly more important anti-inflammatory effect than the leaf essential oil and could be used as a potential therapeutic agent for the treatment of inflammatory diseases.

Volatile Profiling of Magnolia champaca Accessions by Gas Chromatography Mass Spectrometry Coupled with Chemometrics.

Magnolia champaca (L.) Baill. ex Pierre of family Magnoliaceae, is a perennial tree with aromatic, ethnobotanical, and medicinal uses. The M. champaca leaf is reported to have a myriad of therapeutic activities, however, there are limited reports available on the chemical composition of the leaf essential oil of M. champaca. The present study explored the variation in the yield and chemical composition of leaf essential oil isolated from 52 accessions of M. champaca. Through hydrodistillation, essential oil yield was obtained, varied in the range of 0.06 ± 0.003% and 0.31 ± 0.015% (v/w) on a fresh weight basis. GC-MS analysis identified a total of 65 phytoconstituents accounting for 90.23 to 98.90% of the total oil. Sesquiterpene hydrocarbons (52.83 to 65.63%) constituted the major fraction followed by sesquiterpene alcohols (14.71 to 22.45%). The essential oils were found to be rich in ß-elemene (6.64 to 38.80%), γ-muurolene (4.63 to 22.50%), and ß-caryophyllene (1.10 to 20.74%). Chemometrics analyses such as PCA, PLS-DA, sPLS-DA, and cluster analyses such as hierarchical clustering, i.e., dendrogram and partitional clustering, i.e., K-means classified the essential oils of M. champaca populations into three different chemotypes: chemotype I (ß-elemene), chemotype II (γ-muurolene) and chemotype III (ß-caryophyllene). The chemical polymorphism analyzed in the studied populations would facilitate the selection of chemotypes with specific compounds. The chemotypes identified in the M. champaca populations could be developed as promising bio-resources for conservation and pharmaceutical application and further improvement of the taxa.

Application of a Multilayer Perceptron Artificial Neural Network for the Prediction and Optimization of the Andrographolide Content in Andrographis paniculata.

Andrographolide, the principal secondary metabolite of Andrographis paniculata, displays a wide spectrum of medicinal activities. The content of andrographolide varies significantly in the species collected from different geographical regions. Therefore, this study aims at investigating the role of different abiotic factors and selecting suitable sites for the cultivation of A. paniculata with high andrographolide content using a multilayer perceptron artificial neural network (MLP-ANN) approach. A total of 150 accessions of A. paniculata collected from different regions of Odisha and West Bengal in eastern India showed a variation in andrographolide content in the range of 0.28-5.45% on a dry weight basis. The MLP-ANN was trained using climatic factors and soil nutrients as the input layer and the andrographolide content as the output layer. The best topological ANN architecture, consisting of 14 input neurons, 12 hidden neurons, and 1 output neuron, could predict the andrographolide content with 90% accuracy. The developed ANN model showed good predictive performance with a correlation coefficient (R2) of 0.9716 and a root-mean-square error (RMSE) of 0.18. The global sensitivity analysis revealed nitrogen followed by phosphorus and potassium as the predominant input variables influencing the andrographolide content. The andrographolide content could be increased from 3.38% to 4.90% by optimizing these sensitive factors. The result showed that the ANN approach is reliable for the prediction of suitable sites for the optimum andrographolide yield in A. paniculata.

Development of a colloidal gold strip-based immunochromatographic assay for rapid detection of Fusarium oxysporum in ginger.

BACKGROUND: Rhizome rot, caused primarily by Fusarium oxysporum, is one of the most destructive diseases leading to significant loss in ginger worldwide. The loss can be greatly reduced by proper disease management practices steered by accurate and early diagnosis of pathogens. Pathogen detection at an early stage of infection can also reduce the incidence of disease epidemics. Classical methods are often time consuming, relying on culturing the putative pathogens and the availability of expert taxonomic skills for accurate identification, which leads to the delayed application of control measures. The development of a simple, rapid, sensitive and cost-effective point-of-care diagnostic tool is thus one of the major research priorities for rhizome rot. RESULTS: The 65 kDa, immunoreactive protein band was selected as a diagnostic marker and was subjected to MS analysis followed by blastp. Based on blast result, a synthetic antigenic peptide was synthesized, and used to generate pAbs. The peptide-specific antibodies were used to develop a colloidal gold immunochromatographic assay (ICA). The sensitivity, specificity, and accuracy of ICA were 92.59%, 81.25%, and 90%, respectively. The ICA has a visual detection limit of 2.122 µg mL-1 for infected rhizome samples and 5.065 µg mL-1 for leaf samples with optimal detection time within 5 min. Moreover, the ICA also detected early stage infected samples, of which 71.42% (50/70) were true positives. CONCLUSION: Findings from this study indicated that the assay can be utilized as a tool for the investigation of rhizome rot infection in field samples. © 2019 Society of Chemical Industry.

Molecular identification of indigenous manganese solubilising bacterial biodiversity from manganese mining deposits.

Manganese (Mn) ranks twelfth among the most exuberant metal present in the earth's crust and finds its imperative application in the manufacturing steel, chemical, tannery, glass, and battery industries. Solubilisation of Mn can be performed by several bacterial strains which are useful in developing environmental friendly solutions for mining activities. The present investigation aims to isolate and characterize Mn solubilising bacteria from low grade ores from Sanindipur Manganese mine of Sundargh district in Odisha state of India. Four morphologically distinct bacterial strains showing visible growth on Mn supplemented plates were isolated. Mn solubilising ability of the bacterial strains was assessed by visualizing the lightening of the medium appearing around the growing colonies. Three isolates were gram negative and rod shaped while the remaining one was gram positive, coccobacilli. Molecular identification of the isolates was carried out by 16S rRNA sequencing and the bacterial isolates were taxonomically classified as Bacillus anthrasis MSB 2, Acinetobacter sp. MSB 5, Lysinibacillus sp. MSB 11, and Bacillus sp. MMR-1 using BLAST algorithm. The sequences were deposited in NCBI GenBank with the accession number KP635223, KP635224, KP635225 and JQ936966, respectively. Manganese solubilisation efficiency of 40, 96, 97.5 and 48.5% were achieved by MMR-1, MSB 2, MSB 5 and MSB 11 respectively. The efficiency of Mn solubilisation is suggested with the help of a pH variation study. The results are discussed in relation to the possible mechanisms involved in Manganese solubilisation efficiency of bacterial isolates.

Exploring the mechanism of action of Vanda tessellata extract for the treatment of osteoarthritis through network pharmacology, molecular modelling and experimental assays.

The present study employed a comprehensive approach of network pharmacology, molecular dynamic simulation and in-vitro assays to investigate the underlying mechanism of the anti-osteoarthritic potential of Vanda tessellata extract (VTE). Thirteen active compounds of VTE were retrieved from the literature and the IMPPAT database. All of these passed the drug likeness and oral bioavailability parameters. A total of 535 VTE targets and 2577 osteoarthritis related targets were obtained. The compound-target-disease network analysis revealed vanillin, daucosterol, gigantol and syringaldehyde as the core key components. Protein-protein interaction analysis revealed BCL2, FGF2, ICAM 1, MAPK1, MMP1, MMP2, MMP9, COX2, STAT3 and ESR1 as the hub genes. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis revealed AGE-RAGE signalling pathway, HIF-1 signalling pathway and ESR signalling pathway as the major signalling pathway of VTE involved in treating osteoarthritis. Molecular docking analysis showed daucosterol and gigantol to have good binding affinity with BCL2, ESR1 and MMP9, and the results were further confirmed through molecular dynamics simulation analysis. The mechanism predicted by network pharmacology was validated in vitro on IL-1ß-induced SW982 synovial cells. VTE did not show any cytotoxicity and inhibited the migration of SW982 cells. VTE inhibited the expression level of IL-6, IL-8, TNF-α, PGE-2, MMP-2 and MMP-9 in a dose-dependent manner. VTE inhibited nuclear translocation of NF- κß and suppressed phosphorylation of p38, extracellular signal-regulated kinase (ERK), and c-Jun NH2-terminal kinase (JNK) of the mitogen-activated protein kinase (MAPK) signalling pathway. The results showed that VTE exerted an anti-osteoarthritic effect by a multi-target, multi-component and multi-signalling pathway approach.

Exploration of Pharmacological Mechanism of Cinnamomum tamala Essential Oil in Treating Inflammation based on Network Pharmacology, Molecular Modelling, and Experimental Validation.

BACKGROUND: Cinnamomum tamala (Buch.-Ham.) T.Nees & Eberm., also known as Indian Bay leaf, holds a distinctive position in complementary and alternative medicinal systems due to its anti-inflammatory properties. However, the active constituents and key molecular targets by which C. tamala essential oil (CTEO) exerts its anti-inflammatory action remain unclear. OBJECTIVE: The present study used network pharmacology and experimental validation to investigate the mechanism of CTEO in the treatment of inflammation. METHODS: GC-MS analysis was used to identify the constituents of CTEO. The key constituents and core targets of CTEO against inflammation were obtained by network pharmacology. The binding mechanism between the active compounds and inflammatory genes was ascertained by molecular docking and molecular dynamics simulation analysis. The pharmacological mechanism predicted by network pharmacology was verified in lipopolysaccharide-stimulated murine macrophage (RAW 264.7) cell lines. RESULTS: Forty-nine constituents were identified by GC-MS analysis, with 44 constituents being drug-like candidates. A total of 549 compounds and 213 inflammation-related genes were obtained, revealing 68 overlapping genes between them. Compound target network analysis revealed cinnamaldehyde as the core bioactive compound with the highest degree score. PPI network analysis demonstrated Il-1ß, TNF-α, IL8, IL6 and TLR4 as key hub anti-inflammatory targets. KEGG enrichment analysis revealed a Toll-like receptor signalling pathway as the principally regulated pathway associated with inflammation. A molecular docking study showed that cinnamaldehyde strongly interacted with the Il-1ß, TNF-α and TLR-4 proteins. Molecular dynamics simulations and MMPBSA analysis revealed that these complexes are stable without much deviation and have better free energy values. In cellular experiments, CTEO showed no cytotoxic effects on RAW 264.7 murine macrophages. The cells treated with LPS exhibited significant reductions in NO, PGE2, IL-6, TNF-α, and IL-1ß levels following treatment with CTEO. Additionally, CTEO treatment reduced the ROS levels and increased the antioxidant enzymes such as SOD, GSH, GPx and CAT. Immunofluorescence analysis revealed that CTEO inhibited LPS-stimulated NF-κB nuclear translocation. The mRNA expression of TLR4, MyD88 and TRAF6 in the CTEO group decreased significantly compared to the LPS-treated group. CONCLUSION: The current findings suggest that CTEO attenuates inflammation by regulating TLR4/MyD88/NF- κB signalling pathway.

Integrating network pharmacology and experimental verification to decipher the multitarget pharmacological mechanism of Cinnamomum zeylanicum essential oil in treating inflammation.

Inflammatory diseases contribute to more than 50 % of global deaths. Research suggests that network pharmacology can reveal the biological mechanisms underlying inflammatory diseases and drug effects at the molecular level. The aim of the study was to clarify the biological mechanism of Cinnamomum zeylanicum essential oil (CZEO) and predict molecular targets of CZEO against inflammation by employing network pharmacology and in vitro assays. First, the genes related to inflammation were identified from the Genecards and Online Mendelian Inheritance in Man (OMIM) databases. The CZEO targets were obtained from the SwissTargetPrediction and Similarity Ensemble Approach (SEA) database. A total of 1057 CZEO and 526 anti-inflammation targets were obtained. The core hub target of CZEO anti-inflammatory was obtained using the protein-protein interaction network. KEGG pathway analysis suggested CZEO to exert anti-inflammatory effect mainly through Tumor necrosis factor, Toll-like receptor and IL-17 signalling pathway. Molecular docking of active ingredients-core targets interactions was modelled using Pyrx software. Docking and simulation studies revealed benzyl benzoate to exhibit good binding affinity towards IL8 protein. MTT assay revealed CZEO to have non-cytotoxic effect on RAW 264.7 cells. CZEO also inhibited the production of NO, PGE2, IL-6, IL-1ß and TNF-α and promoted the activity of endogenous antioxidant enzymes in LPS-stimulated RAW 264.7 cells. Additionally, CZEO inhibited intracellular ROS generation, NF-kB nuclear translocation and modulated the expression of downstream genes involved in Toll-like receptor signalling pathway. The results deciphered the mechanism of CZEO in treating inflammation and provided a theoretical basis for its clinical application.

Elucidating the anti-cancer potential of Cinnamomum tamala essential oil against non-small cell lung cancer: A multifaceted approach involving GC-MS profiling, network pharmacology, and molecular dynamics simulations.

Cinnamomum tamala (Buch.-Ham.) T.Nees & Eberm., or Indian Bay Leaf, is a well-known traditional ayurvedic medicine used to treat various ailments. However, the molecular mechanism of action of Cinnamomum tamala essential oil (CTEO) against non-small cell lung cancer (NSCLC) remains elusive. The present study aims to decipher the molecular targets and mechanism of CTEO in treating NSCLC. GC-MS analysis detected 49 constituents; 44 successfully passed the drug-likeness screening and were identified as active compounds. A total of 3961 CTEO targets and 4588 anti-NSCLC-related targets were acquired. JUN, P53, IL6, MAPK3, HIF1A, and CASP3 were determined as hub genes, while cinnamaldehyde, ethyl cinnamate and acetophenone were identified as core compounds. Enrichment analysis revealed that targets were mainly involved in apoptosis, TNF, IL17, pathways in cancer and MAPK signalling pathways. mRNA expression, pathological stage, survival analysis, immune infiltrate correlation and genetic alteration analysis of the core hub genes were carried out. Kaplan-Meier overall survival (OS) curve revealed that HIF1A and CASP3 are linked to worse overall survival in Lung Adenocarcinoma (LUAD) cancer patients compared to normal patients. Ethyl cinnamate and cinnamaldehyde showed high binding energy with the MAPK3 and formed stable interactions with MAPK3 during the molecular dynamic simulations for 100 ns. The MM/PBSA analysis revealed that van der Waals (VdW) contributions predominantly account for a significant portion of the compound interactions within the binding pocket of MAPK3. Density functional theory analysis showed cinnamaldehyde as the most reactive and least stable compound. CTEO exhibited selective cytotoxicity by inhibiting the proliferation of A549 cells while sparing normal HEK293 cells. CTEO triggered apoptosis by arresting the cell cycle, increasing ROS accumulation, causing mitochondrial depolarisation, and elevating caspase-3, caspase-8 and caspase-9 levels in A549 cells. The above study provides insights into the pharmacological mechanisms of action of Cinnamomum tamala essential oil against non-small cell lung cancer treatment, suggesting its potential as an adjuvant therapy.

Chemical fingerprinting and multicomponent quantitative analysis for quality control of Cinnamomum tamala collected from Western Himalaya by HPLC-DAD.

Cinnamomum tamala, commonly known as "Indian bay leaf" or "Tejpat", is an economically important plant widely used in medicine, food and cosmetic industries. Growing demand for its leaf and bark in the herbal trade and non-availability of quality materials lead to large-scale species admixture and adulteration in the global market. The present study aims at developing a validated HPLC-DAD (High-performance liquid chromatography coupled with diode array detection) method and multiple markers-based chemical fingerprints for quality evaluation of C. tamala leaf extracts. Five bioactive compounds, viz., coumarin, cinnamyl alcohol, cinnamic acid, cinnamaldehyde and cinnamyl acetate, were identified and quantified in 28 samples collected from the western Himalayan region of India. The chromatographic separation was achieved on Shimadzu Shimpak C18 column (dimension 250 × 4.6 mm, pore size 5 µm) with a gradient elution of mobile phase using acetonitrile and 0.1 percent phosphate buffer and the chromatograms were obtained at a wavelength of 265 nm. The method validation was done by analyzing the linearity, LOD, LOQ, precision, stability, repeatability and recovery rates of standard compounds for quantitative analysis. The values of coefficient of correlation (R2) were found to be close to 1 for linearity and similarity analysis; and standard deviation was less than 3 percent in case of precision, stability, repeatability and recovery rates. The content of target compounds such as coumarin, cinnamyl alcohol, cinnamic acid, cinnamaldehyde and cinnamyl acetate varied in the range of 0-1.09, 0-0.05, 0.07-0.51, 0.39-1.27 and 0-0.27 percent, respectively. In the chemical fingerprint of C. tamala leaves, a total of 13 peaks were assigned as common peaks. The results of the study indicated that the HPLC method now developed combining chemical fingerprint with quantification of analytes could serve as a useful tool for quality evaluation of herbal raw materials of C. tamala and a valuable reference for further study.

Perspectives of genomic diversification and molecular recombination towards R-gene evolution in plants.

Plants are under strong evolutionary pressure in developing new and noble R genes to recognize pathogen avirulence (avr) determinants and bring about stable defense for generation after generations. Duplication, sequence variation by mutation, disparity in the length and structure of leucine rich repeats etc., causes tremendous variations within and among R genes in a plant thereby developing diverse recognitional specificity suitable enough for defense against new pathogens. Recent studies on genome sequencing, diversity and population genetics in different plants have thrown new insights on the molecular evolution of these genes. Tandem and segmental duplication are important factors in R gene abundance as inferred from the distribution of major nucleotide binding site-leucine rich repeats (NBS-LRRs) type R-genes in plant genomes. Likewise, R-gene evolution is also thought to be facilitated by cluster formation thereby causing recombination and sequence exchange and resulting in haplotypic diversity. Population studies have further proven that balancing selection is responsible for the maintenance of allelic diversity in R genes. In this review, we emphasize and discuss on improved perspectives towards the molecular mechanisms and selection pressure responsible for the evolution of NBS-LRR class resistance genes in plants.

Hedychium spicatum rhizome essential oil induces apoptosis in human prostate adenocarcinoma PC-3 cells via mitochondrial stress and caspase activation.

Hedychium spicatum is an essential oil bearing plant extensively used in the traditional system of medicine in several countries. Previous research has revealed H. spicatum essential oil (HSEO) to exhibit anti-tumor activity, although the mechanism of action is still unknown. Therefore, the current study was designed to carry out a comprehensive characterization of HSEO and evaluate the chemotherapeutic potential of HSEO against cancerous cells. The volatile constituents of HSEO was determined by one-dimensional gas chromatography with time-of-flight mass spectrometry (GC-TOFMS) and two-dimensional gas chromatography with time-of-flight mass spectrometry (GCxGC-TOFMS). In total, 193 phytocompounds could be detected, out of which 140 were identified for the first time. The major phytoconstituents detected by GCxGC-TOFMS were ß-pinene (10.94%), eucalyptol (6.45%), sabinene (5.48%) and trans-isolimonene (5.00%). GCxGC-TOFMS analysis showed two and half fold increase in the constituents over GC-TOFMS due to better chromatographic separation of constituents in the 2nd dimensional column. HSEO was tested for in vitro cytotoxic activity against cancerous (PC-3, HCT-116 and A-549) and normal (3T3-L1) cell, with HSEO being most selective for prostate cancer cell (PC-3) over non-tumorigenic fibroblast (3T3-L1) cell. HSEO treatment inhibited the colony formation ability of PC-3 cells. HSEO treatment caused apoptotic cell death and cell cycle arrest at G2/M and S phase in PC-3 cells. HSEO induced apoptosis via intracellular ROS accumulation, mitochondria depolarization and increased caspase-3, 8, and 9 levels in PC-3 cells. Additionally, HSEO treatment led to a decrease of Bcl-2 and Bcl-xL and increase of Bax and Bak protein levels. Overall, results from this study highlighted the anticancer potential of H. spicatum essential oil, which could be considered as a new agent for treating prostate cancer.

Volatile profiling coupled with multivariate analysis, antiproliferative and anti-inflammatory activities of rhizome essential oil of four Hedychium species from India.

ETHNOPHARMACOLOGICAL RELEVANCE: The genus Hedychium of family Zingiberaceae comprises several perennial rhizomatous species widely used in perfumery and traditional folk medicine to treat diseases related to asthma, diarrhoea, nausea, stomach disorders, inflammation and tumours. Several species of Hedychium have remained under-explored with respect to their chemical composition and biological activities. AIM OF THE STUDY: The current research aimed to explore the chemical composition and evaluate the antiproliferative and anti-inflammatory activities of rhizome essential oil from four Hedychium species (H. coccineum, H. gardnerianum, H. greenii and H. griffithianum). MATERIALS AND METHODS: Compound identification was accomplished using a Clarus 580 gas chromatography system in conjunction with mass spectrometry (GC-MS). The multivariate data statistics using chemometrics (PCA, PLS-DA, sPLS-DA) and cluster analysis (Dendrogram, Heat maps, K-means) were used to compare the similarity and relationship among Hedychium metabolomes. MTT assay was employed to visualize the antiproliferative property against MCF7, HepG2 and PC3 cancerous cell lines. The toxicity of essential oils was determined using 3T3-L1 non-tumorigenic/normal cells. Lipopolysaccharide (LPS)-induced RAW 264.7 cells were used to investigate the anti-inflammatory properties of Hedychium essential oils by measuring the production of nitric oxide (NO) using the Griess reagent method. Furthermore, the levels of prostaglandin (PGE2) and pro-inflammatory cytokines (TNF-α, IL-6, IL-1ß) was assessed using the ELISA technique. RESULTS: In total, 82 compounds were identified in four targeted species of Hedychium from which 1,8-cineole (52.71%), ß-pinene (32.83%), α-pinene (19.62%), humulene epoxide II (19.86%) and humulene epoxide I (19.10%) were the major constituents. Monoterpenes (8.5-59.9%) and sesquiterpenes (2.87-54.11%) were the two class of compounds, found as the most prevalent in the extracted essential oils. The multivariate analysis classified the four Hedychium species into three different clusters. Hedychium essential oils exhibited potent antiproliferative activity against MCF7, HepG2 and PC3 cancer cell lines with IC50 values less than 150 µg/mL where H. gardnerianum exhibited the highest selective cytotoxicity against human breast and prostate adenocarcinoma cells with an IC50 value of 44.04 ± 1.07 µg/mL and 56.11 ± 1.44 µg/mL, respectively. The essential oils on normal (3T3-L1) cells displayed no toxicity with higher IC50 values thereby concluding as safe to use for normal human health without causing any side effects. Besides, the essential oils at 100 µg/mL concentration revealed remarkable anti-inflammatory activity in LPS-activated RAW 264.7 murine macrophages by inhibiting the production of inflammatory mediators, with H. greenii exhibiting the maximum anti-inflammation response by significantly suppressing the levels of NO (84%), PGE2 (87%), TNF-α (94.3%), IL-6 (95%) and IL-1ß (85%) as compared to LPS treated group. CONCLUSION: The present findings revealed that the Hedychium species traditionally used in therapeutics could be a potential source of active compounds with antiproliferative and anti-inflammatory properties.

Exploring molecular docking with MM-GBSA and molecular dynamics simulation to predict potent inhibitors of cyclooxygenase (COX-2) enzyme from terpenoid-based active principles of Zingiber species.

Cyclooxygenase 2 (COX-2), the key enzyme involved in prostaglandin (PGs) production, is known to take part in inflammatory and immune responses. Though COX-2 inhibitors are therapeutically effective anti-inflammatory drugs, they deficit anti-thrombotic activity thus leading to increased cardiovascular diseases. Therefore, COX-2 inhibitors with improved therapeutic efficacy and tolerance are still needed. In recent years, traditional medicine systems have paid attention to the essential oil of genus Zingiber, particularly for the treatment of various inflammatory illnesses, with lesser side effects. Thus, the present study aims to explore the anti-inflammatory activity of Zingiber essential oil through computational-biology approaches. In this regard, virtual screening, molecular docking, and simulations were carried out on 53 compounds derived from the essential oil of Zingiber species in order to provide mechanistic insights into COX-2 inhibition and identify the most actively potent anti-inflammatory compounds. Among all the docked ligands, epi-cubenol, δ-cadinene, γ-eudesmol, cubenol, and α-terpineol were found to be powerful bioactive compounds with an increased binding affinity towards COX-2 along with favorable physiochemical properties. Additionally, MD simulation in DPPC lipid bilayers was studied to examine the intrinsic dynamics and adaptability of the chosen ligands and COX-2-complexes. The findings showed that the selected five components interacted steadily with the COX-2 active site residues throughout the simulation via different bondings. The integrative-computational approach showed that the identified natural compounds may be taken into further consideration for potential in vitro and in vivo evaluation as COX-2 inhibitors, which would lead to the development of more potent and efficient anti-inflammatory drugs.Communicated by Ramaswamy H. Sarma.

Anti-proliferative Activity of Piper trioicum Leaf Essential Oil Based on Phytoconstituent Analysis, Molecular Docking and in silico ADMET Approaches.

BACKGROUND: The essential oils isolated from several medicinal plants have been reported to possess anticancer activities. Both the essential oil and extracts of many Piper species (Piperaceae) possess potential cytotoxic effects against cancer cell lines and are being used in traditional systems of medicine for the treatment of cancer. There is a need to evaluate and validate the anticancer properties of essential oils extracted from other wild species of Piper. OBJECTIVE: The current research was undertaken to determine the chemical composition and investigate the anti-proliferative activity of wild-growing Piper trioicum leaf essential oil. The selected five major constituents were subjected to molecular docking to identify possible modes of binding against serine/threonine-protein kinase (MST3) protein. METHODS: The essential oil of leaf of P. trioicum was extracted by hydrodistillation method, and its chemical composition was evaluated by GC-FID and GC-MS. The anti-proliferative activity of the essential oil was evaluated by the MTT assay against normal (3T3-L1) and various cancer (HCT 116, HT-29, PC-3 and HepG2) cell lines. Molecular docking analysis was performed using the AutoDock 4.2 software. The pharmacokinetic and pharmacodynamic properties of the major constituents were determined using absorption, distribution, metabolization, excretion and toxicity (ADMET) analysis. RESULTS: The GC-MS analysis revealed the identification of 45 constituents with δ-cadinene (19.57%), germacrene-D (8.54%), ß-caryophyllene (6.84%), 1-epi-cubenol (4.83%) and α-pinene (4.52%) being predominant constituents in the leaf essential oil of P. trioicum. The highest cytotoxicity of essential oil was observed against HT-29 cells (IC50 value of 33.14 µg/ml). 1-epi-cubenol and δ-cadinene exhibited low binding energy values of -6.25 and -5.92 kcal/mol, respectively. For prediction of in silico pharmacokinetic and drug-like properties of the major compounds, the ADMET prediction tool was used, the results of which were observed to be within the ideal range. CONCLUSION: The present findings demonstrate that P. trioicum essential oil possesses significant anti-proliferative activity and could be effective against cancer treatment.

Assessment of genetic diversity in Zingiberaceae through nucleotide binding site-based motif-directed profiling.

Functional motif-directed profiling was performed with 15 nucleotide binding site (NBS) primer-enzyme combinations to identify and elucidate the phylogenetic relationships among 15 genotypes of the family Zingiberaceae. We retrieved 167 polymorphic bands (24.85 %), with an average of 11.13 bands per primer. Mean polymorphism rates were detected using MseI (26 %), RsaI (21 %), and AluI (28 %) as restriction enzymes. The polymorphism information content (PIC) for each NBS primer-enzyme combination ranged from 0.48 to 0.76 with a mean value of 0.65. The 38 NBS profiling markers had PIC values ranging from 0.3 to 0.6 and exhibited good power to discriminate between genotypes. Comparison of NBS profiling with microsatellite data for the same set of genotypes exhibited a correlation value of 0.78, P ≤ 0.001. Our study suggests that genetic variability assessment could be more efficient if it targeted genes that exhibit functionally relevant variation, rather than random markers.