Search results for: poly(A) tailing assay

Authors: Cheng-Chih Tsai, Yu-Hsuan Liu, Cheng-Ying Ho, Chun-Chin Huang

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The aim of this study evaluated the in vitro and in vivo antimicrobial activity of selected lactic acid bacteria (LAB) against Uropathogenic Escherichia coli (UPEC) for prevention and amelioration of UTIs. We screened LAB strains with antimicrobial effects on UPEC using a well-diffusion assay, bacterial adherence to the uroepithelium cell line SV-HUC-1 (BCRC 60358), and a coculture inhibition assay. The results showed that the 7 LAB strains (Lactobacillus paracasei, L. salivarius, two Pediococcus pentosaceus strains, two L. plantarum strains, and L. crispatus) and the fermented probiotic products produced by these multi-LAB strains exhibited potent zones of inhibition against UPEC. Moreover, the LAB strains and probiotic products adhered strongly to the uroepithelium SV-HUC-1 cell line. The growth of UPEC strains was also markedly inhibited after co-culture with the LAB strains and probiotic products in human urine. In addition, the enhanced levels of IL-6, IL-8 and lactic acid dehydrogenase were significantly decreased by treatments with the LAB strains and probiotic products in UPEC-induced SV-HUC-1 cells. Furthermore, oral administration of probiotic products reduced the number of viable UPEC in the urine of UPEC-challenged BALB/c mice. Taken together, this study demonstrates that probiotic supplementation may be useful as an adjuvant therapy for the treatment of bacterial-induced urinary tract infections.

Keywords: lactic acid bacterium, SV-HUC-1 uroepithelium, urinary tract infection, uropathogenic Escherichia coli, BALB/c mice

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Authors: Sirikwan Ponprateep, Anchalee Tassanakajon, Vichien Rimphanitchayakit

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A Kazal-type serine proteinase inhibitor, SPIPm2, was abundantly expressed in the hemocytes and secreted into shrimp plasma has anti-viral property against white spot syndrome virus (WSSV). To discover the molecular mechanism of antiviral activity, the binding assay showed that SPIPm2 bind to the components of viral particle and shrimp hemocyte. From our previous report, viral target protein of SPIPm2 was identified, namely WSV477 using yeast two-hybrid screening. WSV477 is an early gene product of WSSV and involved in viral propagation. In this study, the co-immunoprecipitation technique and Tandem Mass Spectrometry (LC-MS/MS) was used to identify the target protein of SPIPm2 from shrimp hemocyte. The target protein of SPIPm2 was cyclophilin A. In vertebrate, cyclophilin A or peptidylprolyl isomerase A was reported to be the immune suppressor interacted with cyclosporin A involved in immune defense response. The recombinant cyclophilin A from Penaeus monodon (rPmCypA) was produced in E.coli system and purified using Ni-NTA column to confirm the protein-protein interaction. In vitro pull-down assay showed the interaction between rSPIPm2 and rPmCypA. To study the biological function of these proteins, the expression analysis of immune gene in shrimp defense pathways will be investigated after rPmCypA administration.

Keywords: cyclophilin A, protein-protein interaction, Kazal-type serine proteinase inhibitor, Penaeus monodon

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Authors: Anjani Kumar Tiwari, Neelam Kumari, Anil Mishra

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The 18-kDa translocator protein (TSPO) previously called as peripheral benzodiazepine receptor, is proven biomarker for variety of neuroinflammation. TSPO is tryptophane rich five transmembranal protein found on outer mitochondrial membrane of steroid synthesising and immunomodulatory cells. In case of neuronal damage or inflammation the expression level of TSPO get upregulated as an immunomodulatory response. By utilizing Benzoxazolone as a basic scaffold, series of TSPO ligands have been designed followed by their screening through in silico studies. Synthesis has been planned by employing convergent methodology in six high yielding steps. For the synthesized ligands the ‘in vitro’ assay was performed to determine the binding affinity in term of Ki. On ischemic rat brain, autoradiography studies were also carried to check the specificity and affinity of the designed radiolabelled ligand for TSPO.Screening was performed on the basis of GScore of CADD based schrodinger software. All the modified and better prospective compound were successfully carried out and characterized by spectroscopic techniques (FTIR, NMR and HRMS). In vitro binding assay showed best binding affinity Ki = 6.1+ 0.3 for TSPO over central benzodiazepine receptor (CBR) Ki > 200. ARG studies indicated higher uptake of two analogues on the lesion side compared with that on the non-lesion side of ischemic rat brains. Displacement experiments with unlabelled ligand had minimized the difference in uptake between the two sides which indicates the specificity of the ligand towards TSPO receptor.

Keywords: TSPO, PET, imaging, Acetamidobenzoxazolone

Procedia PDF Downloads 143

Authors: Kang-Hyun Leem, Myung-Gyou Kim, Hye Kyung Kim

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The prickly pear cactus (Opuntia ficus-indica) has a global distribution and have been used for medicinal benefits such as artherosclerosis, diabetes, gastritis, and hyperglycemia. However, very little information is currently available for their mechanism. The prikly pear variety Opuntia ficus-indica var. Saboten (OFS) is widely cultivated in Cheju Island, southwestern region of Korea, and used as a functional food. Present study investigated the effects of OFS on pancreatic β-cell function using pancreatic islet β cells (HIT cell). Alpha-glucosidase inhibition, glucose uptake, insulin secretion, insulin sensitivity, and pancreatic β cell proliferation were determined. The inhibitory effect of ethanol extract of OFS stem on α-glucosidase enzyme was measured in a cell free system. Glucose uptake was determined using fluorescent glucose analogue, 2-NBDG. Insulin secretion was measured by ELISA assay. Cell proliferation was measured by MTT assay. Ethanol extracts of OFS dose-dependently inhibited α-glucosidase activity as well as glucose uptake. Insulinotrophic effect of OFS extract was observed at high glucose media in pancreatic β-islet cells. Furthermore, pancreatic β cell regeneration was also observed.These results suggest that OFS mediates the antidiabetic activity mainly via α-glucosidase inhibition, glucose uptake, and improved insulin sensitivity.

Keywords: prickly pear cactus, Opuntia ficus-indica var. Saboten, pancreatic islet HIT cells, α-glucosidase, glucose uptake, insulinotrophic

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Authors: Kriyapa lairungruang, Arunporn Itharat

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Limonia acidissima L. (LA) (Common name: wood apple, Thai name: ma-khwit) is a medicinal plant which has long been used in Thai traditional medicine. Its bark is used for treatment of diarrhea, abscess, wound healing and inflammation and it is also used in oral cancer. Thus, this research aimed to investigate antioxidant and cytotoxic activities of the LA bark extracts produced by various extraction methods. Different extraction procedures were used to extract LA bark for biological activity testing: boiling in water, maceration with 95% ethanol, maceration with 50% ethanol and water boiling of each the 95% and the 50% ethanolic residues. All extracts were tested for antioxidant activity using DPPH radical scavenging assay, cytotoxic activity against human laryngeal epidermoid carcinoma (HEp-2) cells and human oral epidermoid carcinoma (KB) cells using sulforhodamine B (SRB) assay. The results found that the 95% ethanolic extract of LA bark showed the highest antioxidant activity with EC50 values of 29.76±1.88 µg/ml. For cytotoxic activity, the 50% ethanolic extract showed the best cytotoxic activity against HEp-2 and KB cells with IC50 values of 9.55±1.68 and 18.90±0.86 µg/ml, respectively. This study demonstrated that the 95% ethanolic extract of LA bark showed moderate antioxidant activity and the 50% ethanolic extract provided potent cytotoxic activity against HEp-2 and KB cells. These results confirm the traditional use of LA for the treatment of oral cancer and laryngeal cancer, and also support its ongoing use.

Keywords: antioxidant activity, cytotoxic activity, Laryngeal epidermoid carcinoma, Limonia acidissima L., oral epidermoid carcinoma

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Authors: Manal M. Hamed, Mosad A. Ghareeb, Abdel-Aleem H. Abdel-Aleem, Amal M. Saad, Mohamed S. Abdel-Aziz, Asmaa H. Hadad

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Natural products derived from medicinal plants provide unlimited opportunities for a new medication leads because of the unmatched accessibility of chemical variation. Six compounds were isolated from the n-butanol extract of Callistemon citrinus (Family Myrtaceae), they were identified as; nepetolide (1), callislignan A (2), 6,8-dimethoxy-4,5-dimethyl-3-methyleneisochroman-1-one (3), 3-methyl-7-O-benzoyl-β-D-glucopyranoside (4), 5, 7, 3', 5'-tetrahydroxy-6, 8-di-C-methyl flavanone (5), and (2R,3R,4S,5S)-2,4-bis(4-hydroxyphenyl)-3,5-dihydroxy-tetrahydropyran (6). The isolated compounds were evaluated as antioxidant and antimicrobial agents. The antioxidant activities of the compounds were determined using DPPH-radical scavenging and total antioxidant capacity (TAC) assays. The results indicated that compound (5) was most active in its capacity to scavenge free radicals in the DPPH assay [SC50 value, 4.65 ± 0.74μg/mL] compared to the standard ascorbic acid and exhibited the highest activity in the TAC assay (610.45 ± 1.67mg AAE/g compound). The pure isolates were tested for their antimicrobial activity against four pathogenic microbial strains including Staphylococcus aureus, Methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa and Candida albicans. Also, the GC/MS analysis of its leaves essential oil presented nine identified compounds representing 91% of the total oil constituents. The outcomes got from this study give a reasonable justification for the medicinal uses of Callistemon citrinus plant.

Keywords: Callistemon citrinus, flavanone, antioxidant activity, antimicrobial activity, essential oil, Myrtaceae

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Authors: T. C. Machaba, S. M. Mahlo

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The current study investigates the antifungal properties of crude plant extracts from selected medicinal plant species. Eight plant species used by the traditional healers and local people to treat fungal infections were selected for further phytochemical analysis and biological assay. The selected plant species were extracted with solvent of various polarities such as acetone, methanol, ethanol, hexane, dichloromethane, ethyl acetate and water. Leaf, roots and bark extracts of Maerua juncea Pax, Albuca seineri (Engl & K. Krause) J.C Manning & Goldblatt, Senna italica Mill., Elephantorrhiza elephantina (Burch.) Skeels, Indigofera circinata Benth., Schinus molle L., Asparagus buchananii Bak., were screened for antifungal activity against three animal fungal pathogens (Candida albicans, Aspergillus fumigatus and Cryptococcus neoformans). All plant extracts were active against the tested microorganisms. Acetone, dichloromethane, hexane and ethanol extracts of Senna italica and Elephantorrhiza elephantine had excellent activity against Candida albicans and A. fumigatus with the lowest MIC value of 0.02 mg/ml. Bioautography assay was used to determine the number of antifungal compounds presence in the plant extracts. No active compounds were observed in plant extracts of Indigofera circinnata, Schinus molle and Pentarrhinum insipidum with good antifungal activity against C. albicans and A. fumigatus indicating possible synergism between separated metabolites.

Keywords: antifungal activity, bioautography, ethnobotanical survey, minimum inhibitory concentration

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Authors: Masauso Moses Phiri

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Frequent glucose monitoring is essential to the management of diabetes. Plasmonic enzyme-based glucose biosensors have the advantages of greater specificity, simplicity and rapidity. The aim of this study was to develop a rapid plasmonic colorimetric glucose biosensor based on biocatalytic enlargement of AuNS guided by GOx. Gold nanoparticles of 18 nm in diameter were synthesized using the citrate method. Using these as seeds, a modified seeded method for the synthesis of monodispersed gold nanostars was followed. Both the spherical and star-shaped nanoparticles were characterized using ultra-violet visible spectroscopy, agarose gel electrophoresis, dynamic light scattering, high-resolution transmission electron microscopy and energy-dispersive X-ray spectroscopy. The feasibility of a plasmonic colorimetric assay through growth of AuNS by silver coating in the presence of hydrogen peroxide was investigated by several control and optimization experiments. Conditions for excellent sensing such as the concentration of the detection solution in the presence of 20 µL AuNS, 10 mM of 2-(N-morpholino) ethanesulfonic acid (MES), ammonia and hydrogen peroxide were optimized. Using the optimized conditions, the glucose assay was developed by adding 5mM of GOx to the solution and varying concentrations of glucose to it. Kinetic readings, as well as color changes, were observed. The results showed that the absorbance values of the AuNS were blue shifting and increasing as the concentration of glucose was elevated. Control experiments indicated no growth of AuNS in the absence of GOx, glucose or molecular O₂. Increased glucose concentration led to an enhanced growth of AuNS. The detection of glucose was also done by naked-eye. The color development was near complete in ± 10 minutes. The kinetic readings which were monitored at 450 and 560 nm showed that the assay could discriminate between different concentrations of glucose by ± 50 seconds and near complete at ± 120 seconds. A calibration curve for the qualitative measurement of glucose was derived. The magnitude of wavelength shifts and absorbance values increased concomitantly with glucose concentrations until 90 µg/mL. Beyond that, it leveled off. The lowest amount of glucose that could produce a blue shift in the localized surface plasmon resonance (LSPR) absorption maxima was found to be 10 – 90 µg/mL. The limit of detection was 0.12 µg/mL. This enabled the construction of a direct sensitivity plasmonic colorimetric detection of glucose using AuNS that was rapid, sensitive and cost-effective with naked-eye detection. It has great potential for transfer of technology for point-of-care devices.

Keywords: colorimetric, gold nanostars, glucose, glucose oxidase, plasmonic

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Authors: Hoda Mahgoub, Abeer Hanafy

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Linagliptin (LNG) belongs to dipeptidyl-peptidase-4 (DPP-4) inhibitor class. DPP-4 inhibitors represent a new therapeutic approach for the treatment of type 2 diabetes in adults. The aim of this work was to develop and validate an accurate and reproducible HPLC method for the determination of LNG with high sensitivity in rat plasma. The method involved separation of both LNG and pindolol (internal standard) at ambient temperature on a Zorbax Eclipse XDB C18 column and a mobile phase composed of 75% methanol: 25% formic acid 0.1% pH 4.1 at a flow rate of 1.0 mL.min-1. UV detection was performed at 254nm. The method was validated in compliance with ICH guidelines and found to be linear in the range of 5–1000ng.mL-1. The limit of quantification (LOQ) was found to be 5ng.mL-1 based on 100µL of plasma. The variations for intra- and inter-assay precision were less than 10%, and the accuracy values were ranged between 93.3% and 102.5%. The extraction recovery (R%) was more than 83%. The method involved a single extraction step of a very small plasma volume (100µL). The assay was successfully applied to an in-vivo pharmacokinetic study of LNG in rats that were administered a single oral dose of 10mg.kg-1 LNG. The maximum concentration (Cmax) was found to be 927.5 ± 23.9ng.mL-1. The area under the plasma concentration-time curve (AUC0-72) was 18285.02 ± 605.76h.ng.mL-1. In conclusion, the good accuracy and low LOQ of the bioanalytical HPLC method were suitable for monitoring the full pharmacokinetic profile of LNG in rats. The main advantages of the method were the sensitivity, small sample volume, single-step extraction procedure and the short time of analysis.

Keywords: HPLC, linagliptin, pharmacokinetic study, rat plasma

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Authors: Lubica Muckova, Petr Jost, Jaroslav Pejchal, Daniel Jun

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The development of acetylcholinesterase reactivators, i.e. antidotes against organophosphorus poisoning, is an important goal of defence research. The aim of this study was to compare cytotoxicity and chemical structure of 5 currently available (pralidoxime, trimedoxime, obidoxime, methoxime, and asoxime) and 4 newly developed compounds (K027, K074, K075, and K203). In oximes, there could be at least four important structural factors affecting their toxicity, including the number of oxime groups in the molecule, the position of oxime group(s) on pyridinium ring, the length of carbon linker, and the substitution by oxygen or insertion of the double bond into the connection chain. The cytotoxicity of tested substances was measured using colorimetric 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay (MTT assay) in SH-SY5Y cell line. Toxicity was expressed as toxicological index IC₅₀. The tested compounds showed different cytotoxicity ranging from 1.5 to 27 mM. K027 was the least, and methoxime was the most toxic reactivator. The lowest toxicity was found in a monopyridinium reactivator and bispyridinium reactivators with simple 3C carbon linker. Shortening of connection chain length to 1C, incorporation of oxygen moiety into 3C compounds, elongation of carbon linker to 4C and insertion of a double bond into 4C substances increase AChE reactivators' cytotoxicity. Acknowledgements: This work was supported by a long-term organization development plan Medical Aspects of Weapons of Mass Destruction of the Faculty of Military Health Sciences, University of Defence.

Keywords: acetylcholinesterase, cytotoxicity, organophosphorus poisoning, reactivators of acetylcholinesterase

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Authors: Afzan Mahmad, Santibuana Abd Rahman, Gouri Kumar Dash, Mohd. Syafiq Bin Abdullah

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Antibacterial activity of the methanol extract of fruit peel of Punica granatum Linn (Family: Punicaceae) was evaluated against two Gram positive and two Gram negative bacteria. The Gram positive bacteria included Staphylococcus aureus, Streptococcus pneumoniae and the Gram negative organisms included Escherichia coli and Pseudomonas aeruginosa respectively. The culture media used for antibacterial assay was Mueller Hinton agar for the growth of S. aureus, E. coli, and P. aeruginosa. The media used for the growth of S. pneumoniae was Mueller Hinton blood agar. The antibacterial assay was performed through Disc diffusion technique. The methanol extract was tested at three different concentrations (50, 100 and 200 mg/ml). Standard antibiotic discs containing vancomycin (30 μg) for S. pneumoniae, penicillin (10 units) for S. aureus, ceftriaxone (30 μg) for E. coli and ciprofloxacin (5 μg) for P. aeruginosa were used for the activity comparison. The results of the study revealed that the extract possesses antibacterial activity against S. aureus, S. pneumoniae and P. aeruginosa at all tested concentrations. The maximum zone of inhibition of 19 mm of the extract at 200 mg/ml was observed against S. pneumoniae. However, no zone of inhibition was observed against E. coli at the tested concentrations of the extract. Based on the results obtained in this study, it may be concluded that the fruit peel of P. granatum possess broad spectrum of antibacterial activity against a number bacteria.

Keywords: Punica granatum Linn., methanol extract, antibacterial, zone of inhibition

Procedia PDF Downloads 394

Authors: Ali Essam El Shazly

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The slum survey of 'Sequina' area in Alexandria details the building rooms of twenty-building samples according to the integral measure of space syntax. The essence of room organization sets the most integrative 'visitor' domain between the 'inhabitant' wings of less integrated 'parent' than the 'children' structure with visual ring of 'balcony' space. Despite the collective real relative asymmetry of 'pheno-type' aggregation, the relative asymmetry of individual layouts reveals 'geno-type' structure of spatial diversity. The multifunction of rooms optimizes the integral structure of graph and visibility merge, which contrasts with the deep tailing structure of distinctive social domains. The most integrative layout inverts the geno-type into freed rooms of shallow 'inhabitant' domain against the off-centered 'visitor' space, while the most segregated layout further restricts the pheno-type through isolated 'visitor' from 'inhabitant' domains across the 'staircase' public domain. The catalyst 'kitchen & living' spaces demonstrate multi-structural dimensions among the various social domains. The former ranges from most exposed central integrity to the most hidden 'motherhood' territories. The latter, however, mostly integrates at centrality or at the further ringy 'childern' domain. The study concludes social structure of spatial integrity for redevelopment, which is determined through the micro-level survey of rooms with integral dimensions.

Keywords: Alexandria, Sequina slum, spatial integration, space syntax

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Authors: Perumal Siddhuraju, Arumugam Abirami, Gunasekaran Nagarani, Marimuthu Sangeethapriya

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Aqueous ethanol and aqueous acetone extracts of Moringa oleifera (outer pericarp of immature fruit and flower) and Sesbania grandiflora white variety (flower and leaf) were examined for radical scavenging capacities and antioxidant activities. Ethanol extract of S. grandiflora (flower and leaf) and acetone extract of M. oleifera (outer pericarp of immature fruit and flower) contained relatively higher levels of total dietary phenolics than the other extracts. The antioxidant potential of the extracts were assessed by employing different in vitro assays such as reducing power assay, DPPH˙, ABTS˙+ and ˙OH radical scavenging capacities, antihemolytic assay by hydrogen peroxide induced method and metal chelating ability. Though all the extracts exhibited dose dependent reducing power activity, acetone extract of all the samples were found to have more hydrogen donating ability in DPPH˙ (2.3% - 65.03%) and hydroxyl radical scavenging systems (21.6% - 77.4%) than the ethanol extracts. The potential of multiple antioxidant activity was evident as it possessed antihemolytic activity (43.2 % to 68.0 %) and metal ion chelating potency (45.16 - 104.26 mg EDTA/g sample). The result indicate that acetone extract of M. oleifera (OPIF and flower) and S. grandiflora (flower and leaf) endowed with polyphenols, could be utilized as natural antioxidants/nutraceuticals.

Keywords: antioxidant activity, Moringa oleifera, polyphenolics, Sesbania grandiflora, underutilized vegetables

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Authors: M. A. C. S. Sampath Fernando, James M. Curran, Renate Meyer

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Forensic DNA analysis has received much attention over the last three decades, due to its incredible usefulness in human identification. The statistical interpretation of DNA evidence is recognised as one of the most mature fields in forensic science. Peak heights in an Electropherogram (EPG) are approximately proportional to the amount of template DNA in the original sample being tested. A stutter is a minor peak in an EPG, which is not masking as an allele of a potential contributor, and considered as an artefact that is presumed to be arisen due to miscopying or slippage during the PCR. Stutter peaks are mostly analysed in terms of stutter ratio that is calculated relative to the corresponding parent allele height. Analysis of mixture profiles has always been problematic in evidence interpretation, especially with the presence of PCR artefacts like stutters. Unlike binary and semi-continuous models; continuous models assign a probability (as a continuous weight) for each possible genotype combination, and significantly enhances the use of continuous peak height information resulting in more efficient reliable interpretations. Therefore, the presence of a sound methodology to distinguish between stutters and real alleles is essential for the accuracy of the interpretation. Sensibly, any such method has to be able to focus on modelling stutter peaks. Bayesian nonparametric methods provide increased flexibility in applied statistical modelling. Mixture models are frequently employed as fundamental data analysis tools in clustering and classification of data and assume unidentified heterogeneous sources for data. In model-based clustering, each unknown source is reflected by a cluster, and the clusters are modelled using parametric models. Specifying the number of components in finite mixture models, however, is practically difficult even though the calculations are relatively simple. Infinite mixture models, in contrast, do not require the user to specify the number of components. Instead, a Dirichlet process, which is an infinite-dimensional generalization of the Dirichlet distribution, is used to deal with the problem of a number of components. Chinese restaurant process (CRP), Stick-breaking process and Pólya urn scheme are frequently used as Dirichlet priors in Bayesian mixture models. In this study, we illustrate an infinite mixture of simple linear regression models for modelling stutter ratio and introduce some modifications to overcome weaknesses associated with CRP.

Keywords: Chinese restaurant process, Dirichlet prior, infinite mixture model, PCR stutter

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Authors: Allulu Yousef Alturki, Raghad Abdullah Alshafi, Sara Abdulaziz Alghashem, Sahar Saleh Alghamdi, Rasha Saad Suliman, Zeyad Alehaideb, Rizwan Ali

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Cancer is recognized as a worldwide public health concern. Therefore, there is a continuous quest to discover new effective medications with less side-effects. In recent years, researchers have shown an increased interest in medicinal plants as several plant species have shown promising biological activities. Thus, we seek to investigate three medicinal herbs that are commonly-found in the Middle Easternregion and yet have not been explored in depth, including plants belonging to the Rhamnaceae, Polygonaceae, and Apocynaceaeplant families. Initially, we investigated using three types of cancer cell lines for breast, colorectal, and liver cancers. We performed high Content Imaging (HCI)-Apoptosis Assay and ApoTox-Glo™ Triplex Assay on KAIMRC2 and HCT8 cell lines. The highest activity of HCI-Apoptosis Assay was with Calligonumcomosum and Ziziphusnummularia in ethanol, followed by Calotropis procera and Ziziphusnummularia in ethyl acetate. The IC50values for the families of Rhamnaceae, Polygonaceae, and Apocynaceae in HepG2 and HCT8 cell lines ranged from 0.089 to 9.84mg/mL and 0.080to 15.08mg/mL, respectively. Further screening was conducted on an additional two cell lines, namely the MDA-MB-231 and KAIMRC2, for selected seven extracts with the highest activity having IC50values ranged from 0.058 to0.51mg/mL and 0.029 to0.19mg/mL, respectively. Continuous scientific investigations to isolate and characterize the potent bioactive phytochemical(s) are warranted. Funding: The authors acknowledge financial support from King Abdullah International Medical Research Center (KAIMRC), Ministry of National Guard Health Affairs, Riyadh, Kingdom of Saudi Arabia. Institutional Review Board Statement: The study was approved by the Institutional Review Board of the Institutional Review Board of King Abdullah International Medical Research Center (SP21R/463/12, 24 January 2022). Acknowledgments: The authors want to express their gratitude to the College of Pharmacy (COP) at King Saud bin Abdulaziz University for Health Sciences (KSAU-HS) and King Abdullah International Medical Research Center (KAIMRC) for their continued support.

Keywords: rhamnaceae, polygonaceae, apocynaceae, natural products

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Authors: Aminu Bashir Mohammad, Agwu Ezera, Abdulrazaq G. Habib, Garba Iliyasu

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Background: Drug resistance tuberculosis (DR-TB) continues to be a challenge in developing countries with poor resources. Routine screening for primary DR-TB before commencing treatment is not done in public hospitals in Nigeria, even with the large body of evidence that shows a high prevalence of primary DR-TB. Data on drug resistance and its genetic determinant among follow up TB patients is lacking in Nigeria. Hence the aim of this study was to determine the prevalence and genetic determinant of drug resistance among follow up TB patients in a tertiary hospital in Nigeria. Methods: This was a cross-sectional laboratory-based study conducted on 384 sputum samples collected from consented follow-up tuberculosis patients. Standard microbiology methods (Zeil-Nielsen staining and microscopy) and PCR (Line Probe Assay)] were used to analyze the samples collected. Person’s Chi-square was used to analyze the data generated. Results: Out of three hundred and eighty-four (384) sputum samples analyzed for mycobacterium tuberculosis (MTB) and DR-TB twenty-five 25 (6.5%) were found to be AFB positive. These samples were subjected to PCR (Line Probe Assay) out of which 18(72%) tested positive for DR-TB. Mutations conferring resistance to rifampicin (rpo B) and isoniazid (katG, and or inhA) were detected in 12/18(66.7%) and 6/18(33.3%), respectively. Transmission dynamic of DR-TB was not significantly (p>0.05) dependent on demographic characteristics. Conclusion: There is a need to strengthened the laboratory capacity for diagnosis of TB and drug resistance testing and make these services available, affordable, and accessible to the patients who need them.

Keywords: drug resistance tuberculosis, genetic determinant, intensive phase, Nigeria

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Authors: Alazar Nebyou, Sujata Pandit

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Lactic acid bacteria (LAB) are essential ingredients in probiotic foods, intestinal microflora, and dairy products that are capable of coping up with harsh gastrointestinal tract conditions and are available in a variety of environments. The objective of this study is to evaluate the probiotic property of LAB isolated from bovine milk. Milk samples were collected from local dairy farms. Samples were obtained using sterile test tubes and transported to a laboratory in the icebox for further biochemical characterization. Preliminary physiological and biochemical identification of LAB isolates was conducted by growing on MRS agar after ten-fold serial dilution. Seven of the best isolates were selected for the evaluation of the probiotic property. The LAB isolates were checked for resistance to antibiotics and their antimicrobial activity by disc diffusion assay and agar well diffusion assay respectively. Bile salt hydrolase activity of isolates was studied by growing isolates in a BSH medium with bile salt. Cell surface property of isolates was assayed by studying their autoaggregation and coaggregation percentage with S. aerues. All isolates were found BSH positive. In addition, BCM2 and BGM1 were susceptible to all antibiotic disks except BBM1 which was resistant to all antibiotic disks. BCM1 and BGM1 had the highest autoaggregation and coaggregation potential respectively. Since all LAB isolates showed gastrointestinal tolerance and good cell surface property they could be considered as good potential probiotic candidates for treatment and probiotic starter culture preparation.

Keywords: probiotic, aggregation, lactic acid bacteria, antimicrobial activity

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Authors: Sachin Mulmi Shrestha, Xin Fang, Hui Ye, Lihua Ren, Qinghua Ji, Ruihua Shi

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Background: Esophageal squamous cell carcinoma (ESCC) is a tumor arising from esophageal epithelial cells and is one of the major disease subtype in Asian countries, including China. Esophageal cancer is the 7th highest incidence based on the 2020 data of GLOBOCAN. The pathogenesis of cancer is still not well understood as many molecular and genetic basis of esophageal carcinogenesis has yet to be clearly elucidated. Circular RNAs are RNA molecules that are formed by back-splicing covalently joined 3′- and 5′-endsrather than canonical splicing, and recent data suggest circular RNAs could sponge miRNAs and are enriched with functional miRNA binding sites. Hence, we studied the mechanism of circular RNA, its biological function, and the relationship between microRNA in the carcinogenesis of ESCC. Methods: 4 pairs of normal and esophageal cancer tissues were collected in Zhongda hospital, affiliated to Southeast University, and high-throughput RNA sequencing was done. The result revealed that circ_0032746 was upregulated, and thus we selected circ_0032746 for further study. The backsplice junction of circRNA was validated by sanger sequence, and stability was determined by RNASE R assay. The binding site of circRNA and microRNA was predicted by circinteractome,mirandaand RNAhybrid database. Furthermore, circRNA was silenced by siRNA and then by lentivirus. The regulatory axis of circ0032746/miR4270 was validated by shRNA, mimic, and inhibitor transfection. Then, in vitro experiments were performed to assess the role of circ0032746 on proliferation (CCK-8 assay and colon formation assay), migration and invasion (Transewell assay), and apoptosis of ESCC. Results: The upregulated circ0032746 was validated in 9 pairs of tissues and 5 types of cell lines by qPCR, which showed high expression and was statistically significant (P<0.005) ). Upregulated circ0032746 was silenced by shRNA, which showed significant knockdown in KYSE 30 and TE-1 cell lines expression compared to control. Nuclear and cytoplasmic mRNA fraction experiment displayed the cytoplasmic location of circ0032746. The sponging of miR4270 was validated by co-transfection of sh-circ0032746 and mimic or inhibitor. Transfection with mimic showed the decreased expression of circ_0032746, whereas inhibitor inhibited the result. In vitro experiments showed that silencing of circ_0032746 inhibited the proliferation, migration, and invasion compared to the negative control group. The apoptosis was seen higher in a knockdown group than in the control group. Furthermore, 11 common mircoRNA target mRNAs were predicted by Targetscan, MirTarbase, and miRanda database, which may further play role in the pathogenesis. Conclusion: Our results showed that novel circ_0032746 is upregulated in ESCC and plays role in itsoncogenicity. Silencing of circ_0032746 inhibits the proliferation and migration of ESCC whereas increases the apoptosis of cancer cells. Hence, circ0032746 acts as an oncogene in ESCC by sponging with miR4270 and could be a potential biomarker in the diagnosis of ESCC in the future.

Keywords: circRNA, esophageal squamous cell carcinoma, microRNA, upregulated

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Authors: Richard I. Licayan, Aisle Janne B. Dagpin, Romeo M. Del Rosario, Nenita D. Palmes

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Hiptage benghalensis, Antigonon leptopus, Macroptillium atropurpureum, and Dioscorea bulbifera L. are herbal weeds that have been used by traditional healers in rural communities in the Philippines as medicine. In this study, the basic pharmacological components of the crude secondary metabolites extracted from the four herbal weeds and their in vitro antioxidant properties was investigated to provide baseline data for the possible development of these metabolites in pharmaceutical products. Qualitative screening of the secondary metabolites showed that alkaloids, tannins, saponins, steroids, and flavonoids were present in their leaf extracts. All of the plant extracts showed varied antioxidant activity. The greatest DPPH radical scavenging activity was observed in H. begnhalensis (84.64%), followed by A. leptopus (68.21%), M. atropurpureum (26.62%), and D. bulbifera L. (19.04%). The FRAP assay revealed that H. benghalensis had the highest antioxidant activity (8.32 mg/g) while ABTS assay showed that M. atropurpureum had the strongest scavenging ability of free radicals (0.0842 mg Trolox/g). The total flavonoid content (TFC) analysis showed that D. bulbifera L. had the highest TFC (420.35 mg quercetin per gram-dried material). The total phenolic content (TPC) of the four herbal weeds showed large variations, between 26.56±0.160 and 55.91±0.087 mg GAE/g dried material. The plant leaf extracts arranged in increasing values of TPC are H. benghalensis (26.565) < A. leptopus (37.29) < D. bulbifera L. (46.81) < M. atropurpureum (55.91). The obtained results may support their use in herbal medicine and as baseline data for the development of new drugs and standardized phytomedicines.

Keywords: antioxidant properties, total flavonoids, total phenolics, creeping herbal weeds

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Authors: Duanghathai Saipinta, Tanittian Panyamongkol, Witaya Suriyasathaporn

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Animal management aims to provide a suitable environment for animals allowing maximal productivity in those animals. Prevention of disease is an important part of animal management. Foot-and-mouth disease (FMD) is a highly contagious viral disease in cattle and is an economically important animal disease worldwide. Monitoring the FMD virus in farms is useful management for the prevention of the FMD outbreak. A recent publication indicated collection samples from nasal swabs can be used for monitoring FMD in symptomatic cows. Therefore, the objectives of this study were to determine the FMD virus in asymptomatic dairy cattle using nasal swab samples during the absence of an FMD outbreak. The study was conducted from December 2020 to June 2021 using 185 asymptomatic signs of FMD dairy cattle in Chiang Mai Province, Thailand. By random cow selection, nasal mucosal swabs were used to collect samples from the selected cows and then were to evaluate the presence of FMD viruses using the real-time rt-PCR assay. In total, 4.9% of dairy cattle detected FMD virus, including 2 dairy farms in Mae-on (8 samples; 9.6%) and 1 farm in the Chai-Prakan district (1 sample; 1.2%). Interestingly, both farms in Mae-on were the outbreak of the FMD after this detection for 6 months. This indicated that the FMD virus presented in asymptomatic cattle might relate to the subsequent outbreak of FMD. The outbreak demonstrates the presence of the virus in the environment. In conclusion, monitoring of FMD can be performed by nasal swab collection. Further investigation is needed to show whether the FMD virus presented in asymptomatic FMD cattle could be the cause of the subsequent FMD outbreak or not.

Keywords: cattle, foot-and-mouth disease, nasal swab, real-time rt-PCR assay

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Authors: Salma Baig, Bakrudeen Ali Ahmad, Ainnul Hamidah Syahadah Azizan, Hapipah Mohd Ali, Elham Rouhollahi, Mahmood Ameen Abdulla

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Free radical damage induced by reactive oxygen species (ROS) contributes to etiology of many chronic diseases, cancer being one of them. Recent studies have been successful in ROS targeted therapies via antioxidants using mouse models in cancer therapeutics. The present study was designed to scrutinize anticancer activity, antioxidant activity of 5 different extracts of Thymus serpyllum in MDA-MB-231, MCF-7, HepG2, HCT-116, PC3, and A549. Identification of the phytochemicals present in the most active extract of Thymus serpyllum was conducted using gas chromatography coupled with mass spectrophotometry and antioxidant activity was measured by using DPPH radical scavenging and FRAP assay. Anticancer impact of the extract in terms of IC50 was evaluated using MTT cell viability assay. Results revealed that the hexane extract showed the best anticancer activity in HepG2 (Liver Carcinoma Cell Line) with an IC50 value of 23 ± 0.14 µg/ml followed by 25 µg/ml in HCT-116 (Colon Cancer Cell Line), 30 µm/ml in MCF-7 (Breast Cancer Cell Line), 35 µg/ml in MDA-MB-231 (Breast Cancer Cell Line), 57 µg/ml in PC3 (Prostate Cancer Cell Line) and 60 µg/ml in A549 (Lung Carcinoma Cell Line). GC-MS profile of the hexane extract showed the presence of 31 compounds with carvacrol, thymol and thymoquione being the major compounds. Phenolics such as Vitamin E, terpinen-4-ol, borneol and phytol were also identified. Hence, here we present the first report on cytotoxicity of hexane extract of Thymus serpyllum extract in HepG2 cell line with a robust anticancer activity with an IC50 of 23 ± 0.14 µg/ml.

Keywords: Thymus serpyllum L., hexane extract, GC-MS profile, antioxidant activity, anticancer activity, HepG2 cell line

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Authors: Abbas Jafarizad, Duygu Ekinci

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In this work targeted drug delivery is proposed to decrease adverse effect of drugs with concomitant reduces in consumption and treatment outgoings. Nanoparticles (NPs) can be prepared from a variety of materials such as lipid, biodegradable polymer that prevent the drugs cytotoxicity in healthy cells, etc. One of the most important drugs used in chemotherapy is mitoxantrone (MTX) which prevents cell proliferation by inhibition of topoisomerase II and DNA repair; however, it is not selective and has some serious side effects. In this study, mentioned aim is achieved by using several nanocarriers like magnetite (Fe3O4) and their composites with magnetic graphene oxide (Fe3O4@GO). Also, cytotoxic potential of Fe3O4, Fe3O4-MTX, and Fe3O4@GO-MTX nanocomposite were evaluated on mesenchymal stem cells (MSCs). In this study, we reported the synthesis of monodisperse Fe3O4 NPs and Fe3O4@GO nanocomposite and their structures were investigated by using field emission scanning electron microscope (FESEM), Fourier transform infrared (FTIR) spectra, atomic force microscopy (AFM), Brauneur Emmet Teller (BET) isotherm and contact angle studies. Moreover, we used 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay to evaluate cytotoxic effects of MTX, Fe3O4 NPs, Fe3O4-MTX and Fe3O4@GO-MTX nanocomposite on MSCs. The in-vitro MTT results indicated that all concentrations of MTX and Fe3O4@GO-MTX nanocomposites showed cytotoxic effects while all concentrations of Fe3O4 NPs and Fe3O4-MTX NPs did not show any cytotoxic effect on stem cells. The results from this study indicated that using Fe3O4 NPs as anticancer drug delivery systems could be a trustworthy method for cancer treatment. But for reaching excellent and accurate results, further investigation is necessary.

Keywords: mitoxantrone, magnetite, magnetic graphene oxide, MTT assay, mesenchymal stem cells

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Authors: M. L. Dinis, M. C. Vila, A. Fiúza, A. Futuro, C. Nunes

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The present paper describes the study of paste tailings prepared in laboratory using gold tailings, produced in a Finnish gold mine with the incorporation of coal ash. Natural leaching tests were conducted with the original materials (tailings, fly and bottom ashes) and also with paste mixtures that were prepared with different percentages of tailings and ashes. After leaching, the solid wastes were physically and chemically characterized and the results were compared to those selected as blank – the unleached samples. The tailings and the coal ash, as well as the prepared mixtures, were characterized, in addition to the textural parameters, by the following measurements: grain size distribution, chemical composition and pH. Mixtures were also tested in order to characterize their mechanical behavior by measuring the flexural strength, the compressive strength and the consistency. The original tailing samples presented an alkaline pH because during their processing they were previously submitted to pressure oxidation with destruction of the sulfides. Therefore, it was not possible to ascertain the effect of the coal ashes in the acid mine drainage. However, it was possible to verify that the paste reactivity was affected mostly by the bottom ash and that the tailings blended with bottom ash present lower mechanical strength than when blended with a combination of fly and bottom ash. Surface paste disposal offer an attractive alternative to traditional methods in addition to the environmental benefits of incorporating large-volume wastes (e.g. bottom ash). However, a comprehensive characterization of the paste mixtures is crucial to optimize paste design in order to enhance engineer and environmental properties.

Keywords: coal ash, mine tailings, paste blends, surface disposal

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Authors: Changhan Ouyang, Zhonglin Xie

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In response to proatherosclerotic factors such as oxidized lipids, or to therapeutic interventions such as angioplasty, stents, or bypass surgery, vascular smooth muscle cells (VSMCs) migrate from the media to the intima, resulting in intimal hyperplasia, restenosis, graft failure, or atherosclerosis. These proatherosclerotic factors also activate autophagy in VSMCs. However, the functional role of autophagy in vascular health and disease remains poorly understood. In the present study, we determined the role of autophagy in the regulation of VSMC migration. Autophagy activity in cultured human aortic smooth muscle cells (HASMCs) and mouse carotid arteries was measured by Western blot analysis of microtubule-associated protein 1 light chain 3 B (LC3B) and P62. The VSMC migration was determined by scratch wound assay and transwell migration assay. Ex vivo smooth muscle cell migration was determined using aortic ring assay. The in vivo SMC migration was examined by staining the carotid artery sections with smooth muscle alpha actin (alpha SMA) after carotid artery ligation. To examine the relationship between autophagy and neointimal hyperplasia, C57BL/6J mice were subjected to carotid artery ligation. Seven days after injury, protein levels of Atg5, Atg7, Beclin1, and LC3B drastically increased and remained higher in the injured arteries three weeks after the injury. In parallel with the activation of autophagy, vascular injury-induced neointimal hyperplasia as estimated by increased intima/media ratio. The en face staining of carotid artery showed that vascular injury enhanced alpha SMA staining in the intimal cells as compared with the sham operation. Treatment of HASMCs with platelet-derived growth factor (PDGF), one of the major factors for vascular remodeling in response to vascular injury, increased Atg7 and LC3 II protein levels and enhanced autophagosome formation. In addition, aortic ring assay demonstrated that PDGF treated aortic rings displayed an increase in neovessel formation compared with control rings. Whole mount staining for CD31 and alpha SMA in PDGF treated neovessels revealed that the neovessel structures were stained by alpha SMA but not CD31. In contrast, pharmacological and genetic suppression of autophagy inhibits VSMC migration. Especially, gene silencing of Atg7 inhibited VSMC migration induced by PDGF. Furthermore, three weeks after ligation, markedly decreased neointimal formation was found in mice treated with chloroquine, an inhibitor of autophagy. Quantitative morphometric analysis of the injured vessels revealed a marked reduction in the intima/media ratio in the mice treated with chloroquine. Conclusion: Autophagy activation increases VSMC migration while autophagy suppression inhibits VSMC migration. These findings suggest that autophagy suppression may be an important therapeutic strategy for atherosclerosis and intimal hyperplasia.

Keywords: autophagy, vascular smooth muscle cell, migration, neointimal formation

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Authors: Nadhiya N. Subramony, Jenifer Vaughan, Lesley E. Scott

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In South Africa, the World Health Organisation estimated 454000 new cases of Mycobacterium tuberculosis (M.tb) infection (MTB) in 2015. Disseminated tuberculosis arises from the haematogenous spread and seeding of the bacilli in extrapulmonary sites. The gold standard for the detection of MTB in bone marrow is TB culture which has an average turnaround time of 6 weeks. Histological examinations of trephine biopsies to diagnose MTB also have a time delay owing mainly to the 5-7 day processing period prior to microscopic examination. Adding to the diagnostic delay is the non-specific nature of granulomatous inflammation which is the hallmark of MTB involvement of the bone marrow. A Ziehl-Neelson stain (which highlights acid-fast bacilli) is therefore mandatory to confirm the diagnosis but can take up to 3 days for processing and evaluation. Owing to this delay in diagnosis, many patients are lost to follow up or remain untreated whilst results are awaited, thus encouraging the spread of undiagnosed TB. The Xpert® MTB/RIF (Cepheid, Sunnyvale, CA) is the molecular test used in the South African national TB program as the initial diagnostic test for pulmonary TB. This study investigates the optimisation and performance of the Xpert® MTB/RIF on bone marrow aspirate specimens (BMA), a first since the introduction of the assay in the diagnosis of extrapulmonary TB. BMA received for immunophenotypic analysis as part of the investigation into disseminated MTB or in the evaluation of cytopenias in immunocompromised patients were used. Processing BMA on the Xpert® MTB/RIF was optimised to ensure bone marrow in EDTA and heparin did not inhibit the PCR reaction. Inactivated M.tb was spiked into the clinical bone marrow specimen and distilled water (as a control). A volume of 500mcl and an incubation time of 15 minutes with sample reagent were investigated as the processing protocol. A total of 135 BMA specimens had sufficient residual volume for Xpert® MTB/RIF testing however 22 specimens (16.3%) were not included in the final statistical analysis as an adequate trephine biopsy and/or TB culture was not available. Xpert® MTB/RIF testing was not affected by BMA material in the presence of heparin or EDTA, but the overall detection of MTB in BMA was low compared to histology and culture. Sensitivity of the Xpert® MTB/RIF compared to both histology and culture was 8.7% (95% confidence interval (CI): 1.07-28.04%) and sensitivity compared to histology only was 11.1% (95% CI: 1.38-34.7%). Specificity of the Xpert® MTB/RIF was 98.9% (95% CI: 93.9-99.7%). Although the Xpert® MTB/RIF generates a faster result than histology and TB culture and is less expensive than culture and drug susceptibility testing, the low sensitivity of the Xpert® MTB/RIF precludes its use for the diagnosis of MTB in bone marrow aspirate specimens and warrants alternative/additional testing to optimise the assay.

Keywords: bone marrow aspirate , extrapulmonary TB, low sensitivity, Xpert® MTB/RIF

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Authors: Mohammad K. Parvez, Ahmed H. Arbab, Mohammed S. Al-Dosari, Adnan J. Al-Rehaily

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We investigated ex vivo and in vivo antioxidative and hepatoprotective effect of Aerva javanica. Total ethanol extract of A. javanica aerial parts was prepared, and tested on DCFH-toxicated HepG2 cell in CCl4-injured Wistar rats. MTT-assay was used to determine cell viability, and serum biochemical markers of liver injury as well as histopathology were performed. In vitro DPPH and β-carotene free-radical scavenging assay and phytochemical screening of the extract was done. Furthermore, A. javanica total extract was standardized and validated by HPTLC method. While DCFH-injured cells were recovered to about 56.7% by 100 microg/ml of the extract, a 200 microg/ml dose resulted in hepatocytes recovery by about 90.2%. Oral administration of the extract (100 and 200 mg/kg.bw/day) significantly normalized the serum SGOT, SGPT, GGT, ALP, bilirubin, cholesterol, HDL, LDL, VLDL, TG and MDA levels, including tissue NP-SH and TP in CCl4-injured rats. In addition, the histopathology of dissected liver also revealed that A. javanica cured the tissue lesion compared to reference drug, Silymarin. In vitro assays revealed strong free-radical scavenging ability of the extract and presence of alkaloids, flavonoids, tannins, sterols and saponins where Rutin, a well-known antioxidant flavonoid was identified. Our finding therefore, suggests the therapeutic potential of A. javanica in various liver diseases. However, isolation of the active principles, their mechanism of action and other therapeutic contribution remain to be addressed.

Keywords: Aerva javanica, antioxidant, hepatoprotection, rutin

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Authors: V. Nigam, V. Nambiar

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Aegle Marmelos leaf has been used as a remedy for various gastrointestinal infections and lowering blood sugar level in traditional system of medicine in India due to the presence of various constituents such as flavonoids, tannins and alkaloids (eg. Aegelin, Marmelosin, Luvangetin).The objective of the present study was to evaluate the total antioxidant activity, total and individual phenol content of the wild and cultivated variety of Aegle marmelos leaves to assess the role of this plant in ethanomedicine in India. The methanolic extracts of the leaves were screened for total antioxidant capacity through Ferric Reducing Antioxidant Potential (FRAP) and 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging assay; Total Phenol content (TPC) through spectrophotometric technique based on Folin Ciocalteau assay and for qualitative estimation of phenols, High performance Liquid Chromatography was used. The TPC of wild and cultivated variety was 7.6% and 6.5% respectively whereas HPLC analysis for quantification of individual polyphenol revealed the presence of gallic acid, chlorogenic acid and Ferullic acid in wild variety whereas gallic acid, Ferullic acid and pyrocatechol in cultivated variety. FRAP values and IC 50 value (DPPH) for wild and cultivated variety was 14.65 μmol/l and 11.80μmol/l; 437 μg/ml and 620μg/ml respectively and thus it can be used as potential inhibitor of free radicals. The wild variety was having more antioxidant capacity than the cultivated one it can be exploited further for its therapeutic application. As Aegle marmelos is rich in antioxidant, it can be used as food additives to delay the oxidative deterioration of foods and as nutraceutical in medicinal formulation against degenerative diseases like diabetes.

Keywords: antioxidant activity, aegle marmelos, antidiabetic, nutraceutical

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Authors: Pravaree Phuneerub, Chanida Palanuvej, Nijsiri Ruangrungsi

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Cymbopogon nardus Rendel (Family Gramineae) is commonly known as citronella grass. The dried root of C. nardus is used for antipyretic, anti-inflammation, anti-analgesic and anticancer in traditional Thai medicine. Transverse sectional and pulverized C. nardus root were illustrated. The volatile oil was extracted from oil gland by hydrodistillation and analysed by GC/MS. Cymbopogon nardus root was exhaustively extracted by continuously maceration in ethanol and water respectively. The mutagenic and antimutagenic properties of the ethanol extract and fractionated water extract of C. nardus root were evaluated by Ames assay using the S. typhimurium strains TA98 and TA100 as the models. The result indicated that the anatomical character of root transverse section displayed epidermis, parenchyma, oil gland, phloem, xylem vessel, endodermis and pith. Histological characters of root powder showed parenchyma containing oleoresin, parenchyma in longitudinal view, reticulate vessel, annular vessel, starch granules and fragment of fiber. The root volatile oil was rich in sesquiterpenes dominated by elemol (22.87%) and alpha-eudesmol (16.09%). For mutagenic activity, the both extracts of C. nardus were no mutagenic toward S. typhimurium strains TA98 and TA100. Furthermore, the ethanol extract and fractionated water extract of C. nardus root demonstrated strong antimutagenic effect against of nitrite treated 1-aminopyrene to S. typhimurium strains TA98 and TA100. This present investigation suggested that the dried root extract of C. nardus can be further developed as promising antimutagenic agent.

Keywords: Cymbopogon nardus, volatile oil analysis, mutagenic, antimutagenic effect, Ames Salmonella assay

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Authors: Harukuni Tokuda, Takanari Arai, Xu FengHao, Nobutaka Suzuki

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In our continuous search for anti-tumor promoting, chemopreventive active potency from natural source material, a kind of healthy tea, Gromwell seed (Coix lachryma-jobi) ext., and including compounds Monoolein and Trilinolein have been screened using the in vitro synergistic assay indicated by inhibitory effects on the induction of Epstein-Barr virus early antigen (EBV-EA) by TPA. In assay, Gromwell seed aqueous extract and hot aqueous extract exhibited the potential inhibitory effects on EBV-EA activation without strong cytotoxicity on Raji cells. In our experimental system, the inhibitory effects of both Gromwell extracts and compounds were greater than that of beta-carotene, which is known anti-tumor promoting agent and/or chemopreventive agent. These compounds were evaluated for their in vitro inhibitory effect on EBV-EA activation induced by TPA. The percentages of the inhibition of TPA-induced EBV-EA activation for these materials were 60% and 30% at concentration 100 μg. Based on the results obtained in vitro, we studied the inhibitory effect of compounds, in an in vivo two-stage carcinogenesis test of mouse skin papilloma using DMBA as an initiator and TPA as a potential promoter. The control animals showed a 100% incidence of papilloma at 20 weeks after DMBA-TPA tumor promotion, while treatment with compounds reduced the percentage of number of tumor to 60 % after 20 weeks. Results from in vitro and in vivo studies showing chemopreventive activity against TPA promoting stage and these data support the effective potency of carcinogenic stage in clinical evaluation of integrative oncology.

Keywords: gromwell seed, medicinal plant, chemoprevention, pharmaceutical medicine

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Authors: Yogesh Dalvi, Ruby Varghese, Nibu Varghese, C. K. Krishnan Nair

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Introduction: The Genus Phellinus, locally known as Phansomba is a well-known traditional folk medicine. Especially, in Western Ghats of India, many tribes use several species of Phellinus for various ailments related to teeth, throat, tongue, stomach and even wound healing. It is one of the few mushrooms which play a pivotal role in Ayurvedic Dravyaguna. Aim: The present study focuses on to investigate phytochemical analysis, antioxidant, and antitumor (in vitro and in vivo) potential of Phellinus robinae from South India, Kerala Material and Methods: The present study explores the following: 1. Phellinus samples were collected from Ranni, Pathanamthitta district of Kerala state, India from Artocarpus heterophyllus Lam. and species were identified using rDNA region. 2. The fruiting body was shadow dried, powdered and extracted with 50% alcohol using water bath at 60°C which was further condensed by rotary evaporator and lyophilized at minus 40°C temperature. 3. Secondary metabolites were analyzed by using various phytochemical screening assay (Hager’s Test, Wagner’s Test, Sodium hydroxide Test, Lead acetate Test, Ferric chloride Test, Folin-ciocalteu Test, Foaming Test, Benedict’s test, Fehling’s Test and Lowry’s Test). 4. Antioxidant and free radical scavenging activity were analyzed by DPPH, FRAP and Iron chelating assay. 5. The antitumor potential of Water alcohol extract of Phellinus (PAWE) is evaluated through In vitro condition by Trypan blue dye exclusion method in DLA cell line and In vivo by murine model. Result and Discussion: Preliminary phytochemical screening by various biochemical tests revealed presence of a variety of active secondary molecules like alkaloids, flavanoids, saponins, carbohydrate, protein and phenol. In DPPH and FRAP assay PAWE showed significantly higher antioxidant activity as compared to standard Ascorbic acid. While, in Iron chelating assay, PAWE exhibits similar antioxidant activity that of Butylated Hydroxytoluene (BHT) as standard. Further, in the in vitro study, PAWE showed significant inhibition on DLA cell proliferation in dose dependent manner and showed no toxicity on mice splenocytes, when compared to standard chemotherapy drug doxorubicin. In vivo study, oral administration of PAWE showed dose dependent tumor regression in mice and also raised the immunogenicity by restoring levels of antioxidant enzymes in liver and kidney tissue. In both in vitro and in vivo gene expression studies PAWE up-regulates pro-apoptotic genes (Bax, Caspases 3, 8 and 9) and down- regulates anti-apoptotic genes (Bcl2). PAWE also down regulates inflammatory gene (Cox-2) and angiogenic gene (VEGF). Conclusion: Preliminary phytochemical screening revealed that PAWE contains various secondary metabolites which contribute to its antioxidant and free radical scavenging property as evaluated by DPPH, FRAP and Iron chelating assay. PAWE exhibits anti-proliferative activity by the induction of apoptosis through a signaling cascade of death receptor-mediated extrinsic (Caspase8 and Tnf-α), as well as mitochondria-mediated intrinsic (caspase9) and caspase pathways (Caspase3, 8 and 9) and also by regressing angiogenic factor (VEGF) without any inflammation or adverse side effects. Hence, PAWE serve as a potential antioxidant and antitumor agent.

Keywords: antioxidant, antitumor, Dalton lymphoma ascites (DLA), fungi, Phellinus robinae

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