Search results for: protein isolates
1519 Aminopeptidase P (DAP) Expression Pattern in Drosophila Melanogaster
Authors: Suneeta Gireesh Panicker
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Aim: Aminopeptidase P (APP) is an enzyme that has specificity for proline, can specifically cleave Xaa-Proline peptides and is a metallo-aminopeptidase. The bonds nearby to the imino acid proline are tough to cleave by many peptidases, but APP can specifically break peptide bonds engaged with proline. Membrane-bound form and a cytosolic form are the two forms in which this enzyme exists. The exact physiological function of APP remains unclear and hence the present work attempts to determine it. Methods: In the present study, the expression pattern of cytosolic Aminopeptidase P (DAP) was determined in all the embryonic stages and larval stages of wild-type Drosophila by using polyclonal monospecific antibodies. To show the presence of DAP RNA in embryonic and larval stages, RNA in situ hybridization was performed. DAP promoter-LacZ fusion reporter gene vector was used to construct transgenic embryos to study the regulation pattern of DAP. To study the DAP expression profile, a transgenic fly consisting of a DAP promoter with β-gal and GFP reporter genes in front of it was constructed. Results: DAP protein expression was observed in neuroectodermal cells, posterior midgut primordium, proctodeum, ventral neuroblast and primordial stomatogastric nervous system. It was observed in the ventral cord and midgut in stage 12. The completely developed embryos showed the intense occurrence of it in the ventral cord and gut region. The eye-antennal disc, wing disc and leg disc also showed the presence of DAP protein. LacZ expression in transgenic embryos also showed the same pattern. Conclusion: Similar to various known multiple-functional proteins, DAP could be one with different functions at different stages and in different cells. Data presented here designates DAP functions in the early embryonic and imaginal dics differentiation and development, suggesting that it may be required for the metabolism of proteins like neuropeptides and tachykinins.Keywords: aminopeptidase P, in situ hybridization, transgenic fly, embryonic stages
Procedia PDF Downloads 891518 Evaluation of Anti-inflammatory Activities of Extracts Obtained from Capparis Erythrocarpos In-Vivo
Authors: Benedict Ofori, Kwabena Sarpong, Stephen Antwi
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Background: Medicinal plants are utilized all around the world and are becoming increasingly important economically. The WHO notes that ‘inappropriate use of traditional medicines or practices can have negative or dangerous effects and that future research is needed to ascertain the efficacy and safety of such practices and medicinal plants used by traditional medicine systems. The poor around the world have limited access to palliative care or pain relief. Pharmacologists have been focused on developing safe and effective anti-inflammatory drugs. Most of the issues related to their use have been linked to the fact that numerous traditional and herbal treatments are classified in different nations as meals or dietary supplements. As a result, there is no need for evidence of the quality, efficacy, or safety of these herbal formulations before they are marketed. The fact that access to drugs meant for pain relief is limited in low-income countries means advanced studies should be done on home drugs meant for inflammation to close the gap. Methods: The ethanolic extracts of the plant were screened for the presence of 10 phytochemicals. The Pierce BCA Protein Assay Kit was used for the determination of the protein concentration of the egg white. The rats were randomly selected and put in 6 groups. The egg white was sub-plantar injected into the right-hand paws of the rats to induce inflammation. The animals were treated with the three plant extracts obtained from the root bark, stem, and leaves of the plant. The control groups were treated with normal saline, while the standard groups were treated with standard drugs indomethacin and celecoxib. Plethysmometer was used to measure the change in paw volume of the animals over the course of the experiment. Results: The results of the phytochemical screening revealed the presence of reducing sugars and saponins. Alkaloids were present in only R.L.S (1:1:1), and phytosterols were found in R.L(1:1) and R.L.S (1:1:1). The estimated protein concentration was found to be 103.75 mg/ml. The control group had an observable increase in paw volume, which indicated that inflammation was induced during the 5 hours. The increase in paw volume for the control group peaked at the 1st hour and decreased gradually throughout the experiment, with minimal changes in the paw volumes. The 2nd and 3rd groups were treated with 20 mg/kg of indomethacin and celecoxib. The anti-inflammatory activities of indomethacin and celecoxib were calculated to be 21.4% and 4.28%, respectively. The remaining 3 groups were treated with 2 dose levels of 200mg/kg plant extracts. R.L.S, R.L, and S.R.L had anti-inflammatory activities of 22.3%, 8.2%, and 12.07%, respectively. Conclusions: Egg albumin-induced paw model in rats can be used to evaluate the anti-inflammatory activity of herbs that might have potential anti-inflammatory activity. Herbal medications have potential anti-inflammatory activities and can be used to manage various inflammatory conditions if their efficacy and side effects are well studied. The three extracts all possessed anti-inflammatory activity, with R.L.S having the highest anti-inflammatory activity.Keywords: inflammation, capparis erythrocarpos, anti-inflammatory activity, herbal medicine, paw volume, egg albumin
Procedia PDF Downloads 901517 Identification of the Key Enzyme of Roseoflavin Biosynthesis
Authors: V. Konjik, J. Schwartz, R. Sandhoff, M. Mack
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The rising number of multi-resistant pathogens demands the development of new antibiotics in order to reduce the lethal risk of infections. Here, we investigate roseoflavin, a vitamin B2 analogue which is produced by Streptomyces davawensis and Streptomyces cinnabarinus. We consider roseoflavin to be a 'Trojan horse' compound. Its chemical structure is very similar to riboflavin but in fact it is a toxin. Furthermore, it is a clever strategy with regard to the delivery of an antibiotic to its site of action but also with regard to the production of this chemical: The producer cell has only to convert a vitamin (which is already present in the cytoplasm) into a vitamin analog. Roseoflavin inhibits the activity of Flavin depending proteins, which makes up to 3.5 % of predicted proteins in organisms sequenced so far. We sequentially knocked out gene clusters and later on single genes in order to find the ones which are involved in the roseoflavin biosynthesis. Consequently, we identified the gene rosB, coding for the protein carrying out the first step of roseoflavin biosynthesis, starting form Flavin mononucleotide. Here we show, that the protein RosB has so far unknown features. It is per se an oxidoreductase, a decarboxylase and an aminotransferase, all rolled into one enzyme. A screen of cofactors revealed needs of oxygen, NAD+, thiamine and glutamic acid to carry out its function. Surprisingly, thiamine is not only needed for the decaboxylation step, but also for the oxidation of 8-demethyl-8-formyl Flavin mononucleotide. We had managed to isolate three different Flavin intermediates with different oxidation states, which gave us a mechanistic insight of RosB functionality. Our work points to a so far new function of thiamine in Streptomyces davawensis. Additionally, RosB could be extremely useful for chemical synthesis. Careful engineering of RosB may allow the site-specific replacement of methyl groups by amino groups in polyaromatic compounds of commercial interest. Finally, the complete clarification of the roseoflavin biosynthesis opens the possibility of engineering cost-effective roseoflavin producing strains.Keywords: antibiotic, flavin analogue, roseoflavin biosynthesis, vitamin B2
Procedia PDF Downloads 2441516 Extraction of Phycocyanin from Spirulina platensis by Isoelectric Point Precipitation and Salting Out for Scale Up Processes
Authors: Velasco-Rendón María Del Carmen, Cuéllar-Bermúdez Sara Paulina, Parra-Saldívar Roberto
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Phycocyanin is a blue pigment protein with fluorescent activity produced by cyanobacteria. It has been recently studied to determine its anticancer, antioxidant and antiinflamatory potential. Since 2014 it was approved as a Generally Recognized As Safe (GRAS) proteic pigment for the food industry. Therefore, phycocyanin shows potential for the food, nutraceutical, pharmaceutical and diagnostics industry. Conventional phycocyanin extraction includes buffer solutions and ammonium sulphate followed by chromatography or ATPS for protein separation. Therefore, further purification steps are time-requiring, energy intensive and not suitable for scale-up processing. This work presents an alternative to conventional methods that also allows large scale application with commercially available equipment. The extraction was performed by exposing the dry biomass to mechanical cavitation and salting out with NaCl to use an edible reagent. Also, isoelectric point precipitation was used by addition of HCl and neutralization with NaOH. The results were measured and compared in phycocyanin concentration, purity and extraction yield. Results showed that the best extraction condition was the extraction by salting out with 0.20 M NaCl after 30 minutes cavitation, with a concentration in the supernatant of 2.22 mg/ml, a purity of 3.28 and recovery from crude extract of 81.27%. Mechanical cavitation presumably increased the solvent-biomass contact, making the crude extract visibly dark blue after centrifugation. Compared to other systems, our process has less purification steps, similar concentrations in the phycocyanin-rich fraction and higher purity. The contaminants present in our process edible NaCl or low pHs that can be neutralized. It also can be adapted to a semi-continuous process with commercially available equipment. This characteristics make this process an appealing alternative for phycocyanin extraction as a pigment for the food industry.Keywords: extraction, phycocyanin, precipitation, scale-up
Procedia PDF Downloads 4401515 Combinational Therapeutic Targeting of BRD4 and CDK7 Synergistically Induces Anticancer Effects in Hepatocellular Carcinoma
Authors: Xinxiu Li, Chuqian Zheng, Yanyan Qian, Hong Fan
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Objectives: In hepatocellular carcinoma (HCC), oncogenes are continuously and robustly transcribed due to aberrant expression of essential components of the trans-acting super-enhancers (SE) complex. Preclinical and clinical trials are now being conducted on small-molecule inhibitors that target core-transcriptional components, including as transcriptional bromodomain protein 4 (BRD4) and cyclin-dependent kinase 7 (CDK7), in a number of malignant tumors. This study aims to explore whether co-overexpression of BRD4 and CDK7 is a potential marker of worse prognosis and a combined therapeutic target in HCC. Methods: The expression pattern of BRD4 and CDK7 and their correlation with prognosis in HCC were analyzed by RNA sequencing data and survival data of HCC patients from TCGA and GEO datasets. The protein levels of BRD4 and CDK7 were determined by immunohistochemistry (IHC), and survival data of patients were analyzed using the Kaplan-Meier method. The mRNA expression levels of genes in HCC cell lines were evaluated by quantitative PCR (q-PCR). CCK-8 and colony formation assays were conducted to assess cell proliferation of HCC upon treatment with BRD4 inhibitor JQ1 or/and CDK7 inhibitor THZ1. Results: It was shown that BRD4 and CDK7 were often overexpressed in HCCs and were associated with poor prognosis of HCC by analyzing the TCGA and GEO datasets. BRD4 or CDK7 overexpression was related to a lower survival rate. It's interesting to note that co-overexpression of CDK7 and BRD4 was a worse prognostic factor in HCC. Treatment with JQ1 or THZ1 alone had an inhibitory effect on cell proliferation; however, when JQ1 and THZ1 were combined, there was a more notable suppression of cell growth. At the same time, the combined use of JQ1 and THZ1 synergistically suppresses the expression of HCC driver genes. Conclusion: Our research revealed that BRD4 and CDK7 coupled can be a useful biomarker in HCC prognosis and the combination of JQ1 and THZ1 can be a promising therapeutic therapy against HCC.Keywords: BRD4, CDK7, cell proliferation, combined inhibition
Procedia PDF Downloads 551514 Effect of Yeast Culture (Saccharomyces cerevisiae) Supplementation on Growth Performance, Nutrients Digestibility, and Blood Metabolites in Beetal Male Goats
Authors: Saeed Ahmed, Tamoor Abbas, M. Amir, M. S. Iqbal, D. Hussain
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This study was conducted to evaluate the effect of supplementation of different levels of yeast culture (Saccharomyces cerevisiae) in Beetal male goats diets on growth performance, digestibility of nutrients and selected blood metabolites. Another objective was to determine the inclusion level of yeast culture for optimal growth performance of Beetal male goats. Eighteen (n=18) Beetal male goats were randomly assigned to three total mixed ration treatments (n=6 goats/treatment): T1, T2 and T3 containing 0gm, 3gm and 6gm/day yeast culture (YC) mixed with total mixed ration (TMR). The diets were iso-nitrogenous and iso-caloric having crude protein 15.2% and ME 2.6Mcal/kg. The total duration of the experiment was 8 weeks. Beetal bucks were fed on TMR diets (T1, T2 and T3) having blend of oat silage, Lucerne hay and concentrate mixed with yeast culture (YC). Bucks were housed individually and feed was offered @ 4% of body weight on dry matter basis. Samples of fresh feed and refusal were collected twice weekly of moisture percentage using hot air oven. Data for daily dry matter intake, body weight gain, nutrient digestibility and selected blood metabolites were analyzed through one-way ANOVA technique under Complete randomised design (SAS Institute Inc, 2002-03). Results were declared significant at P≤0.05. Overall, DMI was not affected (P≥0.05) by dietary treatments. Body weight gain, digestibility of crude protein and crude fibre were improved. Blood glucose concentration was detected higher in the group having supplementation of yeast culture (YC) 6gm/day compared to other two dietary treatments. This study suggested the positive impact of inclusion of yeast culture (YC) up to 6gm/day in the TMR diet for optimal growth performance and digestibility of nutrients in Beetal male goats.Keywords: yeast culture, growth performance, digestibility, beetle goat
Procedia PDF Downloads 1971513 Comparative Effects of Resveratrol and Energy Restriction on Liver Fat Accumulation and Hepatic Fatty Acid Oxidation
Authors: Iñaki Milton-Laskibar, Leixuri Aguirre, Maria P. Portillo
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Introduction: Energy restriction is an effective approach in preventing liver steatosis. However, due to social and economic reasons among others, compliance with this treatment protocol is often very poor, especially in the long term. Resveratrol, a natural polyphenolic compound that belongs to stilbene group, has been widely reported to imitate the effects of energy restriction. Objective: To analyze the effects of resveratrol under normoenergetic feeding conditions and under a mild energy restriction on liver fat accumulation and hepatic fatty acid oxidation. Methods: 36 male six-week-old rats were fed a high-fat high-sucrose diet for 6 weeks in order to induce steatosis. Then, rats were divided into four groups and fed a standard diet for 6 additional weeks: control group (C), resveratrol group (RSV, resveratrol 30 mg/kg/d), restricted group (R, 15 % energy restriction) and combined group (RR, 15 % energy restriction and resveratrol 30 mg/kg/d). Liver triacylglycerols (TG) and total cholesterol contents were measured by using commercial kits. Carnitine palmitoyl transferase 1a (CPT 1a) and citrate synthase (CS) activities were measured spectrophotometrically. TFAM (mitochondrial transcription factor A) and peroxisome proliferator-activator receptor alpha (PPARα) protein contents, as well as the ratio acetylated peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α)/Total PGC1α were analyzed by Western blot. Statistical analysis was performed by using one way ANOVA and Newman-Keuls as post-hoc test. Results: No differences were observed among the four groups regarding liver weight and cholesterol content, but the three treated groups showed reduced TG when compared to the control group, being the restricted groups the ones showing the lowest values (with no differences between them). Higher CPT 1a and CS activities were observed in the groups supplemented with resveratrol (RSV and RR), with no difference between them. The acetylated PGC1α /total PGC1α ratio was lower in the treated groups (RSV, R and RR) than in the control group, with no differences among them. As far as TFAM protein expression is concerned, only the RR group reached a higher value. Finally, no changes were observed in PPARα protein expression. Conclusions: Resveratrol administration is an effective intervention for liver triacylglycerol content reduction, but a mild energy restriction is even more effective. The mechanisms of action of these two strategies are different. Thus resveratrol, but not energy restriction, seems to act by increasing fatty acid oxidation, although mitochondriogenesis seems not to be induced. When both treatments (resveratrol administration and a mild energy restriction) were combined, no additive or synergic effects were appreciated. Acknowledgements: MINECO-FEDER (AGL2015-65719-R), Basque Government (IT-572-13), University of the Basque Country (ELDUNANOTEK UFI11/32), Institut of Health Carlos III (CIBERobn). Iñaki Milton is a fellowship from the Basque Government.Keywords: energy restriction, fat, liver, oxidation, resveratrol
Procedia PDF Downloads 2131512 Valorization of Waste and By-products for Protein Extraction and Functional Properties
Authors: Lorena Coelho, David Ramada, Catarina Nobre, Joaquim Gaião, Juliana Duarte
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The development of processes that allows the valorization of waste and by-products generated by industries is crucial to promote symbiotic relationships between different sectors and is mandatory to “close the loop” in the circular economy paradigm. In recent years, by-products and waste from agro-food and forestry sector have attracted attention due to their potential application and technical characteristics. The extraction of bio-based active compounds to be reused is in line with the circular bioeconomy concept trends, combining the use of renewable resources with the process’s circularity, aiming the waste reduction and encouraging reuse and recycling. Among different types of bio-based materials, which are being explored and can be extracted, proteins fractions are becoming an attractive new raw material. Within this context, BioTrace4Leather project, a collaboration between two Technological Centres – CeNTI and CTIC, and a company of Tanning and Finishing of Leather – Curtumes Aveneda, aims to develop innovative and biologically sustainable solutions for leather industry and accomplish the market circularity trends. Specifically, it aims to the valorisation of waste and by-products from the tannery industry through proteins extraction and the development of an innovative and biologically sustainable materials. The achieved results show that keratin, gelatine, and collagen fractions can be successfully extracted from hair and leather bovine waste. These products could be reintegrated into the industrial manufacturing process to attain innovative and functional textile and leather substrates. ACKNOWLEDGEMENT This work has been developed under BioTrace4Leather scope, a project co-funded by Operational Program for Competitiveness and Internationalization (COMPETE) of PORTUGAL2020, through the European Regional Development Fund (ERDF), under grant agreement Nº POCI-01-0247-FEDER-039867.Keywords: leather by-products, circular economy, sustainability, protein fractions
Procedia PDF Downloads 1611511 Impact of Different Ripening Accelerators on the Microbial Load and Proximate Composition of Plantain (Musa paradisiaca) and Banana (Musa sapientum), during the Ripening Process, and the Nutrition Implication for Food Security
Authors: Wisdom Robert Duruji, Oluwasegun Christopher Akinleye
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This study reports on the impact of different ripening accelerators on the microbial load and proximate composition of plantain (Musa paradisiaca) and Banana (Musa sapientum) during the ripening process, and the nutrition implication for food security. The study comprised of four treatments, namely: Calcium carbide, Irvingia gabonensis fruits, Newbouldia laevis leaves and a control, where no ripening accelerator was applied to the fingers of plantain and banana. The unripe and ripened plantain and banana were subjected to microbial analysis by isolating and enumerating their micro flora using pour plate method; and also, their proximate composition was determined using standard methods. The result indicated that the bacteria count of plantain increased from 3.25 ± 0.33 for unripe to 5.31 ± 0.30 log cfu/g for (treated) ripened, and that of banana increased from 3.69 ± 0.11 for unripe to 5.26 ± 0.21 log cfu/g for ripened. Also, the fungal count of plantain increased from 3.20 ± 0.16 for unripe to 4.88 ± 0.22 log sfu/g for ripened; and that of banana increased from 3.61 ± 0.19 for unripe to 5.43 ± 0.26 for ripened. Ripened plantain fingers without any ripening accelerator (control) had significantly (p < 0.05) higher values of crude protein 3.56 ± 0.06%, crude fat 0.42 ± 0.04%, total ash 2.74 ± 0.15 and carbohydrate 31.10 ± 0.20; but with significantly lower value of moisture 62.14 ± 0.07% when compared with treated plantain. The proximate composition trend of treated and banana fingers control is similar to that of treated and plantain control, except that higher moisture content of 75.11 ± 0.07% and lesser protein, crude fat, total ash and carbohydrate were obtained from treated and ripened banana control when the treatments were compared with that of plantain. The study concluded that plantain is more nutritious (mealy) than a banana; also, the ripening accelerators increased the microbial load and reduced the nutritional status of plantain and banana.Keywords: food nutrition, calcium carbide, rvingia gabonensis, newbouldia laevis, plantain, banana
Procedia PDF Downloads 3261510 Quantity, Quality and Water Productivity of Mulberry Leaf Influenced by Different Methods, Levels of Irrigation and Mulching in Eastern Dry Zone of Karnataka, India
Authors: Chengalappa Seenappa, Narayanappa Devkumar, Narayanappa Nagaraja
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Mulberry leaf is the major economic component in sericulture and quality of leaf produced per unit area has a direct effect on quality of cocoon. Among all the agronomical inputs, irrigation water has highest impact on mulberry leaf quantity and quality. The water productivity in sericulture in the country is inadequate and inefficient though India has the largest irrigated area. There is a need of proper irrigation methods and conservation practices to ensure efficiency and economy in water use. Hence, this field experiment was conducted at College of Sericulture, Chintamani, Chickaballapur district, Karnataka, India during 2013 and 2014 to know the quantity, quality and water productivity of mulberry influenced by different methods, levels of irrigation and mulching in Eastern Dry Zone (EDZ) of Karnataka, India. The results revealed that the mulberry leaf quantity, quality and water productivity were significantly influenced by different methods, levels of irrigation and mulching. Subsurface drip irrigation at 0.8 CPE (Cumulative Pan Evaporation) recorded higher leaf yield, chlorophyll, relative water, protein content and water productivity (42857 kg ha-1 yr-1, 8.54, 65.80%, 22.27% and 364.41 kg hacm-1, respectively) than surface drip at 1.0 CPE (38809 kg ha-1 yr-1, 7.34, 62.76%, 17.75% and 264 10 kg hacm-1, respectively) and micro spray jet at 1.0 CPE (39931 kg ha-1 yr-1, 7.96, 63.50%, 19.00%, 35617 kg ha-1 yr-1 and 271.83 kg hacm-1, respectively). Mulching treatment recorded maximum leaf yield, chlorophyll, relative water, protein content and water productivity (38035 kg ha-1 yr-1, 7.12, 62.11%, 16.14% and 330 kg hacm-1, respectively) compared to without mulching. These results clearly indicated that subsurface drip irrigation at lower level of irrigation (0.8 CPE) and mulching increased the quantity, quality and water productivity of mulberry leaf than surface drip and micro spray jet irrigation at higher level of irrigation (1.0 CPE) by saving 20 per cent of water. Therefore, in the coming days subsurface drip irrigation in mulberry cultivation may be more appropriate to realise higher yield, quality and water productivity in EDZ of Karnataka, India.Keywords: subsurface drip irrigation, mulching, water productivity, mulberry
Procedia PDF Downloads 2701509 Adsorptive Membrane for Hemodialysis: Potential, Future Prospection and Limitation of MOF as Nanofillers
Authors: Musawira Iftikhar
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The field of membrane materials is the most dynamic due to the constantly evolving requirements advancement of materials, to address challenges such as biocompatibility, protein-bound uremic toxins, blood coagulation, auto-immune responses, oxidative stress, and poor clearance of uremic toxins. Hemodialysis is a membrane filtration processes that is currently necessary for daily living of the patients with ESRD. Tens of millions of people with ESRD have benefited from hemodialysis over the past 60–70 years, both in terms of safeguarding life and a longer lifespan. Beyond challenges associated with the efficiency and separative properties of the membranes, ensuring hemocompatibility, or the safe circulation of blood outside the body for four hours every two days, remains a persistent challenge. This review explores the ongoing field of metal–Organic Frameworks (MOFs) and their applications in hemodialysis, offering a comprehensive examination of various MOFs employed to address challenges inherent in traditional hemodialysis methodologies. this This review included includes the experimental work done with various MOFs as a filler such as UiO-66, HKUST-1, MIL-101, and ZIF-8, which together lead to improved adsorption capacities for a range of uremic toxins and proteins. Furthermore, this review highlights how effectively MOF-based hemodialysis membranes remove a variety of uremic toxins, including p-cresol, urea, creatinine, and indoxyl sulfate and potential filler choices for the future. Future research efforts should focus on refining synthesis techniques, enhancing toxin selectivity, and investigating the long-term durability of MOF-based membranes. With these considerations, MOFs emerge as transformative materials in the quest to develop advanced and efficient hemodialysis technologies, holding the promise to significantly enhance patient outcomes and redefine the landscape of renal therapy.Keywords: membrane, hemodailysis, metal organic frameworks, seperation, protein adsorbtion
Procedia PDF Downloads 581508 Evaluating Antifungal Potential of Respiratory Inhibitors against Phyto-Pathogenic Fungi
Authors: Sehrish Iftikhar, Ahmad Ali Shahid, Kiran Nawaz, Waheed Anwar
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Discovery and development of new compounds require intense studies in chemistry, biochemistry. Numerous experiments under laboratory-, greenhouse- and field conditions can be performed to select suitable candidates and to understand their full potential. Novel fungicides are fundamental to combat plant diseases. Fusarium solani is important plant pathogen. New broad spectrum foliar fungicides against complex II were designed in this study. Complex II, namely succinate dehydrogenase (SDH), or succinate quinone oxidoreductase (SQR) is a multi-subunit enzyme at the crossroads of TCA and ETC at the inner mitochondrial membrane. The need for new and innovative fungicides is driven by resistance management, regulatory hurdles and increasing customer expectations amongst others. Fungicidal activity was assessed for the effect on mycelial growth and spore germination of the fungi using fungicide amended media assay. In mycelial growth assay compounds C10 and C6 were highly active against all the isolates. The compounds C1 and C10 were found most potent in spore germination test. It fully proved that the SDHIs designed in this paper displayed as good inhibitory effects against Fusarium solani.Keywords: Wilt, Fusarium, SDH, antifungal
Procedia PDF Downloads 2591507 Anticancer and Anti-Apoptotic Potential of Tridham and 1,2,3,4,6-Penta-O-Galloyl-β-D-Glucose in MCF-7 Breast Cancer Cell Line
Authors: R. Stalin, D. Karthick, H. Haseena Banu, T. P. Sachidanandam, P. Shanthi
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Background: Breast cancer is emerging as one of the leading cause of cancer related deaths and hence there arises the need to look out for drugs which are more targets specific with minimal side effects. In recent times, there is a shift towards alternative medicine due to low cost and less side effects. Siddha system of medicine is one the oldest system of medicine practiced against various ailments. Tridham (TD) is a herbal formulation prepared in our laboratory consisting of Terminalia chebula, Elaeocarpus ganitrus and Prosopis cineraria in a definite ratio (TD) and its anticancer potential is evaluated in terms of induction of apoptosis. Objective: The present study was designed to investigate the anti proliferative effect of TD and 1,2,3,4,6-penta-O-galloyl-b-D-glucose (PGG), a pure compound isolated from TD on human mammary carcinoma cell line (MCF-7). Materials and Methods: Cell viability was studied using MTT analysis and trypan blue staining. Mitochondrial membrane potential was studied using DAPI staining. The protein and mRNA expressions of pro-apoptotic and anti- apoptotic markers namely Bax, Bad, Bcl-2 and caspases were also assessed by Western Blotting and RT PCR. Results: Viability studies of TD and PGG treated MCF-7 cells showed an inhibition in cell growth in time and dose dependent manner. The alteration in mitochondrial membrane potential was restored through treatment with TD and PGG which was confirmed by DAPI staining. The protein and mRNA expression of pro-apoptotic markers was found to be significantly increased in TD and PGG treated cells with a concomitant decrease in anti-apoptotic markers. Conclusion: The results of the study suggest that TD and PGG exhibit their anticancer effect through its membrane stabilizing property and activation of apoptotic cascade in MCF-7 cells.Keywords: apoptosis, mammary carcinoma, MCF-7, penta galloyl glucose, Tridham
Procedia PDF Downloads 3141506 Cardiolipin-Incorporated Liposomes Carrying Curcumin and Nerve Growth Factor to Rescue Neurons from Apoptosis for Alzheimer’s Disease Treatment
Authors: Yung-Chih Kuo, Che-Yu Lin, Jay-Shake Li, Yung-I Lou
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Curcumin (CRM) and nerve growth factor (NGF) were entrapped in liposomes (LIP) with cardiolipin (CL) to downregulate the phosphorylation of mitogen-activated protein kinases for Alzheimer’s disease (AD) management. AD belongs to neurodegenerative disorder with a gradual loss of memory, yielding irreversible dementia. CL-conjugated LIP loaded with CRM (CRM-CL/LIP) and that with NGF (NGF-CL/LIP) were applied to AD models of SK-N-MC cells and Wistar rats with an insult of β-amyloid peptide (Aβ). Lipids comprising 1,2-dipalmitoyl-sn-glycero-3- phosphocholine (Avanti Polar Lipids, Alabaster, AL), 1',3'-bis[1,2- dimyristoyl-sn-glycero-3-phospho]-sn-glycerol (CL; Avanti Polar Lipids), 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N- [methoxy(polyethylene glycol)-2000] (Avanti Polar Lipids), 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[carboxy(polyethylene glycol)-2000] (Avanti Polar Lipids) and CRM (Sigma–Aldrich, St. Louis, MO) were dissolved in chloroform (J. T. Baker, Phillipsburg, NJ) and condensed using a rotary evaporator (Panchum, Kaohsiung, Taiwan). Human β-NGF (Alomone Lab, Jerusalem, Israel) was added in the aqueous phase. Wheat germ agglutinin (WGA; Medicago AB, Uppsala, Sweden) was grafted on LIP loaded with CRM for (WGA-CRM-LIP) and CL-conjugated LIP loaded with CRM (WGA-CRM-CL/LIP) using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (Sigma–Aldrich) and N-hydroxysuccinimide (Alfa Aesar, Ward Hill, MA). The protein samples of SK-N-MC cells (American Type Tissue Collection, Rockville, MD) were used for sodium dodecyl sulfate (Sigma–Aldrich) polyacrylamide gel (Sigma–Aldrich) electrophoresis. In animal study, the LIP formulations were administered by intravenous injection via a tail vein of male Wistar rats (250–280 g, 8 weeks, BioLasco, Taipei, Taiwan), which were housed in the Animal Laboratory of National Chung Cheng University in accordance with the institutional guidelines and the guidelines of Animal Protection Committee under the Council of Agriculture of the Republic of China. We found that CRM-CL/LIP could inhibit the expressions of phosphorylated p38 (p-p38), p-Jun N-terminal kinase (p-JNK), and p-tau protein at serine 202 (p-Ser202) to retard the neuronal apoptosis. Free CRM and released CRM from CRM-LIP and CRM-CL/LIP were not in a straightforward manner to effectively inhibit the expression of p-p38 and p-JNK in the cytoplasm. In addition, NGF-CL/LIP enhanced the quantities of p-neurotrophic tyrosine kinase receptor type 1 (p-TrkA) and p-extracellular-signal-regulated kinase 5 (p-ERK5), preventing the Aβ-induced degeneration of neurons. The membrane fusion of NGF-LIP activated the ERK5 pathway and the targeting capacity of NGF-CL/LIP enhanced the possibility of released NGF to affect the TrkA level. Moreover, WGA-CRM-LIP improved the permeation of CRM across the blood–brain barrier (BBB) and significantly reduced the Aβ plaque deposition and malondialdehyde level and increased the percentage of normal neurons and cholinergic function in the hippocampus of AD rats. This was mainly because the encapsulated CRM was protected by LIP against a rapid degradation in the blood. Furthermore, WGA on LIP could target N-acetylglucosamine on endothelia and increased the quantity of CRM transported across the BBB. In addition, WGA-CRM-CL/LIP could be effective in suppressing the synthesis of acetylcholinesterase and reduced the decomposition of acetylcholine for better neurotransmission. Based on the in vitro and in vivo evidences, WGA-CRM-CL/LIP can rescue neurons from apoptosis in the brain and can be a promising drug delivery system for clinical AD therapy.Keywords: Alzheimer’s disease, β-amyloid, liposome, mitogen-activated protein kinase
Procedia PDF Downloads 3321505 Authentication and Traceability of Meat Products from South Indian Market by Species-Specific Polymerase Chain Reaction
Authors: J. U. Santhosh Kumar, V. Krishna, Sebin Sebastian, G. S. Seethapathy, G. Ravikanth, R. Uma Shaanker
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Food is one of the basic needs of human beings. It requires the normal function of the body part and a healthy growth. Recently, food adulteration increases day by day to increase the quantity and make more benefit. Animal source foods can provide a variety of micronutrients that are difficult to obtain in adequate quantities from plant source foods alone. Particularly in the meat industry, products from animals are susceptible targets for fraudulent labeling due to the economic profit that results from selling cheaper meat as meat from more profitable and desirable species. This work presents an overview of the main PCR-based techniques applied to date to verify the authenticity of beef meat and meat products from beef species. We were analyzed 25 market beef samples in South India. We examined PCR methods based on the sequence of the cytochrome b gene for source species identification. We found all sample were sold as beef meat as Bos Taurus. However, interestingly Male meats are more valuable high price compare to female meat, due to this reason most of the markets samples are susceptible. We were used sex determination gene of cattle like TSPY(Y-encoded, testis-specific protein TSPY is a Y-specific gene). TSPY homologs exist in several mammalian species, including humans, horses, and cattle. This gene is Y coded testis protein genes, which only amplify the male. We used multiple PCR products form species-specific “fingerprints” on gel electrophoresis, which may be useful for meat authentication. Amplicons were obtained only by the Cattle -specific PCR. We found 13 market meat samples sold as female beef samples. These results suggest that the species-specific PCR methods established in this study would be useful for simple and easy detection of adulteration of meat products.Keywords: authentication, meat products, species-specific, TSPY
Procedia PDF Downloads 3781504 Development of Low Calorie Jelly with Increased Content of Natural Compounds from Superfoods with No Added Sugar
Authors: Liana C. Salanță, Maria Tofană, Carmen R. Pop, Vlad Mureșan
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The landscape of functional food is expanding very fast, due to the consumer interest for healthy natural products. Consumers nowadays demand healthy products that impart phytonutrients to encourage good health and well-being, prevent diseases, without sacrificing taste and texture. Candies are foodstuffs appreciated by all category of consumers. They are available in a range variety of forms (jellies, marshmallows, caramels, lollipops, etc.). Jelly is characterized by a gummy and chewy texture typically conferred by a hydrocolloid (gelatin, pectin). The purpose of this research was to obtain hypocaloric jelly (no added sugar) enriched with protein powder from acai, chia seeds and hemp, which are considered superfood. Peach and raspberry juice were used for obtaining functional jelly, due to the specific flavour, natural carbohydrate, natural pigments and vitamins (C, B1, PP, etc). Instead of classic hydrocolloids used in Romania for the industry of jelly, agar-agar was used in this study, due to its properties. Agar-agar is able to form gels in the aqueous medium, stronger than other gel-forming agents. High sugar concentrations or an acid environment (as is necessary with pectins) are not needed. In addition to its gelation properties, Agar-agar is considered to have important nutritional benefits, high content of fibre and has low calories. Six prototypes of jellies were obtained and evaluated by physicochemical, microbiological and sensorial analysis. For the textural profile analysis, the Brookfield CT3 Texture Analyzer, equipped with a 10kg load cell, was used. The results revealed that hypocaloric jelly can serve as a good source of bioactive compounds in the diet. The jelly is a convenient way of delivering potential health benefits of protein powder and agar-agar to a wide range of consumers.Keywords: agar-agar, functional food, hypocaloric jelly, superfoods
Procedia PDF Downloads 1321503 Characteristics of Smoked Edible Film Made from Myofibril, Collagen and Carrageenan
Authors: Roike Iwan Montolalu, Henny Adeleida Dien, Feny Mentang, Kristhina P. Rahael, Tomy Moga, Ayub Meko, Siegfried Berhimpon
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In the last 20 years, packaging materials derived from petrochemicals polymers were widely used as packaging materials. This due to various advantages such as flexible, strong, transparent, and the price is relatively cheap. However, the plastic polymer also has various disadvantages, such as the transmission monomer contamination into the material to be packed, and waste is non-biodegradable. Edible film (EF) is an up to date materials, generated after the biodegradable packaging materials. The advantages of the EF materials, is the materials can be eat together with food, and the materials can be applied as a coating materials for a widely kind of foods especially snack foods. The aims of this research are to produce and to analyze the characteristics of smoked EF made from carrageenan, myofibril and collagen of Black Marlin (Makaira indica) industrial waste. Smoked EF made with an addition of 0.8 % smoke liquid. Three biopolymers i.e. carrageenan, myofibril, and collagen were used as treatments, and homogenate for 1 hours at speed of 1500 rpm. The analysis carried out on the pH and physical properties i.e. thickness, solubility, tensile strength, % elongation, and water vapor transmission rate (WVTR), as well as on the sensory characteristics of texture i.e. wateriness, firmness, elasticity, hardness, and juiciness of the coated products. The result shown that the higher the concentration the higher the thickness of EF, where as for myofibril proteins appeared higher than carrageenan and collagen. Both of collagen and myofibril shown that concentration of 6% was most soluble, while for carrageenan were in concentration of 2 to 2.5%. For tensile strength, carrageenan was significantly higher than myofibril and collagen; while for elongation, collagen film more elastic than carragenan and myofibril protein. Water vapor transmission rate, shown that myofibril protein film lower than carrageenan and collagen film. From sensory assessment of texture, carrageenan has a high elasticity and juiciness, while collagen and myofibril have a high in firmness and hardness.Keywords: edible film, collagen, myofibril, carrageenan
Procedia PDF Downloads 4301502 Developing a Set of Primers Targeting Chondroitin Ac Lyase Gene for Specific and Sensitive Detection of Flavobacterium Columnare, a Causative Agent of Freshwater Columnaris
Authors: Mahmoud Mabrok, Channarong Rodkhum
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Flavobacterium columanre is one of the devastating pathogen that causes noticeable economic losses in freshwater cultured fish. Like other filamentous bacteria, F. columanre tends to aggregate and fluctuate to all kind of media, thus revealing obstacles in recognition of its colonies. Since the molecular typing is the only fundamental tool for rapid and precise detection of this pathgen. The present study developed a species-specific PCR assay based on cslA unique gene of F. columnare. The cslA gene sequences of 13 F. columnare, strains retrieved from gene bank database, were aligned to identify a conserved homologous segment prior to primers design. The new primers yielded amplicons of 287 bp from F. columnare strains but not from relevant or other pathogens, unlike to other published set that showed no specificity and cross-reactivity with F. indicum. The primers were sensitive and detected as few as 7 CFUs of bacteria and 3 pg of gDNA template. The sensitivity was reduced ten times when using tissue samples. These primers precisely defined all field isolates in a double-blind study, proposing their applicable use for field detection.Keywords: Columnaris infection, cslA gene, Flavobacterium columnare, PCR
Procedia PDF Downloads 1311501 Characterization of the Lytic Bacteriophage VbɸAB-1 against Drug Resistant Acinetobacter baumannii Isolated from Hospitalized Pressure Ulcers Patients
Authors: M. Doudi, M. H. Pazandeh, L. Rahimzadeh Torabi
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Bedsores are pressure ulcers that occur on the skin or tissue due to being immobile and lying in bed for extended periods. Bedsores have the potential to progress into open ulcers, increasing the possibility of variety of bacterial infection. Acinetobacter baumannii, a pathogen of considerable clinical importance, exhibited a significant correlation with Bedsores (pressure ulcers) infections, thereby manifesting a wide spectrum of antibiotic resistance. The emergence of drug resistance has led researchers to focus on alternative methods, particularly phage therapy, for tackling bacterial infections. Phage therapy has emerged as a novel therapeutic approach to regulate the activity of these agents. The management of bacterial infections greatly benefits from the clinical utilization of bacteriophages as a valuable antimicrobial intervention. The primary objective of this investigation consisted of isolating and discerning potent bacteriophage capable of targeting multi drug-resistant (MDR) and extensively drug-resistant (XDR) bacteria obtained from pressure ulcers. In present study, analyzed and isolated A. baumannii strains obtained from a cohort of patients suffering from pressure ulcers at Taleghani Hospital in Ahvaz, Iran. An approach that included biochemical and molecular identification techniques was used to determine the taxonomic classification of bacterial isolates at the genus and species levels. The molecular identification process was facilitated by using the 16S rRNA gene in combination with universal primers 27 F, and 1492 R. Bacteriophage was obtained through the isolation process conducted on treatment plant sewage located in Isfahan, Iran. The main goal of this study was to evaluate different characteristics of phage, such as their appearance, range of hosts they can infect, how quickly they can enter a host, their stability at varying temperatures and pH levels, their effectiveness in killing bacteria, the growth pattern of a single phage stage, mapping of enzymatic digestion, and identification of proteomics patterns. The findings demonstrated that an examination was conducted on a sample of 50 specimens, wherein 15 instances of A. baumannii were identified. These microorganisms are the predominant Gram-negative agents known to cause wound infections in individuals suffering from bedsores. The study's findings indicated a high prevalence of antibiotic resistance in the strains isolated from pressure ulcers, excluding the clinical strains that exhibited responsiveness to colistin.According to the findings obtained from assessments of host range and morphological characteristics of bacteriophage VbɸAB-1, it can be concluded that this phage possesses specificity towards A. Baumannii BAH_Glau1001 was classified as a member of the Plasmaviridae family. The bacteriophage mentioned earlier showed the strongest antibacterial effect at a temperature of 18 °C and a pH of 6.5. Through the utilization of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis on protein fragments, it was established that the bacteriophage VbɸAB-1 exhibited a size range between 50 and 75 kilodaltons (KDa). The numerous research findings on the effectiveness of phages and the safety studies conducted suggest that the phages studied in this research can be considered as a practical solution and recommended approach for controlling and treating stubborn pathogens in burn wounds among hospitalized patients.Keywords: acinetobacter baumannii, extremely drug- resistant, phage therapy, surgery wound
Procedia PDF Downloads 941500 Broad Protection against Avian Influenza Virus by Using a Modified Vaccinia Ankara Virus Expressing a Mosaic Hemagglutinin
Authors: Attapon Kamlangdee, Brock Kingstad-Bakke, Tavis K. Anderson, Tony L. Goldberg, Jorge E. Osorio
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A critical failure in our preparedness for an influenza pandemic is the lack of a universal vaccine. Influenza virus strains diverge by 1 to 2% per year, and commercially available vaccines often do not elicit protection from one year to the next, necessitating frequent formulation changes. This represents a major challenge to the development of a cross-protective vaccine that can protect against circulating viral antigenic diversity. We have constructed a recombinant modified vaccinia virus Ankara (MVA) that expresses an H5N1 mosaic hemagglutinin (H5M) (MVA-H5M). This mosaic was generated in silico using 2,145 field-sourced H5N1 isolates. A single dose of MVA-H5M provided 100% protection in mice against clade 0, 1, and 2 avian influenza viruses and also protected against seasonal H1N1 virus (A/Puerto Rico/8/34). It also provided short-term (10 days) and long-term (6 months) protection post vaccination. Both neutralizing antibodies and antigen-specific CD4+and CD8+ T cells were still detected at 5 months post vaccination, suggesting that MVA-H5M provides long-lasting immunity.Keywords: modified vaccinia Ankara, MVA, H5N1, hemagglutinin, influenza vaccine
Procedia PDF Downloads 2821499 Community-Based Reference Interval of Selected Clinical Chemistry Parameters Among Apparently Healthy Adolescents in Mekelle City, Tigrai, Northern Ethiopia
Authors: Getachew Belay Kassahun
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Background: Locally established clinical laboratory reference intervals (RIs) are required to interpret laboratory test results for screening, diagnosis, and prognosis. The objective of this study was to establish a reference interval of clinical chemistry parameters among apparently healthy adolescents aged between 12 and 17 years in Mekelle, Tigrai, in the northern part of Ethiopia. Methods: Community-based cross-sectional study was employed from December 2018 to March 2019 in Mekelle City among 172 males and 172 females based on a Multi-stage sampling technique. Blood samples were tested for Fasting blood sugar (FBS), alanine amino transferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), Creatinine, urea, total protein, albumin (ALB), direct and indirect bilirubin (BIL.D and BIL.T) using 25 Bio system clinical chemistry analyzer. Results were analyzed using SPSS version 23 software and based on the Clinical Laboratory Standard Institute (CLSI)/ International Federation of Clinical Chemistry (IFCC) C 28-A3 Guideline which defines the reference interval as the 95% central range of 2.5th and 97.5th percentiles. Mann Whitney U test, descriptive statistics and box and whisker were statistical tools used for analysis. Results: This study observed statistically significant differences between males and females in ALP, ALT, AST, Urea and Creatinine Reference intervals. The established reference intervals for males and females, respectively, were: ALP (U/L) 79.48-492.12 versus 63.56-253.34, ALT (U/L) 4.54-23.69 versus 5.1-20.03, AST 15.7- 39.1 versus 13.3- 28.5, Urea (mg/dL) 9.33-24.99 versus 7.43-23.11, and Creatinine (mg/dL) 0.393-0.957 versus 0.301-0.846. The combined RIs for Total Protein (g/dL) were 6.08-7.85, ALB (g/dL) 4.42-5.46, FBS(mg/dL) 65-110, BIL.D (mg/dL) 0.033-0.532, and BIL.T (mg/dL) 0.106-0.812. Conclusions: The result showed a marked difference between sex and company-derived values for selected clinical chemistry parameters. Thus, the use of age and sex-specific locally established reference intervals for clinical chemistry parameters is recommended.Keywords: reference interval, adolescent, clinical chemistry, Ethiopia
Procedia PDF Downloads 831498 Ring FingerPortein 2 (RNF2) Targeting by miRNAs in Breast Cancer Cell Lines
Authors: Ceyda Okudu, Secil Eroglu, Khandakar A. S. M. Saadat, Sibel O. Balci
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Ring Finger Protein 2 (RNF2) is a member of polycomb repressive complex 1 (PRC1), which is one of the epigenetic regulators in the genome. When RNF2 combines with other PRC1 members, it mediates the mono-ubiquitination of Histon2A (H2A). In breast cancer, RNF2 is commonly overexpressed, and also it promotes metastasis and invasion in other aggressive tumors like melanoma, prostate, and hepatocarcinoma. The role of RNF2 in the metastasis and invasion of breast cancer has not yet been elucidated. Our aim is to observe the role of RNF2 in metastasis and invasion in this study by miRNA mediated RNF2 gene silencing in breast cancer cell lines. We selected miRNAs, targeting to RNF2 by searching online databases. miR-17-5p, miR20a-5p, and miR-106b-5p were transfected to breast cancer cell lines (MCF-7, MDA-MB-231, SK-BR-3, and ZR-75-1), and also we used normal breast epithelial cell line (hTERT-HME1) to compare RNF2 gene expression level. After 48-72 hours post-transfection, mRNAs were isolated from the cells, and gene expressions were measured by RT-qPCR after from cDNA syntheses. We observed that RNF2 was highly expressed in SK-BR-3 and MDA-MB-231 cell lines opposite to MCF-7 and ZR-75-1 cell lines. RNF2 was downregulated 5, 5 and 7 fold by miR17-5p, miR20a-5p and miR106b-5p respectively in MCF-7. However, in SK-BR-3 and ZR-75-1 cell lines, miRNAs did not affect significantly RNF2 gene expression level. miR20a-5p decreased RNF2 3 fold and miR17-5p and miR106b-5p did not affect MDA-MB-231. After gene expression analysis, we performed metastasis and invasion assay in MCF-7 cells. For metastasis, we used both wound healing assay and Transwell Cell Migration Assay, and we used Transwell Cell Invasion Assay for invasion. The data of this assay showed that miR17-5p and miR20a-5p decreased both invasion and metastasis level, but miR106b-5p has no effect. We would like to conclude that RNF2 can be targeted by miR17-5p, miR20a-5p and miR106b-5p in MCF-7 cells and also RNF2, which is one of the upregulated genes in aggressive tumor, can be decreased by using these miRNAs. In future, we would like to confirm these results at the protein level and also whether these miRNAs are direct target of RNF2 or not.Keywords: breast cancer, epigenetic, microRNAs, RNF2
Procedia PDF Downloads 1821497 LaeA/1-Velvet Interplay in Aspergillus and Trichoderma: Regulation of Secondary Metabolites and Cellulases
Authors: Razieh Karimi Aghcheh, Christian Kubicek, Joseph Strauss, Gerhard Braus
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Filamentous fungi are of considerable economic and social significance for human health, nutrition and in white biotechnology. These organisms are dominant producers of a range of primary metabolites such as citric acid, microbial lipids (biodiesel) and higher unsaturated fatty acids (HUFAs). In particular, they produce also important but structurally complex secondary metabolites with enormous therapeutic applications in pharmaceutical industry, for example: cephalosporin, penicillin, taxol, zeranol and ergot alkaloids. Several fungal secondary metabolites, which are significantly relevant to human health do not only include antibiotics, but also e.g. lovastatin, a well-known antihypercholesterolemic agent produced by Aspergillus. terreus, or aflatoxin, a carcinogen produced by A. flavus. In addition to their roles for human health and agriculture, some fungi are industrially and commercially important: Species of the ascomycete genus Hypocrea spp. (teleomorph of Trichoderma) have been demonstrated as efficient producer of highly active cellulolytic enzymes. This trait makes them effective in disrupting and depolymerization of lignocellulosic materials and thus applicable tools in number of biotechnological areas as diverse as clothes-washing detergent, animal feed, and pulp and fuel productions. Fungal LaeA/LAE1 (Loss of aflR Expression A) homologs their gene products act at the interphase between secondary metabolisms, cellulase production and development. Lack of the corresponding genes results in significant physiological changes including loss of secondary metabolite and lignocellulose degrading enzymes production. At the molecular level, the encoded proteins are presumably methyltransferases or demethylases which act directly or indirectly at heterochromatin and interact with velvet domain proteins. Velvet proteins bind to DNA and affect expression of secondary metabolites (SMs) genes and cellulases. The dynamic interplay between LaeA/LAE1, velvet proteins and additional interaction partners is the key for an understanding of the coordination of metabolic and morphological functions of fungi and is required for a biotechnological control of the formation of desired bioactive products. Aspergilli and Trichoderma represent different biotechnologically significant species with significant differences in the LaeA/LAE1-Velvet protein machinery and their target proteins. We, therefore, performed a comparative study of the interaction partners of this machinery and the dynamics of the various protein-protein interactions using our robust proteomic and mass spectrometry techniques. This enhances our knowledge about the fungal coordination of secondary metabolism, cellulase production and development and thereby will certainly improve recombinant fungal strain construction for the production of industrial secondary metabolite or lignocellulose hydrolytic enzymes.Keywords: cellulases, LaeA/1, proteomics, secondary metabolites
Procedia PDF Downloads 2721496 Collagen Hydrogels Cross-Linked by Squaric Acid
Authors: Joanna Skopinska-Wisniewska, Anna Bajek, Marta Ziegler-Borowska, Alina Sionkowska
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Hydrogels are a class of materials widely used in medicine for many years. Proteins, such as collagen, due to the presence of a large number of functional groups are easily wettable by polar solvents and can create hydrogels. The supramolecular network capable to swelling is created by cross-linking of the biopolymers using various reagents. Many cross-linking agents has been tested for last years, however, researchers still are looking for a new, more secure reactants. Squaric acid, 3,4-dihydroxy 3-cyclobutene 1,2- dione, is a very strong acid, which possess flat and rigid structure. Due to the presence of two carboxyl groups the squaric acid willingly reacts with amino groups of collagen. The main purpose of this study was to investigate the influence of addition of squaric acid on the chemical, physical and biological properties of collagen materials. The collagen type I was extracted from rat tail tendons and 1% solution in 0.1M acetic acid was prepared. The samples were cross-linked by the addition of 5%, 10% and 20% of squaric acid. The mixtures of all reagents were incubated 30 min on magnetic stirrer and then dialyzed against deionized water. The FTIR spectra show that the collagen structure is not changed by cross-linking by squaric acid. Although the mechanical properties of the collagen material deteriorate, the temperature of thermal denaturation of collagen increases after cross-linking, what indicates that the protein network was created. The lyophilized collagen gels exhibit porous structure and the pore size decreases with the higher addition of squaric acid. Also the swelling ability is lower after the cross-linking. The in vitro study demonstrates that the materials are attractive for 3T3 cells. The addition of squaric acid causes formation of cross-ling bonds in the collagen materials and the transparent, stiff hydrogels are obtained. The changes of physicochemical properties of the material are typical for cross-linking process, except mechanical properties – it requires further experiments. However, the results let us to conclude that squaric acid is a suitable cross-linker for protein materials for medicine and tissue engineering.Keywords: collagen, squaric acid, cross-linking, hydrogel
Procedia PDF Downloads 3891495 Gene Expression Signature-Based Chemical Genomic to Identify Potential Therapeutic Compounds for Colorectal Cancer
Authors: Yen-Hao Su, Wan-Chun Tang, Ya-Wen Cheng, Peik Sia, Chi-Chen Huang, Yi-Chao Lee, Hsin-Yi Jiang, Ming-Heng Wu, I-Lu Lai, Jun-Wei Lee, Kuen-Haur Lee
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There is a wide range of drugs and combinations under investigation and/or approved over the last decade to treat colorectal cancer (CRC), but the 5-year survival rate remains poor at stages II–IV. Therefore, new, more efficient drugs still need to be developed that will hopefully be included in first-line therapy or overcome resistance when it appears, as part of second- or third-line treatments in the near future. In this study, we revealed that heat shock protein 90 (Hsp90) inhibitors have high therapeutic potential in CRC according to combinative analysis of NCBI's Gene Expression Omnibus (GEO) repository and chemical genomic database of Connectivity Map (CMap). We found that second generation Hsp90 inhibitor, NVP-AUY922, significantly down regulated the activities of a broad spectrum of kinases involved in regulating cell growth arrest and death of NVPAUY922-sensitive CRC cells. To overcome NVP-AUY922-induced upregulation of survivin expression which causes drug insensitivity, we found that combining berberine (BBR), a herbal medicine with potency in inhibiting survivin expression, with NVP-AUY922 resulted in synergistic antiproliferative effects for NVP-AUY922-sensitive and -insensitive CRC cells. Furthermore, we demonstrated that treatment of NVP-AUY922-insensitive CRC cells with the combination of NVP-AUY922 and BBR caused cell growth arrest through inhibiting CDK4 expression and induction of microRNA-296-5p (miR-296-5p)-mediated suppression of Pin1–β-catenin–cyclin D1 signaling pathway. Finally, we found that the expression level of Hsp90 in tumor tissues of CRC was positively correlated with CDK4 and Pin1 expression levels. Taken together, these results indicate that combination of NVP-AUY922 and BBR therapy can inhibit multiple oncogenic signaling pathways of CRC.Keywords: berberine, colorectal cancer, connectivity map, heat shock protein 90 inhibitor
Procedia PDF Downloads 3081494 Phytochemical and Biological Study of Chrozophora oblongifolia
Authors: Al-Braa Kashegari, Ali M. El-Halawany, Akram A. Shalabi, Sabrin R. M. Ibrahim, Hossam M. Abdallah
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Chemical investigation of Chrozophora oblongifolia resulted in the isolation of five major compounds that were identified as apeginin-7-O-glucoside (1), quercetin-3-O-glucuronic acid (2), quercetin-3-O-glacturonic acid (3), rutin (4), and 1,3,6-trigalloyl glucose (5). The identity of isolated compounds was assessed by different spectroscopic methods, including one- and two-dimensional NMR. The isolated compounds were tested for their antioxidant activity using different assays viz., DPPH, FRAP, ABTS, ORAC, and metal chelation effects. In addition, the inhibition of target enzymes involved in the metabolic syndrome, such as alpha-glucosidase and pancreatic lipase, were carried out. Moreover, the effect of the compounds on the advanced glycation end-products (AGEs) as one of the major complications of oxidative stress and hyperglycemia in metabolic syndromes were carried out using BSA‐fructose (bovine serum albumin), BSA-methylglyoxal, and arginine methylglyoxal models. The pure isolates showed a protective effect in metabolic syndromes as well as promising antioxidant activity. The results showed potent activity of compound 5 in all measured parameters meanwhile, none of the tested compounds showed activity against pancreatic lipase.Keywords: Chrozophora oblongifolia, antioxidant, pancreatic lipase, metabolic syndromes
Procedia PDF Downloads 1131493 Swedish–Nigerian Extrusion Research: Channel for Traditional Grain Value Addition
Authors: Kalep Filli, Sophia Wassén, Annika Krona, Mats Stading
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Food security challenge and the growing population in Sub-Saharan Africa centers on its agricultural transformation, where about 70% of its population is directly involved in farming. Research input can create economic opportunities, reduce malnutrition and poverty, and generate faster, fairer growth. Africa is discarding $4 billion worth of grain annually due to pre and post-harvest losses. Grains and tubers play a central role in food supply in the region but their production has generally lagged behind because no robust scientific input to meet up with the challenge. The African grains are still chronically underutilized to the detriment of the well-being of the people of Africa and elsewhere. The major reason for their underutilization is because they are under-researched. Any commitment by scientific community to intervene needs creative solutions focused on innovative approaches that will meet the economic growth. In order to mitigate this hurdle, co-creation activities and initiatives are necessary.An example of such initiatives has been initiated through Modibbo Adama University of Technology Yola, Nigeria and RISE (The Research Institutes of Sweden) Gothenburg, Sweden. Exchange of expertise in research activities as a possibility to create channel for value addition to agricultural commodities in the region under the ´Traditional Grain Network programme´ is in place. Process technologies, such as extrusion offers the possibility of creating products in the food and feed sectors, with better storage stability, added value, lower transportation cost and new markets. The Swedish–Nigerian initiative has focused on the development of high protein pasta. Dry microscopy of pasta sample result shows a continuous structural framework of proteins and starch matrix. The water absorption index (WAI) results showed that water was absorbed steadily and followed the master curve pattern. The WAI values ranged between 250 – 300%. In all aspect, the water absorption history was within a narrow range for all the eight samples. The total cooking time for all the eight samples in our study ranged between 5 – 6 minutes with their respective dry sample diameter ranging between 1.26 – 1.35 mm. The percentage water solubility index (WSI) ranged from 6.03 – 6.50% which was within a narrow range and the cooking loss which is a measure of WSI is considered as one of the main parameters taken into consideration during the assessment of pasta quality. The protein contents of the samples ranged between 17.33 – 18.60 %. The value of the cooked pasta firmness ranged from 0.28 - 0.86 N. The result shows that increase in ratio of cowpea flour and level of pregelatinized cowpea tends to increase the firmness of the pasta. The breaking strength represent index of toughness of the dry pasta ranged and it ranged from 12.9 - 16.5 MPa.Keywords: cowpea, extrusion, gluten free, high protein, pasta, sorghum
Procedia PDF Downloads 2001492 Genetically Modified Organisms
Authors: Mudrika Singhal
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The research paper is basically about how the genetically modified organisms evolved and their significance in today’s world. It also highlights about the various pros and cons of the genetically modified organisms and the progress of India in this field. A genetically modified organism is the one whose genetic material has been altered using genetic engineering techniques. They have a wide range of uses such as transgenic plants, genetically modified mammals such as mouse and also in insects and aquatic life. Their use is rooted back to the time around 12,000 B.C. when humans domesticated plants and animals. At that humans used genetically modified organisms produced by the procedure of selective breeding and not by genetic engineering techniques. Selective breeding is the procedure in which selective traits are bred in plants and animals and then are domesticated. Domestication of wild plants into a suitable cultigen is a well known example of this technique. GMOs have uses in varied fields ranging from biological and medical research, production of pharmaceutical drugs to agricultural fields. The first organisms to be genetically modified were the microbes because of their simpler genetics. At present the genetically modified protein insulin is used to treat diabetes. In the case of plants transgenic plants, genetically modified crops and cisgenic plants are the examples of genetic modification. In the case of mammals, transgenic animals such as mice, rats etc. serve various purposes such as researching human diseases, improvement in animal health etc. Now coming upon the pros and cons related to the genetically modified organisms, pros include crops with higher yield, less growth time and more predictable in comparison to traditional breeding. Cons include that they are dangerous to mammals such as rats, these products contain protein which would trigger allergic reactions. In India presently, group of GMOs include GM microorganisms, transgenic crops and animals. There are varied applications in the field of healthcare and agriculture. In the nutshell, the research paper is about the progress in the field of genetic modification, taking along the effects in today’s world.Keywords: applications, mammals, transgenic, engineering and technology
Procedia PDF Downloads 5991491 Large-Scale Screening for Membrane Protein Interactions Involved in Platelet-Monocyte Interactions
Authors: Yi Sun, George Ed Rainger, Steve P. Watson
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Background: Beyond the classical roles in haemostasis and thrombosis, platelets are important in the initiation and development of various thrombo-inflammatory diseases. In atherosclerosis and deep vein thrombosis, for example, platelets bridge monocytes with endothelium and form heterotypic aggregates with monocytes in the circulation. This can alter monocyte phenotype by inducing their activation, stimulating adhesion and migration. These interactions involve cell surface receptor-ligand pairs on both cells. This list is likely incomplete as new interactions of importance to platelet biology are continuing to be discovered as illustrated by our discovery of PEAR-1 binding to FcεR1α. Results: We have developed a highly sensitive avidity-based assay to identify novel extracellular interactions among 126 recombinantly-expressed platelet cell surface and secreted proteins involved in platelet aggregation. In this study, we will use this method to identify novel platelet-monocyte interactions. We aim to identify ligands for orphan receptors and novel partners of well-known proteins. Identified interactions will be studied in preliminary functional assays to demonstrate relevance to the inflammatory processes supporting atherogenesis. Conclusions: Platelet-monocyte interactions are essential for the development of thromboinflammatory disease. Up until relatively recently, technologies only allow us to limit our studies on each individual protein interaction at a single time. These studies propose for the first time to study the cell surface platelet-monocyte interactions in a systematic large-scale approach using a reliable screening method we have developed. If successful, this will likely to identify previously unknown ligands for important receptors that will be investigated in details and also provide a list of novel interactions for the field. This should stimulate studies on developing alternative therapeutic strategies to treat vascular inflammatory disorders such as atherosclerosis, DVT and sepsis and other clinically important inflammatory conditions.Keywords: membrane proteins, large-scale screening, platelets, recombinant expression
Procedia PDF Downloads 1541490 Clinical and Analytical Performance of Glial Fibrillary Acidic Protein and Ubiquitin C-Terminal Hydrolase L1 Biomarkers for Traumatic Brain Injury in the Alinity Traumatic Brain Injury Test
Authors: Raj Chandran, Saul Datwyler, Jaime Marino, Daniel West, Karla Grasso, Adam Buss, Hina Syed, Zina Al Sahouri, Jennifer Yen, Krista Caudle, Beth McQuiston
Abstract:
The Alinity i TBI test is Therapeutic Goods Administration (TGA) registered and is a panel of in vitro diagnostic chemiluminescent microparticle immunoassays for the measurement of glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase L1 (UCH-L1) in plasma and serum. The Alinity i TBI performance was evaluated in a multi-center pivotal study to demonstrate the capability to assist in determining the need for a CT scan of the head in adult subjects (age 18+) presenting with suspected mild TBI (traumatic brain injury) with a Glasgow Coma Scale score of 13 to 15. TBI has been recognized as an important cause of death and disability and is a growing public health problem. An estimated 69 million people globally experience a TBI annually1. Blood-based biomarkers such as glial fibrillary acidic protein (GFAP) and ubiquitin C-terminal hydrolase L1 (UCH-L1) have shown utility to predict acute traumatic intracranial injury on head CT scans after TBI. A pivotal study using prospectively collected archived (frozen) plasma specimens was conducted to establish the clinical performance of the TBI test on the Alinity i system. The specimens were originally collected in a prospective, multi-center clinical study. Testing of the specimens was performed at three clinical sites in the United States. Performance characteristics such as detection limits, imprecision, linearity, measuring interval, expected values, and interferences were established following Clinical and Laboratory Standards Institute (CLSI) guidance. Of the 1899 mild TBI subjects, 120 had positive head CT scan results; 116 of the 120 specimens had a positive TBI interpretation (Sensitivity 96.7%; 95% CI: 91.7%, 98.7%). Of the 1779 subjects with negative CT scan results, 713 had a negative TBI interpretation (Specificity 40.1%; 95% CI: 37.8, 42.4). The negative predictive value (NPV) of the test was 99.4% (713/717, 95% CI: 98.6%, 99.8%). The analytical measuring interval (AMI) extends from the limit of quantitation (LoQ) to the upper LoQ and is determined by the range that demonstrates acceptable performance for linearity, imprecision, and bias. The AMI is 6.1 to 42,000 pg/mL for GFAP and 26.3 to 25,000 pg/mL for UCH-L1. Overall, within-laboratory imprecision (20 day) ranged from 3.7 to 5.9% CV for GFAP and 3.0 to 6.0% CV for UCH-L1, when including lot and instrument variances. The Alinity i TBI clinical performance results demonstrated high sensitivity and high NPV, supporting the utility to assist in determining the need for a head CT scan in subjects presenting to the emergency department with suspected mild TBI. The GFAP and UCH-L1 assays show robust analytical performance across a broad concentration range of GFAP and UCH-L1 and may serve as a valuable tool to help evaluate TBI patients across the spectrum of mild to severe injury.Keywords: biomarker, diagnostic, neurology, TBI
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