Search results for: recombinant protein purification

Authors: Silvia Gastaldello, Maria Grillo, Luca Tassoni, Claudio Maran, Stefano Balbo

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Antioxidants are nowadays attractive not only for the countless benefits to the human and animal health, but also for the perspective of use as food preservative instead of synthetic chemical molecules. In this study, the radical scavenging activity of six protein extracts from pulse and oleaginous seeds was evaluated. The selected matrices are Pisum sativum (yellow pea from two different origins), Carthamus tinctorius (safflower), Helianthus annuus (sunflower), Lupinus luteus cv Mister (lupin) and Glycine max (soybean), since they are economically interesting for both human and animal nutrition. The seeds were grinded and proteins extracted from 20mg powder with a specific vegetal-extraction kit. Proteins have been quantified through Bradford protocol and scavenging activity was revealed using DPPH assay, based on radical DPPH (2,2-diphenyl-1-picrylhydrazyl) absorbance decrease in the presence of antioxidants molecules. Different concentrations of the protein extract (1, 5, 10, 50, 100, 500 µg/ml) were mixed with DPPH solution (DPPH 0,004% in ethanol 70% v/v). Ascorbic acid was used as a scavenging activity standard reference, at the same six concentrations of protein extracts, while DPPH solution was used as control. Samples and standard were prepared in triplicate and incubated for 30 minutes in dark at room temperature, the absorbance was read at 517nm (ABS30). Average and standard deviation of absorbance values were calculated for each concentration of samples and standard. Statistical analysis using t-students and p-value were performed to assess the statistical significance of the scavenging activity difference between the samples (or standard) and control (ABSctrl). The percentage of antioxidant activity has been calculated using the formula [(ABSctrl-ABS30)/ABSctrl]*100. The obtained results demonstrate that all matrices showed antioxidant activity. Ascorbic acid, used as standard, exhibits a 96% scavenging activity at the concentration of 500 µg/ml. At the same conditions, sunflower, safflower and yellow peas revealed the highest antioxidant performance among the matrices analyzed, with an activity of 74%, 68% and 70% respectively (p < 0.005). Although lupin and soybean exhibit a lower antioxidant activity compared to the other matrices, they showed a percentage of 46 and 36 respectively. All these data suggest the possibility to use undervalued edible matrices as antioxidants source. However, further studies are necessary to investigate a possible synergic effect of several matrices as well as the impact of industrial processes for a large-scale approach.

Keywords: antioxidants, DPPH assay, natural matrices, vegetal proteins

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Authors: Benjaporn Buranrat, Nootchanat Mairuae

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Cratoxylum formosum (Jack) Dyer (CF) has been used for the traditional medicines in South East Asian and Thailand. Normally, northeast Thai vegetables have proven cytotoxic to many cancer cells. Therefore, the present study aims to explore the molecular mechanisms underlying CF-induced cancer cell death and apoptosis on breast and liver cancer cells. The cytotoxicity and antiproliferative effects of CF on the human breast MCF-7 and liver HepG2 cancer cell lines were evaluated using sulforhodamine B assay and colony formation assay. Cell migration assay was measured using wound healing assay. The apoptosis induction mechanisms were investigated through reactive oxygen species formation, caspase 3 activity, and JC-1 activity. Gene expression by real-time PCR and apoptosis related protein levels by Western blot analysis. CF induced MCF-7 and HepG2 cell death by time- and dose-dependent manner. Furthermore, CF had the greater cytotoxic potency on MCF-7 more than HepG2 cells with IC50 values of 85.70+4.52 μM and 219.03±9.96 μM respectively, at 24 h. Treatment with CF also caused a dose-dependent decrease in colony forming ability and cell migration, especially on MCF-7 cells. CF induced ROS formation, increased caspase 3 activities, and decreased the mitochondrial membrane potential, and causing apoptotic body production and DNA fragmentation. CF significantly decreased expression of the cell cycle regulatory protein RAC1 and downstream proteins, cdk6. Additionally, CF enhanced p21 and reduced cyclin D1 protein levels. CF leaf extract induced cell death, apoptosis, antimigration in both of MCF-7 and HepG2 cells. CF could be useful for developing to anticancer drug candidate for breast and liver cancer therapy.

Keywords: cratoxylum formosum (jack) dyer, breast cancer, liver cancer, cell death

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Authors: Camilla Trevor, Matthew K. Higgins, Andrea Gonzalez-Munoz, Mark Carrington

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Trypanosomiasis is a devastating disease affecting both humans and livestock in sub-Saharan Africa. The diseases are caused by infection with African trypanosomes, protozoa transmitted by tsetse flies. Treatment currently relies on the use of chemotherapeutics with ghastly side effects. Here, we describe the development of effective antibody-drug conjugates that target the T. brucei transferrin receptor. The receptor is essential for trypanosome growth in a mammalian host but there are approximately 12 variants of the transferrin receptor in the genome. Two of the most divergent variants were used to generate recombinant monoclonal immunoglobulin G using phage display and we identified cross-reactive antibodies that bind both variants using phage ELISA, fluorescence resonance energy transfer assays and surface plasmon resonance. Fluorescent antibodies were used to demonstrate uptake into trypanosomes in culture. Toxin-conjugated antibodies were effective at killing trypanosomes at sub-nanomolar concentrations. The approach of using antibody-drug conjugates has proven highly effective.

Keywords: antibody-drug conjugates, phage display, transferrin receptor, trypanosomes

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Authors: A. Bara, D. Barkat

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Liquid-liquid extraction is one of the most useful techniques for selective removal and recovery of metal ions from aqueous solutions, applied in purification processes in numerous chemical and metallurgical industries. In this work, The liquid-liquid extraction of cobalt (II) and copper (II) from aqueous solution by capric acid (HL) in chloroform at 25°C has been studied. Our interest in this paper is to study the effect of concentration of capric acid on the extraction of Co(II) and Cu(II) to see the complexes could be formed in the organic phase using various concentration of capric acid. The extraction of cobalt (II) and copper (II) is extracted as the complex CoL2 (HL )2, CuL2 (HL)2.

Keywords: capric acid, Cobalt(II), copper(II), liquid-liquid extraction

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Authors: M. L. Mangena, N. Mokoena, K. Rashamuse, M. G. Tlou

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Tertiary alcohol ester (TAE) hydrolases are a group of esterases (EC 3.1.1.-) that catalyze the kinetic resolution of TAEs and as a result, they are sought-after for the production of optically pure tertiary alcohols (TAs) which are useful as building blocks for number biologically active compounds. What sets these enzymes apart is, the presence of a GGG(A)X-motif in the active site which appears to be the main reason behind their activity towards the sterically demanding TAEs. The genome of Pseudomonas syringae pv. maculicola (Psm) comprises a multitude of genes that encode esterases. We therefore, hypothesize that some of these genes encode TAE hydrolases. In this study, Psm was screened for TAE hydrolase activity using the linalyl acetate (LA) plate assay and a positive reaction was observed. As a result, the genome of Psm was screened for esterases with a GGG(A)X-motif using the motif search tool and two potential TAE hydrolase genes (PsmEST1 and 2, 1100 and 1000bp, respectively) were identified, PsmEST1 was amplified by PCR and the gene sequenced for confirmation. Analysis of the sequence data with the SingnalP 4.1 server revealed that the protein comprises a signal peptide (22 amino acid residues) on the N-terminus. Primers specific for the gene encoding the mature protein (without the signal peptide) were designed such that they contain NdeI and XhoI restriction sites for directional cloning of the PCR products into pET28a. The gene was expressed in E. coli JM109 (DE3) and the clones screened for TAE hydrolase activity using the LA plate assay. A positive clone was selected, overexpressed and the protein purified using nickel affinity chromatography. The activity of the esterase towards LA was confirmed using thin layer chromatography.

Keywords: hydrolases, tertiary alcohol esters, tertiary alcohols, screening, Pseudomonas syringae pv., maculicola genome, esterase activity, linalyl acetate

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Authors: Debananda Mohapatra, Subramanya Badrayyana, Smrutiranjan Parida

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Carbon nano-onions (CNOs) are the spherical graphitic nanostructures composed of concentric shells of graphitic carbon can be hypothesized as the intermediate state between fullerenes and graphite. These are very important members in fullerene family also known as the multi-shelled fullerenes can be envisioned as promising supercapacitor electrode with high energy & power density as they provide easy access to ions at electrode-electrolyte interface due to their curvature. There is still very sparse report concerning on CNOs as electrode despite having an excellent electrodechemical performance record due to their unavailability and lack of convenient methods for their high yield preparation and purification. Keeping all these current pressing issues in mind, we present a facile scalable and straightforward flame synthesis method of pure and highly dispersible CNOs without contaminated by any other forms of carbon; hence, a post processing purification procedure is not necessary. To the best of our knowledge, this is the very first time; we developed an extremely simple, light weight, novel inexpensive, flexible free standing pristine CNOs electrode without using any binder element. Locally available daily used cotton wipe has been used for fabrication of such an ideal electrode by ‘dipping and drying’ process providing outstanding stretchability and mechanical flexibility with strong adhesion between CNOs and porous wipe. The specific capacitance 102 F/g, energy density 3.5 Wh/kg and power density 1224 W/kg at 20 mV/s scan rate are the highest values that ever recorded and reported so far in symmetrical two electrode cell configuration with 1M Na2SO4 electrolyte; indicating a very good synthesis conditions employed with optimum pore size in agreement with electrolyte ion size. This free standing CNOs electrode also showed an excellent cyclic performance and stability retaining 95% original capacity after 5000 charge –discharge cycles. Furthermore, this unique method not only affords binder free - freestanding electrode but also provide a general way of fabricating such multifunctional promising CNOs based nanocomposites for their potential device applications in flexible solar cells and lithium-ion batteries.

Keywords: binder-free, flame synthesis, flexible, carbon nano onion

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Authors: Jae Woong Sull

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Lipid levels are related to the risk of cardiovascular diseases. Cholesterol ester transfer protein (CETP) gene is one of the candidate genes of cardiovascular diseases. A total of 2,304 persons were chosen from a Hospital (N=4,294) in South Korea. Female subjects with the CG/GG genotype had a 2.03 -fold (p=0.0001) higher risk of having abnormal HDL cholesterol levels (<40 mg/dL) than subjects with the CC genotype. Male subjects with the CG/GG genotype had a 1.34 -fold (p=0.0019) higher risk than subjects with the CC genotype. When analyzed by body mass index, the association with CETP was much stronger in male subjects with BMI>=25.69 (OR=1.55, 95% CI: 1.15-2.07, P=0.0037) than in male lean subjects. When analyzed by family history of diabetes, the association with CETP was much stronger in male subjects with positive family history of low physical activity (OR=4.82, 95% CI: 1.86-12.5, P=0.0012) than in male subjects with negative family history of diabetes. This study clearly demonstrates that genetic variants in CETP influence HDL cholesterol levels in Korean adults.

Keywords: CETP, diabetes, obesity, polymorphisms

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Authors: Tansa Nisan Gunerhan

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Dementia comes in different forms, including Alzheimer's disease. The most common dementia diagnosis among elderly individuals is Alzheimer's disease. On average, for patients with Alzheimer’s, life expectancy is around 4-8 years after the diagnosis; however, expectancy can go as high as twenty years or more, depending on the shrinkage of the brain. Normally, along with aging, the brain shrinks at some level but doesn’t lose a vast amount of neurons. However, Alzheimer's patients' neurons are destroyed rapidly; hence problems with loss of memory, communication, and other metabolic activities begin. The toxic changes in the brain affect the stability of the neurons. Beta-amyloid and tau are two proteins that are believed to play a role in the development of Alzheimer's disease through their toxic changes. Beta-amyloid is a protein that is produced in the brain and is normally broken down and removed from the body. However, in people with Alzheimer's disease, the production of beta-amyloid increases, and it begins to accumulate in the brain. These plaques are thought to disrupt communication between nerve cells and may contribute to the death of brain cells. Tau is a protein that helps to stabilize microtubules, which are essential for the transportation of nutrients and other substances within brain cells. In people with Alzheimer's disease, tau becomes abnormal and begins to accumulate inside brain cells, forming neurofibrillary tangles. These tangles disrupt the normal functioning of brain cells and may contribute to their death, forming amyloid plaques which are deposits of a protein called amyloid-beta that build up between nerve cells in the brain. The accumulation of amyloid plaques and neurofibrillary tangles in the brain is thought to contribute to the shrinkage of brain tissue. As the brain shrinks, the size of the brain may decrease, leading to a reduction in brain volume. Brain atrophy in Alzheimer's disease is often accompanied by changes in the structure and function of brain cells and the connections between them, leading to a decline in brain function. These toxic changes that accumulate can cause symptoms such as memory loss, difficulty with thinking and problem-solving, and changes in behavior and personality.

Keywords: Alzheimer, amyloid-beta, brain atrophy, neuron, shrinkage

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Authors: R. Berrached, H. Ait Mahamed, A. Iddou

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Water pollution is nowadays a serious problem, due to the increasing scarcity of water and thus to the impact induced by such pollution on the human health. Various techniques are made use of to deal with water pollution. Among the most used ones, some can be enumerated: the bacterian bed, the activated sludge, lagoons as biological processes and coagulation-flocculation as a physic-chemical process. These processes are very expensive and a decreasing in efficiency treatment with the increase of the initial pollutants concentration. This is the reason why research has been reoriented towards the use of adsorption process as an alternative solution instead of the other traditional processes. In our study, we have tempted to explore the characteristics of hydroxides of Al and Fe to purify contaminated water by two industrial dyes SBL blue and SRL-150 orange. Results have shown the efficiency of the two materials on the blue SBL dye.

Keywords: metallic hydroxydes, dyes, purification, adsorption

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Authors: Shahid-ul-Islam, Faqeer Mohammad

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The present study was conducted to investigate the effect of annatto, teak and flame of the forest natural dyes on color, fastness, and antimicrobial property of protein based textile substrate. The color strength (K/S) of wool samples at various concentrations of dyes were analysed using a Reflective Spectrophotometer. The antimicrobial activity of natural dyes before and after application on wool was tested against common human pathogens Escherichia coli, Staphylococcus aureus, and Candida albicans, by using micro-broth dilution method, disc diffusion assay and growth curve studies. The structural morphology of natural protein fibre (wool) was investigated by Scanning Electron Microscopy (SEM). Annatto and teak natural dyes proved very effective in inhibiting the microbial growth in solution phase and after application on wool and resulted in a broad beautiful spectrum of colors with exceptional fastness properties. The results encourage the search and exploitation of new plant species as source of dyes to replace toxic synthetic antimicrobial agents currently used in textile industry.

Keywords: annatto, antimicrobial agents, natural dyes, green textiles

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Authors: K. Żak, S. Przetocka, R. Kitel, K. Guzik, B. Musielak, S. Malicki, G. Dubin, T. A. Holak

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Studies on tumor genesis revealed a number of factors that may potentially serve as molecular targets for immunotherapies. One of such promising targets are PD1 and PDL1 proteins. PD1 (Programmed cell death protein 1) is expressed by activated T cells and plays a critical role in modulation of the host's immune response. One of the PD1 ligands -PDL1- is expressed by macrophages, monocytes and cancer cells which exploit it to avoid immune attack. The notion of the mechanisms used by cancer cells to block the immune system response was utilized in the development of therapies blocking PD1-PDL1 interaction. Up to date, human PD1-PDL1 complex has not been crystallized and structure of the mouse-human complex does not provide a complete view of the molecular basis of PD1-PDL1 interactions. The purpose of this study is to obtain crystal structure of the human PD1-PDL1 complex which shall allow rational design of small molecule inhibitors of the interaction. In addition, the study presents results of binding small-molecules to PD1 and fragment docking towards PD1 protein which will facilitate the design and development of small–molecule inhibitors of PD1-PDL1 interaction.

Keywords: PD1, PDL1, cancer, small molecule, drug discovery

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Authors: Safa Abusara Mohammed Ali, Mohammed Khair Abdallah, Gurdon A. Brockmann, M. Reissmann

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The aim of the study was the characterization of DGAT1 variants in Sudanese dairy cattle breeds. In this study, we examined 94 Kenana and 91 Butana dairy cattle from two regions of Sudan. We genotyped the DGAT1 sequence variant AJ318490.1:g.10433/10434 AA>GC that leads to the Lysine – Alanine substitution at position 232 (K232A) in the protein and the VNTR polymorphism in the promoter region. Genotyping was performed by allele specific PCR and PCR fragment lengths determination, respectively. In both breeds, the DGAT1 Lysine variant (232K) that is associated with high fat and protein content as well as high fat yield in other breeds is the high frequent allele. The frequencies of the 232K allele were 96.3% and 84.6% in Kenana and Butana breeds, respectively. At the DGAT1 promoter VNTR locus, four alleles containing four to seven repeats of the 18 bp motif were found in both breeds. The highest frequent allele was the VNTR allele 3 containing five repeats with 60.4 % and 57.5 % in Kenana and Butana breeds, respectively. In conclusion, the two examined Sudanese dairy cattle breeds do not differ in allele frequencies at the DGAT1 locus.

Keywords: dairy cattle, DGAT1, Kenana, Butana.

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Authors: Muhammad Zeeshan, Rabia Nazir, Lubna Tahir

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Low cost filter based on iron doped zeolite (Fe-Z) and pottery clay was developed for an effective and efficient treatment of the drinking water contaminated with microbes. Fe-Z was characterized using powder XRD, SEM and EDX and shown to have average particle size of 49 nm with spongy appearance. The simulated samples of water self-contaminated with six microbes (S. typhi, B. subtilus, E. coli, S. aures, K. pneumoniae, and P. aeruginosa) after treatment with Fe-Z indicated effective removal of all the microbes in less than 30 min. Equally good results were obtained when actual drinking water samples, totally unfit for human consumption, were treated with Fe-Z.

Keywords: iron doped zeolite, biological and chemical treatment, drinking water

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Authors: Amar Ranjan, Julieana Durai, Pranay Tanwar

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Background &Introduction: Ovarian cancer is one of the most common malignant tumor in the female. 70% of the cases of ovarian cancer are diagnosed at an advanced stage. The five-year survival rate associated with ovarian cancer is less than 30%. The early diagnosis of ovarian cancer becomes a key factor in improving the survival rate of patients. Presently, CAl25 (carbohydrate antigen125) is used for the diagnosis and therapeutic monitoring of ovarian cancer, but its sensitivity and specificity is not ideal. The introduction of HE4, human epididymis protein 4 has attracted much attention. HE4 has a sensitivity and specificity of 72.9% and 95% for differentiating between benign and malignant adnexal masses, which is better than CA125 detection.  Methods: Serum HE4 and CA -125 were estimated using the chemiluminescence method. Our cases were 40 epithelial ovarian cancer, 9 benign ovarian tumor, 29 benign gynaecological diseases and 13 healthy individuals. This group include healthy woman those who have undergoing family planning and menopause-related medical consultations and they are negative for ovarian mass. Optimal cut off values for HE4 and CA125 were 55.89pmol/L and 40.25U/L respectively (determined by statistical analysis). Results: The level of HE4 was raised in all ovarian cancer patients (n=40) whereas CA125 levels were normal in 6/40 ovarian cancer patients, which were the cases of OC confirmed by histopathology. There is a significant decrease in the level of HE4 with comparison to CA125 in benign ovarian tumor cases. Both the levels of HE4 and CA125 were raised in the nonovarian cancer group, which includes cancer of endometrium and cervix. In the healthy group, HE4 was normal in all patients except in one case of the rudimentary horn, and the reason for this raised HE4 level is due to the incomplete development of uterus whereas CA125 was raised in 3 cases. Conclusions: Findings showed that the serum level of HE4 is an important indicator in the diagnosis of ovarian cancer, and it also distinguishes between benign and malignant pelvic masses. However, a combination of HE4 and CA125 panel will be extremely valuable in improving the diagnostic efficiency of ovarian cancer. These findings of our study need to be validated in the larger cohort of patients.

Keywords: human epididymis protein 4, ovarian cancer, diagnosis, benign lesions

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Authors: Manuel A. Alonso-Tarajano, Roberto Mosca, Patrick Aloy

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Protein kinases participate in a myriad of cellular processes of major biomedical interest. The in vivo substrate specificity of these enzymes is a process determined by several factors, and despite several years of research on the topic, is still far from being totally understood. In the present work, we have quantified the contributions to the kinase substrate specificity of i) the phosphorylation sites and their surrounding residues in the sequence and of ii) the association of kinases to adaptor or scaffold proteins. We have used position-specific scoring matrices (PSSMs), to represent the stretches of sequences phosphorylated by 93 families of kinases. We have found negative correlations between the number of sequences from which a PSSM is generated and the statistical significance and the performance of that PSSM. Using a subset of 22 statistically significant PSSMs, we have identified specificity determinant residues (SDRs) for 86% of the corresponding kinase families. Our results suggest that different SDRs can function as positive or negative elements of substrate recognition by the different families of kinases. Additionally, we have found that human proteins with known function as adaptors or scaffolds (kAS) tend to interact with a significantly large fraction of the substrates of the kinases to which they associate. Based on this characteristic we have identified a set of 279 potential adaptors/scaffolds (pAS) for human kinases, which is enriched in Pfam domains and functional terms tightly related to the proposed function. Moreover, our results show that for 74.6% of the kinase– pAS association found, the pAS colocalize with the substrates of the kinases they are associated to. Finally, we have found evidence suggesting that the association of kinases to adaptors and scaffolds, may contribute significantly to diminish the in vivo substrate crossed- specificity of protein kinases. In general, our results indicate the relevance of several SDRs for both the positive and negative selection of phosphorylation sites by kinase families and also suggest that the association of kinases to pAS proteins may be an important factor for the localization of the enzymes with their set of substrates.

Keywords: kinase, phosphorylation, substrate specificity, adaptors, scaffolds, cellular colocalization

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Authors: Farhat Rashid

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A study was conducted at the Institute of Food Science and Nutrition, University of Sargodha, Sargodha, Pakistan, to evaluate the effect of different concentration of Aloe vera (Aloe barbadensis Mill.) powder on physicochemical and sensory properties of orange marmalade. All treatments (0, 2, 4 6, 8 and 10% Aloe vera powder) were analyzed for titratable acidity, TSS, pH, moisture, fat, fiber and protein contents. The data indicated gradual increase in titratable acidity (0.08 to 0.18%), moisture (0.23 to 0.48%), protein (0.09 to 0.40%) and fiber (0.12 to 1.03%) among all treatments with increasing concentration of Aloe vera powder. However, a decreasing trend in pH (3.81 to 2.74), TSS (68 to 56 °Brix) and fat content (1.1 to 0.08%) was noticed with gradual increase in concentration of Aloe vera powder in orange marmalade. Sensory attributes like color, taste, texture, flavor and overall acceptability were found acceptable among all treatments but T1 (2% Aloe vera powder) was liked most and T5 (10% Aloe vera powder) was least appealing to the judges. It is concluded from present study that the addition of different concentrations of Aloe vera powder in orange marmalade significantly affected the physicochemical and sensory properties of marmalade.

Keywords: orange marmalade, Aloe vera, Aloe barbadensis mill, physicochemical, characteristics, organoleptic properties, Pakistan, treatments, significance

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Authors: Hashaam Akhtar

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IFNλR1 and IL10R2 collectively construct a heterodimer, which is an acknowledged functional receptor for all type III interferons (IFNs). Expression of IFNλR1 is highly tissue specific, which can help in making type III IFNs a drug of choice as comparable to its analogue, type I IFNs, for treating hepatitis C in the near future. Although, expression of IFNλR1 also varies with the concentration of type I IFNs, but in this study it was shown that the expression of IFNλR1 varies with the protein titers of IFN-α, IFN-λ3 and the newly discovered IFN-λ4. High dosage of IFN-α reduces the expression of IFNλR1 in HepG2 cells, which can affect the antiviral activity of type III IFNs in vivo. We premeditated an experimental strategy to differentiate monocytes into dendritic cells (DCs), type I and type II macrophages in vitro and quantified the expression of the IFNλR1 by qPCR. The exposure of newly discovered IFN-λ4 to macrophages and DCs also raised the expression of its own receptor, which shows that expression of IFN-λ4 protein in hepatitis C patient may augment type I treatment and help ease off viral titers. The results of this study may contribute in some understanding towards the mechanisms involved in the selective expression of IFNLR1 and exceptionalities associated with the receptor.

Keywords: IFNLR1, Interferon Lambda 4 (IFN-λ4), Mononuclear Phagocyte Cells (MPCs), expression

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Authors: Laima Silina, Ilze Gramatina, Liga Skudra, Tatjana Rakcejeva

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The aim of the current research was to determine quality parameters changes of dried venison during storage. Protein, fat and moisture content dynamics as well microbiological quality was analyzed. For the experiments the meat (0.02×4.00×7.00 cm) pieces were marinated in “teriyaki sauce” marinade (composition: teriyaki sauce, sweet and sour sauce, taco sauce, soy sauce, American BBQ sauce hickory, sesame oil, garlic, garlic salt, tabasco red pepper sauce) at 4±2°C temperature for 48±1h. Sodium monophosphate (E339) was also added in part of marinade to improve the meat textural properties. After marinating, meat samples were dried in microwave-vacuum drier MUSSON–1, packaged in vacuum pouches made from polymer film (PA/PE) with barrier properties and storage for 4 months at 18±1°C temperature in dark place. Dried venison samples were analyzed after 0, 35, 91 and 112 days of storage. During the storage total plate counts of dried venison samples significantly (p<0.05) increased. No significant differences in the content of protein, fat and moisture were detected when analyzing dried meat samples during storage and comparing them with the chemical parameters of just dried meat.

Keywords: drying, microwave-vacuum drier, quality, venison

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Authors: Christopher R. Triggle, Hong Ding, Khaled Machaca, Gnanapragasam Arunachalam

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The antidiabetic drug, metformin, has been in clinical use for over 50 years and remains the first choice drug for the treatment of type two diabetes. In addition to its effectiveness as an oral anti-hyperglycaemic drug metformin also possesses vasculoprotective effects that are assumed to be secondary to its ability to reduce insulin resistance and control glycated hemoglobin levels; however, recent data from our laboratory indicate that metformin also has direct vasoprotective effects that are mediated, at least in part, via the anti-ageing gene, SIRT1. Diabetes is a major risk factor for the development of cardiovascular disease (CVD) and it is also well established that tobacco use further enhances the risk of CVD; however, it is not known whether treatment with metformin can offset the negative effects of diabetes and tobacco use on cardiac function. The current study was therefore designed to investigate 1: the effects of hyperglycaemia (HG) either alone or in the presence of elevated fatty acids (palmitate) and nicotine on the protein expression levels of the deacetylase sirtuin 1 (the protein product of SIRT1), anti-apoptotic Bcl-2, pro-apoptotic BIM and the pro-apoptotic, tumour suppressor protein, acetylated p53 in cardiomyocytes. 2: the ability of metformin to prevent the detrimental effects of HG, palmitate and nicotine on cardiomyocyte survival. Cell culture protocols were designed using a rat cardiomyocyte cell line, H9c2, either under normal glycaemic (NG) conditions of 5.5mM glucose, or hyperglycaemic conditions (HG) of 25mM glucose with, or without, added palmitate (250μM) or nicotine (1.0mM) for 24h. Immuno-blotting was used to detect the expression of sirtuin 1, Bcl-2, BIM, acetylated (Ac)-p53, p53 with β-actin used as the reference protein. Exposure to HG, palmitate, or nicotine alone significantly reduced expression of sirtuin1, Bcl-2 and raised the expression levels of acetylated p53 and BIM; however, the combination of HG, palmitate and nicotine had a synergistic effect to significantly suppress the expression levels of sirtuin 1 and Bcl-2, but further enhanced the expression of Ac-p53, and BIM. The inclusion of 1000μM, but not 50μM, metformin in the H9c2 cell culture protocol prevented the effects of HG, palmitate and nicotine on the pro-apoptotic pathways. Collectively these data indicate that metformin, in addition to its anti-hyperglycaemic and vasculoprotective properties, also has direct cardioprotective actions that offset the negative effects of hyerglycaemia, elevated free fatty acids and nicotine on cardiac cell survival. These data are of particular significance for the treatment of patients with diabetes who are also smokers as the inclusion of metformin in their therapeutic treatment plan should help reduce cardiac-related morbidity and mortality.

Keywords: apoptosis, cardiac muscle, diabetes, metformin, nicotine

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Authors: Stoyanka Vladeva, Elena Kirilova, Nikola Kirilov

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Background: The creatinine clearance is a widely used value to estimate the GFR. Increased creatinine clearance is often called hyperfiltration and is usually seen during pregnancy, patients with diabetes mellitus preceding the diabetic nephropathy. It may also occur with large dietary protein intake or with plasma volume expansion. Renal injury in lupus nephritis is known to affect the glomerular, tubulointerstitial, and vascular compartment. However high creatinine clearance has not been found in patients with SLE, Target: Follow-up of creatinine clearance values in patients with systemic lupus erythematosus without history of kidney injury. Material and methods: We observed the creatinine, creatinine clearance, GFR and dipstick protein values of 7 women (with a mean age of 42.71 years) with systemic lupus erythematosus. Patients with active lupus have been monthly tested in the period of 13 months. Creatinine clearance has been estimated by Cockcroft-Gault Equation formula in ml/sec. GFR has been estimated by MDRD formula (The Modification of Diet in renal Disease) in ml/min/1.73 m2. Proteinuria has been defined as present when dipstick protein > 1+.Results: In all patients without history of kidney injury we found elevated creatinine clearance levels, but GFRremained within the reference range. Two of the patients were in remission while the other five patients had clinically and immunologically active Lupus. Three of the patients had a permanent presence of high creatinine clearance levels and proteinuria. Two of the patients had periodically elevated creatinine clearance without proteinuria. These results show that kidney disturbances may be caused by the vascular changes typical for SLE. Glomerular hyperfiltration can be result of focal segmental glomerulosclerosis caused by a reduction in renal mass. Probably lupus nephropathy is preceded not only by glomerular vascular changes, but also by tubular vascular changes. Using only the GFR is not a sufficient method to detect these primary functional disturbances. Conclusion: For early detection of kidney injury in patients with SLE we determined that the follow up of creatinine clearance values could be helpful.

Keywords: systemic Lupus erythematosus, kidney injury, elevated creatinine clearance level, normal glomerular filtration rate

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Authors: Abdulrazzaq Hammal

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In this study, Syrian sand was used to create ceramic membranes. The process of preparing the membranes involved several steps, starting with the purification of the studied sand using hydrochloric acid, sorting according to granular size, and mixing the sand with liquid sodium silicates as a binder. Next, the effects of binder ratio, pressure formation, treatment temperature, and sand grain size were studied. Further, nanoparticles of silver were added to the formed membranes to improve their ability to purify bacterially polluted water. Prepared membranes were quite successful in removing bacteria and chemicals from water, and the water's requirements were brought up to level with Syrian drinking water standards.

Keywords: ceramic, membrane, water, wastewater

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Authors: Bea Carmel H. Casiding, Charmaine A. Guy, Funny Jovis P. Malasan, Katrina Chelsea B. Manlutac, Danielle Ann N. Novilla, Marianne R. Oliveros, Magnolia C. Sibulo

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Moon jellyfish, Aurelia aurita, has become a popular research organism for diverse studies. Recent studies have verified the prevention of blood clotting properties of the moon jellyfish tentacle extract through in vitro methods. The purpose of this study was to validate the blood clotting ability of A. aurita tentacle extract using in vivo method of experimentation. The tentacles of A. aurita jellyfish were excised and filtered then centrifuged at 3000xg for 10 minutes. The crude nematocyst extract was suspended in 1:6 ratios with phosphate buffer solution and sonicated for three periods of 20 seconds each at 50 Hz. Protein concentration of the extract was determined using Bradford Assay. Bovine serum albumin was the standard solution used with the following concentrations: 35.0, 70.0, 105.0, 140.0, 175.0, 210.0, 245.0, and 280.0 µg/mL. The absorbance was read at 595 nm. Toxicity testing from OECD guidelines was adapted. The extract suspended in phosphate-buffered saline solution was arbitrarily set into three doses (0.1mg/kg, 0.3mg/kg, 0.5mg/kg) and were administered daily for five days to the experimental groups of five male Sprague-Dawley rats (one dose per group). Before and after the administration period, bleeding time and clotting time tests were performed. The One-way Analysis of Variance (ANOVA) was used to analyze the difference of before and after bleeding time and clotting time from the three treatment groups, time, positive and negative control groups. The average protein concentration of the sonicated crude tentacle extract was 206.5 µg/mL. The highest dose administered (0.5mg/kg) produced significant increase in the time for both bleeding and clotting tests. However, the preceding lower dose (0.3mg/kg) only was significantly effective for clotting time test. The protein contained in the tentacle extract with a concentration of 206.5 mcg/mL and dose of 0.3 mg/kg and 0.5 mg/kg of A. aurita elicited anticoagulating activity.

Keywords: anticoagulant, bleeding time test, clotting time test, moon jellyfish

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Authors: R. Berrached, H. Ait Mahamed, A. Iddou

Abstract:

Water pollution is nowadays a serious problem, due to the increasing scarcity of water and thus to the impact induced by such pollution on the human health. Various techniques are made use of to deal with water pollution. Among the most used ones, some can be enumerated: the bacterian bed, the activated mud, the Lagunage as biological processes and coagulation-floculation as a physic-chemical process. These processes are very expensive and an treatment efficiency which decreases along with the increase of the initial pollutants’ concentration. This is the reason why research has been reoriented towards the use of a process by adsorption as an alternative solution instead of the other traditional processes. In our study, we have tempted to exploit the characteristics of two metallic hydroxides Al and Fe to purify contaminated water by two industrial dyes SBL blue and SRL-150 orange. Results have shown the efficiency of the two materials on the blue SBL dye.

Keywords: Metallic Hydroxydes, industrial dyes, purification, lagunage

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Authors: Isobel Hambleton, Mark Carrington

Abstract:

Trypanosoma brucei, the causal agent of a range of diseases in humans and livestock, evades the mammalian immune system through a population survival strategy based on the expression of a series of antigenically distinct variant surface glycoproteins (VSGs). RNAi mediated knockdown of the active VSG gene triggers a precytokinesis cell cycle arrest. To determine whether this phenotype is the result of reduced VSG transcript or depleted VSG protein, we used morpholino antisense oligonucleotides to block translation of VSG mRNA. The same precytokinesis cell cycle arrest was observed, suggesting that VSG protein abundance is monitored closely throughout the cell cycle. An inducible expression system has been developed to test various GPI-anchored proteins for their ability to rescue this cell cycle arrest. This system has been used to demonstrate that wild-type VSG expressed from a T7 promoter rescues this phenotype. This indicates that VSG expression from one of the specialised bloodstream expression sites (BES) is not essential for cell division. The same approach has been used to define the minimum essential features of a VSG necessary for function.

Keywords: bloodstream expression site, morpholino, precytokinesis cell cycle arrest, variant surface glycoprotein

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Authors: Elene Zhuravliova, Tamar Barbakadze, Irina Kalandadze, Elnari Zaalishvili, Lali Shanshiashvili, David Mikeladze

Abstract:

Background: Multiple sclerosis (MS) is an inflammatory, neurodegenerative disease, which is accompanied by demyelination and autoimmune response to myelin proteins. Among post-translational modifications, which mediate the modulation of inflammatory pathways during MS, methylation is the main one. The methylation of DNA, also amino acids lysine and arginine, occurs in the cell. It was found that decreased trans-methylation is associated with neuroinflammatory diseases. Therefore, abnormal regulation of the methyl cycle could induce demyelination through the action on PAD (peptidyl-arginine-deiminase) gene promoter. PAD takes part in protein citrullination and targets myelin basic protein (MBP), which is affected during demyelination. To determine whether MBP charge isomers are changing the methyl cycle, we have estimated the concentrations of methyl cycle metabolites in MBP-activated primary astrocytes and oligodendrocytes. For this purpose, the action of the citrullinated MBP- C8 and the most cationic MBP-C1 isomers on the primary cells were investigated. Methods: Primary oligodendrocyte and astrocyte cell cultures were prepared from whole brains of 2-day-old Wistar rats. The methyl cycle metabolites, including homocysteine, S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH), were estimated by HPLC analysis using fluorescence detection and prior derivatization. Results: We found that the action of MBP-C8 and MBP-C1 induces a decrease in the concentration of both methyl cycle metabolites, S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), in astrocytes compared to the control cells. As for oligodendrocytes, the concentration of SAM was increased by the addition of MBP-C1, while MBP-C8 has no significant effect. As for SAH, its concentration was increased compared to the control cells by the action of both MBP-C1 and MBP-C8. A significant increase in homocysteine concentration was observed by the action of the MBP-C8 isomer in both oligodendrocytes and astrocytes. Conclusion: These data suggest that MBP charge isomers change the concentration of methyl cycle metabolites. MBP-C8 citrullinated isomer causes elevation of homocysteine in astrocytes and oligodendrocytes, which may be the reason for decreased astrocyte proliferation and increased oligodendrocyte cell death which takes place in neurodegenerative processes. Elevated homocysteine levels and subsequent abnormal regulation of methyl cycles in oligodendrocytes possibly change the methylation of DNA that activates PAD gene promoter and induces the synthesis of PAD, which in turn provokes the process of citrullination, which is the accompanying process of demyelination. Acknowledgment: This research was supported by the SRNSF Georgia RF17_534 grant.

Keywords: myelin basic protein, astrocytes, methyl cycle metabolites, homocysteine, oligodendrocytes

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Authors: Usman Mir Khan, Ishtiaque Ahmad, Saima Inayat, H. M. Arslan Amin, Muhammad Ayaz, Nisar Ahmad

Abstract:

Citrus reticulata Blanco crude flowers extract (CFE) at four different concentration (1, 2, 3 and 4%, v/v) were used as natural milk coagulant instead of rennet to apply for Cheddar cheese making from buffalo milk. The physicochemical properties and nutrition composition of Cheddar cheeses were compared with cheese made with 0.002% (v/v) rennet (control cheese). Physico-chemical of Cheddar cheese showed that cheese made with 1% and 2% of CFE had a crumbly and slightly softer texture of cheese. While, cheeses containing 3 and 4% CFE had semi-hard textural properties of curd similar to rennet added cheese. The CFE made cheese had moisture 37 %, fat 45 % on dry basis similar to rennet made Cheddar cheese. Protein analysis shows that CFE made cheese had significant higher protein content than control. The Cheddar cheese with 3% and 1% CFE were preferred by consumers instead of 2% and 4% CFE for their taste, texture/appearance and overall acceptability. Conclusively, CFE coagulated Cheddar cheese fulfills the nutritional requirement with acceptable organoleptic characteristics and at the same time provides nutritional health benefits.

Keywords: cheddar cheese, Citrus reticulata Blanco, buffalo milk, milk coagulant

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Authors: Ki Woong Lee

Abstract:

The Center for Axion and Precision Physics Research is studying the search for dark matter using 12 tesla superconducting magnets. A dilution refrigerator is being used for search experiments, and superconducting magnets, superconducting cavities. The dilution refrigerator requires a stable cryogenic environment using liquid helium. Accordingly, a cryogenic system for a stable supply of liquid helium is to be established. This cryogenic system includes the liquefying, supply, storage, and purification of liquid helium. This article presents the basic design, construction, and operation plans for building cryogenic systems.

Keywords: cryogenic system, dilution refrigerator, superconducting magnet, helium recovery system

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Authors: Marzieh Eshaghi Koupaei

Abstract:

By cultivating mealworm, we reduce greenhouse gases and avoid the use of transgenic products such as soybeans, and we provide food resources rich in protein, amino acids, minerals, etc. for humans and animals, and it has created employment and entrepreneurship. We serve the environment by producing oil from mealworm in the cosmetic industry, using its waste as organic fertilizer and its powder in bodybuilding, and by breaking down plastic by mealworm. The production and breeding of mealworm requires very little infrastructure and does not require much trouble, and requires very little food, and reproduces easily and quickly, and a mealworm production workshop is noiseless, odorless, and pollution-free And the costs are very low. It is possible to use third grade fruits and unsalable fruits of farmers to feed the mealworms, which is completely economical and cost-effective. Mealworms can break down plastic in their intestines and turn it into carbon dioxide. . This process was done in only 16 days, which is a very short time compared to several centuries for plastic to decompose. By producing mealworm, we have helped to preserve the environment and provided the source of protein needed by humans and animals. This industrial insect has the ability and value of commercialization and creates employment and helps the economy of the society.

Keywords: breeding, production of insects, mealworms, research, animal feed, human feed

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Authors: Jyh-Cheng Chen, Tai-Jing Wang, Yun-Wei Lin

Abstract:

Astaxanthin has been demonstrated to exhibit a wide range of beneficial effects including anti-inflammatory and anti-cancer properties. However, the molecular mechanism of astaxanthin-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination and high levels of Rad51 expression are observed in chemo- or radioresistant carcinomas. In this study, astaxanthin treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1703. Treatment with astaxanthin decreased Rad51 expression and phospho-AKT protein level in a time and dose-dependent manner. Furthermore, expression of constitutively active AKT (AKT-CA) vector significantly rescued the decreased Rad51 protein and mRNA levels in astaxanthin-treated NSCLC cells. Combined treatment with PI3K inhibitors (LY294002 or wortmannin) and astaxanthin further decreased the Rad51 expression in NSCLC cells. Knockdown of Rad51 enhanced astaxanthin-induced cytotoxicity and growth inhibition in NSCLC cells. These findings may have implications for the rational design of future drug regimens incorporating astaxanthin for the treatment of NSCLC.

Keywords: astaxanthin, cytotoxicity, AKT, non-small cell lung cancer, PI3K

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Authors: Muhammad Zubair, Aman Ullah, Jianping Wu

Abstract:

During the last decades, petroleum derived synthetic polymers like polyethylene terephthalate, polyvinylchloride, polyethylene, polypropylene and polystyrene has extensively been used in the field of food packaging and mostly are non-degradable. Biopolymers are a good fit for single-use or short-lived products such as food packaging. Spent hens, a poultry by-product which is of little economic value and their disposal are environmentally harmful. Through current study, we have explored the possibility to transform proteins from spent fowl into green food packaging material. Proteins from spent fowl were extracted within 1 hour using pH shift method with recovery of about 74%. Different plasticizers were tried like glycerol, sorbitol, glutaraldehyde, 1,2 ethylene glycol and 1,2 butanediol. Glycerol was the best plasticizer among all these plasticizers. A naturally occurring and non-toxic cross-linking agent, chitosan, was used to form the chitosan/glycerol/protein blend by casting and compression molding techniques. The mechanical properties were characterized using tensile strength analyzer. The nano-reinforcements with homogeneous dispersion of nanoparticles lead to improved physical properties suggesting that these materials have great potential for food packaging applications.

Keywords: differential scanning calorimetry, dynamic mechanical analysis, scanning electron microscopy, spent hen

Procedia PDF Downloads 276