Search results for: protein based

Authors: Henri El Zakhem, Dona Shalhoub, Elias Atallah, Jessica Koura

Abstract:

Due to the increase of demand on dietetic food and the need for more types of diet food, the production of dietetic food is increasing and improving. This demand on dietetic food has triggered us to study the market in which we found that Kishk (Lebanese dairy product) diet is not available. Production of a low fat product which is diet Kishk was our concern. A strategy was followed to choose the right idea that will satisfy the need of the market. The whole process was studied and explained thoroughly. The percentage of fat was found to be 32.52 % in regular Kishk and 3.84 % in the diet Kishk produced. The new product has the advantage to be high in protein, low in fat.

Keywords: design and industrialization, dietetic, diet Kishk, fat

Procedia PDF Downloads 371

Authors: Adel Alblihy, Reem Ali, Mashael Algethami, Ahmed Shoqafi, Michael S. Toss, Juliette Brownlie, Natalie J. Tatum, Ian Hickson, Paloma Ordonez Moran, Anna Grabowska, Jennie N. Jeyapalan, Nigel P. Mongan, Emad A. Rakha, Srinivasan Madhusudan

Abstract:

Platinum resistance is a clinical challenge in ovarian cancer. Platinating agents induce DNA damage which activate Mre11 nuclease directed DNA damage signalling and response (DDR). Upregulation of DDR may promote chemotherapy resistance. Here we have comprehensively evaluated Mre11 in epithelial ovarian cancers. In clinical cohort that received platinum- based chemotherapy (n=331), Mre11 protein overexpression was associated with aggressive phenotype and poor progression free survival (PFS) (p=0.002). In the ovarian cancer genome atlas (TCGA) cohort (n=498), Mre11 gene amplification was observed in a subset of serous tumours (5%) which correlated highly with Mre11 mRNA levels (p<0.0001). Altered Mre11 levels was linked with genome wide alterations that can influence platinum sensitivity. At the transcriptomic level (n=1259), Mre11 overexpression was associated with poor PFS (p=0.003). ROC analysis showed an area under the curve (AUC) of 0.642 for response to platinum-based chemotherapy. Pre-clinically, Mre11 depletion by gene knock down or blockade by small molecule inhibitor (Mirin) reversed platinum resistance in ovarian cancer cells and in 3D spheroid models. Importantly, Mre11 inhibition was synthetically lethal in platinum sensitive XRCC1 deficient ovarian cancer cells and 3D-spheroids. Selective cytotoxicity was associated with DNA double strand break (DSB) accumulation, S-phase cell cycle arrest and increased apoptosis. We conclude that pharmaceutical development of Mre11 inhibitors is a viable clinical strategy for platinum sensitization and synthetic lethality in ovarian cancer.

Keywords: MRE11; XRCC1, ovarian cancer, platinum sensitization, synthetic lethality

Procedia PDF Downloads 127

Authors: Debamitra Chakravorty, Pratap K. Parida

Abstract:

Cold-adapted proteases enhance wash performance in low-temperature laundry resulting in a reduction in energy consumption and wear of textiles and are also used in the dehairing process in leather industries. Unfortunately, the possible drawbacks of using cold-adapted proteases are their instability at higher temperatures. Therefore, proteases with broad temperature stability are required. Unfortunately, wild-type cold-adapted proteases exhibit instability at higher temperatures and thus have low shelf lives. Therefore, attempts to engineer cold-adapted proteases by protein engineering were made previously by directed evolution and random mutagenesis. The lacuna is the time, capital, and labour involved to obtain these variants are very demanding and challenging. Therefore, rational engineering for cold stability without compromising an enzyme's optimum pH and temperature for activity is the current requirement. In this work, mutations were rationally designed with the aid of high throughput computational methodology of network analysis, evolutionary conservation scores, and molecular dynamics simulations for Savinase from Bacillus lentus with the intention of rendering the mutants cold stable without affecting their temperature and pH optimum for activity. Further, an attempt was made to incorporate a mutation in the most stable mutant rationally obtained by this method to introduce oxidative stability in the mutant. Such enzymes are desired in detergents with bleaching agents. In silico analysis by performing 300 ns molecular dynamics simulations at 5 different temperatures revealed that these three mutants were found to be better in cold stability compared to the wild type Savinase from Bacillus lentus. Conclusively, this work shows that cold adaptation without losing optimum temperature and pH stability and additionally stability from oxidative damage can be rationally designed by in silico enzyme engineering. The key findings of this work were first, the in silico data of H5 (cold stable savinase) used as a control in this work, corroborated with its reported wet lab temperature stability data. Secondly, three cold stable mutants of Savinase from Bacillus lentus were rationally identified. Lastly, a mutation which will stabilize savinase against oxidative damage was additionally identified.

Keywords: cold stability, molecular dynamics simulations, protein engineering, rational design

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Authors: Neelofar, Khursheed Alam, Jamal Ahmad

Abstract:

Protein modification in diabetes mellitus may lead to early glycation products (EGPs) or amadori product as well as advanced glycation end products (AGEs). Early glycation involves the reaction of glucose with N-terminal and lysyl side chain amino groups to form Schiff’s base which undergoes rearrangements to form more stable early glycation product known as Amadori product. After Amadori, the reactions become more complicated leading to the formation of advanced glycation end products (AGEs) that interact with various AGE receptors, thereby playing an important role in the long-term complications of diabetes. Millard reaction or nonenzymatic glycation reaction accelerate in diabetes due to hyperglycation and alter serum protein’s structure, their normal functions that lead micro and macro vascular complications in diabetic patients. In this study, Human Serum Albumin (HSA) with a constant concentration was incubated with different concentrations of glucose at 370C for a week. At 4th day, Amadori product was formed that was confirmed by colorimetric method NBT assay and TBA assay which both are authenticate early glycation product. Conformational changes in native as well as all samples of Amadori albumin with different concentrations of glucose were investigated by various biophysical and biochemical techniques. Main biophysical techniques hyperchromacity, quenching of fluorescence intensity, FTIR, CD and SDS-PAGE were used. Further conformational changes were observed by biochemical assays mainly HMF formation, fructoseamine, reduction of fructoseamine with NaBH4, carbonyl content estimation, lysine and arginine residues estimation, ANS binding property and thiol group estimation. This study find structural and biochemical changes in Amadori modified HSA with normal to hyperchronic range of glucose with respect to native HSA. When glucose concentration was increased from normal to chronic range biochemical and structural changes also increased. Highest alteration in secondary and tertiary structure and conformation in glycated HSA was observed at the hyperchronic concentration (75mM) of glucose. Although it has been found that Amadori modified proteins is also involved in secondary complications of diabetes as AGEs but very few studies have been done to analyze the conformational changes in Amadori modified proteins due to early glycation. Most of the studies were found on the structural changes in Amadori protein at a particular glucose concentration but no study was found to compare the biophysical and biochemical changes in HSA due to early glycation with a range of glucose concentration at a constant incubation time. So this study provide the information about the biochemical and biophysical changes occur in Amadori modified albumin at a range of glucose normal to chronic in diabetes. Although many implicates currently in use i.e. glycaemic control, insulin treatment and other chemical therapies that can control many aspects of diabetes. However, even with intensive use of current antidiabetic agents more than 50 % of diabetic patient’s type 2 suffers poor glycaemic control and 18 % develop serious complications within six years of diagnosis. Experimental evidence related to diabetes suggests that preventing the nonenzymatic glycation of relevant proteins or blocking their biological effects might beneficially influence the evolution of vascular complications in diabetic patients or quantization of amadori adduct of HSA by authentic antibodies against HSA-EGPs can be used as marker for early detection of the initiation/progression of secondary complications of diabetes. So this research work may be helpful for the same.

Keywords: diabetes mellitus, glycation, albumin, amadori, biophysical and biochemical techniques

Procedia PDF Downloads 270

Authors: Karina I. Hidas, Csaba Németh, Anna Visy, Judit Csonka, László Friedrich, Ildikó Cs. Nyulas-Zeke

Abstract:

Egg yolk is a popular ingredient in the food industry due to its gelling, emulsifying, colouring, and coagulating properties. Because of the heat sensitivity of proteins, egg yolk can only be heat treated at low temperatures, so its shelf life, even with the addition of a preservative, is only a few weeks. Freezing can increase the shelf life of liquid egg yolk up to 1 year, but it undergoes gelling below -6 ° C, which is an irreversible phenomenon. The degree of gelation depends on the time and temperature of freezing and is influenced by the process of thawing. Therefore, in our experiment, we examined egg yolks thawed in different ways. In this study, unpasteurized, industrially broken, separated, and homogenized liquid egg yolk was used. Freshly produced samples were frozen in plastic containers at -18°C in a laboratory freezer. Frozen storage was performed for 90 days. Samples were analysed at day zero (unfrozen) and after frozen storage for 1, 7, 14, 30, 60 and 90 days. Samples were thawed in two ways (at 5°C for 24 hours and 30°C for 3 hours) before testing. Calorimetric properties were examined by differential scanning calorimetry, where heat flow curves were recorded. Denaturation enthalpy values were calculated by fitting a linear baseline, and denaturation temperature values were evaluated. Besides, dry matter content of samples was measured by the oven method with drying at 105°C to constant weight. For statistical analysis two-way ANOVA (α = 0.05) was employed, where thawing mode and freezing time were the fixed factors. Denaturation enthalpy values decreased from 1.1 to 0.47 at the end of the storage experiment, which represents a reduction of about 60%. The effect of freezing time was significant on these values, already the enthalpy of samples stored frozen for 1 day was significantly reduced. However, the mode of thawing did not significantly affect the denaturation enthalpy of the samples, and no interaction was seen between the two factors. The denaturation temperature and dry matter content did not change significantly either during the freezing period or during the defrosting mode. Results of our study show that slow freezing and frozen storage at -18°C greatly reduces the amount of protein that can be denatured in egg yolk, indicating that the proteins have been subjected to aggregation, denaturation or other protein conversions regardless of how they were thawed.

Keywords: denaturation enthalpy, differential scanning calorimetry, liquid egg yolk, slow freezing

Procedia PDF Downloads 127

Authors: Adeline Meriaux, Claire Gaiani, Jennifer Burgain, Frantz Fournier, Lionel Muniglia, Jérémy Petit

Abstract:

Spray-drying process is widely used for the production of dairy powders for food and pharmaceuticals industries. It involves the atomization of a liquid feed into fine droplets, which are subsequently dried through contact with a hot air flow. The resulting powders permit transportation cost reduction and shelf life increase but can also exhibit various interesting functionalities (flowability, solubility, protein modification or acid gelation), depending on operating conditions and milk composition. Indeed, particles porosity, surface composition, lactose crystallization, protein denaturation, protein association or crust formation may change. Links between spray-drying conditions and physicochemical and functional properties of powders were investigated by a design of experiment methodology and analyzed by principal component analysis. Quadratic models were developed, and multicriteria optimization was carried out by the use of genetic algorithm. At the time of abstract submission, verification spray-drying trials are ongoing. To perform experiments, milk from dairy farm was collected, skimmed, froze and spray-dried at different air pressure (between 1 and 3 bars) and outlet temperature (between 75 and 95 °C). Dry matter, minerals content and proteins content were determined by standard method. Solubility index, absorption index and hygroscopicity were determined by method found in literature. Particle size distribution were obtained by laser diffraction granulometry. Location of the powder color in the Cielab color space and water activity were characterized by a colorimeter and an aw-value meter, respectively. Flow properties were characterized with FT4 powder rheometer; in particular compressibility and shearing test were performed. Air pressure and outlet temperature are key factors that directly impact the drying kinetics and powder characteristics during spray-drying process. It was shown that the air pressure affects the particle size distribution by impacting the size of droplet exiting the nozzle. Moreover, small particles lead to more cohesive powder and less saturated color of powders. Higher outlet temperature results in lower moisture level particles which are less sticky and can explain a spray-drying yield increase and the higher cohesiveness; it also leads to particle with low water activity because of the intense evaporation rate. However, it induces a high hygroscopicity, thus, powders tend to get wet rapidly if they are not well stored. On the other hand, high temperature provokes a decrease of native serum proteins which is positively correlated to gelation properties (gel point and firmness). Partial denaturation of serum proteins can improve functional properties of powder. The control of air pressure and outlet temperature during the spray-drying process significantly affects the physicochemical and functional properties of powder. This study permitted to better understand the links between physicochemical and functional properties of powder, to identify correlations between air pressure and outlet temperature. Therefore, mathematical models have been developed and the use of genetic algorithm will allow the optimization of powder functionalities.

Keywords: dairy powders, spray-drying, powders functionalities, design of experiment

Procedia PDF Downloads 90

Authors: Dominika Pihikova, Stefan Belicky, Tomas Bertok, Roman Sokol, Petra Kubanikova, Jan Tkac

Abstract:

Since aberrant glycosylation is frequently accompanied by both physiological and pathological processes in a human body (cancer, AIDS, inflammatory diseases, etc.), the analysis of tumor-associated glycan patterns have a great potential for the development of novel diagnostic approaches. Moreover, altered glycoforms may assist as a suitable tool for the specificity and sensitivity enhancement in early-stage prostate cancer diagnosis. In this paper we discuss the construction and optimization of ultrasensitive sandwich biosensor platform employing lectin as glycan-binding protein. We focus on the immunoassay development, reduction of non-specific interactions and final glycoprofiling of human serum samples including both prostate cancer (PCa) patients and healthy controls. The fabricated biosensor was measured by label-free electrochemical impedance spectroscopy (EIS) with further lectin microarray verification. Furthermore, we analyzed different biosensor interfaces with atomic force microscopy (AFM) in nanomechanical mapping mode showing a significant differences in the altitude. These preliminary results revealing an elevated content of α-2,3 linked sialic acid in PCa patients comparing with healthy controls. All these experiments are important step towards development of point-of-care devices and discovery of novel glyco-biomarkers applicable in cancer diagnosis.

Keywords: biosensor, glycan, lectin, prostate cancer

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Authors: Ziaul Islam, Asad Sultan, Sarzamin Khan

Abstract:

Organic acids and micro encapsulated phyto essential oils have revealed great potential as an antibiotic replacement and as an additive to work tremendously for the health maintenance of broiler chicken. To explore more about organic acids, a total of 600 day-old broiler chicks (Cobb-500) were procured from a local hatchery and distributed into 5 treatment groups having 6 replicates of 20 birds each; the duration of the biological trial was of 35 days. Group T1 served as a control group that were fed on corn soy-based diet only. T2 were fed with a diet having 6% poultry by-product meal (PBM) diet, T3, T4, and T5 were served as the same diet as T2 but supplemented with an organic acid, phyto essential oils alone, and a combination, respectively. The findings declared significant improvement (p<0.05) in body weight gain and FCR in groups T3, T4, and T5 while feed intake was not affected. European broiler performance indicators like production efficiency factor (EPEF) and broiler index (EBI) were improved significantly (p<0.05) by the treatments T3, T4, and T5 compared with T1 and T2. Carcass evaluation depicted significantly better (p<0.05) dressed and eviscerated weight along with carcass yield (T3, T4, T5). Broilers fed organic acid and phyto essential oils supplemented diet had significantly lower (p<0.05) Clostridium perfringens, Escherichia coliand Salmonella and increased Lactobacillus counts. Likewise, antibody titer against ND, IB, and IBD were also significantly (p<0.05) improved by the treatments T3, T4 and T5compared with the T1and T2. Litter moisture content was significantly (p<0.05) reduced by treatmentsT3, T4, and T5 on day 28 and 35 compared with the T1 and T2. These findings of the present study revealed that supplementation of organic acids blend and phyto-essential oils as an as an substitute to improve the performance of broilers without the use of feed antibiotics in broilers fed with 6% poultry by-product meal based diet.

Keywords: organic acid, phyto essential oils, growth performance, PBM, gut health, microbiota, immunity

Procedia PDF Downloads 126

Authors: T. C. C. Soo, S. Bhassu

Abstract:

Unlike the common presence of both innate and adaptive immunity in vertebrates, crustaceans, in particular, shrimps, have been discovered to possess only innate immunity. This further emphasizes the importance of innate immunity within shrimps in pathogenic resistance. Under the study of pathogenic immune challenge, different shrimp species actually exhibit varying degrees of immune resistance towards the same pathogen. Furthermore, even within the same shrimp species, different batches of challenged shrimps can have different strengths of immune defence. Several important pathways are activated within shrimps during pathogenic infection. One of them is JAK-STAT pathway that is activated during bacterial, viral and fungal infections by which STAT(Signal Transducer and Activator of Transcription) gene is the core element of the pathway. Based on theory of Central Dogma, the genomic information is transmitted in the order of DNA, RNA and protein. This study is focused in uncovering the important evolutionary patterns present within the DNA (non-coding region) and RNA (coding region). The three shrimp species involved are Macrobrachium rosenbergii, Penaeus monodon and Litopenaeus vannamei which all possess commercial significance. The shrimp species were challenged with a famous penaeid shrimp virus called white spot syndrome virus (WSSV) which can cause serious lethality. Tissue samples were collected during time intervals of 0h, 3h, 6h, 12h, 24h, 36h and 48h. The DNA and RNA samples were then extracted using conventional kits from the hepatopancreas tissue samples. PCR technique together with designed STAT gene conserved primers were utilized for identification of the STAT coding sequences using RNA-converted cDNA samples and subsequent characterization using various bioinformatics approaches including Ramachandran plot, ProtParam and SWISS-MODEL. The varying levels of immune STAT gene activation for the three shrimp species during WSSV infection were confirmed using qRT-PCR technique. For one sample, three biological replicates with three technical replicates each were used for qRT-PCR. On the other hand, DNA samples were important for uncovering the structural variations within the genomic region of STAT gene which would greatly assist in understanding the STAT protein functional variations. The partially-overlapping primers technique was used for the genomic region sequencing. The evolutionary inferences and event predictions were then conducted through the Bayesian Inference method using all the acquired coding and non-coding sequences. This was supplemented by the construction of conventional phylogenetic trees using Maximum likelihood method. The results showed that adaptive evolution caused STAT gene sequence mutations between different shrimp species which led to evolutionary divergence event. Subsequently, the divergent sites were correlated to the differing expressions of STAT gene. Ultimately, this study assists in knowing the shrimp species innate immune variability and selection of disease resistant shrimps for breeding purpose. The deeper understanding of STAT gene evolution from the perspective of both purifying and adaptive approaches not only can provide better immunological insight among shrimp species, but also can be used as a good reference for immunological studies in humans or other model organisms.

Keywords: gene evolution, JAK-STAT pathway, immunology, STAT gene

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Authors: Xiang Wang, Yan Pang, Zhaomiao Liu

Abstract:

Droplet-based microfluidics have been studied extensively with the development of the Micro-Electro-Mechanical System (MEMS) which bears the advantages of high throughput, high efficiency, low cost and low polydispersity. Droplets, worked as versatile carriers, could provide isolated chambers as the internal dispersed phase is protected from the outside continuous phase. Droplets are used to add reagents to start or end bio-chemical reactions, to generate concentration gradients, to realize hydrate crystallization or protein analyses, while droplets coalescence acts as an important control technology. In this paper, deionized water is used as the dispersed phase, and several kinds of oil are used as the continuous phase to investigate the influence of the viscosity ratio of the two phases on the coalescence process. The microchannels are fabricated by coating a polydimethylsiloxane (PDMS) layer onto another PDMS flat plate after corona treatment. All newly made microchannels are rinsed with the continuous oil phase for hours before experiments to ensure the swelling fully developed. High-speed microscope system is used to document the serial videos with a maximum speed of 2000 frames per second. The critical capillary numbers (Ca*) of droplet pairs in various junctions are studied and compared. Ca* varies with different junctions or different liquids within the range of 0.002 to 0.01. However, droplets without extra control would have the problem of synchronism which reduces the coalescence efficiency.

Keywords: coalescence, concentration, critical capillary number, droplet pair, split

Procedia PDF Downloads 249

Authors: Viviane A. O. Silva, Angela M. Costa, Glaucia N. M. Hajj, Ana Preto, Aline Tansini, Martin Roffé, Peter Jordan, Rui M. Reis

Abstract:

Autophagy is a recycling and degradative system suggested to be a major cell death pathway in cancer cells. Autophagy pathway is interconnected with the endocytosis pathways sharing the same ultimate lysosomal destination. Lysosomes are crucial regulators of cell homeostasis, responsible to downregulate receptor signalling and turnover. It seems highly likely that derailed endocytosis can make major contributions to several hallmarks of cancer. WNK2, a member of the WNK (with-no-lysine [K]) subfamily of protein kinases, had been found downregulated by its promoter hypermethylation, and has been proposed to act as a specific tumour-suppressor gene in brain tumors. Although some contradictory studies indicated WNK2 as an autophagy modulator, its role in cancer cell death is largely unknown. There is also growing evidence for additional roles of WNK kinases in vesicular traffic. Aim: To evaluate the role of WNK2 in autophagy and endocytosis on glioma context. Methods: Wild-type (wt) A172 cells (WNK2 promoter-methylated), and A172 transfected either with an empty vector (Ev) or with a WNK2 expression vector, were used to assess the cellular basal capacities to promote autophagy, through western blot and flow-cytometry analysis. Additionally, we evaluated the effect of WNK2 on general endocytosis trafficking routes by immunofluorescence. Results: The re-expression of ectopic WNK2 did not interfere with autophagy-related protein light chain 3 (LC3-II) expression levels as well as did not promote mTOR signaling pathway alteration when compared with Ev or wt A172 cells. However, the restoration of WNK2 resulted in a marked increase (8 to 92,4%) of Acidic Vesicular Organelles formation (AVOs). Moreover, our results also suggest that WNK2 cells promotes delay in uptake and internalization rate of cholera toxin B and transferrin ligands. Conclusions: The restoration of WNK2 interferes in vesicular traffic during endocytosis pathway and increase AVOs formation. This results also suggest the role of WNK2 in growth factor receptor turnover related to cell growth and homeostasis and associates one more time, WNK2 silencing contribution in genesis of gliomas.

Keywords: autophagy, endocytosis, glioma, WNK2

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Authors: Adeline Meriaux, Claire Gaiani, Jennifer Burgain, Frantz Fournier, Lionel Muniglia, Jérémy Petit

Abstract:

Spray-drying process is widely used for the production of dairy powders for food and pharmaceuticals industries. It involves the atomization of a liquid feed into fine droplets, which are subsequently dried through contact with a hot air flow. The resulting powders permit transportation cost reduction and shelf life increase but can also exhibit various interesting functionalities (flowability, solubility, protein modification or acid gelation), depending on operating conditions and milk composition. Indeed, particles porosity, surface composition, lactose crystallization, protein denaturation, protein association or crust formation may change. Links between spray-drying conditions and physicochemical and functional properties of powders were investigated by a design of experiment methodology and analyzed by principal component analysis. Quadratic models were developed, and multicriteria optimization was carried out by the use of genetic algorithm. At the time of abstract submission, verification spray-drying trials are ongoing. To perform experiments, milk from dairy farm was collected, skimmed, froze and spray-dried at different air pressure (between 1 and 3 bars) and outlet temperature (between 75 and 95 °C). Dry matter, minerals content and proteins content were determined by standard method. Solubility index, absorption index and hygroscopicity were determined by method found in literature. Particle size distribution were obtained by laser diffraction granulometry. Location of the powder color in the Cielab color space and water activity were characterized by a colorimeter and an aw-value meter, respectively. Flow properties were characterized with FT4 powder rheometer; in particular, compressibility and shearing test were performed. Air pressure and outlet temperature are key factors that directly impact the drying kinetics and powder characteristics during spray-drying process. It was shown that the air pressure affects the particle size distribution by impacting the size of droplet exiting the nozzle. Moreover, small particles lead to more cohesive powder and less saturated color of powders. Higher outlet temperature results in lower moisture level particles which are less sticky and can explain a spray-drying yield increase and the higher cohesiveness; it also leads to particle with low water activity because of the intense evaporation rate. However, it induces a high hygroscopicity, thus, powders tend to get wet rapidly if they are not well stored. On the other hand, high temperature provokes a decrease of native serum proteins, which is positively correlated to gelation properties (gel point and firmness). Partial denaturation of serum proteins can improve functional properties of powder. The control of air pressure and outlet temperature during the spray-drying process significantly affects the physicochemical and functional properties of powder. This study permitted to better understand the links between physicochemical and functional properties of powder to identify correlations between air pressure and outlet temperature. Therefore, mathematical models have been developed, and the use of genetic algorithm will allow the optimization of powder functionalities.

Keywords: dairy powders, spray-drying, powders functionalities, design of experiment

Procedia PDF Downloads 63

Authors: Leyla Sattarzadeh, Shahin Fathi Molk Kian, Maghsoud Peeri, Mohammadali Azarbaijani, Hasan Matin Homaee

Abstract:

Inflammation by various mechanisms may cause atherosclerosis. Systemic circulating inflammatory markers such as C-reactive protein (CRP), pro-inflammatory cytokines such as Interleukin-6 (IL-6), vascular inflammatory markers as adhesion molecules like Intracellular Adhesion Molecule-1 (ICAM-1) and Vascular Cell Adhesion Molecule-1 (VCAM-1) are the predictors of cardiovascular diseases. Regarding the conflicting results about the effect of different patterns of resistance exercise training on these inflammatory markers, present study aimed to examine the effect of different patterns of eight week resistance exercise training on CRP, IL-6, ICAM-1 and VCAM-1 levels in healthy untrained women. 56 healthy volunteered untrained female university students (aged: 21 ± 3 yr., Body Mass Index: 21.5 ± 3.5 kg/m²) were selected purposefully and divided into four groups. At the end of training protocol and after subject drop during the protocol, upper body exercise training (n=11), lower body (n=12) and whole body resistance exercise training group (n=11) completed the eight weeks of training period although the control group (n=7) did anything. Blood samples gathered pre and post-experimental period and CRP, IL-6, ICAM-1 and VCAM-1 levels were evaluated using special laboratory kits, then the difference of pre and post values of each indices analyzed using one-way analysis of variance (α < 0.05). The results of one way ANOVA for difference of pre and post values of CRP, ICAM-1 and VCAM-1 showed no significant changes due to the exercise training, but there were significant differences between groups about IL-6. Tukey post- hoc test indicated that there is significant difference between the differences of pre and post values of IL-6 between lower body exercise training group and control group, and eight weeks of lower body exercise training lead to significant changes in IL-6 values. There were no changes in anthropometric indices. The findings show that the different patterns of upper, lower and whole body exercise training by involving the different amounts of muscles altered the IL-6 values in lower body exercise training group probably because of engaging the bigger amount of muscles, but showed any significant changes about CRP, ICAM-1 and VCAM-1 probably due to intensity and duration of exercise or the lower levels of these markers at baseline of healthy people.

Keywords: resistance training, C-reactive protein, interleukin-6, intracellular adhesion molecule-1, vascular cell adhesion molecule-1

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Authors: Wenjun Zhang, Yunqing Du, Steven W. Cranford, Ming L. Wang

Abstract:

The beginning of 21st century has witnessed new advancements in the design and use of new materials for biosensing applications, from nano to macro, protein to tissue. Traditional analytical methods lack a complete toolset to describe the complexities introduced by living systems, pathological relations, discrete hierarchical materials, cross-phase interactions, and structure-property dependencies. Materiomics – via systematic molecular dynamics (MD) simulation – can provide structure-process-property relations by using a materials science approach linking mechanisms across scales and enables oriented biosensor design. With this approach, DNA biosensors can be utilized to detect disease biomarkers present in individuals’ breath such as acetone for diabetes. Our wireless sensor array based on single-stranded DNA (ssDNA)-decorated single-walled carbon nanotubes (SWNT) has successfully detected trace amount of various chemicals in vapor differentiated by pattern recognition. Here, we present how MD simulation can revolutionize the way of design and screening of DNA aptamers for targeting biomarkers related to oral diseases and oral health monitoring. It demonstrates great potential to be utilized to build a library of DNDA sequences for reliable detection of several biomarkers of one specific disease, and as well provides a new methodology of creating, designing, and applying of biosensors.

Keywords: biosensor, DNA, biomarker, molecular dynamics simulation

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Authors: Eitan Yefenof

Abstract:

Glucocorticoid (GC) hormones, e.g. Dexamethasone and Prednisone, are widely used in the therapy of leukemia and lymphoma owing to their apoptogenic effect on lymphoid cells. However, the emergence of GC resistant cells during therapy is a major cause for treatment failure, urging the need for novel strategies that maintain leukemia sensitivity to the pro-apoptotic activity of GCs. GCs act by binding to the GC receptor (GR), which, in its inactive state, is sequestered in the cytosol by a multi-subunit complex of heat shock proteins. Upon ligand binding, the complex dissociates, allowing GR activation and translocation to the nucleus, where it regulates transcription of multiple genes. We demonstrated that in addition to gene expression, GR also regulates microRNA (miR) expression. Deep-sequencing analysis revealed 14 miRs that are regulated in GC-sensitive but resistant leukemias upon treatment with GC. GC up-regulates miR-103, miR-15~16 and miR-30e/d, while down-regulates miR-17, mir-18a, miR-19a, miR-19b, miR-20a and miR-92a (members of the miR-17∼92a multi-cistron). Upon transfection, miR-103 confers GC apoptotic sensitivity to otherwise GC-resistant cell. Furthermore, knocking down miR-103 expression reduces the GC apoptotic response of sensitive cells. miR-103 abrogates c-Myc expression, an oncogenic transcription factor which is deregulated in many cancers. In addition, miR-103 up-regulates Bim, a pro-apoptotic protein crucial for GC-induced death. Activated glycogen synthase kinase 3 (GSK3) is also crucial for GC-induced apoptosis. GSK3 is active in GC-sensitive but not in GC-resistant cells. We found that GSK3 associates with the GR multi-subunit complex. Upon GC exposure, it dissociates from the GR and interacts with Bim to enable activation of the mitochondrial apoptosis pathway. miR-103 mediated c-Myc ablation is followed by down-regulation of the multi-cistron miR-17~92a, in particular miR-18a and miR-20a. miR-18a targets GR for degradation whereas miR-20a targets Bim degradation. Hence, miR-103 acts, in concert with Bim and GR, as a "tumor suppressor" that leads to reduced proliferation, cell-cycle arrest and cell death. We suggest that miR-103 can provide a diagnostic tool that predicts the sensitivity of leukemia to GC based therapy. Furthermore, exosomal delivery of miR-103 or up-regulation of the endogenous miR-103 could confer apoptotic sensitivity to resistant cells at the outset, thus becoming a useful therapeutic tool combined with GCs.

Keywords: apoptosis, leukemia, micro-RNA, steroids

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Authors: Sepideh Hassanpour Khodaei

Abstract:

Background/Objectives: The purpose of this study is to evaluate the effect of mummy substances on two issues of proliferation and production of matrix protein synthesis in wound healing. Methods: The methodology used for this aim involves isolating mesenchymal stem cells and human fibroblasts procured at Pastor Institute, Iran. The cells were treated with mummy substances separately and co-cultured between ASCs and WJSCs, and fibroblasts. Proliferation was assessed by Ki67 method in monolayer conditions. Synthesis of components of extracellular matrix (ECM) such as collagen type I, type III, and fibronectin 1 (FN1) was determined by qPCR. Results: The effects of adipocyte stem cells (ASCs), Wharton Jelly Stem Cells (WJSCs), and Mummy material on fibroblast proliferation and migration were evaluated. The present finding underlined the importance of Mummy material, ASCs, and WJSCs in the proliferation and migration of fibroblast cells. Furthermore, the expression of collagen I, III, and FN1 was increased in the presence of the above material and cells. Conclusion: This study presented an effective in vitro method for the healing process. Hence, the prospect of utilizing Mummy material and stem cell-based therapies in wound healing as a therapeutic approach is promising.

Keywords: mummy material, wound healing, adipose tissue, Wharton’s jelly

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Authors: Olga V. Chervyakova, Elmira T. Tailakova, Vitaliy M. Strochkov, Kulyaisan T. Sultankulova, Nurlan T. Sandybayev, Lev G. Nemchinov, Rosemarie W. Hammond

Abstract:

Sheep pox is a highly contagious infection that OIE regards to be one of the most dangerous animal diseases. It causes enormous economic losses because of death and slaughter of infected animals, lower productivity, cost of veterinary and sanitary as well as quarantine measures. To control spread of sheep pox infection the attenuated vaccines are widely used in the Republic of Kazakhstan and other Former Soviet Union countries. In spite of high efficiency of live vaccines, the possible presence of the residual virulence, potential genetic instability restricts their use in disease-free areas that leads to necessity to exploit new approaches in vaccine development involving recombinant DNA technology. Vaccines on the basis of recombinant proteins are the newest generation of prophylactic preparations. The main advantage of these vaccines is their low reactogenicity and this fact makes them widely used in medical and veterinary practice for vaccination of humans and farm animals. The objective of the study is to produce recombinant immunogenic proteins for development of the high-performance means for sheep pox prophylaxis. The SPV proteins were chosen for their homology with the known immunogenic vaccinia virus proteins. Assay of nucleotide and amino acid sequences of the target SPV protein genes. It has been shown that four proteins SPPV060 (ortholog L1), SPPV074 (ortholog H3), SPPV122 (ortholog A33) and SPPV141 (ortholog B5) possess transmembrane domains at N- or C-terminus while in amino acid sequences of SPPV095 (ortholog А 4) and SPPV117 (ortholog А 27) proteins these domains were absent. On the basis of these findings the primers were constructed. Target genes were amplified and subsequently cloned into the expression vector рЕТ26b(+) or рЕТ28b(+). Six constructions (pSPPV060ΔТМ, pSPPV074ΔТМ, pSPPV095, pSPPV117, pSPPV122ΔТМ and pSPPV141ΔТМ) were obtained for expression of the SPV genes under control of T7 promoter in Escherichia coli. To purify and detect recombinant proteins the amino acid sequences were modified by adding six histidine molecules at C-terminus. Induction of gene expression by IPTG was resulted in production of the proteins with molecular weights corresponding to the estimated values for SPPV060, SPPV074, SPPV095, SPPV117, SPPV122 and SPPV141, i.e. 22, 30, 20, 19, 17 and 22 kDa respectively. Optimal protocol of expression for each gene that ensures high yield of the recombinant protein was identified. Assay of cellular lysates by western blotting confirmed expression of the target proteins. Recombinant proteins bind specifically with antibodies to polyhistidine. Moreover all produced proteins are specifically recognized by the serum from experimentally SPV-infected sheep. The recombinant proteins SPPV060, SPPV074, SPPV117, SPPV122 and SPPV141 were also shown to induce formation of antibodies with virus-neutralizing activity. The results of the research will help to develop a new-generation high-performance means for specific sheep pox prophylaxis that is one of key moments in animal health protection. The research was conducted under the International project ISTC # K-1704 “Development of methods to construct recombinant prophylactic means for sheep pox with use of transgenic plants” and under the Grant Project RK MES G.2015/0115RK01983 "Recombinant vaccine for sheep pox prophylaxis".

Keywords: prophylactic preparation, recombinant protein, sheep pox virus, subunit vaccine

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Authors: Sarwan Dhir, Suma Basak, Dipika Parajulee

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Alfalfa is renowned for its nutritional and biopharmaceutical value as a perennial forage legume. However, establishing a rapid plant regeneration protocol using somatic embryogenesis and efficient transformation frequency are the crucial prerequisites for gene editing in alfalfa. This study was undertaken to establish and improve the protocol for somatic embryogenesis and subsequent plant regeneration. The experiments were conducted in response to natural sensitivity using various antibiotics such as cefotaxime, carbenicillin, gentamycin, hygromycin, and kanamycin. Using 3-week-old leaf tissue, somatic embryogenesis was initiated on Gamborg’s B5 basal (B5H) medium supplemented with 3% maltose, 0.9µM Kinetin, and 4.5µM 2,4-D. Embryogenic callus (EC) obtained from the B5H medium exhibited a high rate of somatic embryo formation (97.9%) after 3 weeks when the cultures were placed in the dark. Different developmental stages of somatic embryos and cotyledonary stages were then transferred to Murashige and Skoog’s (MS) basal medium under light, resulting in a 94% regeneration rate of plantlets. Our results indicate that leaf segments can grow (tolerate) up to 450 mg/L of cefotaxime and 400 mg/L of carbenicillin in the culture medium. However, the survival threshold for hygromycin at 12.5 mg/L, kanamycin at 250 mg/L, gentamycin at 50 mg/L, and timentin (300 mg/L). The experiment to improve the protocol for achieving efficient transient gene expression in alfalfa through genetic transformation with the Agrobacterium tumefaciens pCAMBIA1304 vector was also conducted. The vector contains two reporter genes such as β-glucuronidase (GUS) and green fluorescent protein (GFP), along with a selectable hygromycin B phosphotransferase gene (HPT), all driven under the CaMV 35s promoter. Various transformation parameters were optimized using 3-week-old in vitro-grown plantlets. The different parameters such as types of explant, leaf ages, preculture days, segment sizes, wounding types, bacterial concentrations, infection periods, co-cultivation periods, different concentrations of acetosyringone, silver nitrate, and calcium chloride were optimized for transient gene expression. The transient gene expression was confirmed via histochemical GUS and GFP visualization under fluorescent microscopy. The data were analyzed based on the semi-quantitative observation of the percentage and number of blue GUS spots on different days of agro-infection. The highest percentage of GUS positivity (76.2%) was observed in 3-week-old leaf segments wounded using a scalpel blade of 11 size- after 3 days of post-incubation at a bacterial concentration of 0.6, with 2 days of preculture, 30 min of bacterial-leaf segment co-cultivation, with the addition of 150 µM acetosyringone, 4 mM calcium chloride, and 75 µM silver nitrate. Our results suggest that various factors influence T-DNA delivery in the Agrobacterium-mediated transformation of alfalfa. The stable gene expression in the putative transgenic tissue was confirmed using PCR amplification of both marker genes, indicating that gene expression in explants was not solely due to Agrobacterium, but also from transformed cells. The improved protocol could be used for generating transgenic alfalfa plants using genome editing techniques such as CRISPR/Cas9.

Keywords: Medicago sativa l. (Alfalfa), agrobacterium tumefaciens, β-glucuronidase, green fluorescent protein, transient gene

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Authors: Moon Jung Kim, Guil Rhim

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The clinical sensitivity of the “VEUPLEXTM TBI assay”, a clinical trial medical device, in mild traumatic brain injury was 28.6% (95% CI, 19.7%-37.5%), and the clinical specificity was 94.0% (95% CI, 89.3%). -98.7%). In addition, when the results analyzed by marker were put together, the sensitivity was higher when interpreting the two tests together than the two tests, UCHL1 and GFAP alone. Additionally, when sensitivity and specificity were analyzed based on CT results for the mild traumatic brain injury patient group, the clinical sensitivity for 2 CT-positive cases was 50.0% (95% CI: 1.3%-98.7%), and 19 CT-negative cases. The clinical specificity for cases was 68.4% (95% CI: 43.5% - 87.4%). Since the low clinical sensitivity for the two CT-positive cases was not statistically significant due to the small number of samples analyzed, it was judged necessary to secure and analyze more samples in the future. Regarding the clinical specificity analysis results for 19 CT-negative cases, there were a large number of patients who were actually clinically diagnosed with mild traumatic brain injury but actually received a CT-negative result, and about 31.6% of them showed abnormal results on VEUPLEXTM TBI assay. Although traumatic brain injury could not be detected in 31.6% of the CT scans, the possibility of actually suffering a mild brain injury could not be ruled out, so it was judged that this could be confirmed through follow-up observation of the patient. In addition, among patients with mild traumatic brain injury, CT examinations were not performed in many cases because the symptoms were very mild, but among these patients, about 25% or more showed abnormal results in the VEUPLEXTM TBI assay. In fact, no damage is observed with the naked eye immediately after traumatic brain injury, and traumatic brain injury is not observed even on CT. But in some cases, brain hemorrhage may occur (delayed cerebral hemorrhage) after a certain period of time, so the patients who did show abnormal results on VEUPLEXTM TBI assay should be followed up for the delayed cerebral hemorrhage. In conclusion, it was judged that it was difficult to judge mild traumatic brain injury with the VEUPLEXTM TBI assay only through clinical findings without CT results, that is, based on the GCS value. Even in the case of CT, it does not detect all mild traumatic brain injury, so it is difficult to necessarily judge that there is no traumatic brain injury, even if there is no evidence of traumatic brain injury in CT. And in the long term, more patients should be included to evaluate the usefulness of the VEUPLEXTM TBI assay in the detection of microscopic traumatic brain injuries without using CT.

Keywords: brain injury, traumatic brain injury, GFAP, UCHL1

Procedia PDF Downloads 98

Authors: Thanthrige Thiunuwan Priyathilaka, Don Anushka Sandaruwan Elvitigala, Bong-Soo Lim, Hyung-Bok Jeong, Jehee Lee

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Toll like receptors (TLRs) are a phylogeneticaly conserved family of pattern recognition receptors, which participates in the host immune responses against various pathogens and pathogen derived mitogen. TLR21, a non-mammalian type, is almost restricted to the fish species even though those can be identified rarely in avians and amphibians. Herein, this study was carried out to identify and characterize TLR21 from rock bream (Oplegnathus fasciatus) designated as RbTLR21, at transcriptional and genomic level. In this study, the full length cDNA and genomic sequence of RbTLR21 was identified using previously constructed cDNA sequence database and BAC library, respectively. Identified RbTLR21 sequence was characterized using several bioinformatics tools. The quantitative real time PCR (qPCR) experiment was conducted to determine tissue specific expressional distribution of RbTLR21. Further, transcriptional modulation of RbTLR21 upon the stimulation with Streptococcus iniae (S. iniae), rock bream iridovirus (RBIV) and Edwardsiella tarda (E. tarda) was analyzed in spleen tissues. The complete coding sequence of RbTLR21 was 2919 bp in length which can encode a protein consisting of 973 amino acid residues with molecular mass of 112 kDa and theoretical isoelectric point of 8.6. The anticipated protein sequence resembled a typical TLR domain architecture including C-terminal ectodomain with 16 leucine rich repeats, a transmembrane domain, cytoplasmic TIR domain and signal peptide with 23 amino acid residues. Moreover, protein folding pattern prediction of RbTLR21 exhibited well-structured and folded ectodomain, transmembrane domain and cytoplasmc TIR domain. According to the pair wise sequence analysis data, RbTLR21 showed closest homology with orange-spotted grouper (Epinephelus coioides) TLR21with 76.9% amino acid identity. Furthermore, our phylogenetic analysis revealed that RbTLR21 shows a close evolutionary relationship with its ortholog from Danio rerio. Genomic structure of RbTLR21 consisted of single exon similar to its ortholog of zebra fish. Sevaral putative transcription factor binding sites were also identified in 5ʹ flanking region of RbTLR21. The RBTLR 21 was ubiquitously expressed in all the tissues we tested. Relatively, high expression levels were found in spleen, liver and blood tissues. Upon induction with rock bream iridovirus, RbTLR21 expression was upregulated at the early phase of post induction period even though RbTLR21 expression level was fluctuated at the latter phase of post induction period. Post Edwardsiella tarda injection, RbTLR transcripts were upregulated throughout the experiment. Similarly, Streptococcus iniae induction exhibited significant upregulations of RbTLR21 mRNA expression in the spleen tissues. Collectively, our findings suggest that RbTLR21 is indeed a homolog of TLR21 family members and RbTLR21 may be involved in host immune responses against bacterial and DNA viral infections.

Keywords: rock bream, toll like receptor 21 (TLR21), pattern recognition receptor, genomic characterization

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Authors: Hamidreza Miri, Farahnaz Momtazi

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Wheat is one of the most important crops in Fars province, and because of water shortage, there is a great emphasis on its water use efficiency in the production field. A field experiment was conducted in 2021 and 2022 in order to evaluate wheat yield and its components in raised planting system in Arsanjan, Fars province. The experiment was conducted as a split plot with three irrigation treatments (irrigation equal to evapotranspiration, 80% of evapotranspiration irrigation (moderate drought stress), and 60% of evapotranspiration irrigation (severe drought stress)) as the main plot and three planting methods (conventional flat planting, 60 cm raised bed planting and 120 cm raised bed planting) as a subplot. The results indicated that drought stress significantly decreased traits such as plant height, grain yield, ear number, seed number, and biological yield while increasing seed protein. Raised bed planting significantly increased the traits in comparison with conventional flat planting. So that plating with a 120 cm raised bed increased grain yield by 22.1% and 25.9% in the first and second years, respectively. This increase was 17% for biological, 75 for ear number, and 21% for seed number. Planting in raised bed system reduced the adverse effect of drought stress on wheat traits. In conclusion, based on the observed results planting in raised bed system can be adopted as an appropriate planting pattern for improving yield and water productivity in experimental regions and similar climates.

Keywords: wheat, raised bed planting, drought stress, yield, water use

Procedia PDF Downloads 62

Authors: Girgis Younan, Ikram Eltayeb

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Geigeria alata belongs to the family Asteraceae, is an effective plant traditionally used in Sudan as a therapy for hepatic disease and as an antiepileptic, antispasmodic and to treat cough and intestinal complaints.The liver is responsible for many critical functions within the body and any liver disease or injury will result in the loss of those functions leading to significant damage in the body. Liver diseases cause increase in liver enzymes (AST, ALP ALT) and total bilirubin and a decrease in total blood protein level. The objective of this study is to investigate the hepato-protective activity of Geigeria alata leaves ethanolic extract. The plant leaves were extracted using 96% ethanol using Soxhlet apparatus. The hepatoprotective effect was determined using 25 wistar rats, the rats was divided to 5 groups, each group contain 5 rats: [Normal control group] receiving purified water, liver damage was induced in wistar rats by administering a 1:1 (v/v) mixture of CCl4 (1.25 ml/kg) and olive oil once at day four of the experiment [negative control group]. Two doses of extract [400mg/kg and 200mg/kg] was applied daily for 7 days, and standard drug Silymarin (200 mg/kg) were administered daily for 7 days to CCl4-treated rats. The degree of hepato-protective activity was evaluated by determining the hepatic marker enzymes AST, ALP, ALT, total Bilirubin and total proteins (TP). Results have shown that, the extract of G.alata leaves reduced the level of liver enzymes ALT, AST, ALP, total bilirubin and increased the level of total proteins. Since the levels of liver enzymes; bilirubin and total protein are considered as markers of liver function, the extract has proven to reduce the detrimental effects of liver toxicity induced using CCl4. The hepato-protective effect of extract on liver was found to be dose dependent, where the 400mg/kg dose of the extract exhibited higher activity than 200mg/kg dose. In addition, the effect of the higher dose (400mg/kg) of the extract was found to be higher than Silymarin standard drug. The result concludes that, G.alata leaves extract was found to exhibit profound hepato-protective activity, which justifies the traditional use of the plant for the treatment of hepatic diseases.

Keywords: alata, extract, geigeria, hepatoprotective

Procedia PDF Downloads 232

Authors: L. Gómez-Limia, I. Franco, T. Blanco, S. Martínez

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European eels (Anguilla anguilla) belong to Anguilliformes order and Anguillidae family. They are generally classified as warm-water fish. Eels have a great commercial value in Europe and Asian countries. Eels can reach high weights, although their commercial size is relatively low in some countries. The capture of larger eels would facilitate the recovery of the species, as well as having a greater number of either glass eels or elvers for aquaculture. In the last years, the demand and the price of eels have increased significantly. However, European eel is considered critically endangered by the International Union for the Conservation of Nature (IUCN) Red List. The biochemical composition of fishes is an important aspect of quality and affects the nutritional value and consumption quality of fish. In addition, knowing this composition can help predict an individual’s condition for their recovery. Fish is known to be important source of protein rich in essential amino acids. However, there is very little information about changes in amino acids composition of European eels with increase in size. The aim of this study was to evaluate the effect of two different weight categories on the amino acids content in muscle tissue of wild European eels. European eels were caught in River Ulla (Galicia, NW Spain), during winter. The eels were slaughtered in ice water immersion. Then, they were purchased and transferred to the laboratory. The eels were subdivided into two groups, according to the weight. The samples were kept frozen (-20 °C) until their analysis. Frozen eels were defrosted and the white muscle between the head and the anal hole. was extracted, in order to obtain amino acids composition. Thirty eels for each group were used. Liquid chromatography was used for separation and quantification of amino a cids. The results conclude that the eels are rich in glutamic acid, leucine, lysine, threonine, valine, isoleucine and phenylalanine. The analysis showed that there are significant differences (p < 0.05) among the eels with different sizes. Histidine, threonine, lysine, hydroxyproline, serine, glycine, arginine, alanine and proline were higher in small eels. European eels muscle presents between 45 and 46% of essential amino acids in the total amino acids. European eels have a well-balanced and high quality protein source in the respect of E/NE ratio. However, eels with higher weight showed a better ratio of essential and non-essential amino acid.

Keywords: European eels, amino acids, HPLC, body size

Procedia PDF Downloads 101

Authors: A. Zonouzi, M. Auli, M. Javanmard Dakheli, M. A. Hejazi

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Microalgae are tiny photosynthetic plants. Nowadays, microalgae are being used as nutrient-dense foods and sources of fine chemicals. They have significant amounts of lipid, carotenoids, vitamins, protein, minerals, chlorophyll, and pigments. Oil extraction from algae is a hotly debated topic currently because introducing an efficient method could decrease the process cost. This can determine the sustainability of algae-based foods. Scientific research works show that solvent extraction using chloroform/methanol (2:1) mixture is one of the efficient methods for oil extraction from algal cells, but both methanol and chloroform are toxic solvents, and therefore, the extracted oil will not be suitable for food application. In this paper, the effect of two food grade solvents (hexane and hexane/ isopropanol) on oil extraction yield from microalgae Dunaliella sp. was investigated and the results were compared with chloroform/methanol (2:1) extraction yield. It was observed that the oil extraction yield using hexane, hexane/isopropanol (3:2) and chloroform/methanol (2:1) mixture were 5.4, 13.93, and 17.5 (% w/w, dry basis), respectively. The fatty acid profile derived from GC illustrated that the palmitic (36.62%), oleic (18.62%), and stearic acids (19.08%) form the main portion of fatty acid composition of microalgae Dunalliela sp. oil. It was concluded that, the addition of isopropanol as polar solvent could increase the extraction yield significantly. Isopropanol solves cell wall phospholipids and enhances the release of intercellular lipids, which improves accessing of hexane to fatty acids.

Keywords: fatty acid profile‎, microalgae‎, oil extraction‎, polar solvent‎

Procedia PDF Downloads 375

Authors: Alebiosu Olugbenga Shadrak, Victor Victoria

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Honey bees exhibit preferences in their foraging behaviour on pollen and nectar for food and honey production, respectively. Melissopalynology is the study of pollen in honey and other honey products. Several work have been conducted on the palynological studies of honeys from the southern parts of Nigeria but with relatively scant records from the Northern region of the country. This present study aimed at revealing the favourably visited plants by honey bees, Apis melifera var. adansonii, at some apiaries in Northern Nigeria, as well as determining the quality of honeys produced. Honeys were harvested and collected from four apiaries of the region, namely: Sarkin Dawa missionary bee farm, Taraba State; Eleeshuwa Bee Farm, Keffi, Nassarawa State, Bulus Beekeeper Apiaries, Kagarko, Kaduna State and Mai Gwava Bee Farm, Kano State. These honeys were acetolysed for palynological microscopic analysis and subjected to standard treatment methods for the determination of their proximate composition and sugar profiling. Fresh anthers of two dominantly represented plants in the honeys were then collected for the quantification of their pollen protein contents, using the micro-kjeldhal procedure. A total of 30 pollen types were identified in the four honeys, and some of them were common to the honeys. A classification method for expressing pollen frequency class was employed: Senna cf. siamea, Terminalia cf. catappa, Mangifera indica, Parinari curatelifolia, Vitellaria paradoxa, Elaeis guineensis, Parkia biglobosa, Phyllantus muellerianus and Berlina Grandiflora, as “Frequent” (16-45%); while the others are either Rare (3-15%) or Sporadic (less than 3 %). Pollen protein levels of the two abundantly represented plants, Senna siamea (15.90mg/ml) and Terminalia catappa (17.33mg/ml) were found to be considerably lower. The biochemical analyses revealed varying amounts of proximate composition, non-reducing sugar and total sugar levels in the honeys. The results of this study indicate that pollen and nectar of the “Frequent” plants were preferentially foraged by honeybees in the apiaries. The estimated pollen protein contents of Senna same and Terminalia catappa were considerably lower and not likely to have influenced their favourable visitation by honeybees. However, a relatively higher representation of Senna cf. siamea in the pollen spectrum might have resulted from its characteristic brightly coloured and well scented flowers, aiding greater entomophily. Terminalia catappa, Mangifera indica, Elaeis guineensis, Vitellaria paradoxa, and Parkia biglobosa are typical food crops; hence they probably attracted the honeybees owing to the rich nutritional values of their fruits and seeds. Another possible reason for a greater entomophily of the favourably visited plants are certain nutritional constituents of their pollen and nectar, which were not investigated in this study. The nutritional composition of the honeys was observed to fall within the safe limits of international norms, as prescribed by Codex Alimentarius Commission, thus they are good honeys for human consumption. It is therefore imperative to adopt strategic conservation steps in ensuring that these favourably visited plants are protected from indiscriminate anthropogenic activities and also encourage apiarists in the country to establish their bee farms more proximally to the plants for optimal honey yield.

Keywords: honeybees, melissopalynology, preferentially foraged, nutritional, bee farms, proximally

Procedia PDF Downloads 276

Authors: William Kelly, Sorelle Veigne, Xianhua Li, Zuyi Huang, Shyamsundar Subramanian, Eugene Schaefer

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The ambr15™ bioreactor is a single-use microbioreactor for cell line development and process optimization. The ambr system offers fully automatic liquid handling with the possibility of fed-batch operation and automatic control of pH and oxygen delivery. With operating conditions for large scale biopharmaceutical production properly scaled down, micro bioreactors such as the ambr15™ can potentially be used to predict the effect of process changes such as modified media or different cell lines. In this study, gassing rates and dilution rates were varied for a semi-continuous cell culture system in the ambr15™ bioreactor. The corresponding changes to metabolite production and consumption, as well as cell growth rate and therapeutic protein production were measured. Conditions were identified in the ambr15™ bioreactor that produced metabolic shifts and specific metabolic and protein production rates also seen in the corresponding larger (5 liter) scale perfusion process. A Dynamic Flux Balance model was employed to understand and predict the metabolic changes observed. The DFB model-predicted trends observed experimentally, including lower specific glucose consumption when CO₂ was maintained at higher levels (i.e. 100 mm Hg) in the broth. A Computational Fluid Dynamic (CFD) model of the ambr15™ was also developed, to understand transfer of O₂ and CO₂ to the liquid. This CFD model predicted gas-liquid flow in the bioreactor using the ANSYS software. The two-phase flow equations were solved via an Eulerian method, with population balance equations tracking the size of the gas bubbles resulting from breakage and coalescence. Reasonable results were obtained in that the Carbon Dioxide mass transfer coefficient (kLa) and the air hold up increased with higher gas flow rate. Volume-averaged kLa values at 500 RPM increased as the gas flow rate was doubled and matched experimentally determined values. These results form a solid basis for optimizing the ambr15™, using both CFD and FBA modelling approaches together, for use in microscale simulations of larger scale cell culture processes.

Keywords: cell culture, computational fluid dynamics, dynamic flux balance analysis, microbioreactor

Procedia PDF Downloads 280

Authors: S. Stankov, H. Fidan, N. Petkova, M. Stoyanova, Tz. Radoukova, A. Stoyanova

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The aim of this study was to evaluate the chemical composition of ripe Juniperus excelsa M. Bieb. galbuli (female cones) collected from "Izgoraloto Gyune" Reserve in Krichim, Bulgaria. The moisture (36.88%), abs. weight 693.96 g/1000 pcs., and the ash content (10.57%) of ripe galbuli were determined. Lipid fraction (9.12%), cellulose (13.54%), protein (13.64%), and total carbohydrates (31.20%) were evaluated in the ripe galbuli. It was found that the ripe galbuli contained glucose (4.00%) and fructose (4.25%), but disaccharide sucrose was not identified. The main macro elements presented in the sample were K (8390.00 mg/kg), Ca (4596.00 g/kg), Mg (837.72 mg/kg), followed by Na (7.69 mg/kg); while the detected microelements consisted of Zn (8.51 mg/kg), Cu (4.66 mg/kg), Mn (3.65 mg/kg), Fe (3.26 mg/kg), Cr (3.00 mg/kg), Cd (< 0.1 mg/kg), and Pb (0.01 mg/kg).

Keywords: chemical composition, Juniperus excelsa M. Bieb, minerals, ripe galbuli

Procedia PDF Downloads 147

Authors: Valentin Nicorescu, Nicoleta C. Predescu, Camelia Papuc, Iuliana Gajaila, Carmen D. Petcu

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The oxidation of the fresh muscle oxymyoglobin (bright red colour) to metmyoglobin (brown colour) leads to discoloration of red meats. After slaughter, enzymatic systems involved in metmyoglobin reduction are continually depleted as time post-mortem progresses, thus the meat colour is affected. Phenolic compounds are able to scavenge reactive species involved in oxymyoglobin oxidation and to reduce metmyoglobin to oxymyoglobin. The aim of this study was to investigate the effect of polyphenols extracted from hawthorn fruits on the stability of oxymyoglobin and myofibrillar proteins in ground pork subject to refrigeration for 6 days. Hawthorn polyphenols (HP) were added in ground pork in 100, 200 and 300 ppm concentrations. Oxymyoglobin and metmyoglobin were evaluated spectrophotometrically at every 2 days and electrophoretic pattern of myofibrillar proteins was investigated at days 0 and 6 by Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE). For all meat samples, oxymyoglobin concentration significantly decreased during the first 4 days of refrigeration. After 6 days, the significant decrease of oxymyoglobin concentration continued only in the negative control samples. In samples treated with HP and butylated hydroxylanisole (BHA - positive control), oxymyoglobin concentration increased after 6 days of refrigeration, the highest levels complying with the following order: 100 ppm HP > 200 ppm HP > 300 ppm HP > 100 ppm BHA. The increase in metmyoglobin was coincidental with the decrease in oxymyoglobin; metmyoglobin concentration progressively increased during the first 4 days of refrigeration in all meat samples. After 6 days, in meat samples treated with HP and BHA, lower metmyoglobin concentrations were found (compared to day 4), respecting the following order: 100 ppm HP < 200 ppm HP < 300 ppm HP < 100 ppm BHA. These results showed that hawthorn polyphenols and BHA reduced metmyoglobin (MbFe3+) to oxymyoglobin (MbFe2+), and the strongest reducing character was recorded for 100 ppm HP. After 6 days of refrigeration, electrophoretic pattern of myofibrillar proteins showed minor changes compared to day 0, indicating that HP prevent protein degradation as well as synthetic antioxidant BHA. Also, HP did not induce cross-links in the myofibrillar proteins, to form protein aggregates, and no risk of reducing their ability to retain water was identified. The pattern of oxymyoglobin and metmyoglobin concentrations determined in this study showed that hawthorn polyphenols are able to reduce metmyoglobin to oxymyoglobin and to delay oxymyoglobin oxidation, especially when they are added to ground meat in concentration of 100 ppm. This work was carried out through Partnerships in priority areas Program – PN II, implemented with the support of MEN – UEFISCDI (Romania), project nr. 149/2014.

Keywords: Hawthorn polyphenols, metmyoglobin, oxymyoglobin, proteins stability

Procedia PDF Downloads 217

Authors: Pao-Huei Chen, Shu-Mei Lin, Yih-Ming Weng, Zer-Ran Yu, Be-Jen Wang

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Pancreatic α-amylase and intestinal α-glucosidase are the critical enzymes for the breakdown of complex carbohydrates into di- or mono-saccharide, which play an important role in modulating postprandial blood sugars. Angiotensin converting enzyme (ACE) converts inactive angiotensin-I into active angiotensin-II, which subsequently increase blood pressure through triggering vasoconstriction and aldosterone secretion. Thus, inhibition of carbohydrate-digestion enzymes and ACE will help the management of blood glucose and blood pressure, respectively. Studies showed Pleurotus citrinopileatus (PC), an edible mushroom and commonly cultured in oriental countries, exerted anticancer, immune improving, antioxidative, hypoglycemic and hypolipidemic effects. Previous studies also showed various molecular weights (MW) fractioned from extracts may affect biological activities due to varying contents of bioactive components. Thus, the objective of this study is to investigate the in vitro antioxidant, hypoglycemic and hypotenstive effects and distribution of active compounds of various MWs of cold water extract from P. citrinopileatus (CWEPC). CWEPC was fractioned into four various MW fractions, PC-I (＜1 kDa), PC-II (1-3.5 kDa), PC-III (3.5-10 kDa), and PC-IV (＞10 kDa), using an ultrafiltration system. The physiological activities, including antioxidant activities, the inhibition capabilities of pancreatic α-amylase, intestinal α-glucosidase, and hypertension-linked ACE, and the active components, including polysaccharides, protein, and phenolic contents, of CWEPC and four fractions were determined. The results showed that fractions with lower MW exerted a higher antioxidant activity (p<0.05), which was positively correlated to the levels of total phenols. In contrast, the inhibition effects on the activities of α-amylase, α-glucosidase, and ACE of PC-IV fraction were significantly higher than CWEPC and the other three low MW fractions (< 10 kDa), which was more related to protein contents. The inhibition capability of CWEPC and PC-IV on α-amylase activity was 1/13.4 to 1/2.7 relative to that of acarbose (positive control), respectively. However, the inhibitory ability of PC-IV on α-glucosidase (IC50 = 0.5 mg/mL) was significantly higher than acarbose (IC50 = 1.7 mg/mL). Kinetic data revealed that PC-IV fraction followed a non-competitive inhibition on α-glucosidase activity. In conclusion, the distribution of various bioactive components contribute to the functions of different MW fractions on oxidative stress prevention, and blood pressure and glucose modulation.

Keywords: α-Amylase, angiotensin converting enzyme, α-Glucosidase, Pleurotus citrinopileatus

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Authors: Ewelina Musielak, Agnieszka Feliczak-Guzik, Izabela Nowak

Abstract:

Based on the latest data, it is expected that the substances of therapeutic interest used will be as natural as possible. Therefore, active substances with the highest possible efficacy and low toxicity are sought. Among natural substances with therapeutic effects, those of plant origin stand out. Curcumin isolated from the Curcuma longa plant has proven to be particularly important from a medical point of view. Due to its ability to regulate many important transcription factors, cytokines, and protein kinases, curcumin has found use as an anti-inflammatory, antioxidant, antiproliferative, antiangiogenic, and anticancer agent. The unfavorable properties of curcumin, such as low solubility, poor bioavailability, and rapid degradation under neutral or alkaline pH conditions, limit its clinical application. These problems can be solved by combining curcumin with suitable carriers such as hierarchical zeolites. This is a new class of materials that exhibit several advantages. Hierarchical zeolites used as drug carriers enable delayed release of the active ingredient and promote drug transport to the desired tissues and organs. In addition, hierarchical zeolites play an important role in regulating micronutrient levels in the body and have been used successfully in cancer diagnosis and therapy. To apply curcumin to hierarchical zeolites synthesized from commercial FAU zeolite, solutions containing curcumin, carrier and acetone were prepared. The prepared mixtures were then stirred on a magnetic stirrer for 24 h at room temperature. The curcumin-filled hierarchical zeolites were drained into a glass funnel, where they were washed three times with acetone and distilled water, after which the obtained material was air-dried until completely dry. In addition, the effect of piperine addition to zeolite carrier containing a sufficient amount of curcumin was studied. The resulting products were weighed and the percentage of pure curcumin in the hierarchical zeolite was calculated. All the synthesized materials were characterized by several techniques: elemental analysis, transmission electron microscopy (TEM), Fourier transform infrared spectroscopy, Fourier transform infrared (FT-IR), N2 adsorption, and X-ray diffraction (XRD) and thermogravimetric analysis (TGA). The aim of the presented study was to improve the biological activity of curcumin by applying it to hierarchical zeolites based on FAU zeolite. The results showed that the loading efficiency of curcumin into hierarchical zeolites based on commercial FAU-type zeolite is enhanced by modifying the zeolite carrier itself. The hierarchical zeolites proved to be very good and efficient carriers of plant-derived active ingredients such as curcumin.

Keywords: carriers of active substances, curcumin, hierarchical zeolites, incorporation

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