Search results for: molecular differentiation

Authors: Ana M. Lara, Manuela Llano, Felipe Gaitán, Rosa H. Bustos, Ana Maria Perdomo-Arciniegas, Ximena Bonilla

Abstract:

Umbilical cord blood is used as a source of progenitor and stem cells for the regeneration of the hematopoietic and immune system to treat patients with different hematological or non-hematological diseases. This stem cell source represents an advantage over the use of bone marrow or mobilized peripheral blood because it has a lower incidence rate of graft-versus-host disease, probably due to fewer immunological compatibility restrictions. However, its low cellular dose limits its use in pediatric patients. This work proposes the standardization of a cell expansion technique to compensate for the dose of infused cells through the ex-vivo manipulation of hematopoietic progenitor cells from umbilical cord blood before transplantation. The expansion model is carried out through co-cultures with mesenchymal stem cells (MSC) from bone marrow (BM) and less explored fetal tissues such as Wharton's jelly (WJ) and umbilical cord blood (UCB). Initially, a master cell bank of primary mesenchymal stem cells isolated from different sources was established and characterized following International Society of Cell Therapies (ISCT) indications. Additionally, we assessed the effect of a short 25 Gy cycle of gamma irradiation on cell cycle arrest of mesenchymal cells over the support capacity for the expansion of hematopoietic stem cells from umbilical cord blood was evaluated. The results show that co-cultures with MSC from WJ and UCB allow the cellular dose of HSPC to be maximized between 5 and 16 times having a similar support capacity as BM. In addition, was evaluated the hematopoietic stem progenitor cell's HSPC functionality through the evaluation of migration capacity, their differentiation capacity during culture time by flow cytometry to evaluate the expression of membrane markers associated with lineage-committed progenitors, their clonogenic potential, and the evaluation of secretome profile in the expansion process was evaluated. So far, the treatment with gamma irradiation maintains the hematopoietic support capacity of mesenchymal stem cells from the three sources studied compared to treatments without irradiation, favoring the use of fetal tissues that are generally waste to obtain mesenchymal cell lines for ex-vivo expansion systems. With the results obtained, a standardized protocol that will contribute to the development of ex-vivo expansion with MSC on a larger scale will be achieved, enabling its clinical use and expanding its application in adults.

Keywords: ex-vivo expansion, hematopoietic stem cells, hematopoietic stem cell transplantation, mesenchymal stem cells, umbilical cord blood

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Authors: Dilani Wimalasiri, Robert Brkljaca, Sylvia Urban, Terrence Piva, Tien Huynh

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Momordica cochinchinensis (Cucurbitaceae) is used as food and traditional medicine in South East Asia and is commonly known as Red Gac. The fruit aril consists 70 times higher lycopene and 10 times higher β-carotene than all known fruits and vegetables. Despite its nutritional value there is little information available on its genetic variation and its influence on nutritional value. In this study; genetic and nutritional variation (lycopene and β-carotene) was investigated among 47 M. cochinchinensis samples collected from Australia, Thailand and Vietnam using molecular markers (RAPD and ISSR) and HPLC, respectively. UPGMA based cluster analysis of genetic data grouped Northern and Central Vietnam samples together but were separated from Australia, Thailand and Southern Vietnam samples. The concentration of lycopene was significantly higher among the samples collected from Central Vietnam (p<0.05) and the concentration of β-carotene was significantly higher among the samples collected from Northern Vietnam (p<0.05) indicating the existence of best varieties. This study provides vital information in genetic diversity and facilitates the selection and breeding for nutritious M. cochinchinensis varieties.

Keywords: momordica cochinchinensis, lycopene, beta carotene, genetic diversity

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Authors: Amine Mezni, Merfat Algethami, Ali Aldalbahi, Arwa Alrooqi, Abel Santos, Dusan Losic, Sarah Alharthi, Tariq Altalhi

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In situ synthesis of carbon nanotubes featuring different diameters (10-200 nm), lengths (1 to 100 µm) and periodically nanostructured shape was performed in a custom designed chemical vapor deposition (CVD) system using nanoporous anodic alumina (NAA) under different conditions. The morphology of the resulting CNTs/NAA composites and free-standing CNTs were analyzed by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The results confirm that highly ordered arrays of CNTs with precise control of nanotube dimensions in the range 20-200 nm with tube length in the range < 1 µm to > 100 μm and with periodically shaped morphology can be fabricated using nanostructured NAA templates prepared by anodization. This technique allows us to obtain tubes open at one / both ends with a uniform diameter along the pore length without using any metal catalyst. Our finding suggests that this fabrication strategy for designing new CNTs membranes and structures can be significant for emerging applications as molecular separation/transport, optical biosensing, and drug delivery.

Keywords: carbon nanotubes, CVD approach, composites membrane, nanoporous anodic alumina

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Authors: Muhammad Asif Rasheed

Abstract:

Newcastle disease virus (NDV), an avian orthoavulavirus, is a causative agent of Newcastle disease named (NDV) and can cause even the epidemics when the disease is not treated. Previously several vaccines based on attenuated and inactivated viruses have been reported, which are rendered useless with the passage of time due to versatile changes in viral genome. Therefore, we aimed to develop an effective multi-epitope vaccine against the haemagglutinin neuraminidase (HN) protein of 26 NDV strains from Pakistan through a modern immunoinformatic approaches. As a result, a vaccine chimaera was constructed by combining T-cell and B-cell epitopes with the appropriate linkers and adjuvant. The designed vaccine was highly immunogenic, non-allergen, and antigenic; therefore, the potential 3D-structureof multi epitope vaccine was constructed, refined, and validated. A molecular docking study of a multiepitope vaccine candidate with the chicken Toll-like receptor-4 indicated successful binding. An In silico immunological simulation was used to evaluate the candidate vaccine's ability to elicit an effective immune response. According to the computational studies, the proposed multiepitope vaccine is physically stable and may induce immune responses, whichsuggested it a strong candidate against 26 Newcastle disease virus strains from Pakistan. A wet lab study is under process to confirm the results.

Keywords: epitopes, newcastle disease virus, paramyxovirus virus, vaccine

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Authors: Hammed Tanimowo Aiyelabegan, Oluchukwu Franklin Aladi, Mutiu Adewumi Alabi, Raliat Abimbola Aladodo, Emmanuel Oladipupo Ajani, Abdulganiyu Giwa, Esther Owolabi

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The study aimed to evaluate the inhibitory activities of ligands from Enantia chlorantha stem bark on α-amylase and α-glucosidase. In silico pharmacokinetic properties and docking scores were employed to analyse the inhibition using SwissADME and Autodock4.2, respectively. Results revealed that drug-likeness, pharmacokinetics and bioavailability radar of all the ligands except jatrorrhizine and acarbose falls within the radar according to the Lipinski rule of 5. The binding energies of the protein-ligand interactions also show that the ligand fits into the active site. The results obtained from this study show that the chemical constituents from Enantia chlorantha stem bark may bring about positive physiological changes in a patient suffering from diabetes mellitus. Further in vitro studies on diabetes cell lines and in vivo studies on the animal may validate these compounds for diabetes treatment. These phytoconstituents could help in the development of novel anti-diabetic molecules.

Keywords: diabetes mellitus, ?-amylase, ?-glucosidase, in silico, Enantia chlorantha stem bark

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Authors: Aarti Singh, Anees Ahmad

Abstract:

Carotenoids comprise a group of isoprenoid pigments. Carotenes, xanthophylls and their derivatives have been found to play an important role in all living beings through foods, neutraceuticals and pharmaceuticals. α-carotene, β-carotene and β-cryptoxanthin play a vital role in humans to provide vitamin A source for the growth, development and proper functioning of immune system and vision. They are very crucial for plants and humans as they protect from photooxidative damage and are excellent antioxidants quenching singlet molecular oxygen and peroxyl radicals. Diet including more intake of carotenoids results in reduced threat of various chronic diseases such as cancer (lung, breast, prostrate, colorectal and ovarian cancers) and coronary heart diseases. The blue light filtering efficiency of the carotenoids in liposomes have been reported to be maximum in lutein followed by zeaxanthin, β-carotene and lycopene. Lycopene plays a vital role for the protection from CVD. Lycopene in serum is directly related to reduced risk of osteoporosis in postmenopausal women. Carotenoids have major role in the treatment of skin disorders. There is need to identify and isolate novel carotenoids from diverse natural sources for human health benefits.

Keywords: antioxidants, carotenoids, neutraceuticals, osteoporosis, pharmaceuticals

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Authors: Elena Petersen, Inna Kornienko, Svetlana Guryeva, Sergey Simakov

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In vitro organoid cultivation requires the simultaneous provision of necessary vascularization and nutrients perfusion of cells during organoid development. However, many aspects of this problem are still unsolved. The functionality of vascular network intergrowth is limited during early stages of organoid development since a function of the vascular network initiated on final stages of in vitro organoid cultivation. Therefore, a microchannel network should be created in early stages of organoid cultivation in hydrogel matrix aimed to conduct and maintain minimally required the level of nutrients perfusion for all cells in the expanding organoid. The network configuration should be designed properly in order to exclude hypoxic and necrotic zones in expanding organoid at all stages of its cultivation. In vitro vascularization is currently the main issue within the field of tissue engineering. As perfusion and oxygen transport have direct effects on cell viability and differentiation, researchers are currently limited only to tissues of few millimeters in thickness. These limitations are imposed by mass transfer and are defined by the balance between the metabolic demand of the cellular components in the system and the size of the scaffold. Current approaches include growth factor delivery, channeled scaffolds, perfusion bioreactors, microfluidics, cell co-cultures, cell functionalization, modular assembly, and in vivo systems. These approaches may improve cell viability or generate capillary-like structures within a tissue construct. Thus, there is a fundamental disconnect between defining the metabolic needs of tissue through quantitative measurements of oxygen and nutrient diffusion and the potential ease of integration into host vasculature for future in vivo implantation. A model is proposed for growth prognosis of the organoid perfusion based on joint simulations of general nutrient diffusion, nutrient diffusion to the hydrogel matrix through the contact surfaces and microchannels walls, nutrient consumption by the cells of expanding organoid, including biomatrix contraction during tissue development, which is associated with changed consumption rate of growing organoid cells. The model allows computing effective microchannel network design giving minimally required the level of nutrients concentration in all parts of growing organoid. It can be used for preliminary planning of microchannel network design and simulations of nutrients supply rate depending on the stage of organoid development.

Keywords: 3D model, consumption model, diffusion, spheroid, tissue organoid

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Authors: E. Pretorius, A. M. Viljoen, M. van der Bank

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Introduction: The phytochemistry of Agathosma crenulata and A. betulina has been studied extensively, while their molecular analysis through DNA barcoding remains virtually unexplored. This technique can confirm the identity of plant species included in a herbal product, thereby ensuring the efficacy of the herbal product and the accuracy of its label. Materials and methods: Authentic Agathosma reference material of A. betulina (n=16) and A. crenulata (n=10) were obtained. Thirteen commercial products were purchased from various health shops around Johannesburg, South Africa, using the search term “Agathosma” or “Buchu.” The plastid regions matK and ycf1 were used to barcode the Buchu products, and BRONX analysis confirmed the taxonomic identity of the samples. UPLC-MS analyses were also performed. Results: Only (30/60) 60% of the traded samples tested from 13 suppliers contained A. betulina in their herbal products. Similar results were also obtained for the UPLC-MS analysis. Conclusion: In this study, we demonstrate the application of DNA barcoding in combination with phytochemical analysis to authenticate herbal products claiming to contain Agathosma plants as an ingredient in their products. This supports manufacturing efforts to ensure that herbal products that are safe for the consumer.

Keywords: Buchu, substitution, barcoding, BRONX algorithm, matK, ycf1, UPLC-MS

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Authors: Khabat Barkhordari, Ali Mohammad Arabzadeh

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Hepatitis delta virus (HDV) is a RNA virus that needs the function of hepatitis B virus (HBV) for its propagation and assembly. Infection by HDV can occur spontaneously with HBV infection and cause acute hepatitis or develop as secondary infection in HBV suffering patients. Based on genome sequence analysis, HDV has several genotypes which show broad geographic and diverse clinical features. The aim of current study is determine the prevalence of hepatitis delta virus genotype in patients with positive HBsAg in Kerman province of Iran. This cross-sectional study a total of 400 patients with HBV infection attending the clinic center of Besat from 2012 to 2014 were included. We carried out ELISA to detect anti-HDV antibodies. Those testing positive were analyzed further for HDV-RNA and for genotyping using restriction fragment length polymorphism (RFLP) and RT-nested PCR- sequencing. Among 400 patients in this study, 67 cases (16.75 %) were containing anti-HDV antibody which we found HDV RNA in just 7 (1.75%) serum samples. Analysis of these 7 positive HDV showed that all of them have genotype I. According to current study the HDV prevalence in Kerman is higher than the reported prevalence of 6.6% for Iran as a whole and clade 1 (genotype 1) is the predominant clade of HDV in Kerman.

Keywords: genotyping, hepatitis delta virus, molecular epidemiology, Kerman, Iran

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Authors: Sanja O. Podunavac-Kuzmanović, Strahinja Z. Kovačević, Lidija R. Jevrić, Stela Jokić

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The complexes of transition metals with various organic ligands have been extensively studied as models of some important pharmaceutical molecules. It was found that biological properties of different substituted organic molecules are improved when they are complexed by different metals. Therefore, it is of great importance for the development of coordination chemistry to explore the assembly of functional organic ligands with metal ion and to investigate the relationship between the structure and property. In the present work, we have bioassayed the antibacterial potency of benzimidazoles and their metal salts (Cu or Zn) against yeast Sarcina lutea. In order to validate our in vitro study, we performed in silico studies using molecular docking software. The investigated compounds and their metal complexes (Cu, Zn) showed good to moderate inhibitory activity against Sarcina lutea. In silico docking studies of the synthesized compounds suggested that complexed benzimidazoles have a greater binding affinity and improved antibacterial activity in comparison with non-complexed ligands. These results are part of the CMST COST Action No. 1105 "Functional metal complexes that bind to biomolecules".

Keywords: organometallic complexes, benzimidazoles, chemometric design, Sarcina lutea

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Authors: Baharuddin Patandjengi, A. Pabborong, T. Kuswinanti

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Bacterial ring rot caused by a gram-positive Coryneform bacterium Corynebacterium michiganensis subsp. sepedonicus is an important disease on potato crops in the world. The disease still belongs to an A1 quarantine pathogen in Indonesia, although it was found in West Java since 2013. The objective of this study was to know the presence of bacterial ring rot in four potato district areas in South Sulawesi. Infected samples were collected from potato fields and storage warehouses in Enrekang, Gowa, Jeneponto and Bantaeng districts. Potato tuber samples were cut and observed their vasiculer vessels and the bacterial ooze was used for isolation on Nutrient Agar and Nutrient Broth–Yeast extract medium. Bacterial isolates were then morphologically and physiologically characterized. A patogenicity test on eggplant and molecular characterization using PCR with specific primer for Cms (50F and Cms 50 R) was revealed for further identification. The results showed that Cms has become widespread in four districts of South Sulawesi. The bacterial ringrot disease incidence in these districts was reached above 30 %. All of 14 bacterial isolates that identified before using standard methods of EPPO, showed DNA band in size of 224 bp in PCR test, which indicated positively belong to C. michiganensis subsp. sepedonicus.

Keywords: bacterial ring rot, clavibacter michiganensis pv. sepedonicus, PCR, potato

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Authors: Gizachew Belay Adugna

Abstract:

Perovskite solar cells (PSCs) shows rapid development as an emerging photovoltaic material; however, the fast device degradation due to the organic nature, mainly hole transporting material (HTM) and lack of robust and reliable upscaling process for photovoltaic module hindered its commercialization. Herein, HTM molecules with/without fluorine-substituted cyclopenta[2,1-b;3,4-b’]dithiophene derivatives (HYC-oF, HYC-mF, and HYC-H) were developed for PSCs application. The fluorinated HTM molecules exhibited better hole mobility and overall charge extraction in the devices mainly due to strong molecular interaction and packing in the film. Thus, the highest power conversion efficiency (PCE) of 19.64% with improved long stability was achieved for PSCs based on HYC-oF HTM. Moreover, the fluorinated HYC-oF demonstrated excellent film processability in a larger-area substrate (10 cm×10 cm) prepared sequentially with the absorption perovskite underlayer via a scalable bar coating process in ambient air and owned a higher PCE of 18.49% compared to the conventional spiro-OMeTAD (17.51%). The result demonstrates a facile development of HTM towards stable and efficient PSCs for future industrial-scale PV modules.

Keywords: perovskite solar cells, upscaling film coating, power conversion efficiency, solution processing

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Authors: Lidija R. Jevrić, Sanja O. Podunavac-Kuzmanović, Strahinja Z. Kovačević

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In this paper, quantitative structure-retention relationships (QSRR) analysis was applied in order to correlate in silico biological and lipophilicity molecular descriptors with retention values for the set of selected s-triazine herbicides. In silico generated biological and lipophilicity descriptors were discriminated using generalized pair correlation method (GPCM). According to this method, the significant difference between independent variables can be noticed regardless almost equal correlation with dependent variable. Using established multiple linear regression (MLR) models some biological characteristics could be predicted. Established MLR models were evaluated statistically and the most suitable models were selected and ranked using sum of ranking differences (SRD) method. In this method, as reference values, average experimentally obtained values are used. Additionally, using SRD method, similarities among investigated s-triazine herbicides can be noticed. These analysis were conducted in order to characterize selected s-triazine herbicides for future investigations regarding their biodegradability. This study is financially supported by COST action TD1305.

Keywords: descriptors, generalized pair correlation method, pesticides, sum of ranking differences

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Authors: Wenxing Chen, Guangying Chen, Yin Lu, Shile Huang

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Cryptotanshinone (CPT), a fat-soluble tanshinone from Salvia miltiorrhiza Bunge, has been demonstrated to inhibit mTOR pathway, resulting in inhibition of cancer cell proliferation. However, the molecular mechanism how CPT acts on mTOR is unknown. Here, cancer cells expressing rapamycin-resistant mutant mTOR are also sensitive to CPT, while phosphorylation of AMPK and TSC2 was activated, suggesting that CPT inhibition of mTOR maybe due to activating upstream of mTOR, AMPK, but not directly binding to and inhibiting mTOR. Further results indicated that Compound C, inhibitor of AMPK, could partially reversed CPT inhibition effect on cancer cells, and dominant-negative AMPK in cancer cells conferred resistance to CPT inhibition of 4EBP1 and phosphorylation of S6K1, as well as sh-AMPK. Furthermore, compared with MEF cells with AMPK positive, MEF cells with AMPK knock out are less sensitive to CPT by the findings that 4E-BP1 and phosphorylation of S6K1 express comparatively much. Furthermore, downexpression of TSC2 slightly recovered expression of 4EBP1 and phosphorylation of S6K1, while co-immunoprecipitation of TSC2 did not affect expression of TSC1 by CPT. Collectively, the above-mentioned results suggest that CPT inhibited mTOR pathway mostly was due to activation of AMPK-TSC2 pathway rather than specific inhibition of mTOR and then induction of subsequent lethal cellular effect.

Keywords: cryptotanshinone, AMPK, TSC2, mTOR, cancer cells

Procedia PDF Downloads 485

Authors: Aarti Singh, Anees Ahmad

Abstract:

Carotenoids comprise a group of isoprenoid pigments. Carotenes, xanthophylls and their derivatives have been found to play an important role in all living beings through foods, neutraceuticals and pharmaceuticals. α-carotene, β-carotene and β-cryptoxanthin play a vital role in humans to provide vitamin A source for the growth, development and proper functioning of immune system and vision. They are very crucial for plants and humans as they protect from photooxidative damage and are excellent antioxidants quenching singlet molecular oxygen and peroxyl radicals. Diet including more intake of carotenoids results in reduced threat of various chronic diseases such as cancer (lung, breast, prostate, colorectal and ovarian cancers) and coronary heart diseases. The blue light filtering efficiency of the carotenoids in liposomes have been reported to be maximum in lutein followed by zeaxanthin, β-carotene and lycopene. Lycopene play a vital role for the protection from CVD. Lycopene in serum is directly related to reduced risk of osteoporosis in postmenopausal women. Carotenoids have the major role in the treatment of skin disorders. There is a need to identify and isolate novel carotenoids from diverse natural sources for human health benefits.

Keywords: antioxidants, carotenoids, neutraceuticals, osteoporosis, pharmaceuticals

Procedia PDF Downloads 360

Authors: Masoud Sheidaei, Fahimeh Koohdar

Abstract:

Date palms are economically important tree plants with high nutrition and medicinal values. More than 400 date palm cultivars are cultivated in many regions of Iran, but no report is available on landscape genetics and species distribution modeling of these trees from the country. Therefore, the present study provides a detailed insight into the genetic diversity and structure of date palm populations in Iran and investigates the effects of geographical and climatic variables on the structuring of genetic diversity in them. We used different computational methods in the study like, spatial principal components analysis (sPCA), redundancy analysis (RDA), latent factor mixed model (LFMM), and Maxent and Dismo models of species distribution modeling. We used a combination of different molecular markers for this study. The results showed that both global and local spatial features play an important role in the genetic structuring of date palms, and the genetic regions associated with local adaptation and climatic variables were identified. The effects of climatic change on the distribution of these taxa and the genetic regions adaptive to these changes will be discussed.

Keywords: adaptive genetic regions, genetic diversity, isolation by distance, populations divergence

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Authors: Amira Abdelrasoul, Huu Doan, Ali Lohi

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An efficient and well-planned ultrafiltration process is becoming a necessity for monetary returns in the industrial settings. The aim of the present study was to develop a mathematical model for an accurate prediction of ultrafiltration membrane fouling of latex effluent applied to homogeneous and heterogeneous membranes with uniform and non-uniform pore sizes, respectively. The models were also developed for an accurate prediction of power consumption that can handle the large-scale purposes. The model incorporated the fouling attachments as well as chemical and physical factors in membrane fouling for accurate prediction and scale-up application. Both Polycarbonate and Polysulfone flat membranes, with pore sizes of 0.05 µm and a molecular weight cut-off of 60,000, respectively, were used under a constant feed flow rate and a cross-flow mode in ultrafiltration of the simulated paint effluent. Furthermore, hydrophilic ultrafilic and hydrophobic PVDF membranes with MWCO of 100,000 were used to test the reliability of the models. Monodisperse particles of 50 nm and 100 nm in diameter, and a latex effluent with a wide range of particle size distributions were utilized to validate the models. The aggregation and the sphericity of the particles indicated a significant effect on membrane fouling.

Keywords: membrane fouling, mathematical modeling, power consumption, attachments, ultrafiltration

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Authors: Shiraz A. Markarian

Abstract:

Our systematic studies of diethylsulfoxide (DESO), the nearest homologue of dimethylsulfoxide (DMSO), reveal new physicochemical features. DESO has already received worthy biomedical applications: in some cases even are more pronounced compare with DMSO. The several important physicochemical characteristics of DESO including aqueous solutions have been verified and first reported: melting point of pure substance, density, dielectric relaxation data, vapor pressure and volumetric properties. Analysis of the complete vibrational spectra also leads to the conclusion that very strong interactions take place between DESO and water, even stronger than those between DMSO and water. The simultaneous existence of strong DESO-H₂O and DESO-DESO interactions suggest the coexistence of many types of structural molecular aggregates, the presence of which plays a significant role also in diluted water solutions. Our recent investigations have shown that aqueous solution of DESO could provide amorphous, glassy systems, thus avoiding ice crystallization, in a wide range of concentrations and even at very low cooling rates. The ability of DESO to act as an effective cryoprotectant on E. coli survival was also studied and compared with other commonly used cryoprotective agents. The results also confirm that DESO, more than DMSO, is able to penetrate living tissues without causing significant damage.

Keywords: diethylsulfoxide, dimethylsulfoxide, cryoprotectant, properties

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Authors: Zahra Solgi, Hossein Rassi

Abstract:

Colorectal cancer (CRC) is a multi-step disease, and chronic gastric infection with H. pylori could play a role in one or more of the steps in this pathogenic process. Polymorphisms in several miRNAs are considered to increase the risk for the development of CRC by controlling proliferation, apoptosis and H. pylori pathogenesis. Therefore, the aim of this study was to investigate miRNA146a rs2910164 polymorphism and Helicobacter pylori infection in CRC. A total of 65 patients with CRC were divided into 2 groups: 28 patients < 50 years of age and 37 patients ≥ 50 years of age. DNA was extracted from all samples by a standard method and H. pylori cagA and miRNA146a rs2910164 genotypes were determined by PCR method. The results show that there was no significant difference in the frequency of H. pylori cagA gene between the two groups but there was a significant difference in the distribution of rs2910164 genotypes in patients < 50 years of age with the p-value of 0.05 and odds ratio equal to 2.69. On other hand, patients < 50 years of age with genotype CC of miRNA146a showed a significant difference in CRC risk. Furthermore, there was a significant correlation between rs2910164 CC genotype with Helicobacter pylori infection in patients < 50 years of age. The present study suggests that the CC genotype of miRNA146a in combination with H. pylori infection can be effective as risk factors and molecular markers for early diagnosis and treatment of CRC.

Keywords: colorectal cancer, Helicobacter pylori, miRNA146a, rs2910164 polymorphism

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Authors: Adriana Muñoz-Talavera, Ana Rosa Rincón-Sánchez, Abraham Escobedo-Moratilla, María Cristina Islas-Carbajal, Miguel Ángel Gómez-Lim

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The highest rates of diabetes incidence and prevalence worldwide will increase the number of diabetic patients requiring insulin or insulin analogues. Then, current production systems would not be sufficient to meet the future market demands. Therefore, developing efficient expression systems for insulin and insulin analogues are needed. In addition, insulin analogues with better pharmacokinetics and pharmacodynamics properties and without mitogenic potential will be required. SCI-57 (single chain insulin-57) is an insulin analogue having 10 times greater affinity to the insulin receptor, higher resistance to thermal degradation than insulin, native mitogenicity and biological effect. Plants as expression platforms have been used to produce recombinant proteins because of their advantages such as cost-effectiveness, posttranslational modifications, absence of human pathogens and high quality. Immunoglobulin production with a yield of 50% has been achieved by transient expression in Nicotiana benthamiana (Nb). The aim of this study is to produce SCI-57 by transient expression in Nb. Methodology: DNA sequence encoding SCI-57 was cloned in pICH31070. This construction was introduced into Agrobacterium tumefaciens by electroporation. The resulting strain was used to infiltrate leaves of Nb. In order to isolate SCI-57, leaves from transformed plants were incubated 3 hours with the extraction buffer therefore filtrated to remove solid material. The resultant protein solution was subjected to anion exchange chromatography on an FPLC system and ultrafiltration to purify SCI-57. Detection of SCI-57 was made by electrophoresis pattern (SDS-PAGE). Protein band was digested with trypsin and the peptides were analyzed by Liquid chromatography tandem-mass spectrometry (LC-MS/MS). A purified protein sample (20µM) was analyzed by ESI-Q-TOF-MS to obtain the ionization pattern and the exact molecular weight determination. Chromatography pattern and impurities detection were performed using RP-HPLC using recombinant insulin as standard. The identity of the SCI-57 was confirmed by anti-insulin ELISA. The total soluble protein concentration was quantified by Bradford assay. Results: The expression cassette was verified by restriction mapping (5393 bp fragment). The SDS-PAGE of crude leaf extract (CLE) of transformed plants, revealed a protein of about 6.4 kDa, non-present in CLE of untransformed plants. The LC-MS/MS results displayed one peptide with a high score that matches SCI-57 amino acid sequence in the sample, confirming the identity of SCI-57. From the purified SCI-57 sample (PSCI-57) the most intense charge state was 1069 m/z (+6) on the displayed ionization pattern corresponding to the molecular weight of SCI-57 (6412.6554 Da). The RP-HPLC of the PSCI-57 shows the presence of a peak with similar retention time (rt) and UV spectroscopic profile to the insulin standard (SCI-57 rt=12.96 and insulin rt=12.70 min). The collected SCI-57 peak had ELISA signal. The total protein amount in CLE from transformed plants was higher compared to untransformed plants. Conclusions: Our results suggest the feasibility to produce insulin analogue SCI-57 by transient expression in Nicotiana benthamiana. Further work is being undertaken to evaluate the biological activity by glucose uptake by insulin-sensitive and insulin-resistant murine and human cultured adipocytes.

Keywords: insulin analogue, mass spectrometry, Nicotiana benthamiana, transient expression

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Authors: Muskan Parmar, Musthafa Ottakam Thotiyl

Abstract:

The rising need for portable energy sources has led to advancements in direct liquid fuel cells (DLFCs) using various fuels like alcohol, ammonia, hydrazine, and vitamin C. Traditional precious metal catalysts improve reaction speeds but are expensive and prone to poisoning. Our study reveals how non-precious metal organometallic complexes, combined with smartly designed ligands, can significantly boost performance. The key is a unique interaction between the substrate (fuel) and the ligand, which creates a "dragging" effect that enhances reaction rates. By using this approach with a ferricyanide/ferrocyanide half-cell reaction, we developed a vitamin C fuel cell without precious metals. This fuel cell achieves an open circuit voltage of ∼950 mV, a peak power density of ∼97 mW cm⁻², and a peak current density of ∼215 mA cm⁻². Impressively, its performance is about 1.7 times better than traditional precious metal-based DLFCs. This highlights the potential of substrate ligand chemistry in the creation of sustainable DLFCs for efficient energy conversion.

Keywords: molecular electrocatalysts, vitamin C fuel cell, proton charge assembly, ferricyanide half-cell chemistry

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Authors: Saleh H. Salmen, Sulaiman Ali Alharbi, Hany M. Yehia, Mohammad A. Khiyami, Milton Wainwright, Naiyf S. Alharbi, Arunachalam Chinnathambi

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An antifungal protein synthesized by Bacillus cereus has been partially purified by the use of ammonium sulfate precipitation and Sephadex-G-200 column chromatography. The protein was produced from Bacillus cereus grown in potato Dextrose Broth Medium (PDB) at 30 ºC for 3 days at 100 rpm. The protein showed antagonistic effect against some fungi and yeasts. Crude extract from medium and semi-purified protein were tested in vitro against both fungi and yeasts using the disc diffusion method in order to detect the inhibitory effect of the protein. Zones of inhibition of the following diameter were found (mm) were Alternaria alternate (28), Rhodotorula glutinis (20), Fusarium sp. (16), Rhizopus sp. (15), Penicillium digitatum (13), Mucor sp. (13) and Aspergillus niger (10). The isolated protein was found to have a molecular weight of ~35kDa by sodium deodecyl sulfate-poly acrylamide gel electrophoresis. The data showed that the protein of Bacillus cereus has antifungal activity, a fact which points to the possibility of using it as a bio-control agent against some fungi, findings which emphasize the potential role of B. cereus as an important bio-control agent.

Keywords: bacillus cereus, ~35kDa protein, molds, yeasts

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Authors: Wonil Nam, Xiang Ren, Inyoung Kim, Masoud Agah, Wei Zhou

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Despite its ultrasensitive detection capability, surface-enhanced Raman spectroscopy (SERS) faces challenges as a quantitative biochemical analysis tool due to the significant dependence of local field intensity in hotspots on nanoscale geometric variations of plasmonic nanostructures. Therefore, despite enormous progress in plasmonic nanoengineering of high-performance SERS devices, it is still challenging to quantitatively correlate the measured SERS signals with the actual molecule concentrations at hotspots. A significant effort has been devoted to developing SERS calibration methods by introducing internal standards. It has been achieved by placing Raman tags at plasmonic hotspots. Raman tags undergo similar SERS enhancement at the same hotspots, and ratiometric SERS signals for analytes of interest can be generated with reduced dependence on geometrical variations. However, using Raman tags still faces challenges for real-world applications, including spatial competition between the analyte and tags in hotspots, spectral interference, laser-induced degradation/desorption due to plasmon-enhanced photochemical/photothermal effects. We show that electronic Raman scattering (ERS) signals from metallic nanostructures at hotspots can serve as the internal calibration standard to enable quantitative SERS analysis and improve biostatistical analysis. We perform SERS with Au-SiO₂ multilayered metal-insulator-metal nano laminated plasmonic nanostructures. Since the ERS signal is proportional to the volume density of electron-hole occupation in hotspots, the ERS signals exponentially increase when the wavenumber is approaching the zero value. By a long-pass filter, generally used in backscattered SERS configurations, to chop the ERS background continuum, we can observe an ERS pseudo-peak, IERS. Both ERS and SERS processes experience the |E|⁴ local enhancements during the excitation and inelastic scattering transitions. We calibrated IMRS of 10 μM Rhodamine 6G in solution by IERS. The results show that ERS calibration generates a new analytical value, ISERS/IERS, insensitive to variations from different hotspots and thus can quantitatively reflect the molecular concentration information. Given the calibration capability of ERS signals, we performed label-free SERS analysis of living biological systems using four different breast normal and cancer cell lines cultured on nano-laminated SERS devices. 2D Raman mapping over 100 μm × 100 μm, containing several cells, was conducted. The SERS spectra were subsequently analyzed by multivariate analysis using partial least square discriminant analysis. Remarkably, after ERS calibration, MCF-10A and MCF-7 cells are further separated while the two triple-negative breast cancer cells (MDA-MB-231 and HCC-1806) are more overlapped, in good agreement with the well-known cancer categorization regarding the degree of malignancy. To assess the strength of ERS calibration, we further carried out a drug efficacy study using MDA-MB-231 and different concentrations of anti-cancer drug paclitaxel (PTX). After ERS calibration, we can more clearly segregate the control/low-dosage groups (0 and 1.5 nM), the middle-dosage group (5 nM), and the group treated with half-maximal inhibitory concentration (IC50, 15 nM). Therefore, we envision that ERS calibrated SERS can find crucial opportunities in label-free molecular profiling of complicated biological systems.

Keywords: cancer cell drug efficacy, plasmonics, surface-enhanced Raman spectroscopy (SERS), SERS calibration

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Authors: Paula C Arroyo, Norma A Sánchez, Mikhail Tlenkopatchev

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The continuous growth in population has resulted in an increment in road construction. The road construction requires more lasting and resistant pavements. Among the different applications of polymers, the reinforcement of pavements throw the modification of asphalt has demonstrated to be an area of special interest for new polymers. The modified asphalt should exhibit a considerable good performance, good elastic properties and an increment in the performance grade (PG). Some of the current polymers used in asphalt are styrene butadiene styrene (SBS), poly(n-butyl methacrylate)-(glycidyl methacrylate) and ethylene-vinyl acetate EVA. The goal of this study was to synthesize low molecular weight (2,000 – 150,000 D) telechelic polymers to be applied at low concentrations in asphalt in order to modify its rheological properties and make it more resistant and durable. The telechelic polymers were obtained from different molar relationships between tensioned and functionalized olefins by ring opening metathesis polymerization (ROMP) and cross metathesis (CR). The synthesis was carried out under inert conditions with Grubbs second generation catalyst. The reaction efficiency was superior to 96% and telechelic polymers were characterized. The telechelic polymers were used to modify asphalt and the rheological properties of the modified asphalt were evaluated finding that at low concentrations (1%) the PG increased in one or two degrees.

Keywords: asphalt polymers, metathesis polymers, telechelic polymers, modified asphalt

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Authors: Vanessa R. Ralph, Leah J. Scharlott, Megan Y. Deshaye, Ryan L. Stowe

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Given the traditions of chemistry teaching, one may not question whether chemical phenomena play a prominent role. Yet, the role of chemical phenomena in an introductory chemistry course may define the extent to which the course is introductory, chemistry, and equitable. Picture, for example, the classic Ideal Gas Law problem. If one envisions a prompt wherein students are tasked with calculating a missing variable, then one envisions a prompt that relies on chemical phenomena as a context rather than as a model to understand the natural world. Consider a prompt wherein students are tasked with applying molecular models of gases to explain why the vapor pressure of a gaseous solution of water differs from that of carbon dioxide. Here, the chemical phenomenon is not only the context but also the subject of the prompt. Deliveries of general and organic chemistry were identified as ranging wildly in the integration of chemical phenomena. The more incorporated the phenomena, the more equitable the assessment task was for students of varying access to pre-college math and science preparation. How chemical phenomena are integrated may very well define whether courses are chemistry, are introductory, and are equitable. Educators of chemistry are invited colleagues to discuss the role of chemical phenomena in their courses and consider the long-lasting impacts of replicating tradition for tradition’s sake.

Keywords: equitable educational practices, chemistry curriculum, content organization, assessment design

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Authors: Ravinder Singh, Ankita Gurao, Saroj Bandhan, Sudhir Kumar Kashyap

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The Bos taurus Beta defensin 3 is a defensin peptide secreted by neutrophils and epithelial that exhibits anti-microbial activity. It is one of the crucial components forming an innate defense against intra mammary infections in livestock. The beta defensin 3 by virtue of its anti-microbial activity inhibits major mastitis pathogens including Staphylococcus aureus and Pseudomonas aeruginosa etc, which are also responsible for biofilm formation leading to antibiotic resistance phenomenon. Therefore, the defensin may prove as a non-conventional option to treat mastitis. In this study, computational analysis has been performed including sequence comparison among species and homology modeling of Bos taurus beta defensin 3 protein. The assessments of protein structure were done using the protein structure and model assessment tools integrated in Swiss Model server, which employs various local and global quality evaluation parameters. Further, molecular docking was also carried out between the defensin peptide and the components of biofilm to gain insight into various interactions and structural differences crucial for functionality of this protein.

Keywords: beta defensin 3, bos taurus, docking, homology modeling

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Authors: Fatih Topaloğlu

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Although philosophy is inherently a theoretical and intellectual activity, it should not be denied that environmental conditions influence the formation and shaping of philosophical thought. In this context, it should be noted that the Philosophy of Religion has been influential in the debates in the West, especially since the beginning of the 20th century, and that this influence has dimensions that cannot be limited to academic or intellectual fields. The issues and problems that fall within the field of interest of Philosophy of Religion are followed with interest by a significant proportion of society through popular publications. Philosophy of Religion has its share in many social, economic, cultural, scientific, political and ethical developments. Philosophy of Religion, in the most general sense, can be defined as a philosophical approach to religion or a philosophical way of thinking and discussing religion. Philosophy of Religion tries to explain the epistemological foundations of concepts such as belief and faith that shape religious life by revealing their meaning for the individual. Thus, Philosophy of Religion tries to evaluate the effect of beliefs on the individual's values, judgments and behaviours with a comprehensive and critical eye. The Philosophy of Religion, which tries to create new solutions and perspectives by applying the methods of philosophy to religious problems, tries to solve these problems not by referring to the holy book or religious teachings but by logical proofs obtained through the possibilities of reason and evidence filtered through the filter of criticism. Although there is no standard method for doing Philosophy of Religion, it can be said that an approach that can be expressed as thinking about religion in a rational, objective, and consistent way is generally accepted. The evaluations made within the scope of Philosophy of Religion have two stages. The first is the definition stage, and the second is the evaluation stage. In the first stage, the data of different scientific disciplines, especially other religious sciences, are utilized to define the issues objectively. In the second stage, philosophical evaluations are made based on this foundation. During these evaluations, the issue of how the relationship between religion and philosophy should be established is extremely sensitive. The main thesis of this paper is that the Philosophy of Religion, as a branch of philosophy, has been affected by the conditions caused by the historical experience through which it has passed and has differentiated its subjects and the methods it uses to realize its philosophical acts over time under the influence of these conditions. This study will attempt to evaluate the validity of this study based on the "Introduction to Philosophy of Religion" literature, which we assume reflects this differentiation. As a result of this examination will aim to reach some factual conclusions about the nature of both philosophical and religious thought, to determine the phases that the Philosophy of Religion as a discipline has gone through since the day it emerged, and to investigate the possibilities of a holistic view of the field.

Keywords: content analysis, culture, history, philosophy of religion, method

Procedia PDF Downloads 55

Authors: K. J. Keerthi, Vasundhara Kamineni, A. Ravi Shanker, T. Rammurthy, A. Vijaya Lakshmi, Q. Hasan

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Pancreatic β-cells are the predominant insulin-producing cell types within the Islets of Langerhans and insulin is the primary hormone which regulates carbohydrate and fat metabolism. Apoptosis of β-cells or insufficient insulin production leads to Diabetes Mellitus (DM). Current therapy for diabetes includes either medical management or insulin replacement and regular monitoring. Replacement of β- cells is an attractive treatment option for both Type-1 and Type-2 DM in view of the recent paper which indicates that β-cells apoptosis is the common underlying cause for both the Types of DM. With the development of Edmonton protocol, pancreatic β-cells allo-transplantation became possible, but this is still not considered as standard of care due to subsequent requirement of lifelong immunosuppression and the scarcity of suitable healthy organs to retrieve pancreatic β-cell. Fetal pancreatic cells from abortuses were developed as a possible therapeutic option for Diabetes, however, this posed several ethical issues. Hence, in the present study Mesenchymal stem cells (MSCs) were differentiated into insulin producing cells which were isolated from Human Umbilical cord (HUC) tissue. MSCs have already made their mark in the growing field of regenerative medicine, and their therapeutic worth has already been validated for a number of conditions. HUC samples were collected with prior informed consent as approved by the Institutional ethical committee. HUC (n=26) were processed using a combination of both mechanical and enzymatic (collagenase-II, 100 U/ml, Gibco ) methods to obtain MSCs which were cultured in-vitro in L-DMEM (Low glucose Dulbecco's Modified Eagle's Medium, Sigma, 4.5 mM glucose/L), 10% FBS in 5% CO2 incubator at 37°C. After reaching 80-90% confluency, MSCs were characterized with Flowcytometry and Immunocytochemistry for specific cell surface antigens. Cells expressed CD90+, CD73+, CD105+, CD34-, CD45-, HLA-DR-/Low and Vimentin+. These cells were differentiated to β-cells by using H-DMEM (High glucose Dulbecco's Modified Eagle's Medium,25 mM glucose/L, Gibco), β-Mercaptoethanol (0.1mM, Hi-Media), basic Fibroblast growth factor (10 µg /L,Gibco), and Nicotinamide (10 mmol/L, Hi-Media). Pancreatic β-cells were confirmed by positive Dithizone staining and were found to be functionally active as they released 8 IU/ml insulin on glucose stimulation. Isolating MSCs from usually discarded, abundantly available HUC tissue, expanding and differentiating to β-cells may be the most feasible cell therapy option for the millions of people suffering from DM globally.

Keywords: diabetes mellitus, human umbilical cord, mesenchymal stem cells, differentiation

Procedia PDF Downloads 258

Authors: Carlos Amnael Orozco-Diaz, Robert David Moorehead, Gwendolen Reilly, Fiona Gilchrist, Cheryl Ann Miller

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The current gold-standard for maxillofacial reconstruction surgery (MRS) utilizes auto-grafted cancellous bone as a filler. This study was aimed towards developing a polylactic-acid/hydroxyapatite (PLA-HA) composite suitable for fused-deposition 3D printing. Functionalization of the polymer through the addition of HA was directed to promoting bone-regeneration properties so that the material can rival the performance of cancellous bone grafts in terms of bone-lesion repair. This kind of composite enables the production of MRS implants based off 3D-reconstructions from image studies – namely computed tomography – for anatomically-correct fitting. The present study encompassed in-vitro degradation and in-vitro biocompatibility profiling for 3D-printed PLA and PLA-HA composites. PLA filament (Verbatim Co.) and Captal S hydroxyapatite micro-scale HA powder (Plasma Biotal Ltd) were used to produce PLA-HA composites at 5, 10, and 20%-by-weight HA concentration. These were extruded into 3D-printing filament, and processed in a BFB-3000 3D-Printer (3D Systems Co.) into tensile specimens, and were mechanically challenged as per ASTM D638-03. Furthermore, tensile specimens were subjected to accelerated degradation in phosphate-buffered saline solution at 70°C for 23 days, as per ISO-10993-13-2010. This included monitoring of mass loss (through dry-weighing), crystallinity (through thermogravimetric analysis/differential thermal analysis), molecular weight (through gel-permeation chromatography), and tensile strength. In-vitro biocompatibility analysis included cell-viability and extracellular matrix deposition, which were performed both on flat surfaces and on 3D-constructs – both produced through 3D-printing. Discs of 1 cm in diameter and cubic 3D-meshes of 1 cm3 were 3D printed in PLA and PLA-HA composites (n = 6). The samples were seeded with 5000 MG-63 osteosarcoma-like cells, with cell viability extrapolated throughout 21 days via resazurin reduction assays. As evidence of osteogenicity, collagen and calcium deposition were indirectly estimated through Sirius Red staining and Alizarin Red staining respectively. Results have shown that 3D printed PLA loses structural integrity as early as the first day of accelerated degradation, which was significantly faster than the literature suggests. This was reflected in the loss of tensile strength down to untestable brittleness. During degradation, mass loss, molecular weight, and crystallinity behaved similarly to results found in similar studies for PLA. All composite versions and pure PLA were found to perform equivalent to tissue-culture plastic (TCP) in supporting the seeded-cell population. Significant differences (p = 0.05) were found on collagen deposition for higher HA concentrations, with composite samples performing better than pure PLA and TCP. Additionally, per-cell-calcium deposition on the 3D-meshes was significantly lower when comparing 3D-meshes to discs of the same material (p = 0.05). These results support the idea that 3D-printable PLA-HA composites are a viable resorbable material for artificial grafts for bone-regeneration. Degradation data suggests that 3D-printing of these materials – as opposed to other manufacturing methods – might result in faster resorption than currently-used PLA implants.

Keywords: bone regeneration implants, 3D-printing, in vitro testing, biocompatibility, polymer degradation, polymer-ceramic composites

Procedia PDF Downloads 154

Authors: K. Karapetyan, F. Tkhruni, A. Aghajanyan, T. S. Balabekyan, L. Arstamyan

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Introduction: Globalization of the food supply has created the conditions favorable for the emergence and spread of food-borne and especially dangerous pathogens (EDP) in developing countries. The fresh-cut fruit and vegetable industry is searching for alternatives to replace chemical treatments with biopreservative approaches that ensure the safety of the processed foods product. Antimicrobial compounds of lactic acid bacteria (LAB) possess bactericidal or bacteriostatic activity against intestinal pathogens, spoilage organisms and food-borne pathogens such as Listeria monocytogenes, Staphylococcus aureus and Salmonella. Endemic strains of LAB were isolated. The strains, showing broad spectrum of antimicrobial activity against food spoiling microorganisms, were selected. The genotyping by 16S rRNA sequencing, GS-PCR, RAPD PCR methods showed that they were presented by Lactobacillus rhamnosus109, L.plantarum 65, L.plantarum 66 and Enterococcus faecium 64 species. LAB are deposited in "Microbial Depository Center" (MDC) SPC "Armbiotechnology". Methods: LAB strains were isolated from different dairy products from rural households from the highland regions of Armenia. Serially diluted samples were spread on MRS (Merck, Germany) and hydrolyzed milk agar (1,2 % w/v). Single colonies from each LAB were individually inoculated in liquid MRS medium and incubated at 37oC for 24 hours. Culture broth with biomass was centrifuged at 10,000 g during 20 min for obtaining of cell free culture broth (CFC). The antimicrobial substances from CFC broth were purified by the combination of adsorption-desorption and ion-exchange chromatography methods. Separation of bacteriocins was performed using a HPLC method on "Avex ODS" C18 column. Mass analysis of peptides recorded on the device API 4000 in the electron ionization mode. The spot-on-lawn method on the test culture plated in the solid medium was applied. The antimicrobial activity is expressed in arbitrary units (AU/ml). Results. Purification of CFC broth of LAB allowed to obtain partially purified antimicrobial preparations which contains bacteriocins with broad spectrum of antimicrobial activity. Investigation of their main biochemical properties shown, that inhibitory activity of preparations is partially reduced after treatment with proteinase K, trypsin, pepsin, suggesting a proteinaceous nature of bacteriocin-like substances containing in CFC broth. Preparations preserved their activity after heat treatment (50-121 oC, 20 min) and were stable in the pH range 3–8. The results of SDS PAAG electrophoresis show that L.plantarum 66 and Ent.faecium 64 strains have one bacteriocin (BCN) with maximal antimicrobial activity with approximate molecular weight 2.0-3.0 kDa. From L.rhamnosus 109 two BCNs were obtained. Mass spectral analysis indicates that these bacteriocins have peptide bonds and molecular weight of BCN 1 and BCN 2 are approximately 1.5 kDa and 700 Da. Discussion: Thus, our experimental data shown, that isolated endemic strains of LAB are able to produce bacteriocins with high and different inhibitory activity against broad spectrum of microorganisms of different taxonomic group, such as Salmonella sp., Esherichia coli, Bacillus sp., L.monocytogenes, Proteus mirabilis, Staph. aureus, Ps. aeruginosa. Obtained results proved the perspectives for use of endemic strains in the preservation of foodstuffs. Acknowledgments: This work was realized with financial support of the Project Global Initiatives for Preliferation Prevention (GIPP) T2- 298, ISTC A-1866.

Keywords: antimicrobial activity, bacteriocins, endemic strains, food safety

Procedia PDF Downloads 559