Search results for: whey protein isolated

Authors: V. Barbakadze, L. Gogilashvili, L. Amiranashvili, M. Merlani, K. Mulkijanyan

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The high-molecular water-soluble preparations from several species of two genera (Symphytum and Anchusa) of Boraginaceae family Symphytum asperum, S. caucasicum, S.officinale and Anchusa italica were isolated. According to IR, 13C and 1H NMR, APT, 1D NOE, 2D heteronuclear 1H/13C HSQC and 2D DOSY experiments, the main chemical constit¬uent of these preparations was found to be caffeic acid-derived polyether, namely poly[3-(3,4-dihydroxyphenyl)glyceric acid] (PDPGA) or poly[oxy-1-carboxy-2-(3,4-dihydroxyphenyl)ethylene]. Most carboxylic groups of this caffeic acid-derived polymer of A. italica are methylated.

Keywords: Anchusa, poly[3-(3, 4-dihydroxyphenyl)glyceric acid], poly[oxy-1-carboxy-2-(3, 4-dihydroxyphenyl)ethylene], Symphytum

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Authors: Maneesh Kumar, Roshan Kamal Topno, Manas Ranjan Dikhit, Vahab Ali, Ganesh Chandra Sahoo, Bhawana, Major Madhukar, Rishikesh Kumar, Krishna Pandey, Pradeep Das

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Chandipura vesiculovirus is an emerging (-) ssRNA viral entity belonging to the genus Vesiculovirus of the family Rhabdoviridae, associated with fatal encephalitis in tropical regions. The multi-functionally active viral RNA-dependent RNA polymerase (vRdRp) that has been incorporated with conserved amino acid residues in the pathogens, assigned to synthesize distinct viral polypeptides. The lack of proofreading ability of the vRdRp produces many mutated variants. Here, we have performed the evolutionary analysis of 20 viral protein sequences of vRdRp of different strains of Chandipura vesiculovirus along with other viral species from genus Vesiculovirus inferred in MEGA6.06, employing the Neighbour-Joining method. The p-distance algorithmic method has been used to calculate the optimum tree which showed the sum of branch length of about 1.436. The percentage of replicate trees in which the associated taxa are clustered together in the bootstrap test (1000 replicates), is shown next to the branches. No mutation was observed in the Indian strains of Chandipura vesiculovirus. In vRdRp, 1230(His) and 1231(Arg) are actively participated in catalysis and, are found conserved in different strains of Chandipura vesiculovirus. Both amino acid residues were also conserved in the other viral species from genus Vesiculovirus. Many isolates exhibited maximum number of mutations in catalytic regions in strains of Chandipura vesiculovirus at position 26(Ser→Ala), 47 (Ser→Ala), 90(Ser→Tyr), 172(Gly→Ile, Val), 172(Ser→Tyr), 387(Asn→Ser), 1301(Thr→Ala), 1330(Ala→Glu), 2015(Phe→Ser) and 2065(Thr→Val) which make them variants under different tropical conditions from where they evolved. The result clarifies the actual concept of RNA evolution using vRdRp to develop as an evolutionary marker. Although, a limited number of vRdRp protein sequence similarities for Chandipura vesiculovirus and other species. This might endow with possibilities to identify the virulence level during viral multiplication in a host.

Keywords: Chandipura, (-) ssRNA, viral RNA-dependent RNA polymerase, neighbour-joining method, p-distance algorithmic, evolutionary marker

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Authors: F. Saleh, F. Lyall, A. Abdulsid, L. Marks

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Background: Labour in human is a complex biological process that involves interactions of neurological, hormonal and inflammatory pathways, with the placenta being a key regulator of these pathways. It is known that uterine contractions and labour pain cause physiological changes in gene expression in maternal and fetal blood, and in placenta during labour. Oxidative and inflammatory stress pathways are implicated in labour and they may cause alteration of placental gene expression. Additionally, in placental tissues, labour increases the expression of genes involved in placental oxidative stress, inflammatory cytokines, angiogenic regulators and apoptosis. Recently, Hydrogen Sulphide (H2S) has been considered as an endogenous gaseous mediator which promotes vasodilation and exhibits cytoprotective anti-inflammatory properties. The endogenous H2S is synthesised predominantly by two enzymes: cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE). As the H2S pathway has anti-oxidative and anti-inflammatory characteristics thus, we hypothesised that the expression of CBS and CSE in placental tissues would alter during labour. Methods: CBS and CSE expressions were examined in placentas using western blotting and RT-PCR in inner, middle and outer placental zones in placentas obtained from healthy non labouring women who delivered by caesarian section. These were compared with the equivalent zone of placentas obtained from women who had uncomplicated labour and delivered vaginally. Results: No differences in CBS and CSE mRNA or protein levels were found between the different sites within placentas in either the labour or non-labour group. There were no significant differences in either CBS or CSE expression between the two groups at the inner site and middle site. However, at the outer site there was a highly significant decrease in CBS protein expression in the labour group when compared to the non-labour group (p = 0.002). Conclusion: To the best of author’s knowledge, this is the first report to suggest that, CBS is expressed in a spatial manner within the human placenta. Further work is needed to clarify the precise function and mechanism of this spatial regulation although it is likely that inflammatory pathways regulation is a complex process in which this plays a role.

Keywords: anti-inflammatory, hydrogen sulphide, labour, oxidative stress

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Authors: Qi LI, Xu Lin, Nengming Lin

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Objective: The aim of this study was to investigate the role of desmocollin 2 (DSC2) protein in the proliferation, migration and drug resistance of lung cancer cells. Method: CCK-8 assays and colony formation assays were used to evaluate the effect of dsc2 regulation on cancer cell viability and colony formation. Transwell assays and wound healing assays were also performed. Cell flow double staining was used to detect the apoptosis rate of cells with DSC2, which was added cisplatin. Western blot assay was used to detect cell cycle, PI3k/Akt and apoptosis-related proteins. Results: Our data showed that dsc2 is upregulated in clinical lung cancer tissues compared with pericarcinomatous tissues, and it is differentially expressed in lung cancer cell lines. The down-regulation of dsc2 in A549 and H358 lung cancer cells significantly suppressed the cell proliferation, metastasis, and motility. In contrast, the opposite effects were observed in overexpression of dsc2 both in H23 and PC9 cell lines. In addition to lung adenocarcinoma cell lines, we also examined its expression in lung squamous cell lines, such as H226. Western blotting showed that dsc2 could reduce the level of phosphorylated Akt (Ser 473) and p-mTOR. Thus, it is speculated that dsc2 up-regulation promotes proliferation and invasiveness through activation of the PI3K/AKT pathway. Also, knockdown of dsc2 in A549 and H226 could significantly decreased in the levels of cyclinB and wee1 protein. Additionally, flow cytometry showed that dsc2 knockdown combined with cisplatin could significantly enhance cell apoptosis rate. Conclusion: These data suggest that dsc2 promotes the proliferation and migration of lung cancer cells in vitro. Also, the results suggested that dsc2 could affect the cell cycle and apoptosis of lung cells. Furthermore, knockdown of dsc2 could sensitize cisplatin in both lung adenocarcinoma and lung squamous cell lines. Thus we suggested that dsc2 can be used as a therapeutic target for lung cancer.

Keywords: desmocollin 2, cisplatin, lung cancer, PI3K/AKT, lung squamous cell

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Authors: Andrés Quiroz, Emilio Hormazabal, Ana Mutis, Fernando Ortega, Manuel Chacón-Fuentes, Leonardo Parra

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Red clover (Trifolium pratense L.) is an important forage species in temperate regions of the world. The main limitation of this species worldwide is a lack of persistence related to the high mortality of plants due to a complex of biotic and abiotic factors, determining a life span of two or three seasons. Because of the importance of red clover in Chile, a red clover breeding program was started at INIA Carillanca Research Center in 1989, with the main objective of improving the survival of plants, forage yield, and persistence. The main selection criteria for selecting new varieties have been based on agronomical parameters and biotic factors. The main biotic factor associated with red clover mortality in Chile is Hylastinus obscurus (Coleoptera: Curculionidae). Both larval and adults feed on the roots, causing weakening and subsequent death of clover plants. Pesticides have not been successful for controlling infestations of this root borer. Therefore, alternative strategies for controlling this pest are a high priority for red clover producers. Currently, the role of semiochemical in the interaction between H. obscurus and red clover plants has been widely studied for our group. Specifically, from the red clover foliage has been identified limonene is eliciting repellency from the root borer. Limonene is generated in the plant from two independent biosynthetic pathways, the mevalonic acid, and deoxyxylulose pathway. Mevalonate pathway enzymes are localized in the cytosol, whereas the deoxyxylulose phosphate pathway enzymes are found in plastids. In summary, limonene can be determinated by enzymatic bioassay using GPP as substrate and by limonene synthase expression. Therefore, the main objective of this work was to study genetic variation of limonene in material provided by INIA´s Red Clover breeding program. Protein extraction was carried out homogenizing 250 mg of leave tissue and suspended in 6 mL of extraction buffer (PEG 1500, PVP-30, 20 mM MgCl2 and antioxidants) and stirred on ice for 20 min. After centrifugation, aliquots of 2.5 mL were desalted on PD-10 columns, resulting in a final volume of 3.5 mL. Protein determination was performed according to Bradford with BSA as a standard. Monoterpene synthase assays were performed with 50 µL of protein extracts transferred into gas-tight 2 mL crimp seal vials after addition of 4 µL MgCl₂ and 41 µL assay buffer. The assay was started by adding 5 µL of a GPP solution. The mixture was incubated for 30 min at 40 °C. Biosynthesized limonene was quantified in a GC equipped with a chiral column and using synthetic R and S-limonene standards. The enzymatic the production of R and S-limonene from different Superqueli-Carillanca genotypes is shown in this work. Preliminary results showed significant differences in limonene content among the genotypes analyzed. These results constitute an important base for selecting genotypes with a high content of this repellent monoterpene towards H. obscurus.

Keywords: head space, limonene enzymatic determination, red clover, Hylastinus obscurus

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Authors: Minhal Abidi, Moinuddin

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Diabetes mellitus (DM) induced metabolic abnormalities causes oxidative stress which leads to the pathogenesis of complications associated with diabetes like retinopathy, nephropathy periodontitis etc. Combination of glycation and oxidation 'glycoxidation' occurs when oxidative reactions affect the early state of glycation products. Low density lipoprotein (LDL) is prone to glycoxidative attack by sugars and methylglyoxal (MGO) being a strong glycating agent may have severe impact on its structure and consequent role in diabetes. Pro-inflammatory cytokines like IL1β and TNFα produced by the action of gram negative bacteria in periodontits (PD) can in turn lead to insulin resistance. This work discusses modifications to LDL as a result of glycoxidation. The changes in the protein molecule have been characterized by various physicochemical techniques and the immunogenicity of the modified molecules was also evaluated as they presented neo-epitopes. Binding of antibodies present in diabetes patients to the native and glycated LDL has been evaluated. Role of modified epitopes in the generation of antibodies in diabetes and periodontitis has been discussed. The structural perturbations induced in LDL were analyzed by UV–Vis, fluorescence, circular dichroism and FTIR spectroscopy, molecular docking studies, thermal denaturation studies, Thioflavin T assay, isothermal titration calorimetry, comet assay. MALDI-TOF, ketoamine moieties, carbonyl content and HMF content were also quantitated in native and glycated LDL. IL1β and TNFα levels were also measured in the type 2 DM and PD patients. We report increased carbonyl content, ketoamine moieties and HMF content in glycated LDL as compared to native analogue. The results substantiate that in hyperglycemic state MGO modification of LDL causes structural perturbations making the protein antigenic which could obstruct normal physiological functions and might contribute in the development of secondary complications in diabetic patients like periodontitis.

Keywords: advanced glycation end products, diabetes mellitus, glycation, glycoxidation, low density lipoprotein, periodontitis

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Authors: Denis Mustafov, Emmanouil Karteris, Maria Braoudaki

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Glioblastoma multiform (GBM) is a heterogenous primary brain tumour that kills most affected patients. To the authors best knowledge, despite all research efforts there is no early diagnostic biomarker for GBM. MicroRNAs (miRNAs) are short non-coding RNA molecules which are deregulated in many cancers. The aim of this research was to determine miRNAs with a diagnostic impact and to potentially identify promising therapeutic targets for glioblastoma multiform. In silico analysis was performed to identify deregulated miRNAs with diagnostic relevance for glioblastoma. The expression profiles of the chosen miRNAs were then validated in vitro in the human glioblastoma cell lines A172 and U-87MG. Briefly, RNA extraction was carried out using the Trizol method, whilst miRNA extraction was performed using the mirVANA miRNA isolation kit. Quantitative Real-Time Polymerase Chain Reaction was performed to verify their expression. The presence of five target proteins within the A172 cell line was evaluated by Western blotting. The expression of the CORO1C protein within 32 GBM cases was examined via immunohistochemistry. The miRNAs identified in silico included miR-21-5p, miR-34a and miR-128a. These miRNAs were shown to target deregulated GBM genes, such as CDK6, E2F3, BMI1, JAG1, and CORO1C. miR-34a and miR-128a showed low expression profiles in comparison to a control miR-RNU-44 in both GBM cell lines suggesting tumour suppressor properties. Opposing, miR-21-5p demonstrated greater expression indicating that it could potentially function as an oncomiR. Western blotting revealed expression of all five proteins within the A172 cell line. In silico analysis also suggested that CORO1C is a target of miR-128a and miR-34a. Immunohistochemistry demonstrated that 75% of the GBM cases showed moderate to high expression of CORO1C protein. Greater understanding of the deregulated expression of miR-128a and the upregulation of CORO1C in GBM could potentially lead to the identification of a promising diagnostic biomarker signature for glioblastomas.

Keywords: non-coding RNAs, gene expression, brain tumours, immunohistochemistry

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Authors: P. Behera, K. K. Singh, D. K. Saini, M. De

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Implementation of 2D materials in the detection of bio analytes is highly advantageous in the field of sensing because of its high surface to volume ratio. We have designed our sensor array with different cationic two-dimensional MoS₂, where surface modification was achieved by cationic thiol ligands with different functionality. Green fluorescent protein (GFP) was chosen as signal transducers for its biocompatibility and anionic nature, which can bind to the cationic MoS₂ surface easily, followed by fluorescence quenching. The addition of bio-analyte to the sensor can decomplex the cationic MoS₂ and GFP conjugates, followed by the regeneration of GFP fluorescence. The fluorescence response pattern belongs to various analytes collected and transformed to linear discriminant analysis (LDA) for classification. At first, 15 different proteins having wide range of molecular weight and isoelectric points were successfully discriminated at 50 nM with detection limit of 1 nM. The sensor system was also executed in biofluids such as serum, where 10 different proteins at 2.5 μM were well separated. After successful discrimination of protein analytes, the sensor array was implemented for bacteria sensing. Six different bacteria were successfully classified at OD = 0.05 with a detection limit corresponding to OD = 0.005. The optimized sensor array was able to classify uropathogens from non-uropathogens in urine medium. Further, the technique was applied for discrimination of bacteria possessing resistance to different types and amounts of drugs. We found out the mechanism of sensing through optical and electrodynamic studies, which indicates the interaction between bacteria with the sensor system was mainly due to electrostatic force of interactions, but the separation of native bacteria from their drug resistant variant was due to Van der Waals forces. There are two ways bacteria can be detected, i.e., through bacterial cells and lysates. The bacterial lysates contain intracellular information and also safe to analysis as it does not contain live cells. Lysates of different drug resistant bacteria were patterned effectively from the native strain. From unknown sample analysis, we found that discrimination of bacterial cells is more sensitive than that of lysates. But the analyst can prefer bacterial lysates over live cells for safer analysis.

Keywords: array-based sensing, drug resistant bacteria, linear discriminant analysis, two-dimensional MoS₂

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Authors: Vladimíra Kňazovická, Mária Dovičičová, Miroslava Kačániová, Margita Čanigová

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The aim of the study was to test the storage of bee pollen at room temperature and in cold store, and to describe microorganisms originated from it. Fresh bee pollen originating in West Slovakia was collected in May 2010. It was tested for presence of particular microbial groups using dilution plating method, and divided into two parts with different storage (in cold store and at room temperature). Microbial analyses of pollen were repeated after one year of storage. Several bacterial strains were isolated and tested using Gram staining, for catalase and fructose-6-phosphate-phosphoketolase presence, and by rapid ID 32A (BioMérieux, France). Micromycetes were identified at genus level. Fresh pollen contained coliform bacteria, which were not detected after one year of storage in both ways. Total plate count (TPC) of aerobes and anaerobes and of yeasts in fresh bee pollen exceeded 5.00 log CFU/g. TPC of aerobes and anaerobes decreased below 2.00 log CFU/g after one year of storage in both ways. Count of yeasts decreased to 2.32 log CFU/g (at room temperature) and to 3.66 log CFU/g (in cold store). Microscopic filamentous fungi decreased from 3.41 log CFU/g (fresh bee pollen) to 1.13 log CFU/g (at room temperature) and to 1.89 log CFU/g (in cold store). In fresh bee pollen, 12 genera of micromycetes were identified in the following order according to their relative density: Penicillium > Mucor > Absidia > Cladosporium, Fusarium > Alternaria > Eurotium > Aspergillus, Rhizopus > Emericella > Arthrinium and Mycelium sterilium. After one year at room temperature, only three genera were detected in bee pollen (Penicillium > Aspergillus, Mucor) and after one year in cold store, seven genera were detected (Mucor > Penicillium, Emericella > Aspergillus, Absidia > Arthrinium, Eurotium). From the plates designated for anaerobes, eight colonies originating in fresh bee pollen were isolated. Among them, a single yeast isolate occurred. Other isolates were G+ bacteria, with a total of five rod shaped. In three out of these five, catalase was absent and fructose-6-phosphate-phosphoketolase was present. Bacterial isolates originating in fresh pollen belonged probably to genus Bifidobacterium or relative genera, but their identity was not confirmed unequivocally. In general, cold conditions are suitable for maintaining the natural properties of foodstuffs for a longer time. Slight decrease of microscopic fungal number and diversity was recorded in cold temperatures compared with storage at room temperature.

Keywords: bacteria, bee product, microscopic fungi, biosystems engineering

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Authors: Viviana Manzulli, Luigina Serrecchia, Adelia Donatiello, Valeria Rondinone, Sabine Zange, Alina Tscherne, Antonio Parisi, Antonio Fasanella

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Anthrax is a zoonotic disease that affects a wide range of animal species (primarily ruminant herbivores), and can be transmitted to humans through consumption or handling of contaminated animal products. The etiological agent B.anthracis is able to survive in unfavorable environmental conditions by forming endospore which remain viable in the soil for many decades. Furthermore, B.anthracis is considered as one of the most feared agents to be potentially misused as a biological weapon and the importance of the disease and its treatment in humans has been underscored before the bioterrorism events in the United States in 2001. Due to the often fatal outcome of human cases, antimicrobial susceptibility testing plays especially in the management of anthrax infections an important role. In Italy, animal anthrax is endemic (predominantly found in the southern regions and on islands) and is characterized by sporadic outbreaks occurring mainly during summer. Between 2012 and 2017 single human cases of cutaneous anthrax occurred. In this study, 90 diverse strains of B.anthracis, isolated in Italy from 2001 to 2017, were screened to their susceptibility to sixteen clinically relevant antimicrobial agents by using the broth microdilution method. B.anthracis strains selected for this study belong to the strain collection stored at the Anthrax Reference Institute of Italy located inside the Istituto Zooprofilattico Sperimentale of Puglia and Basilicata. The strains were isolated at different time points and places from various matrices (human, animal and environmental). All strains are a representative of over fifty distinct MLVA 31 genotypes. The following antibiotics were used for testing: gentamicin, ceftriaxone, streptomycin, penicillin G, clindamycin, chloramphenicol, vancomycin, linezolid, cefotaxime, tetracycline, erythromycin, rifampin, amoxicillin, ciprofloxacin, doxycycline and trimethoprim. A standard concentration of each antibiotic was prepared in a specific diluent, which were then twofold serial diluted. Therefore, each wells contained: bacterial suspension of 1–5x104 CFU/mL in Mueller-Hinton Broth (MHB), the antibiotic to be tested at known concentration and resazurin, an indicator of cell growth. After incubation overnight at 37°C, the wells were screened for color changes caused by the resazurin: a change from purple to pink/colorless indicated cell growth. The lowest concentration of antibiotic that prevented growth represented the minimal inhibitory concentration (MIC). This study suggests that B.anthracis remains susceptible in vitro to many antibiotics, in addition to doxycycline (MICs ≤ 0,03 µg/ml), ciprofloxacin (MICs ≤ 0,03 µg/ml) and penicillin G (MICs ≤ 0,06 µg/ml), recommend by CDC for the treatment of human cases and for prophylactic use after exposure to the spores. In fact, the good activity of gentamicin (MICs ≤ 0,25 µg/ml), streptomycin (MICs ≤ 1 µg/ml), clindamycin (MICs ≤ 0,125 µg/ml), chloramphenicol(MICs ≤ 4 µg/ml), vancomycin (MICs ≤ 2 µg/ml), linezolid (MICs ≤ 2 µg/ml), tetracycline (MICs ≤ 0,125 µg/ml), erythromycin (MICs ≤ 0,25 µg/ml), rifampin (MICs ≤ 0,25 µg/ml), amoxicillin (MICs ≤ 0,06 µg/ml), towards all tested B.anthracis strains demonstrates an appropriate alternative choice for prophylaxis and/or treatment. All tested B.anthracis strains showed intermediate susceptibility to the cephalosporins (MICs ≥ 16 µg/ml) and resistance to trimethoprim (MICs ≥ 128 µg/ml).

Keywords: Bacillus anthracis, antibiotic susceptibility, treatment, minimum inhibitory concentration

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Authors: Abdessemed Zineb, Atia Yahia, Yeza Salima

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Water is self-purified by activated sludge process which makes its uniqueness. The main goal is the microbial biocenosis study of the input and output water of the waste water treatment system plant Timgad. 89.47% of the identified biocenosis belongs to ɤ-Proteobacteria while the remaining 10.52 % is equally divided between α-Proteobacteria and β-Proteobacteria. The antibiotics susceptibility profiles reveal that over 30 % are wild strains while the penicillinases are often present (11.30-20 %) with also other profiles. This proportion is worrying that the water discharged join the Oued Soltez used for irrigation. This disadvantage involves the installation of a chlorination step.

Keywords: activated sludge, biocenosis, antibiotics profiles, penicillinases, physic-chemical quality

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Authors: Natesan Balasubramanian, Palpandi Pounpandi, Venkatraman Thamil Priya, Vellasamy Shanmugaiah, Karubbiah Balakrishnan, Mandayam Anandam Thirunarayan

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Streptococcus agalactiae is commonly known as Group B Streptococcus (GBS) and it is the most common cause of life-threatening bacterial infection. GBS first considered as a veterinary pathogen causing mastitis in cattle later becomes a human pathogen for severe neonatal infections. In this present study, a total of 20 new clinical isolates of S. agalactiae were collected from male (6) and female patient (14) with different age group. The isolates were from Urinary tract infection (UTI), blood, pus and eye ulcer. All the 20 S. agalactiae isolates has clear hemolysis properties on blood agar medium and were identified by serogrouping and MALTI-TOF-MS analysis. Antibiotic susceptibility/resistance test was performed for 20 S. agalactiae isolates, further phenotypic resistance pattern was observed for tetracycline, vancomycin, ampicillin and penicillin. Genotypically we found two antibiotic resistance genes such as Betalactem antibiotic resistance gene (Tem) (70%) and tetracycline resistance gene Tet(O) 15% in our isolates. Six virulence factors encoding genes were performed by PCR in twenty GBS isolates, cfb gene (100%), followed by, cylE(90.47%), lmp(85.7%), bca(71.42%), rib (38%) and low frequency in bac gene (4.76%) were determined. Most of the S. agalactiae isolates produced strong biofilm in the polystyrene surface (hydrophobic), and low-level biofilm formation was found in glass tube (hydrophilic) surface. lytR is secreted protein and localized in bacterial cell wall, extra cellular membrane, and cytoplasm. In silico docking studies were performed for lytR protein with four antibiofilm compounds, including a peptide (PR39) with the docking study showed peptide has strong interaction followed by ellagic acid and interaction length is 2.95, 2.97 and 2.95 A°. In ligand EGCGO10 and O11 two atoms intract with lytR (Leu271), with binding bond affinity length is 3.24 and 3.14. The aminoacid Leu 271 is act as an impartant aminoacid, since ellagic acid and EGCG interact with same aminoacid.

Keywords: antibiotics, biofilms, clinical isolates, S. agalactiae, virulence

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Authors: S. El Ballal, H. El Sabbagh, M. Abd El Gaber, A. Eisa, A. Al Gamal

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Cadmium is one of the most harmful heavy metals able to induce severe injury. In this study, sixty four male Sprague Dawley rats weighing (70-80 gm) were used. Rats were divided into 4 groups each group of 16 rats. Group A: served as control and received commercial ration and distilled water Group B: cadmium chloride was administered orally in water at dose of 300 ppm cadmium (560 mg/L as CdCl2). Group C: Animals received cadmium in drinking water in addition to administration of N-acetylcysteine (NAC) orally at a dose of 150 mg/kg body weight, equivalent to 1500 ppm in food. Group D: Animals received cadmium in drinking water in addition to administration of alpha lipoic acid (ALA) orally at a dose of 150 mg/kg body weight, equivalent to 1500 ppm in food. The experiment was continued for 2 months. Collection of blood and tissue samples was performed at 2, 4, 6, 8 weeks. Blood sample were collected for serum biochemical analysis including malondialdehyde (MDA), total antioxidants, aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein, albumin, urea and uric acid. Tissue specimens were collected for histopathological examination including liver, kidney, brain and testis. Histopathological examination revealed that cadmium choloride induces pathological alterations which increased in severity with time. The use of NAC and ALA can ameliorate toxic effect of CdCl2. The results showed significant decrease MDA and significant increase total antioxidants in group C and D compared to group B, Liver enzymes include AST and ALT showed significant decrease. Regarding to results of total protein and albumin, they revealed significant increase. Urea and uric acid showed significant decrease. From our study we conclude that NAC and ALA have protective effect against cadmium toxicity.

Keywords: ALA, cadmium, histopathology, NAC

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Authors: Selisha A. Sooklal, Phelelani Mpangase, Shaun Aron, Karl Rumbold

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Organically bound fluorine (C-F bond) is extremely rare in nature. Despite this, the first fluorinated secondary metabolite, fluoroacetate, was isolated from the plant Dichapetalum cymosum (commonly known as Gifblaar). However, the enzyme responsible for fluorination (fluorinase) in Gifblaar was never isolated and very little progress has been achieved in understanding this process in higher plants. Fluorinated compounds have vast applications in the pharmaceutical, agrochemical and fine chemicals industries. Consequently, an enzyme capable of catalysing a C-F bond has great potential as a biocatalyst in the industry considering that the field of fluorination is virtually synthetic. As with any biocatalyst, a range of these enzymes are required. Therefore, it is imperative to expand the exploration for novel fluorinases. This study aimed to gain molecular insights into secondary metabolite biosynthesis in Gifblaar using a high-throughput sequencing-based approach. Mechanical wounding studies were performed using Gifblaar leaf tissue in order to induce expression of the fluorinase. The transcriptome of the wounded and unwounded plant was then sequenced on the Illumina HiSeq platform. A total of 26.4 million short sequence reads were assembled into 77 845 transcripts using Trinity. Overall, 68.6 % of transcripts were annotated with gene identities using public databases (SwissProt, TrEMBL, GO, COG, Pfam, EC) with an E-value threshold of 1E-05. Sequences exhibited the greatest homology to the model plant, Arabidopsis thaliana (27 %). A total of 244 annotated transcripts were found to be differentially expressed between the wounded and unwounded plant. In addition, secondary metabolic pathways present in Gifblaar were successfully reconstructed using Pathway tools. Due to lack of genetic information for plant fluorinases, a transcript failed to be annotated as a fluorinating enzyme. Thus, a local database containing the 5 existing bacterial fluorinases was created. Fifteen transcripts having homology to partial regions of existing fluorinases were found. In efforts to obtain the full coding sequence of the Gifblaar fluorinase, primers were designed targeting the regions of homology and genome walking will be performed to amplify the unknown regions. This is the first genetic data available for Gifblaar. It has provided novel insights into the mechanisms of metabolite biosynthesis and will allow for the discovery of the first eukaryotic fluorinase.

Keywords: biocatalyst, fluorinase, gifblaar, transcriptome

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Authors: Agata M. Kołodziejczyk, Mateusz Harasymczuk, Pierre-Yves Girardin, Lucie Davidová

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Subjective time perception implies connection to cognitive functions, attention, memory and awareness, but a little is known about connections with homeostatic states of the body coordinated by circadian clock. In this paper, we present results from experimental study of subjective time perception in volunteers performing physical activity on treadmill in various phases of their circadian rhythms. Subjects were exposed to several time illusions simulated by programmed timing systems. This study brings better understanding for further improvement of of work quality in isolated areas. 

Keywords: biological clock, light, time illusions, treadmill

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Authors: S. Kudlacik-Kramarczyk, M. Kedzierska, B. Tyliszczak

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Recently, the continuous development of medicine and related sciences has been observed. Particular emphasis is directed on the development of biomaterials, i.e., non-toxic, biocompatible and biodegradable materials that may improve the effectiveness of treatment as well as the comfort of patients. This is particularly important in the case of cancer treatment. Currently, there are many methods of cancer treatment based primarily on chemotherapy and the surgical removal of the tumor, but it is worth noting that these therapies also cause many side effects. Among women, the most common cancer is breast cancer. It may be completely cured, but the consequence of treatment is partial or complete breast mastectomy and radiation therapy, which results in severe skin burns. The skin of the patient after radiation therapy is very burned, and therefore requires intensive care and high frequency of dressing changes. The traditional dressing adheres to the burn wounds and does not absorb adequate amount of exudate from injuries and the patient is forced to change the dressing every 2 hours. Therefore, the main purpose was to develop an innovative combination of dressing material with drug carriers that may be used in anti-cancer therapy. The innovation of this solution is the combination of these two products into one system, i.e., a transdermal system with the possibility of a controlled release of the drug- cytostatic. Besides, the possibility of modifying the hydrogel matrix with aloe vera juice provides this material with new features favorable from the point of view of healing processes of burn wounds resulting from the radiation therapy. In this study, hydrogel materials containing protein spheres with the active substance have been obtained as a result of photopolymerization process. The reaction mixture consisting of the protein (albumin) spheres incorporated with cytostatic, chitosan, adequate crosslinking agent and photoinitiator has been subjected to the UV radiation for 2 minutes. Prepared materials have been subjected to the numerous studies including the analysis of cytotoxicity using murine fibroblasts L929. Analysis was conducted based on the mitochondrial activity test (MTT reduction assay) which involves the determining the number of cells characterized by proper metabolism. Hydrogel materials obtained using different amount of crosslinking agents have been subjected to the cytotoxicity analysis. According to the standards, tested material is defined as cytotoxic when the viability of cells after 24 h incubation with this material is lower than 70%. In the research, hydrogel polymer materials containing protein spheres incorporated with the active substance, i.e. a cytostatic, have been developed. Such a dressing may support the treatment of cancer due to the content of the anti-cancer drug - cytostatic, and may also provide a soothing effect on the healing of the burn wounds resulted from the radiation therapy due to the content of aloe vera juice in the hydrogel matrix. Based on the conducted cytotoxicity studies, it may be concluded that the obtained materials do not adversely affect the tested cell lines, therefore they can be subjected to more advanced analyzes.

Keywords: hydrogel polymers, cytostatics, drug carriers, cytotoxicity

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Authors: Snehal D. Ganjave, Hardik Dodia, Avinash V. Sunder, Swati Madhu, Pramod P. Wangikar

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Batch cultivation of recombinant bacteria in shake flasks results in low cell density due to nutrient depletion. Previous protocols on high cell density cultivation in shake flasks have relied mainly on controlled release mechanisms and extended cultivation protocols. In the present work, we report an optimized fed-batch process for high cell density cultivation of recombinant E. coli BL21(DE3) for protein production. A cybernetic model-based, multi-objective optimization strategy was implemented to obtain the optimum operating variables to achieve maximum biomass and minimized substrate feed rate. A syringe pump was used to feed a mixture of glycerol and yeast extract into the shake flask. Preliminary experiments were conducted with online monitoring of dissolved oxygen (DO) and offline measurements of biomass and glycerol to estimate the model parameters. Multi-objective optimization was performed to obtain the pareto front surface. The selected optimized recipe was tested for a range of proteins that show different extent soluble expression in E. coli. These included eYFP and LkADH, which are largely expressed in soluble fractions, CbFDH and GcanADH , which are partially soluble, and human PDGF, which forms inclusion bodies. The biomass concentrations achieved in 24 h were in the range 19.9-21.5 g/L, while the model predicted value was 19.44 g/L. The process was successfully reproduced in a standard laboratory shake flask without online monitoring of DO and pH. The optimized fed-batch process showed significant improvement in both the biomass and protein production of the tested recombinant proteins compared to batch cultivation. The proposed process will have significant implications in the routine cultivation of E. coli for various applications.

Keywords: cybernetic model, E. coli, high cell density cultivation, multi-objective optimization

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Authors: Gwo-Hwa Wan, Tai-Ho Hung, Fen-Fang Chung, Wan-Ying Lee, Hui-Ching Yang

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Air pollutant exposure results in elevated concentrations of oxidative stress and inflammatory biomarkers in general populations. Increased concentrations of inflammatory biomarkers in pregnant women would be associated with preterm labor and low birth weight. To our best knowledge, the associations between air pollutants exposure and inflammation in pregnant women and fetuses are unknown, as well as their effects on fetal growth. This study aimed to evaluate the influences of outdoor air pollutants in northern Taiwan areas on the inflammatory biomarker (high sensitivity C-reactive protein, hs-CRP) concentration in the blood of healthy pregnant women and how the biomarker impacts fetal growth. In this study, 38 healthy pregnant women who are in their first trimester and live in northern Taiwan area were recruited from the Taipei Chang Gung Memorial Hospital. Personal characteristics and prenatal examination data (e.g., blood pressure) were obtained from recruited subjects. The concentrations of inflammatory mediators, hs-CRP, in the blood of healthy pregnant women were analyzed. Additionally, hourly data of air pollutants (PM10, SO2, NO2, O3, CO) concentrations were obtained from air quality monitoring stations in Taipei area, established by the Taiwan Environmental Protection Administration. The definition of lag 0 and lag 01 are the exposure to air pollutants on the day of blood withdrawal, and the average exposure to air pollutants one day before and on the day of blood withdrawal, respectively. The statistical analyses were conducted using SPSS software version 22.0 (SPSS, Inc., Chicago, IL, USA). This analytical result indicates that the healthy pregnant women aged between 28 and 42 years old. The body mass index before pregnancy averaged 21.51 (sd = 2.51) kg/m2. Around 90% of the pregnant women had never smoking habit, and 28.95% of them had allergic diseases. Approximately around 84% and 5.26% of the pregnant women worked at indoor and outdoor environments, respectively. The mean hematocrit level of the pregnant women was 37.10%, and the hemoglobin levels were ranged between 10.1 and 14.7 g/dL with 12.47 g/dL of mean value. The blood hs-CRP concentrations of healthy pregnant women in the first trimester ranged between 0.32 and 32.5 mg/L with 2.83 (sd = 5.69) mg/L of mean value. The blood hs-CRP concentrations were positively associated with ozone concentrations at lag 0-14 (r = 0.481, p = 0.017) in healthy pregnant women. Significant lag effects were identified in ozone at lag 0-14 with a positive excess concentration of blood hs-CRP.

Keywords: air pollutant, hs-CRP, pregnant woman, ozone, first trimester

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Authors: Gordana Ćetković, Vesna Tumbas Šaponjac, Jasna Čanadanović-Brunet, Sonja Đilas, Slađana Stajčić, Jelena Vulić, Mirjana Jakišić

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Despite the recommended amount of daily intake of fruits, the consumption in modern age remains very low. Therefore there is a need for delivering valuable phytochemicals into the human body through different foods by developing functional food products fortified with natural bioactive compounds from plant sources. Recently, a growing interest rises in exploiting the fruit and vegetable by-products as sources of phytochemicals such as polyphenols, carotenoids, vitamins etc. Cherry contain high amounts of polyphenols, which are known to display a wide range of biological activities like antioxidant, anti-inflammatory, antimicrobial or anti-carcinogenic activities, improvement of vision, induction of apoptosis and neuroprotective effects. Also, cherry pomace, a by-product in juice processing, can also be promising source of phenolic compounds. However, the application of polyphenols as food additives is limited because after extraction these compounds are susceptible to degradation. Microencapsulation is one of the alternative approaches to protect bioactive compounds from degradation during processing and storage. Freeze-drying is one of the most used microencapsulation methods for the protection of thermosensitive and unstable molecules. In this study sour cherry pomace was extracted with food-grade solvent (50% ethanol) to be suitable for application in products for human use. Extracted polyphenols have been concentrated and stabilized on whey (WP) and soy (SP) proteins. Encapsulation efficiency in SP was higher (94.90%), however not significantly (p<0.05) from the one in WP (90.10%). Storage properties of WP and SP encapsulate in terms of total polyphenols, anthocyanins and antioxidant activity was tested for 6 weeks. It was found that the retention of polyphenols after 6 weeks in WP and SP (67.33 and 69.30%, respectively) was similar. The content of anthocyanins has increased in WP (for 47.97%), while their content in SP has very slightly decreased (for 1.45%) after 6-week storage period. In accordance with anthocyanins the decrease in antioxidant activity in WP (87.78%) was higher than in SP (43.02%). According to the results obtained in this study, the technique reported herewith can be used for obtaining quality encapsulates for their further use as functional food additives, and, on the other hand, for fruit waste valorization.

Keywords: cherry pomace, microencapsulation, polyphenols, storage

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Authors: Talita Kato, Moises Grespan, Elza I. Ida, Massami Shimokomaki, Adriana L. Soares

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White Striping (WS) is becoming a concern for the poultry industry, as it affects the look of breast broiler chicken meat leading it to rejection by the consumers. It is characterized by the appearance of varying degrees of white striations on the Pectoralis major muscle surface following the direction of the muscle fiber. The etiology of this myopathy is still unknown, however it is suggested to be associated with increased weight gain rate and age of the bird, attributing the phenomenon to the genetically bird’s selection for efficiently higher meat production. The aim of this study was to evaluate the occurrence of Pectoralis major WS in a commercial plant in southern Brazil and its chemical characterization. The breast meat samples (n=660) from birds of 47 days of age, were classified as: Normal NG (no apparent white striations), Moderate MG (when the fillets present thin lines <1 mm) and Severe SG (white striations present ˃1 mm thick covering a large part of the fillet surface). Thirty samples (n = 10 for each level of severity) were analyzed for pH, color (L*, a*, b*), proximate chemical composition (moisture, protein, ash and lipids contents) and hydroxyproline in order to determine the collagen content. The results revealed the occurrence for NG group was 16.97%, 51.67% for MG group and 31.36% for SG group. Although the total protein content did not differ significantly, the collagen index was 42% higher in favor to SG in relation to NG. Also the lipid fraction was 27% higher for SG group. The NG presented the lowest values of the parameters L* and a* (P ≤ 0.05), as there was no white striations on its surface and highest b* value in SG, because of the maximum lipid contents. These results indicate there was a contribution of the SG muscle cells to oversynthesize connective tissue components on the muscle fascia. In conclusion, this study revealed a high incidence of White Striping on broiler commercial line in Brazil thus, there is a need to identify the causes of this abnormality in order to diminish or to eliminate it.

Keywords: collagen content, commercial line, pectoralis major muscle, proximate composition

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Authors: Magdy G. Asaad

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Solitons are among the most beneficial solutions for science and technology for their applicability in physical applications including plasma, energy transport along protein molecules, wave transport along poly-acetylene molecules, ocean waves, constructing optical communication systems, transmission of information through optical fibers and Josephson junctions. In this talk, we will apply the bilinear technique to generate a class of soliton solutions to the (3+1)-dimensional nonlinear soliton equation of Jimbo-Miwa type. Examples of the resulting soliton solutions are computed and a few solutions are plotted.

Keywords: Pfaffian solutions, N-soliton solutions, soliton equations, Jimbo-Miwa

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Authors: Myung-Gon Yoon, Jung-Ho Moon, Tae Kwon Ha

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This paper considers a robust stabilization problem of a single agent in a multi-agent consensus system composed of identical agents, when the network topology of the system is completely unknown. It is shown that the transfer function of an agent in a consensus system can be described as a multiplicative perturbation of the isolated agent transfer function in frequency domain. Applying known robust stabilization results, we present sufficient conditions for a robust stabilization of an agent against unknown network topology.

Keywords: single agent control, multi-agent system, transfer function, graph angle

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Authors: Tariku Nefo Duke

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Embelia is a genus of climbing shrubs in the family Myrsinaceae. Embelia schimperi is as important in traditional medicine as the other species in the genus. The plant has been much known as a local medicine for the treatment of tapeworms. In this project, extraction, phytochemical screening tests, isolation, and characterization of berries of the Embelia schimperi plant have been conducted. The chemical investigations of methanol and ethyl acetate (1:1) ratio extracts of the berries lead to the isolation of three new compounds. The compounds were identified to be alkaloids coded as AD, AN, and AG. Structural elucidations of the isolated compounds were accomplished using spectroscopic methods (IR, UV, ¹H NMR, ¹³C NMR, DEPT and 2D NMR, HPLC, and LC-MS). The alkaloid coded as (AN) has a wide MIC range of 6.31-25.46 mg/mL against all tested bacteria strains.

Keywords: Embelia schimper, HPLC, alkaloids, 2D NMR, MIC

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Authors: Shivani Khare

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It is of paramount concern for economists to uncover the factors that determine human capital development, considered now to be one of the major factors behind economic growth and development. However, no human action is isolated but rather works within the set-up of the society. In recent years, a new field of investigation has come up that analyses the relationships that exist between social and human capital. Along these lines, this paper explores the effect of social capital on the indicators of human capital development – life expectancy at birth, mean years of schooling, and per capita income. The applied part of the analysis is performed using a panel data model for OECD countries and by using a series of chronological periods that within the 2005–2020 time frame.

Keywords: social capital, human capital development, trust, social networks, socioeconomics

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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

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Authors: Shabnam Abdi, Behzad Toosi

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Background: Melanoma is the most lethal type of malignant skin cancer in humans and dogs since it spreads rapidly throughout the body. Despite significant advances in treatment, cancer at an advanced stage has a poor prognosis. Hence, more effective treatments are needed to enhance outcomes with fewer side effects. Erythropoietin-producing hepatocellular receptors are the largest family of receptor tyrosine kinases and are divided into two subfamilies, EphA and EphB, both of which play a significant role in disease, especially cancer. Due to their association with proliferation and invasion in many aggressive types of cancer, Eph receptor tyrosine kinases (Eph RTKs) are promising cancer therapy molecules. Because these receptors have not been studied in canine melanoma, we investigated how EphA2 influences survival and tumorigenicity of melanoma cells. Methods: Expression of EphA2 protein in canine melanoma cell lines and human melanoma cell line was evaluated by Western blot. Melanoma cells were transduced with lentiviral particles encoding Eph-targeting shRNAs or non-silencing shRNAs (control) for silencing the expression of EphA2 receptor, and silencing was confirmed by Western blotting and immunofluorescence. The effect of siRNA treatment on cellular proliferation, colony formation, tumorsphere assay, invasion was analyzed by Resazurin assay Matrigel invasion assay, respectively. Results: Expression of EphA2 was detected in canine and human melanoma cell lines. Moreover, stably silencing EphA2 by specific shRNAs significantly and consistently decreased the expression of EphA2 protein in both human and canine melanoma cells. Proliferation, colony formation, tumorsphere and invasion of melanoma cells were significantly decreased in EphA2 siRNA-treated cells compared to control. Conclusion: Our data provide the first functional evidence that the EphA2 receptor plays a critical role in the malignant cellular behavior of melanoma in both human and dogs.

Keywords: ephA2, targeting, melanoma, human, canine

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Authors: Sushil Kumar Singh, Kumar Rahul, Vivek Tewarson, Sarvesh Kumar, Shobhit Kumar

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Objective: Rheumatic mitral stenosis continues to be a major public health problem in developing countries. When the left atrium (LA) is unable to fill the left ventricle (LV) at normal LA pressures due to impaired relaxation and impaired compliance, diastolic dysfunction occurs. The assessment of left ventricular (LV) diastolic function and filling pressures is of clinical importance to identify underlying cardiac disease, its treatment, and to assess prognosis. 2D echocardiography can detect diastolic dysfunction with excellent sensitivity and minimal risk when compared to the gold standard of invasive pressure-volume measurements. Material and Method: This was a one-year study consisting of twenty-nine patients of isolated rheumatic severe mitral stenosis. Data was analyzed preoperative and post operative (at one month follow-up). Transthoracic 2D echocardiographic parameters of the diastolic function are transmitral flow, pulmonary venous flow, mitral annular tissue doppler, and color M-mode doppler. In our study, mitral valve orifice area, ejection fraction, deceleration time, E/A-wave, E/E’-wave, myocardial performance index of left ventricle (Tei index ), and Mitral inflow propagation velocity were included for echocardiographic evaluation. The statistical analysis was performed on SPSS Version 15.0 statistical analysis software. Result: Twenty-nine patients underwent successful closed mitral commissurotomy for isolated mitral stenosis. The outcome measures were observed pre-operatively and at one-month follow-up. The majority of patients were in NYHA grade III (69.0%) in the preoperative period, which improved to NYHA grade I (48.3%) after closed mitral commissurotomy. Post-surgery mitral valve area increased from 0.77 ± 0.13 to 2.32 ± 0.26 cm, ejection fraction increased from 61.38 ± 4.61 to 64.79 ± 3.22. There was a decrease in deceleration time from 231.55 ± 49.31 to 168.28 ± 14.30 ms, E/A ratio from 1.70 ± 0.54 from 0.89 ± 0.39, E/E’ ratio from 14.59 ± 3.34 to 8.86 ± 3.03. In addition, there was improvement in TIE index from 0.50 ± 0.03 to 0.39 ± 0.06 and mitral inflow propagation velocity from 47.28 ± 3.71 to 57.86 ± 3.19 cm/sec. In peri-operative and follow-up, there was no incidence of severe mitral regurgitation (MR). There was no thromboembolic incident and no mortality.

Keywords: closed mitral valvotomy, mitral stenosis, open mitral commissurotomy, balloon mitral valvotomy

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Authors: Supriya Chatla, Anjana Male, Srikala Kamireddy

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L-asparaginase (E.C.3.5.1.1) is an anti-cancer enzyme that has been purified and characterized for decades to study and evaluate its anti-carcinogenic activity against Hodgkin’s lymphoma. The present investigation deals with screening, isolation and optimization of L-asparaginase giving fungal strain of soil samples from different areas of AP, India. L-Aspariginase activity was estimated on the basis of the pink color surrounding the growing colony. A total of 132 colonies were screened and isolated from different samples. Based on the zone diameter, L-asparaginase activity is determined, L- asparaginase activity is optimized at 28oc and Immobilized Aspariginase had more potency than the free enzymes.

Keywords: aspariginase, anticancer enzyme, Isolation, optimization

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Authors: Irena Kostovska, Katerina Tosheska Trajkovska, Svetlana Cekovska, Julijana Brezovska Kavrakova, Hristina Ampova, Sonja Topuzovska, Ognen Kostovski, Goce Spasovski, Danica Labudovic

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Background: Proteinuria is an important feature of secondary nephropathies. The quantitative and qualitative analysis of proteinuria plays an important role in determining the types of proteinuria (glomerular, tubular and mixed), in the diagnosis and prognosis of secondary nephropathies. The damage of the glomerular basement membrane is responsible for a proteinuria characterized by the presence of large amounts of protein with high molecular weights such as albumin (69 kilo Daltons-kD), transferrin (78 kD) and immunoglobulin G (150 kD). An insufficiency of proximal tubular function is the cause of a proteinuria characterized by the presence of proteins with low molecular weight (LMW), such as retinol binding protein (21 kD) and α1-microglobulin (31 kD). In some renal diseases, a mixed glomerular and tubular proteinuria is frequently seen. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) is the most widely used method of analyzing urine proteins for clinical purposes. The main aim of the study is to determine the type of proteinuria in the most common secondary nephropathies such as diabetic, hypertensive nephropathy and preeclampsia. Material and methods: In this study were included 90 subjects: subjects with diabetic nephropathy (n=30), subjects with hypertensive nephropahty (n=30) and pregnant women with preeclampsia (n=30). We divided all subjects according to UM/CR into three subgroups: macroalbuminuric (UM/CR >300 mg/g), microalbuminuric (UM/CR 30-300 mg/g) and normolabuminuric (UM/CR<30 mg/g). In all subjects, we measured microalbumin and creatinine in urine with standard biochemical methods. Separation of urinary proteins was performed by SDS-PAGE, in several stages: linear gel preparation (4-22%), treatment of urinary samples before their application on the gel, electrophoresis, gel fixation, coloring with Coomassie blue, and identification of the separated protein fractions based on standards with exactly known molecular weight. Results: According to urinary microalbumin/creatinin ratio in group of subject with diabetic nephropathy, nine patients were macroalbuminuric, while 21 subject were microalbuminuric. In group of subjects with hypertensive nephropathy, we found macroalbuminuria (n=4), microalbuminuria (n=20) and normoalbuminuria (n=6). All pregnant women with preeclampsia were macroalbuminuric. Electrophoretic separation of urinary proteins showed that in macroalbuminric patients with diabetic nephropathy 56% have mixed proteinuria, 22% have glomerular proteinuria and 22% have tubular proteinuria. In subgroup of subjects with diabetic nephropathy and microalbuminuria, 52% have glomerular proteinuria, 8% have tubular proteinuria, and 40% of subjects have normal electrophoretic findings. All patients with maroalbuminuria and hypertensive nephropathy have mixed proteinuria. In subgroup of patients with microalbuminuria and hypertensive nephropathy, we found: 32% with mixed proteinuria, 27% with normal findings, 23% with tubular, and 18% with glomerular proteinuria. In all normoalbuminruic patiens with hypertensive nephropathy, we detected normal electrophoretic findings. In group of subjects pregnant women with preeclampsia, we found: 81% with mixed proteinuria, 13% with glomerular, and 8% with tubular proteinuria. Conclusion: By SDS PAGE method, we detected that in patients with secondary nephropathies the most common type of proteinuria is mixed proteinuria, indicating both loss of glomerular permeability and tubular function. We can conclude that SDS PAGE is high sensitive method for detection of renal impairment in patients with secondary nephropathies.

Keywords: diabetic nephropathy, preeclampsia, hypertensive nephropathy, SDS PAGE

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Authors: Nawras Kurzom, Avi Mendelsohn

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The effects of background music on learning and memory are inconsistent, partly due to the intrinsic complexity and variety of music and partly to individual differences in music perception and preference. A prominent musical feature that is known to elicit strong emotional responses is musical tension. Musical tension can be brought about by building anticipation of rhythm, harmony, melody, and dynamics. Delaying the resolution of dominant-to-tonic chord progressions, as well as using dissonant harmonics, can elicit feelings of tension, which can, in turn, affect memory formation of concomitant information. The aim of the presented studies was to explore how forming declarative memory is influenced by musical tension, brought about within continuous music as well as in the form of isolated chords with varying degrees of dissonance/consonance. The effects of musical tension on long-term memory of declarative information were studied in two ways: 1) by evoking tension within continuous music pieces by delaying the release of harmonic progressions from dominant to tonic chords, and 2) by using isolated single complex chords with various degrees of dissonance/roughness. Musical tension was validated through subjective reports of tension, as well as physiological measurements of skin conductance response (SCR) and pupil dilation responses to the chords. In addition, music information retrieval (MIR) was used to quantify musical properties associated with tension and its release. Each experiment included an encoding phase, wherein individuals studied stimuli (words or images) with different musical conditions. Memory for the studied stimuli was tested 24 hours later via recognition tasks. In three separate experiments, we found positive relationships between tension perception and physiological measurements of SCR and pupil dilation. As for memory performance, we found that background music, in general, led to superior memory performance as compared to silence. We detected a trade-off effect between tension perception and memory, such that individuals who perceived musical tension as such displayed reduced memory performance for images encoded during musical tension, whereas tense music benefited memory for those who were less sensitive to the perception of musical tension. Musical tension exerts complex interactions with perception, emotional responses, and cognitive performance on individuals with and without musical training. Delineating the conditions and mechanisms that underlie the interactions between musical tension and memory can benefit our understanding of musical perception at large and the diverse effects that music has on ongoing processing of declarative information.

Keywords: musical tension, declarative memory, learning and memory, musical perception

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