Search results for: crystal column

Authors: Utpal Das, Sona Das

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InAs/GaSb type-II superlattice (T2SL) Mid-wave infrared (MWIR) focal plane arrays (FPAs) have recently seen rapid development. However, in small pixel size large format FPAs, the occurrence of high mesa sidewall surface leakage current is a major constraint necessitating proper surface passivation. A simple pixel isolation technique in InAs/GaSb T2SL detector arrays without the conventional mesa etching has been proposed to isolate the pixels by forming a more resistive higher band gap material from the SL, in the inter-pixel region. Here, a single step femtosecond (fs) laser anneal of the T2SL structure of the inter-pixel T2SL regions, have been used to increase the band gap between the pixels by QW-intermixing and hence increase isolation between the pixels. The p-i-n photodiode structure used here consists of a 506nm, (10 monolayer {ML}) InAs:Si (1x10¹⁸cm⁻³)/(10ML) GaSb SL as the bottom n-contact layer grown on an n-type GaSb substrate. The undoped absorber layer consists of 1.3µm, (10ML)InAs/(10ML)GaSb SL. The top p-contact layer is a 63nm, (10ML)InAs:Be(1x10¹⁸cm⁻³)/(10ML)GaSb T2SL. In order to improve the carrier transport, a 126nm of graded doped (10ML)InAs/(10ML)GaSb SL layer was added between the absorber and each contact layers. A 775nm 150fs-laser at a fluence of ~6mJ/cm² is used to expose the array where the pixel regions are masked by a Ti(200nm)-Au(300nm) cap. Here, in the inter-pixel regions, the p+ layer have been reactive ion etched (RIE) using CH₄+H₂ chemistry and removed before fs-laser exposure. The fs-laser anneal isolation improvement in 200-400μm pixels due to spatially selective quantum well intermixing for a blue shift of ~70meV in the inter-pixel regions is confirmed by FTIR measurements. Dark currents are measured between two adjacent pixels with the Ti(200nm)-Au(300nm) caps used as contacts. The T2SL quality in the active photodiode regions masked by the Ti-Au cap is hardly affected and retains the original quality of the detector. Although, fs-laser anneal of p+ only etched p-i-n T2SL diodes show a reduction in the reverse dark current, no significant improvement in the full RIE-etched mesa structures is noticeable. Hence for a 128x128 array fabrication of 8μm square pixels and 10µm pitch, SU8 polymer isolation after RIE pixel delineation has been used. X-n+ row contacts and Y-p+ column contacts have been used to measure the optical response of the individual pixels. The photo-response of these 8μm and other 200μm pixels under a 2ns optical pulse excitation from an Optical-Parametric-Oscillator (OPO), shows a peak responsivity of ~0.03A/W and 0.2mA/W, respectively, at λ~3.7μm. Temporal response of this detector array is seen to have a fast response ~10ns followed typical slow decay with ringing, attributed to impedance mismatch of the connecting co-axial cables. In conclusion, response times of a few ns have been measured in 8µm pixels of a 128x128 array. Although fs-laser anneal has been found to be useful in increasing the inter-pixel isolation in InAs/GaSb T2SL arrays by QW inter-mixing, it has not been found to be suitable for passivation of full RIE etched mesa structures with vertical walls on InAs/GaSb T2SL.

Keywords: band-gap blue-shift, fs-laser-anneal, InAs/GaSb T2SL, Inter-pixel isolation, ns-Response, photodiode array

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Authors: Renata Gerhardt, Detlev Belder

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Raman spectroscopy has attracted much attention as a structurally descriptive and label-free detection method. It is particularly suited for chemical analysis given as it is non-destructive and molecules can be identified via the fingerprint region of the spectra. In this work possibilities are investigated how to integrate Raman spectroscopy as a detection method for chip-based chromatography, making use of a droplet interface. A demanding task in lab-on-a-chip applications is the specific and sensitive detection of low concentrated analytes in small volumes. Fluorescence detection is frequently utilized but restricted to fluorescent molecules. Furthermore, no structural information is provided. Another often applied technique is mass spectrometry which enables the identification of molecules based on their mass to charge ratio. Additionally, the obtained fragmentation pattern gives insight into the chemical structure. However, it is only applicable as an end-of-the-line detection because analytes are destroyed during measurements. In contrast to mass spectrometry, Raman spectroscopy can be applied on-chip and substances can be processed further downstream after detection. A major drawback of Raman spectroscopy is the inherent weakness of the Raman signal, which is due to the small cross-sections associated with the scattering process. Enhancement techniques, such as surface enhanced Raman spectroscopy (SERS), are employed to overcome the poor sensitivity even allowing detection on a single molecule level. In SERS measurements, Raman signal intensity is improved by several orders of magnitude if the analyte is in close proximity to nanostructured metal surfaces or nanoparticles. The main gain of lab-on-a-chip technology is the building block-like ability to seamlessly integrate different functionalities, such as synthesis, separation, derivatization and detection on a single device. We intend to utilize this powerful toolbox to realize Raman detection in chip-based chromatography. By interfacing on-chip separations with a droplet generator, the separated analytes are encapsulated into numerous discrete containers. These droplets can then be injected with a silver nanoparticle solution and investigated via Raman spectroscopy. Droplet microfluidics is a sub-discipline of microfluidics which instead of a continuous flow operates with the segmented flow. Segmented flow is created by merging two immiscible phases (usually an aqueous phase and oil) thus forming small discrete volumes of one phase in the carrier phase. The study surveys different chip designs to realize coupling of chip-based chromatography with droplet microfluidics. With regards to maintaining a sufficient flow rate for chromatographic separation and ensuring stable eluent flow over the column different flow rates of eluent and oil phase are tested. Furthermore, the detection of analytes in droplets with surface enhanced Raman spectroscopy is examined. The compartmentalization of separated compounds preserves the analytical resolution since the continuous phase restricts dispersion between the droplets. The droplets are ideal vessels for the insertion of silver colloids thus making use of the surface enhancement effect and improving the sensitivity of the detection. The long-term goal of this work is the first realization of coupling chip based chromatography with droplets microfluidics to employ surface enhanced Raman spectroscopy as means of detection.

Keywords: chip-based separation, chip LC, droplets, Raman spectroscopy, SERS

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Authors: Zakia Fatima, Liu Lu, Donghao Li

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The analysis of biological fluid metabolites holds significant importance in various areas, such as medical research, food science, and public health. Investigating the levels and distribution of nutrients and their metabolites in biological samples allows researchers and healthcare professionals to determine nutritional status, find hypovitaminosis or hypervitaminosis, and monitor the effectiveness of interventions such as dietary supplementation. Moreover, analysis of nutrient metabolites provides insight into their metabolism, bioavailability, and physiological processes, aiding in the clarification of their health roles. Hence, the exploration of a distinct, efficient, eco-friendly, and simpler methodology is of great importance to evaluate the metabolic content of complex biological samples. In this work, a green and rapid analytical method based on an automated online two-dimensional microscale carbon fiber/activated carbon fiber fractionation system and time-of-flight mass spectrometry (2DμCFs-TOF-MS) was used to evaluate metabolites of urine samples after oral administration of food supplements. The automated 2DμCFs instrument consisted of a microcolumn system with bare carbon fibers and modified carbon fiber coatings. Carbon fibers and modified carbon fibers exhibit different surface characteristics and retain different compounds accordingly. Three kinds of mobile-phase solvents were used to elute the compounds of varied chemical heterogeneities. The 2DμCFs separation system has the ability to effectively separate different compounds based on their polarity and solubility characteristics. No complicated sample preparation method was used prior to analysis, which makes the strategy more eco-friendly, practical, and faster than traditional analysis methods. For optimum analysis results, mobile phase composition, flow rate, and sample diluent were optimized. Water-soluble vitamins, fat-soluble vitamins, and amino acids, as well as 22 vitamin metabolites and 11 vitamin metabolic pathway-related metabolites, were found in urine samples. All water-soluble vitamins except vitamin B12 and vitamin B9 were detected in urine samples. However, no fat-soluble vitamin was detected, and only one metabolite of Vitamin A was found. The comparison with a blank urine sample showed a considerable difference in metabolite content. For example, vitamin metabolites and three related metabolites were not detected in blank urine. The complete single-run screening was carried out in 5.5 minutes with the minimum consumption of toxic organic solvent (0.5 ml). The analytical method was evaluated in terms of greenness, with an analytical greenness (AGREE) score of 0.72. The method’s practicality has been investigated using the Blue Applicability Grade Index (BAGI) tool, obtaining a score of 77. The findings in this work illustrated that the 2DµCFs-TOF-MS approach could emerge as a fast, sustainable, practical, high-throughput, and promising analytical tool for screening and accurate detection of various metabolites, pharmaceuticals, and ingredients in dietary supplements as well as biological fluids.

Keywords: metabolite analysis, sustainability, carbon fibers, urine.

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Authors: S. Mamoucha, N.Tsafantakis, T. Ioannidis, S. Chatzipanagiotou, C. Nikolaou, L. Skaltsounis, N. Fokialakis, N. Christodoulakis

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Introduction: Pistacia lentiscus L. (well known as Mastic tree) is an evergreen sclerophyllous shrub that extensively thrives in the eastern Mediterranean area yet only the trees cultivated in the southern region of the Greek island Chios produces mastic resin. Different parts of P. lentiscus L. var. chia have been used in folk medicine for various purposes, such as tonic, aphrodisiac, antiseptic, antihypertensive and management of dental, gastrointestinal, liver, urinary, and respiratory tract disorders. Several studies have focused on the antibacterial activity of its resin (gum) and its essential oil. However, there is no study combining anatomy of the plant organs, phytochemical profile, and antibacterial screening of the plant. In our attempt to discover novel bioactive metabolites from the mastic tree, we screened its antibacterial activity not only against ATCC strains but also against clinical, resistant strains. Materials-methods: Leaves were investigated using Transmission (ΤΕΜ) and Scanning Εlectron Microscopy (SEM). Histochemical tests were performed on fresh and fixed tissue. Extracts prepared from dried, powdered leaves using 3 different solvents (DCM, MeOH and H2O) the waste water obtained after a hydrodistillation process for essential oil production were screened for their phytochemical content and antibacterial activity. Μetabolite profiling of polar and non-polar extracts was recorded by GC-MS and LC-HRMS techniques and analyzed using in-house and commercial libraries. The antibacterial screening was performed against Staphylococcus aureus ATCC25923, Escherichia coli ATCC25922, Pseudomonas aeruginosa ATCC27853 and against clinical, resistant strains Methicillin-resistant S. aureus (MRSA), Carbapenem-Resistant Metallo-β-Lactamase (carbapenemase) P. aeruginosa (VIM), Klebsiella pneumoniae carbapenemases (KPCs) and Acinetobacter baumanii resistant strains. The antibacterial activity was tested by the Kirby Bauer and the Agar Well Diffusion method. The zone of inhibition (ZI) of each extract was measured and compared with those of common antibiotics. Results: Leaf is compact with inosclereids and numerous idioblasts containing a globular, spiny crystal. The major nerves of the leaf contain a resin duct. Mesophyll cells showed accumulation of osmiophillic metabolites. Histochemical treatments defined secondary metabolites in subcellular localization. The phytochemical investigation revealed the presence of a large number of secondary metabolites, belonging to different chemical groups, such as terpenoids, phenolic compounds (mainly myricetin, kaempferol and quercetin glycosides), phenolic, and fatty acids. Among the extracts, the hydrostillation wastewater achieved the best results against most of the bacteria tested. MRSA, VIM and A. baumanii were inhibited. Conclusion: Extracts from plants have recently been of great interest with respect to their antimicrobial activity. Their use emerged from a growing tendency to replace synthetic antimicrobial agents with natural ones. Leaves of P. lentiscus L. var. chia showed a high antimicrobial activity even against drug - resistant bacteria. Future prospects concern the better understanding of mode of action of the antibacterial activity, the isolation of the most bioactive constituents and the clarification if the activity is related to a single compound or to the synergistic effect of several ones.

Keywords: antibacterial screening, leaf anatomy, phytochemical profile, Pistacia lentiscus var. chia

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Authors: Emilie Gout, Mathias Monnot, Olivier Boutin, Pierre Vanloot, Philippe Moulin

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Water is a precious resource. Treating industrial wastewater remains a considerable technical challenge of our century. The effluent considered for this study is landfill leachate treated by reverse osmosis (RO). Nowadays, in most developed countries, sanitary landfilling is the main method to deal with municipal solid waste. Rainwater percolates through solid waste, generating leachates mostly comprised of organic and inorganic matter. Whilst leachate ages, its composition varies, becoming more and more bio-refractory. RO is already used for landfill leachates as it generates good quality permeate. However, its mains drawback is the production of highly polluted concentrates that cannot be discharged in the environment or reused, which is an important industrial issue. It is against this background that the study of coupling RO with wet air oxidation (WAO) was set to intensify and optimize processes to meet current regulations for water discharge in the environment. WAO is widely studied for effluents containing bio-refractory compounds. Oxidation consists of a destruction reaction capable of mineralizing the recalcitrant organic fraction of pollution into carbon dioxide and water when complete. WAO process in subcritical conditions requires a high-energy consumption, but it can be autothermic in a certain range of chemical oxygen demand (COD) concentrations (10-100 g.L⁻¹). Appropriate COD concentrations are reached in landfill leachate RO concentrates. Therefore, the purpose of this work is to report the performances of mineralization during WAO on RO concentrates. The coupling of RO/WAO has shown promising results in previous works on both synthetic and real effluents in terms of organic carbon (TOC) reduction by WAO and retention by RO. Non-catalytic WAO with air as oxidizer was performed in a lab-scale stirred autoclave (1 L) on landfill leachates RO concentrates collected in different seasons in a sanitary landfill in southern France. The yield of WAO depends on operating parameters such as total pressure, temperature, and time. Compositions of the effluent are also important aspects for process intensification. An experimental design methodology was used to minimize the number of experiments whilst finding the operating conditions achieving the best pollution reduction. The simulation led to a set of 18 experiments, and the responses to highlight process efficiency are pH, conductivity, turbidity, COD, TOC, and inorganic carbon. A 70% oxygen excess was chosen for all the experiments. First experiments showed that COD and TOC abatements of at least 70% were obtained after 90 min at 300°C and 20 MPa, which attested the possibility to treat RO leachate concentrates with WAO. In order to meet French regulations and validate process intensification with industrial effluents, some continuous experiments in a bubble column are foreseen, and some further analyses will be performed, such as biological oxygen demand and study of gas composition. Meanwhile, other industrial effluents are treated to compare RO-WAO performances. These effluents, coming from pharmaceutical, petrochemical, and tertiary wastewater industries, present different specific pollutants that will provide a better comprehension of the hybrid process and prove the intensification and feasibility of the process at an industrial scale. Acknowledgments: This work has been supported by the French National Research Agency (ANR) for the Project TEMPO under the reference number ANR-19-CE04-0002-01.

Keywords: hybrid process, landfill leachates, process intensification, reverse osmosis, wet air oxidation

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Authors: Abderrazak Bannari

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Acid Mine Drainage (AMD) from mine wastes and contaminations of soils and water with metals are considered as a major environmental problem in mining areas. It is produced by interactions of water, air, and sulphidic mine wastes. This environment problem results from a series of chemical and biochemical oxidation reactions of sulfide minerals e.g. pyrite and pyrrhotite. These reactions lead to acidity as well as the dissolution of toxic and heavy metals (Fe, Mn, Cu, etc.) from tailings waste rock piles, and open pits. Soil and aquatic ecosystems could be contaminated and, consequently, human health and wildlife will be affected. Furthermore, secondary minerals, typically formed during weathering of mine waste storage areas when the concentration of soluble constituents exceeds the corresponding solubility product, are also important. The most common secondary mineral compositions are hydrous iron oxide (goethite, etc.) and hydrated iron sulfate (jarosite, etc.). The objectives of this study focus on the detection and mapping of MIOP in the soil using Hyperion EO-1 (Earth Observing - 1) hyperspectral data and constrained linear spectral mixture analysis (CLSMA) algorithm. The abandoned Kettara mine, located approximately 35 km northwest of Marrakech city (Morocco) was chosen as study area. During 44 years (from 1938 to 1981) this mine was exploited for iron oxide and iron sulphide minerals. Previous studies have shown that Kettara surrounding soils are contaminated by heavy metals (Fe, Cu, etc.) as well as by secondary minerals. To achieve our objectives, several soil samples representing different MIOP classes have been resampled and located using accurate GPS ( ≤ ± 30 cm). Then, endmembers spectra were acquired over each sample using an Analytical Spectral Device (ASD) covering the spectral domain from 350 to 2500 nm. Considering each soil sample separately, the average of forty spectra was resampled and convolved using Gaussian response profiles to match the bandwidths and the band centers of the Hyperion sensor. Moreover, the MIOP content in each sample was estimated by geochemical analyses in the laboratory, and a ground truth map was generated using simple Kriging in GIS environment for validation purposes. The acquired and used Hyperion data were corrected for a spatial shift between the VNIR and SWIR detectors, striping, dead column, noise, and gain and offset errors. Then, atmospherically corrected using the MODTRAN 4.2 radiative transfer code, and transformed to surface reflectance, corrected for sensor smile (1-3 nm shift in VNIR and SWIR), and post-processed to remove residual errors. Finally, geometric distortions and relief displacement effects were corrected using a digital elevation model. The MIOP fraction map was extracted using CLSMA considering the entire spectral range (427-2355 nm), and validated by reference to the ground truth map generated by Kriging. The obtained results show the promising potential of the proposed methodology for the detection and mapping of mine iron oxide pollution in the soil.

Keywords: hyperion eo-1, hyperspectral, mine iron oxide pollution, environmental impact, unmixing

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Authors: Adela Diepoltova, Klara Konecna, Ondrej Jandourek, Petr Nachtigal

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A loss of control over antibiotic-resistant pathogens has become a global issue due to severe and often untreatable infections. This state is reflected in complicated treatment, health costs, and higher mortality. All these factors emphasize the urgent need for the discovery and development of new anti-infectives. One of the most common pathogens mentioned in the phenomenon of antibiotic resistance are bacteria of the genus Staphylococcus. These bacterial agents have developed several mechanisms against the effect of antibiotics. One of them is biofilm formation. In staphylococci, biofilms are associated with infections such as endocarditis, osteomyelitis, catheter-related bloodstream infections, etc. To author's best knowledge, no validated and standardized methodology evaluating candidate compound activity against staphylococcal biofilms exists. However, a variety of protocols for in vitro drug activity testing has been suggested, yet there are often fundamental differences. Based on our experience, a key methodological step that leads to credible results is to form a robust biofilm with appropriate attributes such as firm adherence to the substrate, a complex arrangement in layers, and the presence of extracellular polysaccharide matrix. At first, for the purpose of drug antibiofilm activity evaluation, the focus was put on various conditions (supplementation of cultivation media by human plasma/fetal bovine serum, shaking mode, the density of initial inoculum) that should lead to reproducible and robust in vitro staphylococcal biofilm formation in microtiter plate model. Three model staphylococcal reference strains were included in the study: Staphylococcus aureus (ATCC 29213), methicillin-resistant Staphylococcus aureus (ATCC 43300), and Staphylococcus epidermidis (ATCC 35983). The total biofilm biomass was quantified using the Christensen method with crystal violet, and results obtained from at least three independent experiments were statistically processed. Attention was also paid to the viability of the biofilm-forming staphylococcal cells and the presence of extracellular polysaccharide matrix. The conditions that led to robust biofilm biomass formation with attributes for biofilms mentioned above were then applied by introducing an alternative method analogous to the commercially available test system, the Calgary Biofilm Device. In this test system, biofilms are formed on pegs that are incorporated into the lid of the microtiter plate. This system provides several advantages (in situ detection and quantification of biofilm microbial cells that have retained their viability after drug exposure). Based on our preliminary studies, it was found that the attention to the peg surface and substrate on which the bacterial biofilms are formed should also be paid to. Therefore, further steps leading to the optimization were introduced. The surface of pegs was coated by human plasma, fetal bovine serum, and L-polylysine. Subsequently, the willingness of bacteria to adhere and form biofilm was monitored. In conclusion, suitable conditions were revealed, leading to the formation of reproducible, robust staphylococcal biofilms in vitro for the microtiter model and the system analogous to the Calgary biofilm device, as well. The robustness and typical slime texture could be detected visually. Likewise, an analysis by confocal laser scanning microscopy revealed a complex three-dimensional arrangement of biofilm forming organisms surrounded by an extracellular polysaccharide matrix.

Keywords: anti-biofilm drug activity screening, in vitro biofilm formation, microtiter plate model, the Calgary biofilm device, staphylococcal infections, substrate modification, surface coating

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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: Sajeli A. Begum, Ameer Basha, Kirti Hira, Rukaiyya Khan

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Cyclic dipeptides are simple diketopiperazine derivatives being investigated by several scientists for their biological effects which include anticancer, antimicrobial, haematological, anticonvulsant, immunomodulatory effect, etc. They are potentially active microbial metabolites having been synthesized too, for developing into drug candidates. Cultures of Pseudomonas species have earlier been reported to produce cyclic dipeptides, helping in quorum sensing signals and bacterial–host colonization phenomena during infections, causing cell anti-proliferation and immunosuppression. Fluorescing Pseudomonas species have been identified to secrete lipid derivatives, peptides, pyrroles, phenazines, indoles, aminoacids, pterines, pseudomonic acids and some antibiotics. In the present work, results of investigation on the cyclic dipeptide metabolites secreted by the culture broth of Pseudomonas species as potent pro-inflammatory cytokine inhibitors are discussed. The bacterial strain was isolated from the rhizospheric soil of groundnut crop and identified as Pseudomonas aeruginosa by 16S rDNA sequence (GenBank Accession No. KT625586). Culture broth of this strain was prepared by inoculating into King’s B broth and incubating at 30 ºC for 7 days. The ethyl acetate extract of culture broth was prepared and lyophilized to get a dry residue (EEPA). Lipopolysaccharide (LPS)-induced ELISA assay proved the inhibition of tumor necrosis factor-alpha (TNF-α) secretion in culture supernatant of RAW 264.7 cells by EEPA (IC50 38.8 μg/mL). The effect of oral administration of EEPA on plasma TNF-α level in rats was tested by ELISA kit. The LPS mediated plasma TNF-α level was reduced to 45% with 125 mg/kg dose of EEPA. Isolation of the chemical constituents of EEPA through column chromatography yielded ten cyclic dipeptides, which were characterized using nuclear magnetic resonance and mass spectroscopic techniques. These cyclic dipeptides are biosynthesized in microorganisms by multifunctional assembly of non-ribosomal peptide synthases and cyclic dipeptide synthase. Cyclo (Gly-L-Pro) was found to be more potentially (IC50 value 4.5 μg/mL) inhibiting TNF-α production followed by cyclo (trans-4-hydroxy-L-Pro-L-Phe) (IC50 value 14.2 μg/mL) and the effect was equal to that of standard immunosuppressant drug, prednisolone. Further, the effect was analyzed by determining mRNA expression of TNF-α in LPS-stimulated RAW 264.7 macrophages using quantitative real-time reverse transcription polymerase chain reaction. EEPA and isolated cyclic dipeptides demonstrated diminution of TNF-α mRNA expression levels in a dose-dependent manner under the tested conditions. Also, they were found to control the expression of other pro-inflammatory cytokines like IL-1β and IL-6, when tested through their mRNA expression levels in LPS-stimulated RAW 264.7 macrophages under LPS-stimulated conditions. In addition, significant inhibition effect was found on Nitric oxide production. Further all the compounds exhibited weak toxicity to LPS-induced RAW 264.7 cells. Thus the outcome of the study disclosed the effectiveness of EEPA and the isolated cyclic dipeptides in down-regulating key cytokines involved in pathophysiology of autoimmune diseases.In another study led by the investigators, microbial cyclic dipeptides were found to exhibit excellent antimicrobial effect against Fusarium moniliforme which is an important causative agent of Sorghum grain mold disease. Thus, cyclic dipeptides are emerging small molecular drug candidates for various autoimmune diseases.

Keywords: cyclic dipeptides, cytokines, Fusarium moniliforme, Pseudomonas, TNF-alpha

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Authors: Juergen Orasche, Vesta Kohlmeier, George C. Dragan, Gert Jakobi, Patricia Forbes, Ralf Zimmermann

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Underground mining poses a difficult environment for both man and machines. At more than 1000 meters underneath the surface of the earth, ores and other mineral resources are still gained by conventional and motorised mining. Adding to the hazards caused by blasting and stone-chipping, the working conditions are best described by the high temperatures of 35-40°C and high humidity, at low air exchange rates. Separate ventilation shafts lead fresh air into a mine and others lead expended air back to the surface. This is essential for humans and machines working deep underground. Nevertheless, mines are widely ramified. Thus the air flow rate at the far end of a tunnel is sensed to be close to zero. In recent years, conventional mining was supplemented by mining with heavy diesel machines. These very flat machines called Load Haul Dump (LHD) vehicles accelerate and ease work in areas favourable for heavy machines. On the other hand, they emit non-filtered diesel exhaust, which constitutes an occupational hazard for the miners. Combined with a low air exchange, high humidity and inorganic dust from the mining it leads to 'black smog' underneath the earth. This work focuses on the air quality in mines employing LHDs. Therefore we performed personal sampling (samplers worn by miners during their work), stationary sampling and aethalometer (Microaeth MA200, Aethlabs) measurements in a platinum mine in around 1000 meters under the earth’s surface. We compared areas of high diesel exhaust emission with areas of conventional mining where no diesel machines were operated. For a better assessment of health risks caused by air pollution we applied a separated gas-/particle-sampling tool (or system), with first denuder section collecting intermediate VOCs. These multi-channel silicone rubber denuders are able to trap IVOCs while allowing particles ranged from 10 nm to 1 µm in diameter to be transmitted with an efficiency of nearly 100%. The second section is represented by a quartz fibre filter collecting particles and adsorbed semi-volatile organic compounds (SVOC). The third part is a graphitized carbon black adsorber – collecting the SVOCs that evaporate from the filter. The compounds collected on these three sections were analyzed in our labs with different thermal desorption techniques coupled with gas chromatography and mass spectrometry (GC-MS). VOCs and IVOCs were measured with a Shimadzu Thermal Desorption Unit (TD20, Shimadzu, Japan) coupled to a GCMS-System QP 2010 Ultra with a quadrupole mass spectrometer (Shimadzu). The GC was equipped with a 30m, BP-20 wax column (0.25mm ID, 0.25µm film) from SGE (Australia). Filters were analyzed with In-situ derivatization thermal desorption gas chromatography time-of-flight-mass spectrometry (IDTD-GC-TOF-MS). The IDTD unit is a modified GL sciences Optic 3 system (GL Sciences, Netherlands). The results showed black carbon concentrations measured with the portable aethalometers up to several mg per m³. The organic chemistry was dominated by very high concentrations of alkanes. Typical diesel engine exhaust markers like alkylated polycyclic aromatic hydrocarbons were detected as well as typical lubrication oil markers like hopanes.

Keywords: diesel emission, personal sampling, aethalometer, mining

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Authors: Olga Kononova, Sergey Lyapin

Abstract:

Global information society requirements are transparency and reliability of data, as well as ability to manage information resources independently; particularly to search, to analyze, to evaluate information, thereby obtaining new expertise. Moreover, it is satisfying the society information needs that increases the efficiency of the enterprise management and public administration. The study of structurally organized thematic and semantic contexts of different types, automatically extracted from unstructured data, is one of the important tasks for the application of information technologies in education, science, culture, governance and business. The objectives of this study are the contextual knowledge typologization, selection or creation of effective tools for extracting and analyzing contextual knowledge. Explication of various kinds and forms of the contextual knowledge involves the development and use full-text search information systems. For the implementation purposes, the authors use an e-library 'Humanitariana' services such as the contextual search, different types of queries (paragraph-oriented query, frequency-ranked query), automatic extraction of knowledge from the scientific texts. The multifunctional e-library «Humanitariana» is realized in the Internet-architecture in WWS-configuration (Web-browser / Web-server / SQL-server). Advantage of use 'Humanitariana' is in the possibility of combining the resources of several organizations. Scholars and research groups may work in a local network mode and in distributed IT environments with ability to appeal to resources of any participating organizations servers. Paper discusses some specific cases of the contextual knowledge explication with the use of the e-library services and focuses on possibilities of new types of the contextual knowledge. Experimental research base are science texts about 'e-government' and 'computer games'. An analysis of the subject-themed texts trends allowed to propose the content analysis methodology, that combines a full-text search with automatic construction of 'terminogramma' and expert analysis of the selected contexts. 'Terminogramma' is made out as a table that contains a column with a frequency-ranked list of words (nouns), as well as columns with an indication of the absolute frequency (number) and the relative frequency of occurrence of the word (in %% ppm). The analysis of 'e-government' materials showed, that the state takes a dominant position in the processes of the electronic interaction between the authorities and society in modern Russia. The media credited the main role in these processes to the government, which provided public services through specialized portals. Factor analysis revealed two factors statistically describing the used terms: human interaction (the user) and the state (government, processes organizer); interaction management (public officer, processes performer) and technology (infrastructure). Isolation of these factors will lead to changes in the model of electronic interaction between government and society. In this study, the dominant social problems and the prevalence of different categories of subjects of computer gaming in science papers from 2005 to 2015 were identified. Therefore, there is an evident identification of several types of contextual knowledge: micro context; macro context; dynamic context; thematic collection of queries (interactive contextual knowledge expanding a composition of e-library information resources); multimodal context (functional integration of iconographic and full-text resources through hybrid quasi-semantic algorithm of search). Further studies can be pursued both in terms of expanding the resource base on which they are held, and in terms of the development of appropriate tools.

Keywords: contextual knowledge, contextual search, e-library services, frequency-ranked query, paragraph-oriented query, technologies of the contextual knowledge extraction

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Authors: Rowan Moore, Kai Scheffler, Eugen Damoc, Jennifer Sutton, Aaron Bailey, Stephane Houel, Simon Cubbon, Jonathan Josephs

Abstract:

Purpose: MS analysis of monoclonal antibodies (mAbs) at the protein and peptide levels is critical during development and production of biopharmaceuticals. The compositions of current generation therapeutic proteins are often complex due to various modifications which may affect efficacy. Intact proteins analyzed by MS are detected in higher charge states that also provide more complexity in mass spectra. Protein analysis in native or native-like conditions with zero or minimal organic solvent and neutral or weakly acidic pH decreases charge state value resulting in mAb detection at higher m/z ranges with more spatial resolution. Methods: Three commercially available mAbs were used for all experiments. Intact proteins were desalted online using size exclusion chromatography (SEC) or reversed phase chromatography coupled on-line with a mass spectrometer. For streamlined use of the LC- MS platform we used a single SEC column and alternately selected specific mobile phases to perform separations in either denaturing or native-like conditions: buffer A (20 % ACN, 0.1 % FA) with Buffer B (100 mM ammonium acetate). For peptide analysis mAbs were proteolytically digested with and without prior reduction and alkylation. The mass spectrometer used for all experiments was a commercially available Thermo Scientific™ hybrid Quadrupole-Orbitrap™ mass spectrometer, equipped with the new BioPharma option which includes a new High Mass Range (HMR) mode that allows for improved high mass transmission and mass detection up to 8000 m/z. Results: We have analyzed the profiles of three mAbs under reducing and native conditions by direct infusion with offline desalting and with on-line desalting via size exclusion and reversed phase type columns. The presence of high salt under denaturing conditions was found to influence the observed charge state envelope and impact mass accuracy after spectral deconvolution. The significantly lower charge states observed under native conditions improves the spatial resolution of protein signals and has significant benefits for the analysis of antibody mixtures, e.g. lysine variants, degradants or sequence variants. This type of analysis requires the detection of masses beyond the standard mass range ranging up to 6000 m/z requiring the extended capabilities available in the new HMR mode. We have compared each antibody sample that was analyzed individually with mixtures in various relative concentrations. For this type of analysis, we observed that apparent native structures persist and ESI is benefited by the addition of low amounts of acetonitrile and formic acid in combination with the ammonium acetate-buffered mobile phase. For analyses on the peptide level we analyzed reduced/alkylated, and non-reduced proteolytic digests of the individual antibodies separated via reversed phase chromatography aiming to retrieve as much information as possible regarding sequence coverage, disulfide bridges, post-translational modifications such as various glycans, sequence variants, and their relative quantification. All data acquired were submitted to a single software package for analysis aiming to obtain a complete picture of the molecules analyzed. Here we demonstrate the capabilities of the mass spectrometer to fully characterize homogeneous and heterogeneous therapeutic proteins on one single platform. Conclusion: Full characterization of heterogeneous intact protein mixtures by improved mass separation on a quadrupole-Orbitrap™ mass spectrometer with extended capabilities has been demonstrated.

Keywords: disulfide bond analysis, intact analysis, native analysis, mass spectrometry, monoclonal antibodies, peptide mapping, post-translational modifications, sequence variants, size exclusion chromatography, therapeutic protein analysis, UHPLC

Procedia PDF Downloads 361