A Comprehensive Metabolic Profile of Cultured Astrocytes Using Isotopic Transient Metabolic Flux Analysis and 13C-Labeled Glucose

Metabolic models have been used to elucidate important aspects of brain metabolism in recent years. This work applies for the first time the concept of isotopic transient 13C metabolic flux analysis (MFA) to estimate intracellular fluxes in primary cultures of astrocytes. This methodology comprehensively explores the information provided by 13C labeling time-courses of intracellular metabolites after administration of a 13C-labeled substrate. Cells were incubated with medium containing [1-13C]glucose for 24 h and samples of cell supernatant and extracts collected at different time points were then analyzed by mass spectrometry and/or high performance liquid chromatography. Metabolic fluxes were estimated by fitting a carbon labeling network model to isotopomer profiles experimentally determined. Both the fast isotopic equilibrium of glycolytic metabolite pools and the slow labeling dynamics of TCA cycle intermediates are described well by the model. The large pools of glutamate and aspartate which are linked to the TCA cycle via reversible aminotransferase reactions are likely to be responsible for the observed delay in equilibration of TCA cycle intermediates. Furthermore, it was estimated that 11% of the glucose taken up by astrocytes was diverted to the pentose phosphate pathway. In addition, considerable fluxes through pyruvate carboxylase [PC; PC/pyruvate dehydrogenase (PDH) ratio = 0.5], malic enzyme (5% of the total pyruvate production), and catabolism of branched-chained amino acids (contributing with ∼40% to total acetyl-CoA produced) confirmed the significance of these pathways to astrocytic metabolism. Consistent with the need of maintaining cytosolic redox potential, the fluxes through the malate–aspartate shuttle and the PDH pathway were comparable. Finally, the estimated glutamate/α-ketoglutarate exchange rate (∼0.7 μmol mg prot−1 h−1) was similar to the TCA cycle flux. In conclusion, this work demonstrates the potential of isotopic transient MFA for a comprehensive analysis of energy metabolism

Integrated system for macroalgae production and conversion into biogas

Recently research on energy from macroalgae is being reconsidered driven by the following factors: fuel price increase, CO2 mitigation policies and interest on renewable energy sources after the Kyoto protocol, and need for energy crops not competing with land for food production. However, the commercial expansion of this energy source is limited by its economic feasibility. In this presentation we analyse the development of integrated systems that promote synergies between macroalgae/biogas production and activities such as aquaculture and urban wastewater treatment. The recycling of nutrients and CO2 by macroalgae can be an opportunity to reduce the biomass-biogas production cost. Other advantage is the proximity between biomass production, conversion into energy and its consumption, thus avoiding energy losses and pollution in transportation. Experimental work is underway and includes batch tests to evaluate biodegradability of Ulva spp. and Gracilaria sp. and co-digestion of these macroalgae with sewage sludge from Beirolas wastewater treatment plant (WWTP). Furthermore, continuous co-digestion experiments are planned to be carried out in a 30 L experimental reactor to be placed in the same WWTP. The objectives are to reproduce sludge digestion in Beirolas WWTP and evaluate algae/sludge co-digestion performance. Preliminary results indicate methane yields in accordance with similar batch experiments, whereby our CH4 yields per volatile solids added range between 0.14-0.20 m3 CH4 kg-1 VSadded against 0.16-0.27 m3 CH4 kg-1 VSadded [1,2] depending on the algae species and pretreatment. Overall, Ulva sp. shows a better performance over Gracilaria sp. as also confirmed in other experiments [1]

Inhalable antitubercular therapy mediated by locust bean gum microparticles

Tuberculosis remains a major global health problem and alternative therapeutic approaches are needed. Considering the high prevalence of lung tuberculosis (80% of cases), the pulmonary delivery of antitubercular drugs in a carrier system capable of reaching the alveoli, being recognised and phagocytosed by alveolar macrophages (mycobacterium hosts), would be a significant improvement to current oral drug regimens. Locust bean gum (LBG) is a polysaccharide composed of galactose and mannose residues, which may favour specific recognition by macrophages and potentiate phagocytosis. LBG microparticles produced by spray-drying are reported herein for the first time, incorporating either isoniazid or rifabutin, first-line antitubercular drugs (association efficiencies >82%). Microparticles have adequate theoretical properties for deep lung delivery (aerodynamic diameters between 1.15 and 1.67 μm). The cytotoxic evaluation in lung epithelial cells (A549 cells) and macrophages (THP-1 cells) revealed a toxic effect from rifabutin-loaded microparticles at the highest concentrations, but we may consider that these were very high comparing with in vivo conditions. LBG microparticles further evidenced strong ability to be captured by macrophages (percentage of phagocytosis >94%). Overall, the obtained data indicated the potential of the proposed system for tuberculosis therapy

O depósito de machados do Bronze Final de Cobidalto, Areosa (Viana do Castelo) : novos dados para a sua contextualização e interpretação

Pretende-se com este artigo dar a conhecer os resultados dos trabalhos arqueológicos levados a cabo no monu-mento funerário de Laceiras do Côvo 3, estrutura tumular enquadrável, provavelmente, na Idade do Bronze, localizada na freguesia de Arões, concelho de Vale de Cambra. Este monumento revela-se de particular inte-resse pela policromia intencional na matéria usada na sua construção e pela disposição cuidada dos elementos litológicos do tumulus, em que nada parece estar ao acaso. Tais características sugerem a intenção de manter o espaço dos mortos visível, assim como uma interligação simbólica entre rochas e minerais e as práticas relacio-nadas com a morte. Na linha de Tim Ingold (2000) consideramos, ainda, que a matéria do monumento seria portadora de determinadas propriedades, revelando-se como entidade viva e atuante na construção de sentidos e memórias associadas ao mundo dos mortos.Fundação para a Ciência e a Tecnologia (FCT

Inhibition studies with 2-bromoethanesulfonate reveal a novel syntrophic relationship in anaerobic oleate degradation

Degradation of long-chain fatty acids (LCFAs) in methanogenic environments is a syntrophic process involving the activity of LCFA-degrading bacteria and hydrogen-utilizing methanogens. If methanogens are inhibited, other hydrogen scavengers are needed to achieve complete LCFA degradation. In this work, we developed two different oleate (C18:1 LCFA)-degrading anaerobic enrichment cultures, one methanogenic (ME) and another in which methanogenesis was inhibited (IE). Inhibition of methanogens was attained by adding a solution of 2-bromoethanesulfonate (BrES), which turned out to consist of a mixture of BrES and isethionate. Approximately 5 times faster oleate degradation was accomplished by the IE culture compared with the ME culture. A bacterium closely related to Syntrophomonas zehnderi (99\% 16S rRNA gene identity) was the main oleate degrader in both enrichments, in syntrophic relationship with hydrogenotrophic methanogens from the genera Methanobacterium and Methanoculleus (in ME culture) or with a bacterium closely related to Desulfovibrio aminophilus (in IE culture). A Desulfovibrio species was isolated, and its ability to utilize hydrogen was confirmed. This bacterium converted isethionate to acetate and sulfide, with or without hydrogen as electron donor. This bacterium also utilized BrES but only after 3 months of incubation. Our study shows that syntrophic oleate degradation can be coupled to desulfonation.IMPORTANCE In anaerobic treatment of complex wastewater containing fat, oils, and grease, high long-chain fatty acid (LCFA) concentrations may inhibit microbial communities, particularly those of methanogens. Here, we investigated if anaerobic degradation of LCFAs can proceed when methanogens are inhibited and in the absence of typical external electron acceptors, such as nitrate, iron, or sulfate. Inhibition studies were performed with the methanogenic inhibitor 2-bromoethanesulfonate (BrES). We noticed that, after autoclaving, BrES underwent partial hydrolysis and turned out to be a mixture of two sulfonates (BrES and isethionate). We found out that LCFA conversion proceeded faster in the assays where methanogenesis was inhibited, and that it was dependent on the utilization of isethionate. In this study, we report LCFA degradation coupled to desulfonation. Our results also showed that BrES can be utilized by anaerobic bacteria.Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of the UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145-FEDER-006684) and BioTecNorte operation (NORTE-01-0145-FEDER-000004), funded by the European Regional Development Fund under the scope of Norte2020—Programa Operacional Regional do Norte. We also acknowledge Project MultiBiorefinery (SAICTPAC/0040/2015 [POCI-01-0145-FEDER-016403]), funded by Sistema de Apoio à Investigação Científica e Tecnológica (SAICT), Programas de Atividades Conjuntas (PAC), and the financial support of the European Research Council under the European Union Seventh Framework Programme (FP/2007-2013)/ERC (grant agreement 323009)info:eu-repo/semantics/publishedVersio

Effect of sulfate and iron (III) on LCFA degradation by a methanogenic community

[Excerpt] Under anaerobic conditions long chain fatty acids (LCFA) can be converted to methane by syntrophic bacteria and methanogenic archaea. LCFA degradation was also reported in the presence of alternative hydrogenotrophic partners, such as sulfate-reducing bacteria (SRB) and iron-reducing bacteria (IRB), which generally show higher affinity for H2 than methanogens and are more resistant to LCFA [1,2,3]. Their presence in a microbial culture degrading LCFA can be advantageous to reduce LCFA toxicity towards methanogens, although high concentrations of external electron acceptor (EEA) can lead to outcompetition of methanogens and cease methane production. In this work, we tested the effect of adding sub-stoichiometric concentrations of sulfate and iron(III) to methanogenic communities degrading LCFA. (...

Anaerobic co-digestion of cork based oil sorbent and cow manure or sludge

Cork, a material with great economic, social and environmental importance in Portugal, is also a good oil sorbent that can be used in the remediation of oil spills. The oil-impregnated cork can be easily removed, but requires further treatment. In the case of vegetable oil spills, anaerobic digestion may be a potential solution. This study aims to evaluate the effect of adding cork contaminated with sunflower oil as co-substrate in anaerobic digestion processes. Biodegradability assays were prepared with cow manure or sludge from a wastewater treatment plant, in the presence of five concentrations of oil-contaminated cork, between 200 and 1000 mg· L-1 as COD. Maximum cumulative methane production increased with the amount of oily cork up to 41 % and 101 % in the assays with manure and sludge, respectively. Sporadic addition of cork contaminated with vegetable oil during anaerobic digestion of manure or sludge increases significantly the methane production of these processes.Programa Operacional Regional do Norte (ON.2 - 0 Novo Norte), QREN, FEDERPortuguese Foundation for Science and Technology (FCT, in the frame of projects FCOMPO 1-0124-FEDER-014784 (FCT: PTDC/EBBEBI/114364/2009

Ciprofloxacin removal catalysed by conductive carbon materials

Current wastewater treatment technologies are not effective in the removal of pharmaceuticals. In anaerobic bioreactors, the electrons produced during the oxidation of organic matter can potentially be used for the biological reduction of pharmaceuticals. However, these reactions occur generally at a slow rate, due to electron transfer limitations, and might be accelerated through redox mediators (RM). Carbon nanomaterials (CM) have been effective RM in the biological reduction of other pollutants. For instance, CNT@2%Fe were found to increase 76-fold the biological reduction of Acid Orange 10. The magnetic properties of those composites allow their easier recover after the process by using a magnetic field. In this study, CNT and CNT@2%Fe were studied in the anaerobic removal of Ciprofloxacin (CIP). Furthermore, the potential contribution of adsorption and biodegradation processes for CIP removal was assessed. Toxicity assessment is highly important as it is desired that the products formed after the process are not more toxic than the initial compound. Moreover, the evaluation of the possible contribution of nanomaterials used in the process for the final toxic effect of threated solution, is crucial. In this sense, the detoxification of the treated solutions was evaluated towards Vibrio fischeri.This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2019 unit and BioTecNorte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte.info:eu-repo/semantics/publishedVersio