ISEL - Eng. Quim. Biol. - Comunicações
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- Biodegradable nanoparticles of alginate and chitosan as non-viral DNA oral delivery systemPublication . Gonçalves, Lídia M. D.; Cadete, Ana; Figueiredo, Lara; Calado, Cecília; Almeida, António J.The delivery of nucleic acids via the oral route involves overcoming barriers such as degradation of nucleic acids by low pH in the stomach, enzymatic degradation by DNases in the gut, crossing the physical barrier imposed by the mucus layer, cellular uptake, intracellular trafficking and nuclear uptake. As an oral drug carrier system chitosan nanoparticles are ideal, being mucoadhesive, interacting with the anionic sialic acid residues in mucin. In this study, plasmid DNA expressing a humanized secreted Gaussia Luciferase as reporter gene was encapsulated in alginate and chitosan nanoparticles, via a mild ionotropic gelation procedure with sodium tripolyphosphate as a counterion. The nanoparticle system here developed shows effective transfection of different human gastric epithelial cell lines with distinct cell differention. That was confirmed by the expression of luciferase in the different tested conditions, particularly the amount of encapsulated pGLuc.
- Monitoring bacterial processes by Fourier transform infrared spectroscopy: Helicobacter pylori drug inactivation and plasmid bioproduction in recombinant Escherichia coli culturesPublication . Scholz, Teresa; Lopes, Vítor V.; Calado, CecíliaFourier transform infrared (FTIR) spectroscopy is evaluated as a tool to monitor two bacterial processes: strain discrimination and drug inactivation studies with the gastric pathogen Helicobacter pylori and the plasmid production process based on high-density cultures of recombinant Escherichia coli. Results show, that after evaluation of different incubation conditions of H.pylori with the drug model, the application of principal component analysis to the FTIR spectra assembles the samples into clusters which can be related with the minimal inhibitory concentration. Morever, the same methodology applied to FTIR spectra from 12 different strains can be used to distinguish them. For the E.coli cultures it is possible to estimate the concentration of relevant bioprocess monitoring variables, such as plasmid, biomass, and carbon sources (glucose, glycerol, acetate) by using partial least squares (PLS) models based on FTIR spectra.
- Industry energy optimization: A case study in a biodiesel production sitePublication . Palmeira, José; Silva, João; Matos, Henrique A. S.This paper presents a case study of heat exchanger network (HEN) retrofit with the objective to reduce the utilities consumption in a biodiesel production process. Pinch analysis studies allow determining the minimum duty utilities as well the maximum of heat recovery. The existence of heat exchangers for heat recovery already running in the process causes a serious restriction for the implementation of grassroot HEN design based on pinch studies. Maintaining the existing HEN, a set of alternatives with additional heat exchangers was created and analysed using some industrial advice and selection criteria. The final proposed solution allows to increase the actual 18 % of recovery heat of the all heating needs of the process to 23 %, with an estimated annual saving in hot utility of 35 k(sic)/y.
- Solution enthalpies of hydroxylic compoundsPublication . Reis, Marina; Moreira, Luís; Nunes, Nélson; Elvas Leitao, Ruben; Martins, FilomenaSolution enthalpies of adamantan-1-ol, 2-methyl- butan-2-ol, and 3-methylbutan-1-ol have been measured at 298.15 K, in a set of 16 protogenic and non-protogenic solvents. The identification and quantification of solvent effects on the solution processes under study were performed using quantitative-structure property relationships. The results are discussed in terms of solute-solvent-solvent interactions and also in terms of the influence of compound's size and position of its hydroxyl group.
- Is it possible to remove polymeric nanoparticles from aqueous paints during the activated sludge treatment?Publication . Nobre, A.; Barreiros, A. M.; Piçarra, S.The market for emulsion polymers (latexes) is large and growing at the expense of other manufacturing processes that emit higher amounts of volatile organic solvents. The paint industry is not an exception and solvent-borne paints have been gradually substituted by aqueous paints. In their life-cycle, much of the aqueous paint used for architectural or decorative purposes will eventually be discharged into wastewater treatment facilities, where its polymeric nanoparticles (mainly acrylic and styrene-acrylic) can work as xenobiotics to the microbial communities present in activated sludge. It is well established that these materials are biocompatible at macroscopic scale. But is their behaviour the same at nanoscale? What happens to the polymeric nanoparticles during the activated sludge process? Do nanoparticles agregate and are discharged together with the sludge or remain in emulsion? How do microorganisms interact with these nanoparticles? Are nanoparticles degradated by them? Are they adsorbed? Are these nanoparticles toxic to the microbial community? To study the influence of these xenobiotics in the activated sludge process, an emulsion of cross-linked poly(butyl methacrylate) nanoparticles of ca. 50 nm diameter was produced and used as model compound. Activated sludge from a wastewater treatment plant was tested by the OCDE’s respiration inhibition test using several concentrations of PBMA nanoparticles. Particle aggregation was followed by Dynamic Light Scattering and microorganism surfaces were observed by Atomic Force Microscopy. Using sequential batch reactors (SBRs) and continuous reactors, both inoculated with activated sludge, the consumption of carbon, ammonia, nitrite and nitrate was monitored and compared, in the presence and absence of nanoparticles. No particles were detected in all treated waters by Dynamic Light Scattering. This can either mean that microorganisms can efficiently remove all polymer nanoparticles or that nanoparticles tend to aggregate and be naturally removed by precipitation. Nevertheless respiration inhibition tests demonstrated that microorganisms consume more oxygen in the presence of nanoparticles, which suggests a stress situation. It was also observed a slight decrease in the efficiency of nitrification in the presence of nanoparticles. AFM images showed that while the morphology of some organisms remained the same both in the presence and absence of nanoparticles, others assumed a rough surface with hilly like shapes of ca. 50 nm when exposed to nanoparticles. Nanoparticles are thus likely to be either incorporated or adsorbed at the surface of some organisms, increasing the overall respiration rate and decreasing nitrification efficiency. Thus, despite its biocompatibility at macroscopic scale, PBMA is likely to be no longer innocuous at nanoscale.
- Protein and DNA nanoparticulate multiantigenic vaccines against H. pylori: In vivo evaluationPublication . Figueiredo, Lara; Calado, Cecília; Almeida, António J.; Gonçalves, Lídia M. D.Immunisation against H. pylori is an attractive option for antibiotic resistance and reinfection situations. Strain genetic heterogeneity, and low immunogenicity of protein antigens and DNA alone are nonetheless obstacles to this approach. We developed multigenic H. pylori DNA-nanoparticle and protein-nanoparticle vaccines based on pathogenic relevance. Six antigens were chosen for the vaccine construction: CagA, VacA, HpaA, UreB, HomB and GroEL. Different combinations of CS/DS and CS/Alg /TPP nanoparticles with DNA and chimeric proteins were produced as vaccine systems. Immune responses were evaluated after i.m. and oral immunisation of BALB/c mice. Oral vaccination successfully stimulated mucosal immunity while i.m. immunisation efficiently elicited a more equilibrated cellular/humoral immune response.
- The importance of Helicobacter pylori's genetic variability for the construction of an efficient vaccinePublication . Vaz, Filipa; Couto, Andreia; Calado, CecíliaHelicobacter pylori is responsible for several gastric diseases. The main constraints of vaccine trials against this pathogen are mainly due to the bacterium high antigenic variability and to down-regulation of the host immune responses. To counteract these factors we propose a DNA vaccine able to induce a balanced humoural and citotoxic specific immune responses, based on multi-antigens. The selection of the antigens NapA, HpaA, VacA and HomB were conducted based on immunoproteomic data and the protein role on infection and pathogenesis. A fragment of each target-antigen was selected by in silico methods based on the maximization of the gene conservation and antigenicity. The set of these small fragments will be presented as a vaccine based on several conserved epitopes of multi-antigenic targets, and consequently representative of the bacterium antigenic variability.
- Modelling, monitoring and control of plasmid bioproduction in Escherichia coli culturesPublication . Lopes, Marta B.; Scholtz, Teresa; Silva, Daniel; Santos, Inês; Silva, Tito; Sampaio, Pedro; Couto, Andreia; Lopes, Vitor V.; Calado, CecíliaAn integrated approach for modelling, monitoring and control the plasmid bioproduction in Escherichia coli cultures is presented. In a first stage, by the implementation of a kinetic model for E. coli cultures, a better bioprocess understanding was reached, concerning the availability of nutrients and products along the bioprocess, and their effects on the plasmid production. Results presented may provide significant help for future modelling and monitoring implementation. In a second stage, FTIR spectroscopy coupled with chemometrics, namely PLS regression, shows its potential as a high-throughput technique for simultaneously estimating the key variables involved in the plasmid production process by E. coli cultures run under distinct conditions. Finally, owing to online monitoring and process control, an NIR fibre optic probe and chemometrics provided promising results concerning the control of biomass and carbon sources in E. coli cultures.
- Real-time plasmid monitoring of batch and fed-batch Escherichia coli cultures by NIR spectroscopyPublication . Lopes, Marta B.; Sales, Kevin C.; Lopes, Vítor V.; Calado, CecíliaThe development of in-situ monitoring techniques enabling the real-time acquisition of information concerning the key variables over different Escherichia coli cultivation conditions and strategies is a crucial step towards the optimization of a plasmid bioproduction process. This work shows the use of a Near-InfraRed (NIR) fiber optic probe immersed in the culture broth for the real-time acquisition of NIR spectra along different E. coli cultures conducted with mixtures of the carbon sources glucose and glycerol, and performed in both batch and fed-batch modes. Accurate partial least squares models based on the acquired spectral data over such different cultivation conditions were built, yielding a R 2 = 0.97 for biomass and plasmid productions and a RMSEP of 0.34 and 7.52, respectively; a R 2 of 0.93 and a RMSEP of 0.46 and 0.33 was obtained for glucose and glycerol, respectively; the acetate model produced a R 2 of 0.96 and a RMSEP of 0.32.
- The role of teaching decision analysis for sustainability in engeneering schoolsPublication . João, Isabel; Quadrado, José Carlos LourençoThis paper addresses the role that decision analysis plays in helping engineers to gain a greater understanding of the problems they face. The need of structured decision analysis is highlighted as well as the use of multiple criteria decision analysis to tackle sustainability issues with emphasis in the use of MACBETH approach. Some insights from a Portuguese Summer Course on engineering for sustainable development are presented namely the students 'and teacher perceptions about the module of decision analysis for sustainability.