In situ NIR spectroscopy monitoring of plasmid production processes effect of producing strain, medium composition and the cultivation strategy

BACKGROUNDWhile the pharmaceutical industry keeps an eye on plasmid DNA production for new generation gene therapies, real-time monitoring techniques for plasmid bioproduction are as yet unavailable. This work shows the possibility of in situ monitoring of plasmid production in Escherichia coli cultures using a near infrared (NIR) fiber optic probe. RESULTSPartial least squares (PLS) regression models based on the NIR spectra were developed for predicting bioprocess critical variables such as the concentrations of biomass, plasmid, carbon sources (glucose and glycerol) and acetate. In order to achieve robust models able to predict the performance of plasmid production processes, independently of the composition of the cultivation medium, cultivation strategy (batch versus fed-batch) and E. coli strain used, three strategies were adopted, using: (i) E. coliDH5 cultures conducted under different media compositions and culture strategies (batch and fed-batch); (ii) engineered E. coli strains, MG1655endArecApgi and MG1655endArecA, grown on the same medium and culture strategy; (iii) diverse E. coli strains, over batch and fed-batch cultivations and using different media compositions. PLS models showed high accuracy for predicting all variables in the three groups of cultures. CONCLUSIONNIR spectroscopy combined with PLS modeling provides a fast, inexpensive and contamination-free technique to accurately monitoring plasmid bioprocesses in real time, independently of the medium composition, cultivation strategy and the E. coli strain used.

Monitoring the ex-vivo expansion of human mesenchymal stem/stromal cells in xeno-free microcarrier-based reactor systems by MIR spectroscopy

Human mesenchymal stem/stromal cells (MSCs) have received considerable attention in the field of cell-based therapies due to their high differentiation potential and ability to modulate immune responses. However, since these cells can only be isolated in very low quantities, successful realization of these therapies requires MSCs ex-vivo expansion to achieve relevant cell doses. The metabolic activity is one of the parameters often monitored during MSCs cultivation by using expensive multi-analytical methods, some of them time-consuming. The present work evaluates the use of mid-infrared (MIR) spectroscopy, through rapid and economic high-throughput analyses associated to multivariate data analysis, to monitor three different MSCs cultivation runs conducted in spinner flasks, under xeno-free culture conditions, which differ in the type of microcarriers used and the culture feeding strategy applied. After evaluating diverse spectral preprocessing techniques, the optimized partial least square (PLS) regression models based on the MIR spectra to estimate the glucose, lactate and ammonia concentrations yielded high coefficients of determination (R2 ≥ 0.98, ≥0.98, and ≥0.94, respectively) and low prediction errors (RMSECV ≤ 4.7%, ≤4.4% and ≤5.7%, respectively). Besides PLS models valid for specific expansion protocols, a robust model simultaneously valid for the three processes was also built for predicting glucose, lactate and ammonia, yielding a R2 of 0.95, 0.97 and 0.86, and a RMSECV of 0.33, 0.57, and 0.09 mM, respectively. Therefore, MIR spectroscopy combined with multivariate data analysis represents a promising tool for both optimization and control of MSCs expansion processes.

Some pioneers of the applications of fractional calculus

In the last decades fractional calculus (FC) became an area of intensive research and development. This paper goes back and recalls important pioneers that started to apply FC to scientific and engineering problems during the nineteenth and twentieth centuries. Those we present are, in alphabetical order: Niels Abel, Kenneth and Robert Cole, Andrew Gemant, Andrey N. Gerasimov, Oliver Heaviside, Paul Lévy, Rashid Sh. Nigmatullin, Yuri N. Rabotnov, George Scott Blair.

Comparative analysis of near and mid-infrared spectroscopy to monitor recombinant cyprosin production

Infrared spectroscopy, either in the near and mid (NIR/MIR) region of the spectra, has gained great acceptance in the industry for bioprocess monitoring according to Process Analytical Technology, due to its rapid, economic, high sensitivity mode of application and versatility. Due to the relevance of cyprosin (mostly for dairy industry), and as NIR and MIR spectroscopy presents specific characteristics that ultimately may complement each other, in the present work these techniques were compared to monitor and characterize by in situ and by at-line high-throughput analysis, respectively, recombinant cyprosin production by Saccharomyces cerevisiae. Partial least-square regression models, relating NIR and MIR-spectral features with biomass, cyprosin activity, specific activity, glucose, galactose, ethanol and acetate concentration were developed, all presenting, in general, high regression coefficients and low prediction errors. In the case of biomass and glucose slight better models were achieved by in situ NIR spectroscopic analysis, while for cyprosin activity and specific activity slight better models were achieved by at-line MIR spectroscopic analysis. Therefore both techniques enabled to monitor the highly dynamic cyprosin production bioprocess, promoting by this way more efficient platforms for the bioprocess optimization and control.

Creating and optimizing client-server applications on mobile devices

Mobile devices are embedded systems with very limited capacities that need to be considered when developing a client-server application, mainly due to technical, ergonomic and economic implications to the mobile user. With the increasing popularity of mobile computing, many developers have faced problems due to low performance of devices. In this paper, we discuss how to optimize and create client-server applications for in wireless/mobile environments, presenting techniques to improve overall performance.

Dual-phase inkjet printed electrochromic layers based on PTA and WOX/TiO2 nanoparticles for electrochromic applications

Thesis submitted in Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa for the degree of Master in Materials Engineering

Processing and characterization of 3D dense chitosan pieces, for orthopedic applications, by adding plasticizers

In this work, plasticizer agents were incorporated in a chitosan based formulation, as a strategy to improve the fragile structure of chitosan based-materials. Three different plasticizers: ethylene glycol, glycerol and sorbitol, were blended with chitosan to prepare 3D dense chitosan specimens. The properties of the obtained structures were assessed for mechanical, microstructural, physical and biocompatibility behavior. The results obtained revealed that from the different specimens prepared, the blend of chitosan with glycerol has superior mechanical properties and good biological behavior, making this chitosan based formulation a good candidate to improve robust chitosan structures for the construction of bioabsorbable orthopedic implants.

Core language for web applications

Trabalho apresentado no âmbito do Mestrado em Engenharia Informática, como requisito parcial para obtenção do grau de Mestre em Engenharia Informática

CMOS indoor light energy harvesting system for wireless sensing applications

This book discusses in detail the CMOS implementation of energy harvesting. The authors describe an integrated, indoor light energy harvesting system, based on a controller circuit that dynamically and automatically adjusts its operation to meet the actual light circumstances of the environment where the system is placed. The system is intended to power a sensor node, enabling an autonomous wireless sensor network (WSN). Although designed to cope with indoor light levels, the system is also able to work with higher levels, making it an all-round light energy harvesting system. The discussion includes experimental data obtained from an integrated manufactured prototype, which in conjunction with a photovoltaic (PV) cell, serves as a proof of concept of the desired energy harvesting system. © 2016 Springer International Publishing. All rights are reserved.

Glass in Cultural Heritage – An Introduction

Proceedings of the Chemistry and Conservation Science

Downstream processing development of enveloped viruses for clinical applications: innovative tools for rational process optimization

Dissertation presented to obtain a Ph.D. degree in Engineering and Technology Sciences, Biotechnology at the Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa

Personal learning environments: integration of Web 2.0 applications and content management systems

This paper presents an ongoing project that implements a platform for creating personal learning environments controlled by students, integrating Web 2.0 applications and content management systems, enabling the safe use of content created in Web 2.0 applications, allowing its publication in the infrastructure controlled by the HEI. Using this platform, students can develop their personal learning environment (PLE) integrated with the Learning Management System (LMS) of the HEI, enabling the management of their learning and, simultaneously, creating their e-portfolio with digital content developed for Course Units (CU). All this can be maintained after the student completes his academic studies, since the platform will remain accessible to students even after they leave the HEI and lose access to its infrastructure. The platform will enable the safe use of content created in Web 2.0 applications, allowing its protected publication in the infrastructure controlled by HEI, thus contributing to the adaptation of the L&T paradigm to the Bologna process.

Teaching and learning with social software in higher education: content management systems integrated with Web-based applications as a key factor for success

The change of paradigm imposed by the Bologna process, in which the student will be responsible for their own learning, and the presence of a new generation of students with higher technological skills, represent a huge challenge for higher education institutions. The use of new Web Social concepts in teaching process, supported by applications commonly called Web 2.0, with which these new students feel at ease, can bring benefits in terms of motivation and the frequency and quality of students' involvement in academic activities. An e-learning platform with web-based applications as a complement can significantly contribute to the development of different skills in higher education students, covering areas which are usually in deficit.

Optimização e Validação da Produção de [13n]-Nh3 para Estudos Clínicos de Tomografia por Emissão de Positrões na Avaliação da Perfusão Miocárdica

A par das patologias oncológicas, as doenças do foro cardíaco, em particular a doença arterial coronária, são uma das principais causas de morte nos países industrializados, devido sobretudo, à grande incidência de enfartes do miocárdio. Uma das formas de diagnóstico e avaliação desta condição passa pela obtenção de imagens de perfusão miocárdica com radionuclídeos, realizada por Tomografia por Emissão de Positrões (PET). As soluções injectáveis de [15O]-H2O, [82Rb] e [13N]-NH3 são as mais utilizadas neste tipo de exame clínico. No Instituto de Ciências Nucleares Aplicadas à Saúde (ICNAS), a existência de um ciclotrão tem permitido a produção de uma variedade de radiofármacos, com aplicações em neurologia, oncologia e cardiologia. Recentemente, surgiu a oportunidade de iniciar exames clínicos com [13N]-NH3 para avaliação da perfusão miocárdica. É neste âmbito que surge a oportunidade do presente trabalho, pois antes da sua utilização clínica é necessário realizar a optimização da produção e a validação de todo o processo segundo as normas de Boas Práticas Radiofarmacêuticas. Após uma fase de optimização do processo, procedeu-se à avaliação dos parâmetros físico-químicos e biológicos da preparação de [13N]-NH3, de acordo com as indicações da Farmacopeia Europeia (Ph. Eur.) 8.2. De acordo com as normas farmacêuticas, foram realizados 3 lotes de produção consecutivos para validação da produção de [13N]-NH3. Os resultados mostraram um produto final límpido e ausente de cor, com valores de pH dentro do limite especificado, isto é, entre 4,5 e 8,5. A pureza química das amostras foi verificada, uma vez que relativamente ao teste colorimétrico, a tonalidade da cor da solução de [13N]-NH3 não era mais intensa que a solução de referência. As preparações foram identificadas como sendo [13N]-NH3, através dos resultados obtidos por cromatografia iónica, espectrometria de radiação gama e tempo de semi-vida. Por examinação do cromatograma obtido com a solução a ser testada, observou-se que o pico principal possuia um tempo de retenção aproximadamente igual ao pico do cromatograma obtido para a solução de referência. Além disso, o espectro de radiação gama mostrou um pico de energia 0,511 MeV e um outro adicional de 1,022 MeV para os fotões gama, característico de radionuclídeos emissores de positrões. O tempo de semi-vida manteve-se dentro do intervalo indicado, entre 9 e 11 minutos. Verificou-se, igualmente, a pureza radioquímica das amostras, correspondendo um mínimo de 99% da radioactividade total ao [13N], bem como a pureza radionuclídica, observando-se uma percentagem de impurezas inferiores a 1%, 2h após o fim da síntese. Os testes realizados para verificação da esterilidade e determinação da presença de endotoxinas bacterianas nas preparações de [13N]-NH3 apresentaram-se negativos.Os resultados obtidos contribuem, assim, para a validação do método para a produção de [13N]-NH3, uma vez que cumprem os requisitos especificados nas normas europeias, indicando a obtenção de um produto seguro e com a qualidade necessária para ser administrado em pacientes para avaliação da perfusão cardíaca por PET.

Upper Ontology for Multi-Agent Energy Systems’ Applications

Energy systems worldwide are complex and challenging environments. Multi-agent based simulation platforms are increasing at a high rate, as they show to be a good option to study many issues related to these systems, as well as the involved players at act in this domain. In this scope the authors research group has developed three multi-agent systems: MASCEM, which simulates the electricity markets; ALBidS that works as a decision support system for market players; and MASGriP, which simulates the internal operations of smart grids. To take better advantage of these systems, their integration is mandatory. For this reason, is proposed the development of an upper-ontology which allows an easier cooperation and adequate communication between them. Additionally, the concepts and rules defined by this ontology can be expanded and complemented by the needs of other simulation and real systems in the same areas as the mentioned systems. Each system’s particular ontology must be extended from this top-level ontology.