Design of bio-inspired ionic liquids for protein stabilisation

Ionic Liquids (ILs) consist in organic salts that are liquid at/or near room temperature. Since ILs are entirely composed of ions, the formation of ion pairs is expected to be one essential feature for describing solvation in ILs. In recent years, protein - ionic liquid (P-IL) interactions have been the subject of intensive studies mainly because of their capability to promote folding/unfolding of proteins. However, the ion pairs and their lifetimes in ILs in P-IL thematic is dismissed, since the action of ILs is therefore the result of a subtle equilibrium between anion-cation interaction, ion-solvent and ion-protein interaction. The work developed in this thesis innovates in this thematic, once the design of ILs for protein stabilisation was bio-inspired in the high concentration of organic charged metabolites found in cell milieu. Although this perception is overlooked, those combined concentrations have been estimated to be ~300 mM among the macromolecules at concentrations exceeding 300 g/L (macromolecular crowding) and transient ion-pair can naturally occur with a potential specific biological role. Hence the main objective of this work is to develop new bio-ILs with a detectable ion-pair and understand its effects on protein structure and stability, under crowding environment, using advanced NMR techniques and calorimetric techniques. The choline-glutamate ([Ch][Glu]) IL was synthesized and characterized. The ion-pair was detected in water solutions using mainly the selective NOE NMR technique. Through the same technique, it was possible to detect a similar ion-pair promotion under synthetic and natural crowding environments. Using NMR spectroscopy (protein diffusion, HSQC experiments, and hydrogen-deuterium exchange) and differential scanning calorimetry (DSC), the model protein GB1 (production and purification in isotopic enrichment media) it was studied in the presence of [Ch][Glu] under macromolecular crowding conditions (PEG, BSA, lysozyme). Under dilute condition, it is possible to assert that the [Ch][Glu] induces a preferential hydration by weak and non-specific interactions, which leads to a significant stabilisation. On the other hand, under crowding environment, the [Ch][Glu] ion pair is promoted, destabilising the protein by favourable weak hydrophobic interactions , which disrupt the hydration layer of the protein. However, this capability can mitigates the effect of protein crowders. Overall, this work explored the ion-pair existence and its consequences on proteins in conditions similar to cell milieu. In this way, the charged metabolites found in cell can be understood as key for protein stabilisation.

An evolutionary multi-objective optimization system for earthworks

Earthworks involve the levelling or shaping of a target area through the moving or processing of the ground surface. Most construction projects require earthworks, which are heavily dependent on mechanical equipment (e.g., excavators, trucks and compactors). Often, earthworks are the most costly and time-consuming component of infrastructure constructions (e.g., road, railway and airports) and current pressure for higher productivity and safety highlights the need to optimize earthworks, which is a nontrivial task. Most previous attempts at tackling this problem focus on single-objective optimization of partial processes or aspects of earthworks, overlooking the advantages of a multi-objective and global optimization. This work describes a novel optimization system based on an evolutionary multi-objective approach, capable of globally optimizing several objectives simultaneously and dynamically. The proposed system views an earthwork construction as a production line, where the goal is to optimize resources under two crucial criteria (costs and duration) and focus the evolutionary search (non-dominated sorting genetic algorithm-II) on compaction allocation, using linear programming to distribute the remaining equipment (e.g., excavators). Several experiments were held using real-world data from a Portuguese construction site, showing that the proposed system is quite competitive when compared with current manual earthwork equipment allocation.

Comparison of single and multi-objective evolutionary algorithms for robust link-state routing

Traffic Engineering (TE) approaches are increasingly impor- tant in network management to allow an optimized configuration and resource allocation. In link-state routing, the task of setting appropriate weights to the links is both an important and a challenging optimization task. A number of different approaches has been put forward towards this aim, including the successful use of Evolutionary Algorithms (EAs). In this context, this work addresses the evaluation of three distinct EAs, a single and two multi-objective EAs, in two tasks related to weight setting optimization towards optimal intra-domain routing, knowing the network topology and aggregated traffic demands and seeking to mini- mize network congestion. In both tasks, the optimization considers sce- narios where there is a dynamic alteration in the state of the system, in the first considering changes in the traffic demand matrices and in the latter considering the possibility of link failures. The methods will, thus, need to simultaneously optimize for both conditions, the normal and the altered one, following a preventive TE approach towards robust configurations. Since this can be formulated as a bi-objective function, the use of multi-objective EAs, such as SPEA2 and NSGA-II, came nat- urally, being those compared to a single-objective EA. The results show a remarkable behavior of NSGA-II in all proposed tasks scaling well for harder instances, and thus presenting itself as the most promising option for TE in these scenarios.

Evaluation of antibacterial activity of caffeic acid encapsulated by -cyclodextrins

Context: Caffeic acid is described as antibacterial, but this bioactive molecule has some issues regarding solubility and stability to environmental stress. Thus, encapsulation devices are required. Objective: The aim of this work was to study the effect of the caffeic acid encapsulation by cyclodextrins on its antibacterial activity. Materials and methods: The interactions between the caffeic acid and three cyclodextrins (-cyclodextrin (CD), 2-hydroxypropyl--cyclodextrin (HPCD) and methyl--cyclodextrin were study. Results and discussion: The formation of an aqueous soluble inclusion complex was confirmed for CD and HPCD with a 1:1 stoichiometry. The CD/caffeic acid complex showed higher stability than HPCD/caffeic acid. Caffeic acid antibacterial activity was similar at pH 3 and pH 5 against the three bacteria (K. pneumoniae, S. epidermidis and S. aureus). Conclusions: The antibacterial activity of the inclusion complexes was described here for the first time and it was shown that the caffeic acid activity was remarkably enhanced by the cyclodextrins encapsulation.

Cationic Liposomes as antigenic delivery systems: development of an immunoprotective strategy against Candida albicans

Tese de Doutoramento Biologia Molecular e Ambiental - Especialidade em Biologia Celular e Saúde

A new empirical system for rock slope stability analysis in exploitation stage

An empirical system was developed to obtain a quality index for rock slopes in road infrastructures, named Slope Quality Index (SQI), and it was applied to a set of real slopes.The SQI is supported in nine factors affecting slope stability that contemplate the evaluation of different parameters. Consequently, each factor is classified by the degree of importance and influence by assigned weights. These weights were established through a statistical analysis of replies to a survey that was distributed to several experienced professionals in the field. The proposed SQI varies between1 and 5, corresponding to slopes in very good and very bad condition state, respectively. Besides the advantage linked to a quantitative and qualitative evaluation of slopes, theSQI also allows identifying the most critical factors on the slope stability, which is a fundamental issue for an efficient management of the slope network in the road infrastructure, namely in the planning of conservation and maintenance operations.

Rényi entropy and the thermodynamic stability of black holes

Thermodynamic stability of black holes, described by the Rényi formula as equilibrium compatible entropy function, is investigated. It is shown that within this approach, asymptotically flat, Schwarzschild black holes can be in stable equilibrium with thermal radiation at a fixed temperature. This implies that the canonical ensemble exists just like in anti-de Sitter space, and nonextensive effects can stabilize the black holes in a very similar way as it is done by the gravitational potential of an anti-de Sitter space. Furthermore, it is also shown that a Hawking–Page-like black hole phase transition occurs at a critical temperature which depends on the q-parameter of the Rényi formula.

Combining non-dominance, objective-order and spread metric to extend firefly algorithm to multi-objective optimization

In this paper, we propose an extension of the firefly algorithm (FA) to multi-objective optimization. FA is a swarm intelligence optimization algorithm inspired by the flashing behavior of fireflies at night that is capable of computing global solutions to continuous optimization problems. Our proposal relies on a fitness assignment scheme that gives lower fitness values to the positions of fireflies that correspond to non-dominated points with smaller aggregation of objective function distances to the minimum values. Furthermore, FA randomness is based on the spread metric to reduce the gaps between consecutive non-dominated solutions. The obtained results from the preliminary computational experiments show that our proposal gives a dense and well distributed approximated Pareto front with a large number of points.

Evaluating balance, stability and gait symmetry of stroke patients using kinematic and dynamic gait lab data

Dissertação de mestrado integrado em Engenharia Biomédica

Investigation on the homogeneity and stability of aqueous nano cellulose suspensions prepared using pluronic F-127

This paper presents the results of experimental investigation on the aqueous dispersion behaviour of micro crystalline cellulose (MCC) prepared using Pluronic F-127. For this purpose, different concentrations (0.5-3.0 wt.%) of MCC were dispersed in water with the help of ultrasonication technique using various concentrations of Pluronic F-127. The homogeneity of the suspensions and agglomerations were characterized by optical and transmission electron microscopy and the concentration of well dispersed MCC was measured using UV-Vis spectroscopy. Also, the suspensions were subjected to high speed ultracentrifugation at 3000 rpm and observed visually for sedimentation and subsequently, concentration was calculated using UV-Vis, in order to assess the long term stability of the suspensions. Based on these experiments, optimum concentration of Pluronic to disperse different MCC concentrations has been suggested.

The impact of urban noise on primary schools: perceptive evaluation and objective assessment

This paper aims to assess the impact of environmental noise in the vicinity of primary schools and to analyze its influence in the workplace and in student performance through perceptions and objective evaluation. The subjective evaluation consisted of the application of questionnaires to students and teachers, and the objective assessment consisted of measuring in situ noise levels. The survey covered nine classes located in three primary schools. Statistical Package for Social Sciences was used for data processing and to draw conclusions. Additionally, the relationship of the difference between environmental and background noise levels of each classroom and students with difficulties in hearing the teacherâ s voice was examined. Noise levels in front of the school, the schoolyard, and the most noise-exposed classrooms (occupied and unoccupied) were measured. Indoor noise levels were much higher than World Health Organization (WHO) recommended values: LAeq,30min averaged 70.5 dB(A) in occupied classrooms, and 38.6 dB(A) in unoccupied ones. Measurements of indoor and outdoor noise suggest that noise from the outside (road, schoolyard) affects the background noise level in classrooms but in varying degrees. It was concluded that the façades most exposed to road traffic noise are subjected to values higher than 55.0 dB(A), and noise levels inside the classrooms are mainly due to the schoolyard, students, and the road traffic. The difference between background (LA95,30min) and the equivalent noise levels (LAeq,30min) in occupied classrooms was 19.2 dB(A), which shows that studentsâ activities are a significant source of classroom noise.

Brazilian norms for the International Affective Picture System (IAPS): comparison of the affective ratings for new stimuli between Brazilian and North-American subjects

OBJECTIVE: The study presents the Brazilian norms for 240 new stimuli from International Affective Picture System (IAPS), a database of affective images widely used in research, compared to the North-American normative ratings. METHODS: The participants were 448 Brazilian university students from several courses (269 women and 179 men) with mean age of 24.2 (SD = 7.8), that evaluated the IAPS pictures in the valence, arousal and dominance dimensions by the Self-Assessment Manikin (SAM) scales. Data were compared across the populations by Pearson linear correlation and Student's t-tests. RESULTS: Correlations were highly significant for all dimensions; however, Brazilians' averages for arousal were higher than North-Americans'. CONCLUSIONS: The results show stability in relation to the first part of the Brazilian standardization and they are also consistent with the North-American standards, despite minor differences relating to interpretation of the arousal dimension, demonstrating that IAPS is a reliable instrument for experimental studies in the Brazilian population.

Metabolic engineering with multi-objective optimization of kinetic models

Kinetic models have a great potential for metabolic engineering applications. They can be used for testing which genetic and regulatory modifications can increase the production of metabolites of interest, while simultaneously monitoring other key functions of the host organism. This work presents a methodology for increasing productivity in biotechnological processes exploiting dynamic models. It uses multi-objective dynamic optimization to identify the combination of targets (enzymatic modifications) and the degree of up- or down-regulation that must be performed in order to optimize a set of pre-defined performance metrics subject to process constraints. The capabilities of the approach are demonstrated on a realistic and computationally challenging application: a large-scale metabolic model of Chinese Hamster Ovary cells (CHO), which are used for antibody production in a fed-batch process. The proposed methodology manages to provide a sustained and robust growth in CHO cells, increasing productivity while simultaneously increasing biomass production, product titer, and keeping the concentrations of lactate and ammonia at low values. The approach presented here can be used for optimizing metabolic models by finding the best combination of targets and their optimal level of up/down-regulation. Furthermore, it can accommodate additional trade-offs and constraints with great flexibility.

Avaliação do desempenho ambiental do aglomerado negro de cortiça

Dissertação de mestrado integrado em Engenharia Civil

Development of Monoolein-based lipofection vectors for therapeutic siRNA delivery

Tese de Doutoramento em Biologia Molecular e Ambiental - Especialidade em Biologia Celular e Saúde