Development of electrospun Ion jelly fibers® for drug delivery

Dissertação para obtenção do Grau de Mestre em Biotecnologia

Development of molecularly imprinted polymers using supercritical fluid technology

Dissertação para obtenção do Grau de Doutor em Química Sustentável

Selective synthesis under microwave irradiation of new monomers for potential applications in PDLC films

Dissertation to obtain the degree of master in Bioorganic

Edible coatings based on chitosan-beeswax emulsions

Dissertação para obtenção do Grau de Mestre em Engenharia Química e Bioquímica

Non-clinical isolates bring new findings on enterococcal virulence

Dissertation presented to obtain the PhD degree in Biology

Study of membrane emulsification process as a pre-step for the microencapsulation of lipid compounds by spray drying

Dissertation presented to Faculdade de Ciências e Tecnologia, Universidade Nova de Lisboa for obtaining the master degree in Membrane Engineering

Deeper insights into SRB-driven biocorrosion mechanisms

Dissertação para obtenção do Grau de Doutor em Química Sustentável

Porous structures for the purification of biopharmaceuticals

This work aimed at the development of a (bio)polymeric monolithic support for biopharmaceuticals purification and/or capture. For that, it was assured that functional groups on its surface were ready to be involved in a plethora of chemical reactions for incorporation of the desired and most suitable ligand. Using cryogelation as preparation method a screening on multiple combinations of materials was performed in order to create a potentially efficient support with the minimal footprint, i.e. a monolithic support with reasonable mechanical properties, highly permeable, biocompatible, ready to use, with gravitational performance and minimal unspecific interactions towards the target molecules, but also biodegradable and produced from renewable materials. For the pre-selection all monoliths were characterized physico-chemically and morphologically; one agarose-based and two chitosan-based monoliths were then subjected to further characterizations before and after their modification with magnetic nanoparticles. These three specimens were finally tested towards adenovirus and the recovery reached 84% for the chitosan-GMA plain monolith prepared at -80°C. Monoliths based on chitosan and PVA were prepared in the presence and absence of magnetic particles, and tested for the isolation of GFP directly from crude cellular extracts. The affinity ligand A4C7 previously selected for GFP purification was synthesized on the monolith. The results indicated that the solid-phase synthesis of the ligand directly onto the monolith might require optimization and that the large pores of the monoliths are unsuitable for the purification of small proteins, such as GFP.

Role of surfactants in filtration and fouling of colloidal silica

The EM3E Master is an Education Programme supported by the European Commission, the European Membrane Society (EMS), the European Membrane House (EMH), and a large international network of industrial companies, research centres and universities

Writing/erasing 3D micro and nano wrinkles in flexible elastomers for volatile organic compounds sensor

Micro/nano wrinkled patterns on cross-linked urethane/urea polymeric flexible free standing films with two soft segments, polypropylene oxide and polybutadiene, can be induced by UV-irradiation. The ability to write/erase these 3D structures, in a controlled manner, is the main focus of this work. The imprinting of the wrinkled structures was accomplished by swelling in an appropriate solvent followed by drying the membranes after the cross-linking process and UV irradiation. The surface tailoring of the elastomeric membranes was imaged by optical microscopy, scanning electronic microscopy and by atomic force microscopy. To erase the wrinkled structures the elastomers were swollen. The swelling as well as the sol/gel fraction and the UV radiation were tuned in order to control the wrinkles characteristics. It was found that the wrinkles wavelength, in the order of microns (1±0,25μm), was stamped by the UV radiation intensity and exposure time while the wrinkles' amplitude, in the order of nanometers (150-450 nm), was highly dependent on the swelling and sol/gel fraction. A prototype for volatile organic compounds detection was developed taking advantage of the unique 3D micro/nano wrinkles features.

Biological effects of acrylic engineered particulate-systems

Polymeric particulate-systems are of great relevance due to their possible biomedical applications, among them as carriers for the nano- or microencapsulation of drugs. However, due to their unique specific properties, namely small size range, toxicity issues must be discarded before allowing its use on health-related applications. Several polymers, as poly(methyl methacrylate) (PMMA), have proved to be suitable for the preparation of particulate-systems. However, a major drawback of its use refers to incomplete drug release from particles matrix. Recent strategies to improve PMMA release properties mention the inclusion of other acrylic polymers as Eudragit (EUD) on particles formulation. Though PMMA and EUD are accepted by the FDA as biocompatible, their safety on particle composition lacks sufficient toxicological data. The main objective of this thesis was to evaluate the biological effects of engineered acrylic particulate-systems. Preparation, physicochemical characterization and in vitro toxicity evaluation were assessed on PMMA and PMMA-EUD (50:50) particles. The emulsification-solvent evaporation methodology allowed the preparation of particles with spherical and smooth surfaces within the micrometer range (±500 nm), opposing surface charges and different levels of hydrophobicity. It was observed that particles physicochemical properties (size and charge) were influenced by biological media composition, such as serum concentration, ionic strength or pH. In what concerns to the in vitro toxicological studies, particle cellular uptake was observed on different cell lines (macrophages, osteoblasts and fibroblasts). Cytotoxicity effects were only found after 72 h of cells exposure to the particles, while no oxidative damage was observed neither on osteoblasts nor fibroblasts. Also, no genotoxicity was found in fibroblast using the comet assay to assess DNA damage. This observation should be further confirmed with other validated genotoxicity assays (e.g. Micronucleus Assay). The present study suggests that the evaluated acrylic particles are biocompatible, showing promising biological properties for potential use as carriers in drug-delivery systems.

Influence of the molecular weight in mechanical properties and degradation kinetics of chitosan inverted colloidal crystals

Tissue engineering arises from the need to regenerate organs and tissues, requiring the development of scaffolds, which can provide an optimum environment for tissue growth. In this work, chitosan with different molecular weights was used to develop biodegradable 3D inverted colloidal crystals (ICC) structures for bone regeneration, exhibiting uniform pore size and interconnected network. Moreover, in vitro tests were conducted by studying the influence of the molecular weight in the degradation kinetics and mechanical properties. The production of ICC included four major stages: fabrication of microspheres; assembly into a cohesive structure, polymeric solution infiltration and microsphere removal. Chitosan’s degree of deacetylation was determined by infrared spectroscopy and molecular weight was obtained via capillary viscometry. In order to understand the effect of the molecular weight in ICC structures, the mass loss and mechanical properties were analyzed after degradation with lysozyme. Structure morphology observation before and after degradation was performed by scanning electron microscopy. Cellular adhesion and proliferation tests were carried out to evaluate ICC in vitro response. Overall, medium molecular weight ICC revealed the best balance in terms of mechanical properties, degradation rate, morphology and biological behaviour.

Design and characterization of novel biopolymeric structure using biocompatible ionic liquids

The main objective of this thesis was the development of polymeric structures from the dissolution of FucoPol, a bacterial exopolysaccharide (EPS), in a biocompatible ionic liquid, choline acetate. The FucoPol was produced by the bacteria Enterobacter A47 using glycerol as carbon source at controlled temperature and pH (30ºC and 7, respectively). At the end of 3 days it was produced 7 g/L of FucoPol. The net yield of Fucopol in glycerol (YP/S) was 0.22 g/g and the maximum productivity 2.37 g/L.d This polymer was characterized about its composition in sugars and acyl groups (by High-Performance Liquid Chromatography - HPLC), containing fucose (35 % mol), galactose (21 % mol), glucose (29 % mol), rhamnose (3% mol) and glucuronic acid (12% mol) as well as acetate (14.28 % mol), pyruvate (2.15 % mol) and succinate (1.80 % mol). Its content of water and ash was 15% p/p and 2% p/p, respectively, and the chemical bonds (determined by Infrared Spectroscopy - FT-IR) are consistent to the literature reports. However, due to limitations in Differential Scanning Calorimetry (DSC) equipment it was not possible to determine the glass transition temperature. In turn, the ionic liquid showed the typical behavior of a Newtonian fluid, glass transition temperature (determined by DSC) -98.03ºC and density 1.1031 g/cm3. The study of chemical bonds by FT-IR showed that amount of water (8.80%) influenced the visualization of the bands predicted to in view of their chemical structure. After the dissolution of the FucoPol in the ionic liquid at different temperatures (50, 60, 80 and 100 ° C) it was promoted the removal of this by the phase inversion method using deionized water as a solvent, followed by drying in an oven at 70 ° C. The mixtures before and after the phase inversion method were characterized through the studies mentioned above. In order to explore possible application field’s biocompatibility assays and collage on balsa wood tests were performed. It was found that the process of washing with water by the phase inversion method was not totally effective in removing the biocompatible ionic liquid, since all FucoPol – IL mixtures still contained ionic liquid in their composition as can be seen by the DSC results and FT-IR. In addition, washing the mixtures with water significantly altered the composition of FucoPol. However, these mixtures, that developed a viscous behavior typical of a non-Newtonian fluid (shear-thinning), have the potential to be applied in the biomedical field as well as biological glues.

Nanocomposite polymer beads for cell detection

Circulating tumor cells (CTCs) may induce metastases when detached from the primary tumor. The numbers of these cells in blood offers a valuable prognostic indication. Magnetoresistive sensing is an attractive option for CTC counting. In this technique, cells are labeled with nancomposite polymer beads that provide the magnetic signal. Bead properties such as size and magnetic content must be optimized in order to be used as a detection tool in a magnetoresistive platform. Another important component of the platform is the magnet required for proper sensing. Both components are addressed in this work. Nanocomposite polymer beads were produced by nano-emulsion and membrane emulsification. Formulations of the oil phase comprising a mixture of aromatic monomers and iron oxide were employed. The effect of emulsifier (surfactant) concentration on bead size was studied. Formulations of polydimethilsiloxane (PDMS) with different viscosities were also prepared with nano-emulsion method resulting in colloidal beads. Polycaprolactone (PCL) beads were also synthetized by the membrane emulsification method. The beads were characterized by different techiques such as dynamic light scattering (DLS), thermogravimetric analysis (TGA) and scanning electron microscopy (SEM). Additionally, the magnet dimensions of the platform designed to detect CTCs were optimized through a COMSOL multiphysics simulation.

Production and characterization of electrospun composite fibers: confinement of thermosensitive microgels

Materials engineering focuses on the assembly of materials´ properties to design new products with the best performance. By using sub-micrometer size materials in the production of composites, it is possible to obtain objects with properties that none of their compounds show individually. Once three-dimensional materials can be easily customized to obtain desired properties, much interest has been paid to nanostructured poly-mers in order to build biocompatible devices. Over the past years, the thermosensitive microgels have become more common in the framework of bio-materials with potential applicability in therapy and/or diagnostics. In addition, high aspect ratio biopolymers fibers have been produced using the cost-effective method called electrospinning. Taking advantage of both microgels and electrospun fibers, surfaces with enhanced functionalities can be obtained and, therefore employed in a wide range of applications. This dissertation reports on the confinement of stimuli-responsive microgels through the colloidal electro-spinning process. The process mainly depends on the composition, properties and patterning of the precur-sor materials within the polymer jet. Microgels as well as the electrospun non-woven mats were investigated to correlate the starting materials with the final morphology of the composite fibers. PNIPAAm and PNIPAAm/Chitosan thermosensitive microgels with different compositions were obtained via surfactant free emulsion polymerization (SFEP) and characterized in terms of chemical structure, morphology, thermal sta-bility, swelling properties and thermosensitivity. Finally, the colloidal electrospinning method was carried out from spinning solutions composed of the stable microgel dispersions (up to a concentration of about 35 wt. % microgels) and a polymer solution of PEO/water/ethanol mixture acting as fiber template solution. The confinement of microgels was confirmed by Scanning Electron Microscopy (SEM). The electrospinning process was statistically analysed providing the optimum set of parameters aimed to minimize the fiber diameter, which give rise to electrospun nanofibers of PNIPAAm microgels/PEO with a mean fiber diameter of 63 ± 25 nm.