Information systems used in archaeological research have several needs that can be summarized as follows: interoperability among diverse, heterogeneous systems, making information available without significant delay, providing a sustainable approach to long-term preservation of data, and providing a suite of services to users of the system. In this thesis, we describe how digital library techniques can be employed to provide solutions to these problems and describe our experiences in creating a prototype for ETANA-DL. ETANA-DL is a model-based, componentized, extensible, archaeological Digital Library that manages complex information sources using the client-server paradigm of the Open Archives Initiative Protocol for Metadata Harvesting (OAI-PMH). We have designed and developed the prototype system with the following main goals: 1) to achieve information sharing between different heterogeneous archaeological systems, 2) to make primary archaeological data rapidly available to users, 3) to provide useful services to users of the DL, 4) to elicit requirements that users of the system will have beyond the services that it supports, and 5) to provide a sustainable solution to long-term preservation of valuable archaeological data. Consequently, we describe our approach to handling heterogeneous archaeological information from disparate sources; suggest an architecture for ETANA-DL, to be validated through prototyping; and show that given a pool of components that implement common DL services, a prototype DL can be rapidly created that supports several useful services over integrated data. Further, and most fundamentally, we note that understanding complex information systems is a difficult task. Finally, therefore, we describe our efforts to model complex archaeological information systems using the 5S framework, and show how we have used the resulting partial models to implement ETANA-DL with cross-collection searching and browsing capabilities. / Master of Science

The discovery that negatively charged aggregates of C60 fullerene are stable in aqueous environments has elicited concerns regarding the potential environmental and health effects of these aggregates. Although many previous studies have used aggregates synthesized using intermediate organic solvents, this work primarily employed an aggregate production method that more closely emulates the fate of C60 upon accidental release into the environment — extended mixing in water. The aggregates formed via this method (aqu/nC60) differ from those produced using the more common solvent exchange methods. The aqu/nC60 aggregates are heterogeneous in size (20 nm and larger) and shape (angular to round), negatively charged, and crystalline in structure — exhibiting a face centered cubic (fcc) system. Solution characteristics such as aqu/nC60 aggregate size and concentration were found to be dependant upon preparation variables such as stirring time, initial C60 concentration, and initial particle size. Additional experiments indicate that aggregate charge, structure, and stability are highly dependant upon the identity of co-solutes (NaCl, CaCl2, sodium citrate) and their concentrations. Citrate concentrations greater than 0.5 mM resulted in the formation of very small (< 20 nm) spherical aqu/nC60 particles. At moderate citrate concentrations (~ 1 mM) a more negative surface charge was observed, which may be an indication of increased nC60 stability. In contrast, high concentrations of monovalent and divalent electrolytes result in aggregation and sedimentation of nC60 out of solution. Our research describes the effect that solution composition has on aggregate formation and stability, and suggests that C60 fate and transport will be a function of solution composition. / Master of Science

A hemoglobina extracelular de Glossoscolex paulistus (HbGp) possui estrutura oligomérica composta por 144 cadeias globínicas e 36 cadeias linkers, que não possuem heme, formando uma bicamada hexagonal. Estudos mostraram que a HbGp possui uma alta estabilidade á variação de pH e presença de agentes desnaturantes, tais como, surfactantes e ureia, a 25&deg;C. Com esses conhecimentos prévios, o presente estudo tem por objetivo avaliar a estabilidade térmica da HbGp 0,5-3,0 mg/mL, nas formas oxi-, meta- e cianometa-, em diferentes valores de pH. O efeito do SDS na estabilidade térmica da oxi-HbGp 0,5 e 3,0 mg/mL, em função do pH, será investigado também. Esses estudos foram realizados usando as técnicas de absorção óptica, dicroísmo circular (CD), espalhamento de luz dinâmico (DLS) e espalhamento de raios X a baixo ângulo (SAXS). Os resultados de absorção óptica e CD revelam que o processo de desnaturação da oxi- e cianometa-HbGp, no pH 7,0, envolve a formação das espécies oxidadas aquo-meta-HbGp e hemicromo. O processo de desnaturação é próximo ao modelo de dois estados, com uma temperatura crítica (Tc) de 58-59 &deg;C. No pH ácido, a proteína agrega a partir de 58 &deg;C. A cinética de agregação da oxi-HbGp, no pH 7,0, é dependente da concentração de proteína e da temperatura. Os dados de DLS mostram que a meta- e cianometa-HbGp 0,5 mg/mL, pH 7,0, desnaturam a 48 &plusmn; 1 e 56 &plusmn; 1 &deg;C, respectivamente. Em pH alcalino, a proteína dissocia parcialmente antes de desnaturar, e o aumento da concentração de proteína faz aumentar o valor de Tc para a cianometa-HbGp. Dados de SAXS mostram que a oxi- e meta-HbGp, pH 7,0, desnaturam a 60 &deg;C, apresentando valores de Rg=143&plusmn;1 &Aring; e Dmax=450&plusmn;1 &Aring;, enquanto que a cianometa-HbGp se mantém estável, com valores de Rg=107&plusmn;1 &Aring; e Dmax=300&plusmn;1 &Aring;. As análises das curvas p(r) mostram uma porcentagem crescente de dodecâmero e tetrâmero em solução, em relação à fração de protreína íntegra e de subunidades maiores, com o aumento do pH e da temperatura. As análises baseadas no programa OLIGOMER são similares às baseadas na função p(r). A presença do SDS induz a dissociação da oxi-HbGp 0,5 mg/mL pH 7,0. Entretanto, com 3,0 mg/mL de proteína a dissociação é parcial, com a sobreposição dos processos de dissociação, desnaturação e agregação, com o aumento da temperatura. No pH 5,0, o SDS promove a agregação da oxi-HbGp em temperaturas menores. As constantes cinéticas de dissociação da oxi-HbGp 0,5 mg/mL, pH 7,0 aumentam de (0,53&plusmn;0,07)x10-4 s-1 para (2,1&plusmn;0,2)x10-4 s-1 na presença de 0,4 e 0,6 mmol/L de SDS a 40 &deg;C, respectivamente. Na temperatura de 42 &deg;C a constante aumenta 2,6 vezes, com 0,6 mmol/L de SDS, comparada a 40 &deg;C. A oxi-HbGp 3,0 mg/ml na presença de 0,6 mmol/L de SDS, dissocia parcialmente em tempos menores com o aumento da temperatura antes de agregar. Portanto, esses estudos mostram que a estabilidade térmica da HbGp é sensível ao aumento de pH e presença de SDS. A ordem de estabilidade térmica em pH alcalino é dado por: cianometa->oxi->meta-HbGp. Alem disso, o processo de desnaturação é governado pelo valor do pH e dependente da concentração de proteína em solução. / The extracellular hemoglobin Glossoscolex paulistus of (HbGp) has an oligomeric structure composed by 144 globin chains and 36 non globin chains (named linkers), forming a hexagonal bilayer. HbGp presents a high stability reagarding pH variation and the presence of denaturing agents, such as, for example, urea and surfactant, at 25&deg;C. In this way, the present studies aim to evaluate the thermal stability for oxy-, meta- and cyanomet-HbGp 0.5-3.0 mg/ml, at different pH values. The SDS effect on the thermal stability of oxy-HbGp 0.5 and 3.0 mg / mL is also investigated. Optical absorption, circular dichroism (CD), dynamic light scattering (DLS) and small angle X-ray scattering (SAXS) techniques were emplayed for these studies. The results based on the optical absorption and CD spectroscopies show that the denaturation process for oxy- and cyanomet-HbGp, at pH 7.0, involves the formation of oxidized species, such as aquo-met-HbGp and hemichrome. This denaturation process is very close to a two-state model, with a critical temperature (Tc) of 58-59 &deg;C. However, in the acidic pH, the aggregation of protein occurs at 58 &deg;C. The aggregation process kinetics for oxy-HbGp, pH 7.0, is dependent on the protein concentration and temperature. DLS data show that meta- and cyanomet-HbGp, 0.5 mg/mL, pH 7.0, undergo denaturation at 48 &plusmn; 1 and 56 &plusmn; 1 &deg; C, respectively. At alkaline pH, two HbGp forms undergo partial dissociation before denaturation, and at higher protein concentration, an increase of Tc values for cyanomet-HbGp is observed. SAXS results show that the denaturation of oxy-and met-HbGp occur at 60 &deg;C, presenting Rg=143&plusmn;1 &Aring; and Dmax=450&plusmn;15 &Aring;, while cyanomet-HbGp remains stable with Rg =107&plusmn;1 &Aring; and Dmax= 300&plusmn;10&Aring;, at this temperature. The p(r) curves analysis show the increase of dodecamer and tetramer percentages in solution, with increase of pH and temperature. The results using the OLIGOMER program are similar to the p(r) data analysis. For oxy-HbGp 0.5 mg/mL pH 7.0, in the presence of SDS, oligomeric dissociation before denaturation is observed. However, with 3.0 mg/ml of protein the dissociation process is slower, showing an overlap of the dissociation, denaturation and aggregation processes in the system, with increase of temperature. At pH 5.0, SDS promotes the aggregation of oxy-HbGp at lower temperatures, as compared to the absence of surfactant. The kinetic dissociation constant values for oxy-HbGp 0.5 mg/mL increase from (0.53 &plusmn; 0.07) x10-4 s-1 to (2.1 &plusmn; 0.2) x10-4 s-1, in the presence of 0.4 and 0.6 mmol/L SDS at 40 &deg; C, respectively. At 42 &deg;C the dissociation constant value increases 2.6-fold, with 0.6 mmol/L SDS, as compared to 40 &deg;C. For oxy-HbGp 3.0 mg/ml, in the presence of 0.6 mmol/L SDS, the oligomeric dissociation is smallest occurring in shorter times with increasing temperature before aggregation. Therefore, these studies show that the thermal stability of HbGp is sensitive to the pH variation and the presence of SDS. At alkaline pH, the order of thermal stability is the following: cyanomet->oxy->met-HbGp. Furthermore, the denaturation process is governed by the pH value, being dependent on the protein concentration in solution.

Na primeira parte deste trabalho foi estudada a interação da porfirina catiônica meso-tetrakis N-metil-4-piridil (TMPyP) e a porfirina aniônica meso-tetrakis 4-fenilsulfonato (TPPS4) nas formas de base livre com sistemas modelos de membrana biológica (micelas iônicas, micelas mistas e vesículas de fosfolipídios) em soluções aquosas, através das técnicas de absorção ótica, espalhamento de luz ressonante (RLS do inglês \"resonante light scattering), fluorescência e SAXS, do inglês \"Small Angle X-Ray Scattering\". As curvas de SAXS das micelas catiônicas de CTAC (cloreto de cetiltrimetilamônio) foram ajustadas como um elipsóide prolato na ausência e na presença de 2-10 mM de TPPS4. Os dados de SAXS mostraram que a presença da porfirina TPPS4 modifica o centro hidrofóbico micelar, levando a formação de micelas menores. Através das análises dos dados de SAXS das micelas de SDS (dodecilsulfato de sódio) observamos que a forma da micela na ausência e na presença de 2-10 mM TMPyP apresenta a forma de um elipsóide prolato sem mudanças. Entretanto, o coeficiente de ionização, diminuiu com o aumento da concentração de porfirina, sugerindo a \"blindagem\" da carga aniônica do SDS pela porfirina catiônica. A supressão de fluorescência da TPPS4 e TMPyP foi estudada na ausência e na presença de diferentes micelas de surfactantes, tais como as de SDS, CTAC, HPS (N-hexadecil-N,N,dimetil-3-amônio-1-propano sulfato) e TX-100 (t-octil-fenoxi-polietoxi-etanol). O iodeto de potássio (KI) foi utilizado como supressor. Os gráficos de Stern-Volmer dos dados de fluorescência no estado estacionário foram ajustados pela equação quadrática, incluindo a supressão dinâmica (KD) e estática (KS). Os valores de KS são muito menores do que os valores de KD. Os resultados da TMPyP são consistentes com as constantes de ligação reportadas na literatura: uma redução significativa de supressão acontece para a TMPyP na presença de SDS, e uma redução moderada é observada para o sistema TMPyP-HPS e quase nenhuma mudança é vista para a TMPyP na presença de TX-100. Para o sistema CTAC-TPPS4 um aumento na supressão foi observado quando comparada com a TPPS4 em tampão puro. Isto provavelmente é associado ao acúmulo de iodeto na interface da micela catiônica. A atração entre a cabeça polar do CTAC e I-, e a repulsão entre SDS e I-, aumenta e reduz a supressão de fluorescência, respectivamente, das porfirinas que se localizam na interface micelar. A pequena supressão da TPPS4 em TX-100 é coerente com a forte ligação entre a TPPS4-TX-100 reportada na literatura. A TPPS4 e a TMPyP na presença de concentrações baixas dos surfactantes CTAC e SDS, respectivamente, apresentaram formação de agregados pré-micelares. A adição de surfactante neutro, TX-100, reduziu o efeito de agregação, acompanhada pelas várias técnicas espectroscópicas utilizadas neste trabalho. Portanto, sob condições onde temos a máxima formação de agregados (porfirina-surfactante), a titulação da TPPS4 com micelas de 40%CTAC-60%TX-100 e a TMPyP com micelas de 80%SDS-20%TX-100 não foi suficiente para eliminar a agregação, apesar da diminuição significativa do efeito de supressão de fluorescência e da intensidade de luz espalhada. A interação da TMPyP com 1-Palmitoil-2-Oleoil-sn-Glicero-3-fosfocolina (POPC), 1-Palmitoil-2-Oleoil-sn-Glicero-3-[Fosfo-rac-(1-glicero)] (POPG) e a mistura POPC + POPG é predominantemente devido à contribuição de eletrostática. O aumento da carga negativa, devido à adição de POPG, favorece a interação das vesículas com a porfirina catiônica. Na segunda parte deste trabalho foram estudados os efeitos de três surfactantes na estrutura oligomérica da hemoglobina extracelular gigante de Glossoscolex paulistus (HbGp) na sua forma oxi. O estudo com o SDS, CTAC e HPS permitiu diferenciar os efeitos de cargas opostas da cabeça polar dos surfactantes na dissociação da estrutura oligomérica e na autoxidação da hemoglobina. A interação do HPS com HbGp foi claramente menos intensa que a interação desta hemoglobina com os surfactantes catiônico (CTAC) e aniônico (SDS). Provavelmente, esta menor interação da HbGp com HPS, quando comparada com o SDS e o CTAC, é devido a menor atração eletrostática entre o HPS e os sítios iônicos da proteina. Dados espectroscópicos foram discutidos e comparados com os da literatura, afim de compreender a interação hemoglobina-surfactante, e como o ponto isoelétrico ácido (pI) pode influenciar na relação da estrutura-atividade das hemoglobinas gigantes extracelulares. As amostras de HbGp foram estudadas por espalhamento de luz dinâmico (DLS do inglês \"Dynamic light scattering\"). Na faixa de pH 6.0 a 8.0, HbGp é bastante estável e a distribuição de tamanho das partículas é monodispersa com um diâmetro hidrodinâmico médio (Dh) de 27 nm. O aumento dos valores de pH (pH>9.0) induziu um processo de dissociação irreversível, resultando num valor do Dh menor (10 nm). A diminuição do Dh sugere uma dissociação completa da hemoglobina. Em pH>9,0 a cinética de dissociação é lenta, com um mínimo 24 h para ser completada. As constantes cinéticas de dissociação aumentam progressivamente, com o aumento do valor do pH. As curvas de melting point para HbGp apresentaram dissociação oligomérica e desnaturação da proteina em função do pH. Os processos de autoxidação e dissociação estão intimamente relacionados, de modo que a dissociação da proteina oligomérica promove um aumento na velocidade de autoxidação e vice-versa. / In the first part of this work interactions of the cationic meso tetrakis (4-N-methilpyridil) porphyrin (TMPyP) and meso-tetrakis (4-sulfonatophenyl) porphyrin (TPPS4) in the free base forms with membrane model systems (ionic micelles, mixed micelles and phospholipids vesicles) in aqueous solutions, have been investigated by optical absorption, resonance light scattering (RLS), fluorescence and SAXS (Small Angle X-Ray Scattering). The best-fit SAXS curves were obtained assuming for cetyltrimethylammonium chloride (CTAC) micelle a prolate ellipsoidal shape in the absence and upon incorporation of 2-10 mM TPPS4. SAXS results show that the presence of TPPS4 impacts on micellar hydrophobic core, leading to a micellar reassembling into smaller micelles. SAXS data analysis demonstrated a prolate ellipsoidal shape for sodium dodecyl sulfate (SDS) micelles; no significant changes in shape and size were observed for SDS-TMPyP co-micelles. Moreover, the ionization coefficient, &alpha;, decreases with the increase of the porphyrin concentration, suggesting the \"screening\" of the anionic charge of SDS by the cationic porphyrin. These results are consistent with optical absorption, fluorescence and RLS spectroscopies data, allowing to conclude that neutral surfactants present a smaller interaction with the cationic porphyrin as compared with ionic surfactants. Fluorescence quenching of TPPS4 and TMPyP is studied in aqueous solution and upon addition of micelles of SDS, CTAC, N-hexadecyl-N,N-dimethyl-3-ammonio-1- propanesulfonate (HPS) and t-octylphenoxypolyethoxyethanol (Triton X-100). Potassium iodide (KI) was used as quencher. Steady-state Stern-Volmer plots were best fitted by a quadratic equation, including dynamic (KD) and static (KS) quenching. KS was significantly smaller than KD. For TMPyP quenching results are consistent with reported binding constants: a significant reduction of quenching takes place for SDS, a moderate reduction is observed for HPS and almost no change is seen for Triton X-100. For CTAC-TPPS4 system an enhancement of quenching was observed as compared to pure buffer. This is probably associated to accumulation of iodide at the cationic micellar interface. The attraction between CTAC headgroups and I-, and repulsion between SDS and I-, enhances and reduces the fluorescence quenching, respectively, of porphyrins located at the micellar interface. The small quenching of TPPS4 in Triton X-100 is consistent with strong binding as reported in the literature. Anionic TPPS4 and cationic TMPyP in the presence of low concentrations of the surfactants CTAC and SDS, respectively, showed formation of aggregates, monitored by optical absorption, fluorescence and resonance light scattering intensity (RLS). The addition of nonionic surfactant, Triton X-100, reduced the effect of aggregation monitored by the various techniques used in the present work. Therefore, under conditions for the maximum of aggregate formation (porphyrin-surfactant), apparently, the CTAC: TX-100 ratio equal to 40:60 and SDS:TX-100 ratio equal to 80:20 are not sufficient to eliminate aggregation, despite the significant decrease of the quenching effect of fluorescence and of the light scattering intensity. The interaction of TMPyP with 1-Palmitoyl-2-Oleoyl-sn- Glycero-3-Phosphocholine (POPC), 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-[Phospho-rac-(1- glycerol)] (POPG) and the mixture POPC+POPG is predominantly due to the electrostatic contribution. The increase of the negative charge, due to addition of POPG, favors the interaction of vesicles with the cationic porphyrin. On the second part of this work the effects of three surfactants upon the oligomeric structure of the giant extracellular hemoglobin of Glossoscolex paulistus (HbGp) in the oxy - form was studied. The use of SDS, CTAC and HPS has allowed to differentiate the effects of opposite headgroup charges on the oligomeric structure dissociation and hemoglobin autoxidation. Furthermore, the interaction of HPS with HbGp was clearly less intense than the interaction of this hemoglobin with cationic (CTAC) and anionic (SDS) surfactants. Probably, this lower interaction with HPS is due to the lower electrostatic attraction between the HPS surfactant and the protein surface ionic sites when compared to the electrostatic interaction between HbGp and cationic and anionic surfactants. Spectroscopic data are discussed and compared with the literature in order to improve the understanding of hemoglobin-surfactant interaction as well as the acid isoelectric point (pI) influence of the giant extracellular hemoglobins on its structure-activity relationship. HbGp samples were studied by dynamic light scattering (DLS). In the pH from range 6.0 to 8.0, HbGp is stable and a monodisperse size distribution with a z-average hydrodynamic diameter (Dh) of 27±1 nm is observed. More alkaline pH (pH>9.0) induced an irreversible dissociation process, resulting in smaller Dh of 10±1 nm. Dh decrease suggests a complete hemoglobin dissociation. At pH 9.0 the dissociation kinetics is slow, taking a minimum of 24 h to be completed. Dissociation rate constants progressively increase at higher pH. Melting curves for HbGp showed oligomeric dissociation and protein denaturation as a function of pH. Autoxidation and dissociation processes are intimately related, so that oligomeric protein dissociation promotes the increase of autoxidation rate and vice-versa.

L'utilisation massive des nanomatériaux pose de réels enjeux sanitaires et environnementaux. C'est pourquoi ils sont désormais soumis à une réglementation qui prévoit une traçabilité de ceux-ci depuis leur fabrication jusqu'à leur distribution et l'établissement d'une fiche d'identité de la substance (composition, taille, état d'agglomération, forme, etc.). Une routine de caractérisation de nanoparticules en suspension a ainsi été développée. La Microscopie Électronique en Transmission (MET) a permis d'établir une majorité des paramètres de la fiche d'identité, en combinant à la fois imagerie et spectroscopie (analyses chimiques). La préparation, dont dépendra la qualité des observations, nécessite un développement pour chaque matériau analysé. Pour cela, trois techniques ont été mises au point : le dépôt en voie sèche qui permet une observation directe et simple, la cryogénie qui permet de fixer l'état de la suspension et l'in-situ liquide qui permet d'observer directement la suspension sans changement d'état. Les analyses MET étant locales, une comparaison avec des techniques indirectes a été effectuée par Diffusion Statique (MALS) et Dynamique (DLS) de la Lumière avec et sans fractionnement par couplage flux-force (FFF). Deux matériaux modèles ont été choisis. Le premier est une nanoémulsion de lipides stabilisés par des surfactants, servant de vecteurs à des principes actifs. Une étude de vieillissement par interaction avec des protéines a été menée et de légères variations de taille ont été obtenues. Le second matériau sélectionné est une poudre de nanoparticules de dioxyde de titane, remises en suspension, utilisée dans les crèmes solaires en tant que filtres UV. Ces particules ont été observées avant et après passage en enceinte climatique afin d'observer les effets des rayons UV sur celles-ci. Ceci a confirmé la stabilité des particules. Les protocoles de caractérisation développés au cours de cette thèse peuvent ainsi servir de supports à l'étude d'autres nanoparticules en suspension. / The extensive use of nanomaterials has raised awareness about health issues and their fate in the environment. That is why they are now subject to regulation that has imposed their traceability from their manufacturing to their distribution as the establishment of their characteristics (chemical composition, size, agglomeration state, shape ...). A characterization routine for nanoparticles in suspension was developed. Transmission Electron Microscopy (TEM) fulfills most of the criteria cited before by combining imaging and spectroscopy techniques. Three sample preparation methods were optimized to ensure high quality results : a dry process, rapid freezing to vitrify the sample and the use of an textit{in-situ} liquid TEM holder to prevent any preparation artefact (no phase change). To obtain quantitative analysis, a comparison was made between Dynamic Light Scattering (DLS), Multi-Angle Light Scattering (MALS), with and without a fractionation system (AF4), and TEM. To support this work, two nanomaterials were analyzed. The first one is a nanoemulsion composed of lipid nanoparticles stabilized by surfactants used as nanocarriers for drug delivery. Their stability after protein interaction was investigate and some size variations were observed. The second material is a powder composed of titanium dioxide nanoparticles used as UV filters in sunscreens. These nanoparticles were analyzed before and after interaction with UV radiation in a climatic chamber to confirm their stability. The different protocols developed in this PhD may be used for the analysis of other nanomaterials.

A hemoglobina extracelular de Glossoscolex paulistus (HbGp) possui estrutura oligomérica composta por 144 cadeias globínicas e 36 cadeias linkers, que não possuem heme, formando uma bicamada hexagonal. Estudos mostraram que a HbGp possui uma alta estabilidade á variação de pH e presença de agentes desnaturantes, tais como, surfactantes e ureia, a 25&deg;C. Com esses conhecimentos prévios, o presente estudo tem por objetivo avaliar a estabilidade térmica da HbGp 0,5-3,0 mg/mL, nas formas oxi-, meta- e cianometa-, em diferentes valores de pH. O efeito do SDS na estabilidade térmica da oxi-HbGp 0,5 e 3,0 mg/mL, em função do pH, será investigado também. Esses estudos foram realizados usando as técnicas de absorção óptica, dicroísmo circular (CD), espalhamento de luz dinâmico (DLS) e espalhamento de raios X a baixo ângulo (SAXS). Os resultados de absorção óptica e CD revelam que o processo de desnaturação da oxi- e cianometa-HbGp, no pH 7,0, envolve a formação das espécies oxidadas aquo-meta-HbGp e hemicromo. O processo de desnaturação é próximo ao modelo de dois estados, com uma temperatura crítica (Tc) de 58-59 &deg;C. No pH ácido, a proteína agrega a partir de 58 &deg;C. A cinética de agregação da oxi-HbGp, no pH 7,0, é dependente da concentração de proteína e da temperatura. Os dados de DLS mostram que a meta- e cianometa-HbGp 0,5 mg/mL, pH 7,0, desnaturam a 48 &plusmn; 1 e 56 &plusmn; 1 &deg;C, respectivamente. Em pH alcalino, a proteína dissocia parcialmente antes de desnaturar, e o aumento da concentração de proteína faz aumentar o valor de Tc para a cianometa-HbGp. Dados de SAXS mostram que a oxi- e meta-HbGp, pH 7,0, desnaturam a 60 &deg;C, apresentando valores de Rg=143&plusmn;1 &Aring; e Dmax=450&plusmn;1 &Aring;, enquanto que a cianometa-HbGp se mantém estável, com valores de Rg=107&plusmn;1 &Aring; e Dmax=300&plusmn;1 &Aring;. As análises das curvas p(r) mostram uma porcentagem crescente de dodecâmero e tetrâmero em solução, em relação à fração de protreína íntegra e de subunidades maiores, com o aumento do pH e da temperatura. As análises baseadas no programa OLIGOMER são similares às baseadas na função p(r). A presença do SDS induz a dissociação da oxi-HbGp 0,5 mg/mL pH 7,0. Entretanto, com 3,0 mg/mL de proteína a dissociação é parcial, com a sobreposição dos processos de dissociação, desnaturação e agregação, com o aumento da temperatura. No pH 5,0, o SDS promove a agregação da oxi-HbGp em temperaturas menores. As constantes cinéticas de dissociação da oxi-HbGp 0,5 mg/mL, pH 7,0 aumentam de (0,53&plusmn;0,07)x10-4 s-1 para (2,1&plusmn;0,2)x10-4 s-1 na presença de 0,4 e 0,6 mmol/L de SDS a 40 &deg;C, respectivamente. Na temperatura de 42 &deg;C a constante aumenta 2,6 vezes, com 0,6 mmol/L de SDS, comparada a 40 &deg;C. A oxi-HbGp 3,0 mg/ml na presença de 0,6 mmol/L de SDS, dissocia parcialmente em tempos menores com o aumento da temperatura antes de agregar. Portanto, esses estudos mostram que a estabilidade térmica da HbGp é sensível ao aumento de pH e presença de SDS. A ordem de estabilidade térmica em pH alcalino é dado por: cianometa->oxi->meta-HbGp. Alem disso, o processo de desnaturação é governado pelo valor do pH e dependente da concentração de proteína em solução. / The extracellular hemoglobin Glossoscolex paulistus of (HbGp) has an oligomeric structure composed by 144 globin chains and 36 non globin chains (named linkers), forming a hexagonal bilayer. HbGp presents a high stability reagarding pH variation and the presence of denaturing agents, such as, for example, urea and surfactant, at 25&deg;C. In this way, the present studies aim to evaluate the thermal stability for oxy-, meta- and cyanomet-HbGp 0.5-3.0 mg/ml, at different pH values. The SDS effect on the thermal stability of oxy-HbGp 0.5 and 3.0 mg / mL is also investigated. Optical absorption, circular dichroism (CD), dynamic light scattering (DLS) and small angle X-ray scattering (SAXS) techniques were emplayed for these studies. The results based on the optical absorption and CD spectroscopies show that the denaturation process for oxy- and cyanomet-HbGp, at pH 7.0, involves the formation of oxidized species, such as aquo-met-HbGp and hemichrome. This denaturation process is very close to a two-state model, with a critical temperature (Tc) of 58-59 &deg;C. However, in the acidic pH, the aggregation of protein occurs at 58 &deg;C. The aggregation process kinetics for oxy-HbGp, pH 7.0, is dependent on the protein concentration and temperature. DLS data show that meta- and cyanomet-HbGp, 0.5 mg/mL, pH 7.0, undergo denaturation at 48 &plusmn; 1 and 56 &plusmn; 1 &deg; C, respectively. At alkaline pH, two HbGp forms undergo partial dissociation before denaturation, and at higher protein concentration, an increase of Tc values for cyanomet-HbGp is observed. SAXS results show that the denaturation of oxy-and met-HbGp occur at 60 &deg;C, presenting Rg=143&plusmn;1 &Aring; and Dmax=450&plusmn;15 &Aring;, while cyanomet-HbGp remains stable with Rg =107&plusmn;1 &Aring; and Dmax= 300&plusmn;10&Aring;, at this temperature. The p(r) curves analysis show the increase of dodecamer and tetramer percentages in solution, with increase of pH and temperature. The results using the OLIGOMER program are similar to the p(r) data analysis. For oxy-HbGp 0.5 mg/mL pH 7.0, in the presence of SDS, oligomeric dissociation before denaturation is observed. However, with 3.0 mg/ml of protein the dissociation process is slower, showing an overlap of the dissociation, denaturation and aggregation processes in the system, with increase of temperature. At pH 5.0, SDS promotes the aggregation of oxy-HbGp at lower temperatures, as compared to the absence of surfactant. The kinetic dissociation constant values for oxy-HbGp 0.5 mg/mL increase from (0.53 &plusmn; 0.07) x10-4 s-1 to (2.1 &plusmn; 0.2) x10-4 s-1, in the presence of 0.4 and 0.6 mmol/L SDS at 40 &deg; C, respectively. At 42 &deg;C the dissociation constant value increases 2.6-fold, with 0.6 mmol/L SDS, as compared to 40 &deg;C. For oxy-HbGp 3.0 mg/ml, in the presence of 0.6 mmol/L SDS, the oligomeric dissociation is smallest occurring in shorter times with increasing temperature before aggregation. Therefore, these studies show that the thermal stability of HbGp is sensitive to the pH variation and the presence of SDS. At alkaline pH, the order of thermal stability is the following: cyanomet->oxy->met-HbGp. Furthermore, the denaturation process is governed by the pH value, being dependent on the protein concentration in solution.

Na primeira parte deste trabalho foi estudada a interação da porfirina catiônica meso-tetrakis N-metil-4-piridil (TMPyP) e a porfirina aniônica meso-tetrakis 4-fenilsulfonato (TPPS4) nas formas de base livre com sistemas modelos de membrana biológica (micelas iônicas, micelas mistas e vesículas de fosfolipídios) em soluções aquosas, através das técnicas de absorção ótica, espalhamento de luz ressonante (RLS do inglês \"resonante light scattering), fluorescência e SAXS, do inglês \"Small Angle X-Ray Scattering\". As curvas de SAXS das micelas catiônicas de CTAC (cloreto de cetiltrimetilamônio) foram ajustadas como um elipsóide prolato na ausência e na presença de 2-10 mM de TPPS4. Os dados de SAXS mostraram que a presença da porfirina TPPS4 modifica o centro hidrofóbico micelar, levando a formação de micelas menores. Através das análises dos dados de SAXS das micelas de SDS (dodecilsulfato de sódio) observamos que a forma da micela na ausência e na presença de 2-10 mM TMPyP apresenta a forma de um elipsóide prolato sem mudanças. Entretanto, o coeficiente de ionização, diminuiu com o aumento da concentração de porfirina, sugerindo a \"blindagem\" da carga aniônica do SDS pela porfirina catiônica. A supressão de fluorescência da TPPS4 e TMPyP foi estudada na ausência e na presença de diferentes micelas de surfactantes, tais como as de SDS, CTAC, HPS (N-hexadecil-N,N,dimetil-3-amônio-1-propano sulfato) e TX-100 (t-octil-fenoxi-polietoxi-etanol). O iodeto de potássio (KI) foi utilizado como supressor. Os gráficos de Stern-Volmer dos dados de fluorescência no estado estacionário foram ajustados pela equação quadrática, incluindo a supressão dinâmica (KD) e estática (KS). Os valores de KS são muito menores do que os valores de KD. Os resultados da TMPyP são consistentes com as constantes de ligação reportadas na literatura: uma redução significativa de supressão acontece para a TMPyP na presença de SDS, e uma redução moderada é observada para o sistema TMPyP-HPS e quase nenhuma mudança é vista para a TMPyP na presença de TX-100. Para o sistema CTAC-TPPS4 um aumento na supressão foi observado quando comparada com a TPPS4 em tampão puro. Isto provavelmente é associado ao acúmulo de iodeto na interface da micela catiônica. A atração entre a cabeça polar do CTAC e I-, e a repulsão entre SDS e I-, aumenta e reduz a supressão de fluorescência, respectivamente, das porfirinas que se localizam na interface micelar. A pequena supressão da TPPS4 em TX-100 é coerente com a forte ligação entre a TPPS4-TX-100 reportada na literatura. A TPPS4 e a TMPyP na presença de concentrações baixas dos surfactantes CTAC e SDS, respectivamente, apresentaram formação de agregados pré-micelares. A adição de surfactante neutro, TX-100, reduziu o efeito de agregação, acompanhada pelas várias técnicas espectroscópicas utilizadas neste trabalho. Portanto, sob condições onde temos a máxima formação de agregados (porfirina-surfactante), a titulação da TPPS4 com micelas de 40%CTAC-60%TX-100 e a TMPyP com micelas de 80%SDS-20%TX-100 não foi suficiente para eliminar a agregação, apesar da diminuição significativa do efeito de supressão de fluorescência e da intensidade de luz espalhada. A interação da TMPyP com 1-Palmitoil-2-Oleoil-sn-Glicero-3-fosfocolina (POPC), 1-Palmitoil-2-Oleoil-sn-Glicero-3-[Fosfo-rac-(1-glicero)] (POPG) e a mistura POPC + POPG é predominantemente devido à contribuição de eletrostática. O aumento da carga negativa, devido à adição de POPG, favorece a interação das vesículas com a porfirina catiônica. Na segunda parte deste trabalho foram estudados os efeitos de três surfactantes na estrutura oligomérica da hemoglobina extracelular gigante de Glossoscolex paulistus (HbGp) na sua forma oxi. O estudo com o SDS, CTAC e HPS permitiu diferenciar os efeitos de cargas opostas da cabeça polar dos surfactantes na dissociação da estrutura oligomérica e na autoxidação da hemoglobina. A interação do HPS com HbGp foi claramente menos intensa que a interação desta hemoglobina com os surfactantes catiônico (CTAC) e aniônico (SDS). Provavelmente, esta menor interação da HbGp com HPS, quando comparada com o SDS e o CTAC, é devido a menor atração eletrostática entre o HPS e os sítios iônicos da proteina. Dados espectroscópicos foram discutidos e comparados com os da literatura, afim de compreender a interação hemoglobina-surfactante, e como o ponto isoelétrico ácido (pI) pode influenciar na relação da estrutura-atividade das hemoglobinas gigantes extracelulares. As amostras de HbGp foram estudadas por espalhamento de luz dinâmico (DLS do inglês \"Dynamic light scattering\"). Na faixa de pH 6.0 a 8.0, HbGp é bastante estável e a distribuição de tamanho das partículas é monodispersa com um diâmetro hidrodinâmico médio (Dh) de 27 nm. O aumento dos valores de pH (pH>9.0) induziu um processo de dissociação irreversível, resultando num valor do Dh menor (10 nm). A diminuição do Dh sugere uma dissociação completa da hemoglobina. Em pH>9,0 a cinética de dissociação é lenta, com um mínimo 24 h para ser completada. As constantes cinéticas de dissociação aumentam progressivamente, com o aumento do valor do pH. As curvas de melting point para HbGp apresentaram dissociação oligomérica e desnaturação da proteina em função do pH. Os processos de autoxidação e dissociação estão intimamente relacionados, de modo que a dissociação da proteina oligomérica promove um aumento na velocidade de autoxidação e vice-versa. / In the first part of this work interactions of the cationic meso tetrakis (4-N-methilpyridil) porphyrin (TMPyP) and meso-tetrakis (4-sulfonatophenyl) porphyrin (TPPS4) in the free base forms with membrane model systems (ionic micelles, mixed micelles and phospholipids vesicles) in aqueous solutions, have been investigated by optical absorption, resonance light scattering (RLS), fluorescence and SAXS (Small Angle X-Ray Scattering). The best-fit SAXS curves were obtained assuming for cetyltrimethylammonium chloride (CTAC) micelle a prolate ellipsoidal shape in the absence and upon incorporation of 2-10 mM TPPS4. SAXS results show that the presence of TPPS4 impacts on micellar hydrophobic core, leading to a micellar reassembling into smaller micelles. SAXS data analysis demonstrated a prolate ellipsoidal shape for sodium dodecyl sulfate (SDS) micelles; no significant changes in shape and size were observed for SDS-TMPyP co-micelles. Moreover, the ionization coefficient, &alpha;, decreases with the increase of the porphyrin concentration, suggesting the \"screening\" of the anionic charge of SDS by the cationic porphyrin. These results are consistent with optical absorption, fluorescence and RLS spectroscopies data, allowing to conclude that neutral surfactants present a smaller interaction with the cationic porphyrin as compared with ionic surfactants. Fluorescence quenching of TPPS4 and TMPyP is studied in aqueous solution and upon addition of micelles of SDS, CTAC, N-hexadecyl-N,N-dimethyl-3-ammonio-1- propanesulfonate (HPS) and t-octylphenoxypolyethoxyethanol (Triton X-100). Potassium iodide (KI) was used as quencher. Steady-state Stern-Volmer plots were best fitted by a quadratic equation, including dynamic (KD) and static (KS) quenching. KS was significantly smaller than KD. For TMPyP quenching results are consistent with reported binding constants: a significant reduction of quenching takes place for SDS, a moderate reduction is observed for HPS and almost no change is seen for Triton X-100. For CTAC-TPPS4 system an enhancement of quenching was observed as compared to pure buffer. This is probably associated to accumulation of iodide at the cationic micellar interface. The attraction between CTAC headgroups and I-, and repulsion between SDS and I-, enhances and reduces the fluorescence quenching, respectively, of porphyrins located at the micellar interface. The small quenching of TPPS4 in Triton X-100 is consistent with strong binding as reported in the literature. Anionic TPPS4 and cationic TMPyP in the presence of low concentrations of the surfactants CTAC and SDS, respectively, showed formation of aggregates, monitored by optical absorption, fluorescence and resonance light scattering intensity (RLS). The addition of nonionic surfactant, Triton X-100, reduced the effect of aggregation monitored by the various techniques used in the present work. Therefore, under conditions for the maximum of aggregate formation (porphyrin-surfactant), apparently, the CTAC: TX-100 ratio equal to 40:60 and SDS:TX-100 ratio equal to 80:20 are not sufficient to eliminate aggregation, despite the significant decrease of the quenching effect of fluorescence and of the light scattering intensity. The interaction of TMPyP with 1-Palmitoyl-2-Oleoyl-sn- Glycero-3-Phosphocholine (POPC), 1-Palmitoyl-2-Oleoyl-sn-Glycero-3-[Phospho-rac-(1- glycerol)] (POPG) and the mixture POPC+POPG is predominantly due to the electrostatic contribution. The increase of the negative charge, due to addition of POPG, favors the interaction of vesicles with the cationic porphyrin. On the second part of this work the effects of three surfactants upon the oligomeric structure of the giant extracellular hemoglobin of Glossoscolex paulistus (HbGp) in the oxy - form was studied. The use of SDS, CTAC and HPS has allowed to differentiate the effects of opposite headgroup charges on the oligomeric structure dissociation and hemoglobin autoxidation. Furthermore, the interaction of HPS with HbGp was clearly less intense than the interaction of this hemoglobin with cationic (CTAC) and anionic (SDS) surfactants. Probably, this lower interaction with HPS is due to the lower electrostatic attraction between the HPS surfactant and the protein surface ionic sites when compared to the electrostatic interaction between HbGp and cationic and anionic surfactants. Spectroscopic data are discussed and compared with the literature in order to improve the understanding of hemoglobin-surfactant interaction as well as the acid isoelectric point (pI) influence of the giant extracellular hemoglobins on its structure-activity relationship. HbGp samples were studied by dynamic light scattering (DLS). In the pH from range 6.0 to 8.0, HbGp is stable and a monodisperse size distribution with a z-average hydrodynamic diameter (Dh) of 27±1 nm is observed. More alkaline pH (pH>9.0) induced an irreversible dissociation process, resulting in smaller Dh of 10±1 nm. Dh decrease suggests a complete hemoglobin dissociation. At pH 9.0 the dissociation kinetics is slow, taking a minimum of 24 h to be completed. Dissociation rate constants progressively increase at higher pH. Melting curves for HbGp showed oligomeric dissociation and protein denaturation as a function of pH. Autoxidation and dissociation processes are intimately related, so that oligomeric protein dissociation promotes the increase of autoxidation rate and vice-versa.

Ce travail de thèse s’inscrit dans une approche fondamentale expérimentale de l’étude de suspensions céramiques, au travers de la mise en place d’un banc de mesure de diffusion dynamique de la lumière (DLS) sur une suspension en écoulement. L’étude a été effectuée sur des suspensions céramiques concentrées de silice nanométrique et de laponite dans une géométrie de Couette. La technique de mesure par DLS permet d’obtenir des informations locales telles que le taux de cisaillement et la vitesse locale des particules. À travers ces mesures et d’observation vidéo, les profils de vitesse ont permis de comprendre l’évolution du comportement rhéologique de ces suspensions concentrées. De plus, les suspensions concentrées en laponite forment un gel lorsque celles-ci sont au repos. Si ce gel subit un cisaillement, il se déstructure pour redevenir une suspension « liquide ». Cependant, du gel apparait sous cisaillement et sa formation a également été observée par DLS et par vidéo. / This thesis is part of a fundamental experimental approach to the study of ceramics suspensions, through the establishment of a panel of dynamic light scattering (DLS) measurement to a flowing suspension. The study was performed on ceramic concentrated suspensions of nanometric silica and laponite in Couette geometry. The measurement technique by DLS provides local information such as the shear rate and the local velocity of the particles. Through these measurements and video observation, velocity profiles have allowed to understand the evolution of the rheological behavior of concentrated suspensions. Moreover, concentrated suspensions of laponite form a gel when these are at rest. If this gel undergoes shear, this one deconstructs to become a « liquid » suspension. However, the gel appears under shear and its formation was also observed by DLS and camera.

[pt] Os materiais compósitos foram desenvolvidos para substituir o uso de ligas metálicas em aplicações de alto desempenho, se tornando materiais muito utilizados devido a uma ampla combinação de propriedades mecânicas. O setor de oleodutos, com o passar dos anos, tem enfrentado problemas nas tubulações devido ao envelhecimento e deterioração causados, principalmente, pela corrosão. Para solucionar esses problemas, normalmente, são utilizados reparos convencionais feitos com materiais metálicos tendo a finalidade de substituir a parte danificada. Porém, esse processo é muito trabalhoso e o setor de oleodutos tem visto como alternativa o uso de materiais compósitos poliméricos reforçados com fibra como reparo. As principais vantagens em relação aos reparos convencionais são a alta relação resistência-peso, a baixa concentração de tensão e a fácil aplicação. Entretanto, muitos estudos já mostraram que quando expostos a ambientes agressivos como temperatura, umidade e radiação ultravioleta, os materiais compósitos podem se deteriorar, ou seja, perdem suas propriedades mecânicas. Com isso, é importante analisar o efeito desses ambientes com o intuito de prever o comportamento mecânico, a vida útil e as possíveis falhas do reparo de juntas coladas. Com a finalidade de entender o comportamento mecânico, a adesão e a vida útil da junta adesiva, o objetivo desse trabalho foi analisar o envelhecimento de reparos em uma atmosfera salina. Foram utilizadas juntas Double-Lap Shear – DLS de compósito polimérico reforçado com fibra de vidro sobre um substrato de aço. As juntas foram fabricadas com quatro tipos tratamentos superficiais diferentes: I) Sem Silano / lixamento manual; II) Sem Silano / máquina Monti; III) Com Silano / lixamento manual; IV) Com Silano / máquina Monti. Os reparos de juntas coladas foram analisados em relação aos efeitos de temperatura (35 graus C, 55 graus C e 70 graus C) e da exposição a ambientes de névoa salina por um tempo de envelhecimento de até 6830h. O efeito do envelhecimento e a variação das propriedades mecânicas foram avaliados pelo ensaio destrutivo DLS e foi utilizado o ensaio não destrutivo de ultrassom para detectar o tamanho e localização dos defeitos. Como resultado, observou-se que quanto maior a temperatura de envelhecimento mais rápida foi a degradação das propriedades mecânicas e que as amostras que possuem silano, mantiveram maior resistência ao longo do tempo de envelhecimento. Também foi observado que o modo de falha de junta mudou com o passar do tempo e da temperatura tornando a falha adesiva a mais comum. Em relação ao ensaio de ultrassom, foi observado que a técnica de ultrassom foi eficiente para localizar e dimensionar o tamanho dos defeitos. Porém, não foi possível fazer uma comparação do aumento da área de defeitos com a variação das propriedades mecânicas. / [en] Composite materials have been developed to replace the use of metal alloys in high-performance applications, becoming widely used materials due to their broad combination of mechanical properties. Over the years, the pipeline sector has faced problems in pipelines due to aging and deterioration caused mainly by corrosion. To solve these problems, conventional repairs made with metallic materials are usually used to replace the damaged part. However, this process is very laborious, and the pipeline sector has seen the use of fiber-reinforced polymer composite materials as an alternative for repair. The main advantages compared to conventional repairs are high strength-to-weight ratio, low stress concentration, and easy application. However, many studies have shown that when exposed to aggressive environments such as temperature, humidity, and ultraviolet radiation, composite materials may deteriorate, meaning they lose their mechanical properties. Therefore, it is important to analyze the effect of these environments to predict the mechanical behavior, service life, and possible failures of adhesive joint repairs. In order to understand the mechanical behavior, adhesion, and service life of the adhesive joint, the aim of this work was to analyze the aging of repairs in a saline atmosphere. Double-Lap Shear (DLS) joints made of fiberglass-reinforced polymer composite on a steel substrate were used. The joints were made with four different surface treatments: I) Without Silane / manual sanding; II) Without Silane / Monti machine; III) With Silane / manual sanding; IV) With Silane / Monti machine. The bonded joint repairs were analyzed for the effects of temperature (35 degrees C, 55 degrees C, and 70 degrees C) and exposure to salt spray environments for an aging time of up to 6830h. The effect of aging and variation of mechanical properties were evaluated by destructive DLS testing, and non-destructive ultrasonic testing was used to detect the size and location of defects. As a result, it was observed that the higher the aging temperature, the faster the degradation of mechanical properties, and that samples with silane maintained greater resistance over the aging time. It was also observed that the joint failure mode changed over time and temperature, making adhesive failure more common. Regarding ultrasonic testing, it was observed that the technique was effective in locating and sizing defects. However, it was not possible to compare the increase in defect area with the change of the mechanical properties.

Les travaux de recherche présentés ici avaient pour objectif principal la synthèse de copolymères statistiques à base d’éthylène et d’acide acrylique (AA). Pour cela, la déprotection des groupements esters d’un copolymère statistique précurseur, le poly(éthylène-co-(tert-butyl)acrylate), a été effectuée par hydrolyse à l’aide d’iodure de triméthylsilyle. La synthèse de ce précurseur est réalisée par polymérisation catalytique en présence d’un système à base de Palladium (Pd). Le deuxième objectif a été d’étudier et de caractériser des polymères synthétisés à l’état solide et en suspension colloïdale. Plusieurs copolymères précurseurs comprenant différents pourcentages molaires en tert-butyl acrylate (4 à 12% molaires) ont été synthétisés avec succès, puis déprotégés par hydrolyse pour obtenir des poly(éthylène-coacide acrylique) (pE-co-AA) avec différentes compositions. Seuls les copolymères comprenant 10% molaire ou plus de AA sont solubles dans le Tétrahydrofurane (THF) et uniquement dans ce solvant. De telles solutions peuvent être dialysées dans l’eau, ce qui conduit à un échange lent entre cette dernière et le THF, et l’autoassemblage du copolymère dans l’eau peut ensuite être étudié. C’est ainsi qu’ont pu être observées des nanoparticules stables dans le temps dont le comportement est sensible au pH et à la température. Les polymères synthétisés ont été caractérisés par Résonance Magnétique Nucléaire (RMN) ainsi que par spectroscopie Infra-Rouge (IR), avant et après déprotection. Les pourcentages molaires d’AA ont été déterminés par combinaison des résultats de RMN et ii de titrages conductimètriques. A l’état solide, les échantillons ont été analysés par Calorimétrie différentielle à balayage (DSC) et par Diffraction des rayons X. Les solutions colloïdales des polymères pE-co-AA ont été caractérisées par Diffusion dynamique de la lumière et par la DSC-haute sensibilité. De la microscopie électronique à transmission (TEM) a permis de visualiser la forme et la taille des nanoparticules. / The first objective of this research is to synthesize random linear copolymers of ethylene and acrylic acid (AA). The synthesis relies on the deprotection of the functional groups in the copolymer’s precursor, which is represented by poly(ethylene-co-tertbutyl acrylate). The synthesis of the precursor was realized by the catalytic approach, where Pd-based catalytic systems are frequently utilized nowadays. The deprotection was carried out by hydrolysis of the ester functionality using trimethylsilyl iodide agent. The second objective is to investigate and characterize the synthesized polymers in the bulk and in colloidal solution. A set of different precursor polymers with various degrees of molar incorporation of tertbutyl acrylate (from 4 to 12 mol %) was successfully synthesized and deprotected. The resulting poly(ethylene-co-acrylic acid) copolymers were found to be soluble in tetrahydrofuran THF, when the molar incorporation of AA reaches the value of 12 and more. This aspect gave the possibility to study the self-assembly of this copolymer in aqueous medium by slow THF to water exchange (dialysis). It was found that the copolymers self-assemble into nano-sized structures and these nanoparticles remain stable in colloidal solution for extended periods of time. Moreover, it was shown that the nanoparticles formed by the discussed copolymer possess thermo- and pH-responsive behaviour. The polymers synthesized were characterized by nuclear magnetic resonance (NMR) and infrared spectroscopies (IR) before and after deprotection. The bulk samples were analyzed by conventional differential scanning calorimetry and by X-ray diffraction iv technique. The molar percentages of AA were determined using a combination of NMR and conductimetric titration. Colloidal solutions of pE-co-AA copolymers were analyzed by dynamic light scattering and high-sensitivity differential scanning calorimetry techniques. The nanoparticles formed were visualized and characterized by transmission electron microscopy.