This dissertation has focused on studying the effect of four glycine derivatives on the solid state properties of mannitol, glycine, and sucrose when freeze dried into blended mixtures. The primary goal was to assess their value for use in the stabilisation of vaccines in the solid state, by examining key physical and chemical characteristics, which have been documented to be beneficial to the stabilisation of biopharmaceutical formulations. The novel excipients; dimethyl glycine, and trimethyl glycine, were shown to retard the crystallisation and increase the overall glass transition temperature, of mannitol, when freeze dried as evidenced by DSC and Powder X-ray diffraction. Mannitol’s glass transition temperature increased from 100C to 12.650C and 13.610C when mixed with methyl-glycine and dimethyl glycine respectively. The glycine derivatives did not show the same effect on sucrose which remained amorphous regardless of the concentration of the other excipient. The different behaviour with the sucrose system was thought to be due to relatively high glass transition temperature of sucrose. Conversely glycine remained highly crystalline due it’s relatively low glass transition temperature. The novel excipient formulations were also assessed for their effect on the aggregation of the adjuvant aluminium hydroxide when freeze dried by Dynamic Light Scattering (DLS).The formulations containing the glycine derivatives all caused a decrease in the aggregation size of the adjuvant from ~26 μm, to 185 nm in the presence of methyl glycine. The effects of lysozyme and viral antigen on the adjuvants were also examined showing that the addition of the virus did not affect the size of the aggregates formed, however lysozyme showed significant decreases in the aggregates formed. Examination of the freezing method were also made showing that faster freezing rates produced smaller aggregates of the adjuvant. When investigating the rate at which the excipients lost water during secondary drying there was evidence of the formation of hydrates of glycine, trimethyl glycine, and mannitol has shown that the glycine derivatives have attributes which would be beneficial in stabilising vaccines in the solid state when freeze dried.

Um polieletrólito catiônico (poliamina), com baixo peso molecular e elevada densidade de carga, normalmente aplicado como agente coagulante do lixo aniônico, foi usado para estudos de retenção e drenagem na fabricação de papel. O uso do carboximentil celulose de sódio para simulação do teor de lixo aniônico e seu efeito na retenção de cargas minerais foi uma característica importante para este trabalho. Pode ser observado que o aumento da dosagem do polímero catiônico tanto melhora a retenção de cargas minerais, avaliada pela turbidez do filtrado, quanto melhora a drenagem do sistema, avaliada pela velocidade de escoamento. Entretanto, elevadas dosagens deste polímero comprometeram os resultados destes parâmetros devido à inversão de carga do sistema. Pode também ser confirmado que forças de cisalhamento excessivas prejudicam a retenção de cargas minerais. Ademais, um polianfótero, com peso molecular e densidade de carga elevados, contendo grupos positivo (N-[3-(N,N-dimetilamino)propil]acrilamida), negativo (ácido metileno butanodióico) e nulo (acrilamida) na mesma cadeia, foi testado como agente de resistência a seco do papel. Todos os estudos em nível molecular sobre o comportamento do polianfótero em solução e o seu comportamento de adsorção sobre superfícies modelos carregadas, em diferentes condições de pH e de força iônica, foram importantes para explicar tanto dos fenômenos de adsorção, envolvendo fibras celulósicas e polianfótero, quanto o seu efeito na resistência mecânica do papel. Foi observado que a solubilidade do polímero aumenta à medida que o pH se distancia do seu ponto isoelétrico, pHPIE 7,3, e reduz para valores de pH próximos ao pHPIE. O tamanho das estruturas do polianfótero depende do pH do meio de dispersão. As características de tamanho do polianfótero tanto sob a forma de cadeias individuais ou quanto sob a forma de agregados, foram medidas através da técnica de espalhamento dinâmico de luz. As propriedades viscoelásticas das camadas adsorvidas e a quantidade de polímero adsorvida foram medidas através da técnica da balança microgravimétrica com dissipação de energia. Estas duas determinações, associadas às imagens no microscópio de força atômica, foram importantes para o entendimento dos resultados práticos do uso do polianfótero como agente de resistência a seco do papel. Maiores resultados de resistência do papel, avaliada através da resistência à tração, foram alcançados para valores de pH próximos ao ponto isoelétrico onde foram encontrados o seguinte: (1) maiores tamanhos para as estruturas do polímero em solução, (2) maior quantidade de massa nas camadas adsorvidas e (3) a formação de camadas mais viscoelásticas. O fenômeno de separação de fases, associado à mudança da solubilidade do polímero em solução devido ao balanço dos grupos positivos e negativos ionizados ao longo da faixa de pH estudada, foi considerado o principal aspecto para a variação em tamanho dos agregados. Embora este polímero tenha apresentado comportamento antipolieletrólito devido à expansão da sua cadeia e ao aumento da densidade de carga com o aumento da força iônica, considerando o efeito da força iônica para pH 4,3, o comportamento de adsorção do polianfótero foi avaliado como o comportamento de um polieletrólito monocarregado de alta densidade de carga. Maiores e menores quantidades de massas adsorvidas foram encontradas para valores intermediários e extremos de força iônica, respectivamente. As interações eletrostáticas foram consideradas as principais responsáveis pela adsorção do polímero sobre superfícies carregadas. Entretanto, a blindagem de cargas foi considerada a explicação para os baixos valores de massa adsorvida para valores mais elevados de força iônica. / A cationic polyelectrolyte (polyamine), with low molecular weight and high charge density, usually applied as anionic trash coagulant, was used for the retention and drainage studies in the papermaking. The use of sodium carboxymethyl cellulose to simulate the anionic trash content and its effect on the filler retention was an important feature of the work. It could be noted that the increasing of the cationic polymer dosage improves both the filler retention, evaluated by the turbidity of the filtrate, and the system drainage, evaluated by the flow speed. However, high dosages of this polymer compromised the results of these parameters due to the reversal of the system charge. It can also be confirmed that excessive shear forces affect the filler retention. Furthermore, a polyampholyte, with high molecular weight and charge density, containing positive (N-[3-(N,N- dimethylamino)propyl]acrylamide), negative (methylene butanedioic acid), and neutral (acrylamide) groups in the same chain, was tested as a dry strength agent. All of the studies at molecular level concerning to the polyampholyte behavior in the solution and its adsorption behavior on charged model surfaces at different conditions of pH and ionic strength, were important to explain both the adsorption phenomena, involving cellulosic fibers and polyampholyte, and its impact on the paper strength. It was observed that the polymer solubility increases as the pH moves away from its isoelectric point, pHIEP 7.3, and decreases when the pH approaches close to pHIEP. The sizes of the structures of the polyampholytes depend on the pH of the dispersion medium. Also the size characteristics of polyampholyte, both in individual and aggregated forms, were measured by dynamic light scattering technique. The viscoelastic properties of adsorbed layers, as well as the amount of the adsorbed polymer, were measured by quartz crystal microbalance technique with energy dissipation. These two measurements, associated with the atomic force microscopy images, were important to understand the practical results of polyampholyte usage as a dry strength agent. Best results of paper strength, evaluated by paper strength index, were achieved at pH close to the isoelectric point on which one were found the following features: (1) larger sizes of the polymer structures in solution, (2) higher amount of mass in the adsorbed layers, and (3) the formation of more viscoelastic layers. The phase separation phenomenon, associated with the change in the solubility of the polymer due to the balance of the positive and negative groups throughout the studied pH range, was considered the main aspect for the variation in size of the aggregates. Although this polymer shows antipolyelectrolyte behavior due to the expansion of the its chain and the increasing in charge density with the ionic strength, considering the effect of ionic strength at pH 4.3, the adsorption behavior of polyampholyte was evaluated as a monocharged polyelectrolyte behavior with high charge density. Major and minor amounts of adsorbed masses were found for intermediates and extremes values of ionic strength, respectively. The electrostatic interactions were considered the main cause of the adsorption on charged surfaces. However, the electrostatic screening was considered the explanation for the low values of adsorbed mass at higher values of ionic strength.

Ces travaux de recherche portent sur le développement d'un système original de fusion de données de diffusion électromagnétique et optique par des milieux denses et complexes. La méthode, à la fois théorique, numérique et expérimentale, permet la fusion des signatures de diffusion hyperspectrales, polarimétriques et angulaires d'un milieu d'étude. Un système expérimental multi-capteurs comprenant une source laser supercontinuum est présenté pour mesurer les signatures de diffusion de différentes cibles. Des modèles directs de simulation physique ont aussi été développés via : (i) une approche dite « top-down » qui modélise les signatures à partir de paramètres macroscopiques (ex. rugosité, indices optiques effectifs) ou (ii) une approche dite « bottom-up » qui modélise les signatures à partir de paramètres microscopiques (ex. distribution en taille, géométrie, concentration, indices optiques et structuration des diffuseurs) en résolvant soit l'équation de transfert radiatif ou directement les équations de Maxwell. Des méthodes inverses appliquées sur les signatures mesurées sont développées pour retrouver simultanément les paramètres d’intérêt du milieu analysé. Les avancées de ces travaux permettent une amélioration de la compréhension des phénomènes de diffusion électromagnétiques et optiques par des milieux denses et complexes tels que les surfaces rugueuses, les revêtements, les nanomatériaux, les suspensions colloïdales ou les agrégats fractals d'aérosols ultrafins. Les domaines d'applications de ces travaux sont l'aéronautique (ex. peintures d'aéronefs), l'imagerie aéroportée ou satellite (ex. imagerie active hyperspectrale ou polarimétrique), la sécurité et la défense (ex. matériaux pour la furtivité) ou bien les sciences de l'atmosphère (ex. systèmes LiDAR, suivi de pollution, suies), l'industrie chimique (ex. suspensions colloïdales) ou le biomédical (ex. diagnostic de tumeurs et mélanomes). / This work reports the development of an original data fusion system dedicated to electromagnetic and light scattering by dense and complex media. The dissertation encompasses the theoretical, numerical and experimental studies. The output of the data fusion system is a fused hyperspectral, polarimetric and angular scattering signature. An experimental multi-sensor and supercontinuum laser-based system is presented to measure the scattering signatures for various targets. Direct physical simulation models were developed using a two-level modelling scheme: (i) a top-down approach is used to model signatures from macro-physical parameters, e.g. the surface roughness or the effective refractive index and, (ii) a bottom-up approach is used to model signatures from microphysical parameters, e.g. the size distribution, the geometry, the concentration, the refractive index and the structuration of the scatterers, by solving the radiative transfer equation or directly the Maxwell's equations. Inversion schemes are deployed to retrieve these parameters by inverting the experimental signatures. The advancements described throughout this dissertation will serve to improve understanding of electromagnetic and light scattering by dense and complex media such as rough surfaces, coatings, nanomaterials, colloidal suspension and fractal aggregates of ultrafine aerosols. This study has relevant applications in fields as diverse as aeronautics (e.g. aircraft paint coatings), remote-sensing (e.g. hyperspectral, polarimetric, active or passive imaging), security and defense (e.g. furtive materials), atmospheric science (e.g. black carbon or soot characterization, LiDAR systems), chemical engineering (e.g. colloidal suspensions), or biomedical (e.g. tumor and melanoma diagnostic).

The mechanochemical polycondensation between a diamine and a dialdehyde constitutes a sustainable alternative to classical solvent-based polymerization reactions. This process not only allows for a higher conversion and a shorter reaction time as compared to standard solvent-based syntheses of this conjugated polymer, but the reaction can also be adjusted by the energy introduced via the ball mill.

The presented doctoral thesis is focused on preparation, characterization and application of organic micro- and nanoparticles as transport systems for active components and some their complex natural sources. Active component were packed into liposomes and polysaccharide particles. As active components were used caffeine, some drugs – clotrimazole and ibuprofen, further antioxidants and vitamins. Antimicrobial herbs and spices extract, antimicrobial peptides lysozyme, nisin and other antimicrobial ingredients were encapsulated too. Encapsulation of selected hydrolytic enzymes was tested, too. Particles were also used for encapsulation of probiotic strains Bifidobacterium breve and Lactobacillus acidophilus and prebiotic components. These prebiotics were co-encapsulated into capsules with probiotic cells. Natural extracts were encapsulated e.g. extracts of guarana, ginseng, goji, green barley, propolis, black, green and white tea, coffee, fruit and vegetable extracts. The efficiency of encapsulation was determined by HPLC/PDA and by spectrophotometry. Long-term stability of particles and amount of released component in model/real foods, in model cosmetic conditions and in a model physiological environment were monitored too. Size of prepared liposomes and polysaccharide particles was determined by dynamic light scattering and by light microscopy and electron microscopy, respectively. Stability of the particles was measured using a zeta potential. Also, analytical centrifugation was used to measurement of sedimentation velocity and stability of the prepared particles. The antimicrobial activity were tested using two Gram-positive (Bacillus subtilis, Micrococcus luteus), two Gram-negative (Escherichia coli, Serratia marcescens) bacteria and one fungal strains (Candida glabrata). For determining the antimicrobial properties of active component and prepared particles two the most widely used methods were used - agar diffusion method and broth dilution method. The viability of probiotic strains were performed using flow cytometry and fluorescence microscopy. Encapsulation of active component was successful in all types of particles. Liposome showed a very good long-term stability mainly in water conditions with neutral pH and polysaccharide particles were stable in acidic conditions. Prepared particles showed a very good stability in model stomach environment, while in model intestines environments particles were disintegrated and active component were released. Prepared particles with encapsulated caffeine as well as other tested antioxidants and vitamins could be used to modern types of energy drinks, food supplements and also for some cosmetics applications. Encapsulated antimicrobial components could be used for food application as well as for cosmetics and pharmaceutical application like antimicrobial wound formulation. Encapsulated enzymes can be used for controlled release of proteases in wound healing, as delivery systems in digestive tract and as a part of pharmaceutical preparative and food supplements for enzyme therapy. The study revealed that encapsulation of probiotics and also co-encapsulation of probiotics with prebiotics exhibited longer stability of particles and survival bacterial cells. So, prepared particles are suitable for use to food product with beneficial effects on the human body.

Předkládaná diplomová práce se zabývá extrakcí nanočástic oxidu titaničitého z produktů osobní péče, konkrétně opalovacích krémů a následnou charakterizací těchto částic. Počet komerčně dostupných produktů s obsahem nanočástic TiO2 se neustále zvyšuje a to se sebou přináší potřebu vyhodnotit potenciální osud a nepřímou expozici TiO2 nanošástic o různých velikostí a tvarů a zkoumat jejich celý životní cyklus. Bylo zkoumáno použití ultrafiltrace a ultracentrifugace jako extrakční metody. Dvě metody pro extrakci TiO2 nanočástic byly vyvinuty a aplikovány na vzorky opalovacích krémů. Extrahované částice mohou být použity pro ekotoxikologické studie, případně experimenty v mesokosmu. Velikost částic byla stanovena pomocí metody dynamického rozptylu světla a transmisní elektronové mikroskopie.

Le phénomène de givrage qui se produit durant les vols d’aéronefs, lors de leurs traversées de la troposphère (zone dans laquelle la température peut descendre jusqu’à -60°C) se manifeste par une accrétion de glace sur différentes parties de l’appareil (voilures, réacteurs, sondes de mesure…) mettant en péril la sécurité de celui-ci. Ce phénomène constitue alors une problématique majeure pour la sécurité de l’aviation civile, c’est pourquoi il est nécessaire de développer de nouvelles techniques de mesure afin de détecter et d’éviter les zones à risques. Notre intérêt s’est porté sur l’imagerie interférométrique en défaut de mise au point, une technique optique offrant de nombreux avantages (large champ de mesure, gamme de tailles étudiée étendue [50 μm : quelques millimètres], distance particule/appareil de mesure de plusieurs dizaines de centimètres…). Au cours de ces travaux de thèse, nous avons développé un dispositif d’Imagerie Interférométrique de Particules (IIP) multi-vues permettant de caractériser les cristaux de glace en suspension dans latroposphère. En effet, par comparaison avec des mesures obtenues sur des images nettesenregistrées simultanément, nous avons validé l’utilisation de l’IIP multi-vues pour l’estimation dedimensions de particules de glace avec un taux d’erreur inférieur à 20%. Nous avons égalementproposé différentes approches permettant d’estimer les volumes des particules de glace et mis enévidence la « signature » typique d’une gouttelette en cours de givrage à partir d’images d’IIP.Dans une seconde partie, nous avons validé l’utilisation de l’IIP multi-vues pour le cas complexe oùles interférogrammes d’une paire de particules de glace proches l’une de l’autre se recouvrent etdiscuté des phénomènes de Moiré qui peuvent apparaître et perturber nos mesures. En outre, nousavons étendu le domaine d’utilisation de l’IIP à des milieux moins dilués. Enfin, dans une dernière partie, nous avons développé un dispositif expérimental innovant nous permettant d’effectuer des mesures d’IIP expérimentales à partir de particules « programmées » sur une matrice de micro-miroirs (DMD). / The icing phenomenon that occurs during aircraft flights during their troposphere crossings (an area where the temperature can drop to -60 ° C) is manifested by an accretion of ice on different parts of the apparatus (wings, reactors, measurement probes ...) endangering the safety of this latter. This phenomenon is therefore a major problem for the safety of civil aviation, that's why it is necessary to develop new measurement techniques to detect and avoid risk's areas. Our interest has been on interferometric out-of-focus imaging, an optical technique offering many advantages (wide measurement field, range of studied sizes extended [50 μm: a few millimeters], distance particle / measuring device several tens of centimeters ...). During this thesis, we have developed a multi-view Interferometric Particle Imaging (IPI) device to characterize suspended ice crystals in the troposphere. Indeed, by comparison with measurements obtained on in-focus images recorded simultaneously, we have validated the use of multi-view IIP for the estimation of ice particle dimensions with an error rate lower than 20%. We also proposed different approaches to estimate ice particle volumes and highlighted the typical "signature" of a droplet during icing from IIP images.In a second part, we validated the use of multi-view IIP for the complex case where the interferograms of a pair of ice particles close to each other overlap and discuss about Moiré phenomena that may appear and disrupt our measurements. In addition, we have extended the field of use of IIP to less diluted media. Finally, in the last part, we have developed an innovative experimental device allowing us to perform experimental IIP measurements from particles "programmed" on a matrix of micro-mirrors (DMD : Digital Micromirror Device).

<div>Large protein molecules are abundant in biological cells but are very difficult to study in physiological conditions due to molecular disorder. For large proteins, most structural information is obtained in crystalline states which can be achieved in certain conditions at very low temperature. X-ray and neutron crystallography methods can then be used for determination of crystalline structures at atomic level. However, in solution at room or physiological temperatures such highly resolved descriptions cannot be obtained except in very few cases. Scattering methods that can be used to study this type of structures at room temperature include small-angle x-ray and neutron scattering. These methods are used here to study two distinct proteins that are both classified as glycoproteins, which are a large class of proteins with diverse biological functions. In this study, two specific plasma glycoproteins were used: Fibrinogen (340 kDa) and Alpha 1-Antitrypsin or A1AT (52 kDa). These proteins have been chosen based on the fact that they have a propensity to form very large molecular aggregates due to their tendency to polymerize. One goal of this project is to show that for such complex structures, a combination of scattering methods that include SAXS, SANS, and DLS can address important structural and interaction questions despite the fact that atomic resolution cannot be obtained as in crystallography. A1AT protein has been shown to have protective roles of lung cells against emphysema, while fibrinogen is a major factor in the blood clotting process. A systematic approach to study these proteins interactions with lipid membranes and other proteins, using contrast-matching small-angle neutron scattering (SANS), small angle x-ray scattering (SAXS) and dynamic light scattering (DLS), is presented here. A series of structural reference points for each protein in solution were determined by performing measurements under osmotic stress controlled by the addition of polyethylene glycol-1,500 MW (PEG 1500) in the samples. Osmotic pressure changes the free energy of the molecular mixture and has consequences on the structure and the interaction of molecular aggregates. In particular, the measured radius of gyration (Rg) for A1AT shows a sharp structural transition when the concentration of PEG 1500 is between 33 wt\% and 36 wt\%. Similarly, a significant structural change was observed for fibrinogen when the concentration of PEG 1500 was above 40 wt\%. This analysis is applied to a study of A1AT interacting with lipid membranes and to a study of fibrinogen polymerization in the presence of the enzyme thrombin, which catalyzes the formation of blood clots. The experimental approach presented here and the applications to specific questions show that an appropriate combination of scattering methods can produce useful information on the behavior and the interactions of large protein systems in physiological conditions despite the lower resolution compared to crystallography.</div>

Title: Self-assembly of polymers with amphiphilic compounds (surfactants) in aqueous solutions Abstract: This PhD Thesis is devoted to the co-assembly in systems containing electrically charged polymers (polyelectrolytes and block copolymers containing polyelectrolyte sequences). I studied the interactions between block copolymers and oppositely charged surfactants in aqueous solutions, and the structure and properties of co-assembled nanoparticles by a combination of several experimental methods. I found that the spontaneous formation, solubility and stability of complex nanoparticles depend not only on the electrostatic attractive forces but also on the hydrophobic effects. In a major part of my Thesis, I studied the interaction of polyelectrolytes with oppositely charged gemini surfactants (containing two charged head-groups interconnected by a short linker and two hydrophobic tails) which is a relatively new topic - much less studied than the co-assembly with conventional single tail surfactants. Better understanding of the formation and properties of complexes containing gemini surfactants and polymers provides knowledge that should lead to novel tailor-made nanoparticles with desired properties for applications in medicine and new technologies (including nano-technologies). We have shown that the...

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