Les réactions pallado-catalysées permettant la formation de liaisons carbone-carbone trouvent de nombreuses applications en synthèse organique et constituent l’étape clé de la synthèse d’un grand nombre de molécules. La première partie de cette thèse décrit la préparation et l’étude de catalyseurs hétérogènes et réutilisables comportant du palladium supporté sur un polymère portant des groupements phosphinés, polymère dérivé soit d'une résine de Merrifield soit d’une résine Rasta. De très bons résultats ont été obtenus pour leur utilisation dans les couplages croisés de Hiyama, Heck et Suzuki et dans chaque cas la possibilité de réutilisation jusqu’à 4 fois du catalyseur a été vérifiée. Notre travail constitue une des premières utilisations d’un catalyseur réutilisable dans le couplage de Hiyama. Nous avons aussi mis au point des conditions permettant d’effectuer le couplage de Heck en présence d’une quantité infime de palladium. Notre étude sur les résines Rasta constitue la première application de ces supports en pallado-catalyse. La deuxième partie de la thèse décrit l’étude de la préparation de catalyseurs où le palladium serait supporté sur des nanoparticules superparamagnétiques et qui pourraient donc être récupérés de tout milieu réactionnel grâce à un champ magnétique externe. Des nanoparticules de cobalt ont été préparées puis recouvertes de pyrocarbone par dépôt chimique en phase vapeur. Des groupements organiques ont été fixés sur la coque de carbone, ce qui permet l’introduction de ligands phosphinés. La structure de ces particules a été étudiée par microscopie électronique en transmission et leur préparation optimisée en fonction des résultats structuraux. / Palladium-catalyzed reactions forming carbon-carbon bonds have found widespread applications in organic synthesis as they represent the key step of many important syntheses in various domains. The first part of this work describes the preparation and study of heterogeneous, reusable, catalysts containing palladium supported on a polymer bearing phosphino groups derived from either a Merrifield resin or a Rasta resin. Very good results have been obtained for the use of these catalysts in Hiyama, Heck and Suzuki cross-couplings and, in each case, the possibility of reuse of the catalyst at least 4 times has been verified. Our work constitutes one of the first use of a reusable catalyst in a Hiyama coupling. We have also found experimental conditions allowing to run the Heck coupling in the presence of only a minimal amount of palladium. Our work constitutes the first use of Rasta resins to prepare heterogeneous palladium catalysts. The second part of the thesis describes the study of the preparation of catalysts where palladium would be supported on superparamagnetic nanoparticles and which could therefore be recovered from any reaction medium by using a magnetic field. Cobalt nanoparticles were prepared and then covered by pyrocarbon by Chemical Vapor Deposition. Organic groups, allowing the subsequent introduction of phosphino ligands, were then grafted on the carbon shells. The structure of the particles was determined by Transmission Electron Microscopy and their preparation optimized according to these structural results.

Les limites des nanovecteurs commerciaux ou actuellement en développement ont motivé l’élaboration de nouvelles nanoparticules mésoporeuses de silice (MSNP), hybrides et multiphasées, pour le contrôle de la délivrance d’actifs à application théranostique. Ainsi, de nouvelles MSNP ont été conçues pour la pénétration intracellulaire (diamètre entre 30 et 60 nm, taille des pores de 2,8 nm). Afin de les rendre hémocompatibles et de contrôler la cinétique de délivrance de principes actifs encapsulés, ces MSNP ont été enrobées d’une bicouche lipidique (MSNP+@SLB-). La composition lipidique s’inspire des membranes asymétriques des globules rouges ciblés par la présente étude.La technologie MSNP+@SLB- ayant montré des limites avec une cinétique de libération trop élevée de la calcéine et trop lente de la rhodamine B, deux améliorations majeures ont été apportées :1- Le recouvrement des SLB par un nanogel d’alginate, permettant un excellent contrôle de la libération d’actifs.2- L’insertion de nanoparticules magnétiques dans le coeur des MSNP, déclenchant la libération de l’actif par hyperthermie.Ces nouvelles architectures de nanovecteurs permettent de moduler les cinétiques de délivrance d’actifs, renforçant et élargissant ainsi le champ d’applications des vecteurs silicés dans les domaines biomédical ( Voie orale et intraveineuse) et dermato-cosmétique (Voie topique). / The limitations of commercial nanovectors or currently under development have motivated the development of new hybrid and core shell mesoporous silica nanoparticles (MSNP) for the control of molecular delivery.Therefore, new MSNP were designed for intracellular penetration (diameter between 30 and 60 nm, pore size of 2.8 nm). In order to make them hemocompatible and to control the kinetics of delivery of encapsulated active ingredients, these MSNP were coated with a lipid bilayer (MSNP+@SLB-). The lipid composition is inspired by the asymmetric membranes of the red blood cells.Since the MSNP+@SLB- technology has shown some limitations associated to the release of payloads which can be too fast (in the case of calcein) or to slow (case of rhodamine B), two major improvements have been made:1- The coating of SLB by an alginate nanogel, allowing an excellent control of the release of active molecules.2- Insertion of magnetic nanoparticles in the MSNP core, triggering the release of the active ingredient by hyperthermia.These new nanovector architectures enable the fine tuning of active ingredient delivery kinetics, reinforcing and expanding the applications of silicated vectors in the fields of biomedicine (oral and intravenous) and dermato-cosmetics (topical).

This thesis deals with the studies of magnetization and electrical transport properties of iron nanoparticles embedded in the carbon matrix. The synthesis and characteristics of the nanoparticle systems studied, are also presented. Carbon-iron (C-Fe) based systems are of growing interest due to their improved magnetic properties as well as in their potential application as sensors, catalysts, and in various other applications. In particular, nanocomposites of iron carbide, such as the cementite phase Fe3C, are further suited to diverse technological exploitations due to their enhanced mechanical properties and importance in ferrous metallurgy. The recent interest in magnetic nanostructures lies in the emergence of novel magnetic and transport properties with the reduction of size. As the dimension approaches the nanometer length scale, interesting size-dependent properties like enhanced coercivity, enhanced magnetic moment, super paramagnetism etc. are seen. Thermal assisted chemical vapour deposition (CVD) is used to decompose and chemically react the introduced precursors, maleic anhydride and ferrocene. This method provides relative size control over the individual particles by varying C/Fe concentration in precursors and the pyrolysis temperature during the co-deposition process. Ferrocene has been used actively for the production of nanoparticle composites and in the production of nanostructured carbon. The temperature of preparation, reaction rate, and the time duration of annealing directly effects the nanoparticle compositions. The catalytic effect of transitional elements are well documented in literature. This thesis is an effort to understand the growth of ferromagnetic nanocrystallites in carbon matrix, which undergo partial graphitization due to the catalytic effect of transitional elements. The effect of transitional metal on the degree of graphitization of the carbon matrix, morphology of the nanoparticle and the carbon matrix are studied. The phase of the ferromagnetic iron nanoparticles and the structural investigation forms part of the study. Here X-Ray diffraction (XRD) is employed to study the presence of different phases of iron in the partially graphitized carbon matrix. The matrix morphology and the particle size distribution were studied using Transmission Electron Microscopy (TEM) and High-Resolution TEM (HRTEM). The ferromagnetic states of the iron nanoparticles are investigated using Mossbauer spectroscopy. The results from these studies, are used to correlated the macroscopic properties to the microscopic studies. The enhanced magnetization, coercivity and the temperature dependence of the magnetization value is understood within the frame work of ferromagnetic Bloch law and surrounding carbon spins. The logarithmic temperature dependence of conductivity of the nanoparticle composites is analyzed in the framework of interference models as well as the many-body Kondo interaction effect. This thesis contains seven chapters: In chapter 1, a brief introduction to mesoscopic physics and the size-dependent phenomenon are given. Special attention is paid to magnetic nanoparticle and its composites, and the various finite-size effects exhibited by them are discussed in detail. The relevance of carbon matrix and its importance on the growth of iron nanoparticles with high thermal stability is also discussed. The ballistic and diffusive transport phenomena observed in low-dimensional systems are briefly discussed. The interplay of localization and various interaction effects at nanoscale are examined. In disordered metals the low temperature conductivity is dominated by the interference effects. A brief discussion is made on the conductivity in disorder systems, with the presence of magnetic impurities and how the classic many-body Kondo problem, is effected by various interactions. Chapter 2, mainly deals with the experimental techniques employed in the thesis. The thermal-assisted chemical vapour deposition setup used to decompose and chemically react the introduced organometallic precursors, for the preparation of C:Fe composites are discussed and its advantage over other preparation methods are emphasized. The method is optimized to provide relative size control over the nanoparticles composites and the phase compositions by varying C/Fe concentration in precursors and the pyrolysis temperature, during the co-deposition process. The various structural characterization tools used in the present study are summed up concisely in this chapter. The SQUID magnetometer system; its working principle and the various protocol used for the low temperature magnetization measurements are elaborated. Further, details regarding superconducting magnetic cryostat, utilized for the low temperature conductivity and magneto resistance measurements, are discussed. Films of C:Fe composites are grown on substrates to study the effect of disorder and sample size on the conductivity behaviour of the composites at low temperature. Chapter 3, presents the outcome of the structural studies undertaken on the C:Fe composites using XRD, TEM, and HRTEM. X-ray diffraction measurements performed on the powder composites reveal that, in addition to the presence of sharp diffraction peak from nanographite, peaks corresponding to the different phases of Fe are also seen. The effect of preparation temperature on the matrix morphology is revealed from the estimation of degree of graphitization. Iron carbide is the predominant phase in all the prepared composites. For low concentration of iron, iron carbide alone is present but as the percentage of iron in the samples increased other phases of iron are also seen. The microscopic studies on the prepared compositions revealed the presence of nanosized iron particles well embedded in the partially graphitized matrix. Here again, with the increase in iron percentage, agglomeration of ferromagnetic nanoparticles are seen. The kinetics of the particle growth and the filamentous nature of the carbon matrix are also discussed. Mossbauer investigation on C:Fe composites are presented in chapter 4. The measurements revealed the iron atom occupation in the crystal lattice. In the lower Fe concentration samples, the room temperature Mossbauer spectrum revealed the presence of sextet from Fe3C (cementite) phase. As the percentage of iron increased, sextet from α-Fe, Fe3O4 are also seen in some of the prepared compositions. Effect of carbon atoms on the structure and magnetic properties of the nanoparticle species are obvious from the isomer shift measurements. Chapter 5 comprises of the various magnetic properties and interactions present in small particle system such as magnetic anisotropy, coercivity, enhanced magnetization, inter-and intra-particle interactions etc. Magnetization measurements carried out in SQUID magnetometer on the C:Fe composites and carbon flakes (prepared from organic precursor, maleic anhydride alone) are presented. The enhanced magnetic properties of the nanoparticle assembly is discussed in detail. The hysteresis loops trace, with a finite coercivity at room temperature, indicates the ferromagnetic nature of the samples. At room temperature the magnetization value saturates at high magnetic field, indicating negligible effect from super paramagnetic particles on the hysteresis loop. The squareness ratio, saturation magnetization, coercivity and remanence magnetization values are analyzed in detail. The temperature dependence of magnetization shows a combination of Bloch law and Curie-Weiss behaviour, consistent with the picture of ferromagnetic clusters embedded in a carbon matrix. The Bloch’s constant is found to be larger by an order of magnitude compared to the bulk value, implying stronger dependence of magnetization with temperature. Effort to understand the enhanced magnetic moment in the light of magnetism in carbon was taken up. The proximity effect of ferromagnetic metal on the carbon and the hydrogen bonding with the dangling bonds, both studied in detail in literature, in connection with the induced magnetic moments in carbon, are invoked. In chapter 6, the different conductivity regimes are identified, to study the conduction mechanisms in composites and films. For the transport measurements pelletized samples are used for the resistivity and magneto resistance measurements. The conductivity data are analyzed based on the interplay of localization and Kondo effect in the ferromagnetic disordered system. In order to understand the effect of disorder and thickness on the Kondo problem, transport measurements are carried on thin films of C:Fe composites grown on quartz and alumina substrate. Disorder induced metal-insulator transition is observed in the prepared samples. The zero-field conductivity and magneto resistance data is fitted to variable range hopping (VRH) in strong localization regime. Chapter 7 summarizes the thesis and presents some perspectives for the future.

Cette étude concerne le développement de nanoparticules hybrides offrant de nouvelles stratégies pour la thérapie et le diagnostic médical. Elles sont constituées d’un cœur magnétique jouant le rôle d’agent de contraste pour l’IRM et d’inducteur de chaleur par hyperthermie, d’une couronne de polymère thermosensible permettant d’encapsuler des principes actifs et de peptides de reconnaissance biologique. Une grande partie de l’étude a consisté à étudier les processus d'adsorption de copolymères poly(éther)-b-poly(L-lysine) de composition variable sur les particules magnétiques et à comprendre le rôle de la conformation des chaînes polymère à la surface des particules sur la stabilité des colloïdes en milieu physiologique. Un agent antitumoral a été encapsulé puis libéré de façon contrôlée sous l’effet d’un champ magnétique alternatif en exploitant le caractère thermosensible des blocs polyéthers. Des séquences peptidiques ciblant les zones d’inflammation de la barrière hémato-encéphalique ont été greffées sur les copolymères. L’efficacité du ciblage a été validée par IRM et fluorescence sur un modèle animal démontrant ainsi la multifonctionnalité des nanoparticules. / This work deals with the development of hybrid nanoparticles that could offer new strategies for therapy and diagnostic. These are based on a magnetic core which can play the role of contrast agent for MRI as well as heat inductor in AC magnetic field. This inorganic core is surrounded by a thermo-responsive polymeric brush that controls the loading and the release of drugs, and can be functionalized by specific ligands ensuring the targeting specificity. A large part of this work consists in studying the adsorption mechanism of poly(ether)-b-poly(L-lysine) based block copolymers onto magnetic particle and to better understand the influence of the polymer chain conformation at particles surface on the colloidal stability under physiological conditions. An anticancer drug has been loaded and released in a controlled manner under alternative magnetic field by taking advantage from the thermosensitivity of the polyether block. Targeting peptides specific of inflammation sites at the blood brain barrier have been grafted onto copolymers. The targeting specificity has been demonstrated by MRI and fluorescence imaging in rats attesting the multifunctionality of such nanoparticles.

Ce travail concerne la modification de surface de nanoparticules magnétiques (MNP) par des copolymères réactifs renfermant des cycles azlactone, aux fins de l’élaboration de nano-supports destinés à l’immobilisation de biomolécules. Trois stratégies basées sur des techniques de polymérisation radicalaire contrôlée ont été mises en œuvre.Dans la première, un copolymère poly(méthacrylate de poly(éthylène glycol)-stat-2-vinyl-4,4-diméthylazlactone) (poly(PEGMA-stat-VDM)) a été préparé par polymérisation radicalaire par transfert d’atome (ATRP) selon la technique « grafting from » à partir des MNP et utilisé pour la bioconjugaison de thymine peptide nucleic acid (PNA). La présence de l’écorce polymère et l’immobilisation du PNA ont été confirmées par différentes techniques complémentaires (FTIR, VSM).La deuxième stratégie est basée sur l’élaboration de MNP greffées pour la bioconjugaison de l’acide folique, via l’ARTP du PEGMA et de la VDM. L’analyse par microscopie électronique à transmission (TEM) a montré qu’après bioconjugaison les MNP possèdent une très bonne aptitude à la dispersion en milieu aqueux.La troisième stratégie met en œuvre la technique «grafting onto » de copolymères poly(oxyde d’éthylène)-bloc-poly(2-vinyl-4,4-dimethylazlactone) (PEO-b-PVDM) pour la préparation de nanosupports magnétiques recyclables. Des copolymères à blocs PEO-b-PVDM ont été préparés par la technique de polymérisation RAFT puis greffés sur des MNP fonctionnalisées amino-silane. Les analyses en TEM et par spectroscopie de corrélation de photons ont révélé l’aptitude à la dispersion aqueuse et à la formation de nanoclusters. Les clusters ainsi obtenus ont été utilisés en tant que nanosupports magnétiques recyclables pour l’adsorption d’anticorps. / We herein report the surface modification of magnetite nanoparticle (MNP) with copolymers containing active azlactone rings via a grafting ‘from’ and grafting ‘onto’ controlled radical polymerization (CRP) for use as a nano-solid support for immobilization with biomolecules. Three different approaches were presented as following. First, synthesis of poly(poly(ethylene glycol) methyl ether methacrylate-stat-2-vinyl-4,4-dimethylazlactone) (PEGMA-stat-VDM)-grafted MNP via a grafting ‘from’ atom transfer radical polymerization (ATRP) and its application as a platform for conjugating thymine peptide nucleic acid (PNA) monomer were presented. The presence of polymeric shell and the immobilization of thymine PNA on MNP core were confirmed by fourier transform infrared spectroscopy (FTIR) and vibrating sample magnetometry (VSM) techniques. The second strategy is based on the synthesis of MNP grafted with PEGMA and VDM via ATRP for conjugation with folic acid (FA). The existence of PEGMA and VDM in the structure was characterized by FTIR, TGA and VSM. After the FA conjugation, Transmission Electron Microscopy (TEM) results indicated that the FA-conjugated MNP having high VDM content exhibited good dispersibility in water.Third, the synthesis of MNP grafted with poly(ethylene oxide)-block-poly(2-vinyl-4,4-dimethylazlactone) (PEO-b-PVDM) block copolymer via a grafting ‘onto’ strategy and its application as recyclable magnetic nano-support for adsorption with antibody were studied. PEO-b-PVDM diblock copolymers were first synthesized using reversible addition-fragmentation chain transfer (RAFT) polymerization and then grafted onto amino-functionalized MNP. TEM images and photo correlation spectroscopy (PCS) indicated an improvement in the particle dispersibility in water after coating with the copolymers. The nanoclusters with PEO-b-PVDM copolymer coating were used as recyclable magnetic nano-supports for adsorption with antibody.

L'industrie alimentaire est de loin l'industrie la plus grande consommatrice d'eau potable et elle rejette environ 500 millions de m3 d'eaux usées par an contenant une charge organique très élevée. Un simple traitement de ce flux par des technologies conventionnelles échoue souvent en raison de facteurs de coûts. Aussi, récemment, l'accent a été largement mis sur la valorisation de ces effluents par récupération des éléments d'intérêt et la production d'eau de bonne qualité en utilisant des procédés à membrane intégrés. Les procédés membranaires couvrent pratiquement toutes les opérations unitaires utiles et nécessaires qui sont utilisés dans les usines de traitement des eaux usées. Ils apportent souvent des avantages comme la simplicité, la modularité, le caractère innovant, la compétitivité et le respect de l'environnement. Ainsi, l'objectif principal de cette thèse est le développement d'un procédé à membrane intégré comprenant microfiltration (MF), osmose directe (FO), ultrafiltration (UF) et nanofiltration (NF) pour la valorisation des eaux usées d'origine agro-alimentaire dans une logique de " zéro effluent liquide ". Nous avons pris les eaux de végétation provenant de la production d'huile d'olive comme support d'étude. Les défis associés au traitement des eaux usées de végétation sont: la variabilité des charges hydrauliques ou organiques, la présence de composés bio phénoliques, le colmatage des membranes et le rejet périodique de grands volumes d'eaux usées. En particulier, la présence de composés bio phénoliques rend ces eaux usées nocives pour l'environnement. Toutefois, la récupération de ces composés phytotoxiques peut également apporter une valeur ajoutée, car ils ont des activités biologiques intéressantes qui peuvent être exploitées dans les industries cosmétique, alimentaire et pharmaceutique. / The food industry is by far the largest potable water consuming industry that releases about 500 million m3 of wastewater per annum with very high organic loading. Simple treatment of this stream using conventional technologies often fails due to cost factors overriding their pollution abating capacity. Hence, recently the focus has been largely centered on valorization through combinatorial recovery of valuable components and reclaiming good quality water using integrated membrane process. Membrane processes practically cover all existing and needed unit operations that are used in wastewater treatment facilities. They often come with advantages like simplicity, modularity, process or product novelty, improved competitiveness, and environmental friendliness. Thus, the main focus of this PhD thesis is development of integrated membrane process comprising microfiltration (MF), forward osmosis (FO), ultrafiltration (UF) and nanofiltration (NF) for valorization of food based wastewater within the logic of zero liquid discharge. As a case study, vegetation wastewater coming from olive oil production was taken. Challenges associated with the treatment of vegetation wastewater are: absence of unique hydraulic or organic loadings, presence of biophenolic compounds, sever membrane fouling and periodic release of large volume of wastewater. Especially presence of biophenolic compounds makes the wastewater detrimental to the environment. However, recovering these phytotoxic compounds can also add economic benefit to the simple treatment since they have interesting bioactivities that can be exploited in the food, pharmaceutical and cosmetic industries.

Le travail de thèse est consacré à l'étude numérique de nanoparticules (NPs) magnétiques core@shell Fe3O4@CoO présentant des propriétés d'échange bias (EB) en utilisant la méthode Monte Carlo (MC). En particulier, nous nous sommes concentrés sur l'étude de l'effet des réponses collectives (interactions inter-particules telles que les interactions dipolaires (ID)) sur les propriétés magnétiques de ces structures. Des résultats expérimentaux préliminaires, montrant l'existence d'une relation entre le décalage du cycle d'hystérésis et l'interaction entre NPs, ont motivé le travail numérique mené dans le cadre de cette thèse.La première partie de ce mémoire est une étude méthodologique visant à trouver les conditions optimales pour simuler les cycles d'hystérésis d'une façon correcte par MC.Les résultats révèlent une dépendance linéaire entre le champ coercitif Hc et la constante d'anisotropie effective pour des conditions non biaisées (algorithme libre, algorithme du cône, algorithme mixte). La deuxième partie est consacrée à l'étude, à l'échelle atomique, des nanostructures présentant l'EB dont nous avons reproduit les deux caractéristiques (un décalage du cycle d'hystérésis, une augmentation importante de Hc).Nous avons également proposé une méthode permettant l'évaluation de la valeur de l'anisotropie effective.En passant à l'échelle d'une assemblée de NPs, plusieurs modèles furent étudiés. Nous arrivons à interpréter les résultats expérimentaux selon le degré d'agrégation des NPs. Nous montrons que l'agrégation (interactions d'échanges entre les NPs) a un effet direct sur le champ d'échange bias, mais le rôle d'ID sur le champ d'échange mérite des études complémentaires. / This thesis is dedicated to the numerical study by means of Monte Carlo (MC) simulations of core@shell Fe3O4@CoO magnetic nanoparticles (NPs) presenting exchange bias properties (EB). In particular, we focused our study on the effect of collective responses (inter-particle interactions as dipolar interactions (DI)) on the magnetic properties of these structures. Our numerical work is motivated by some preliminary experimental results showing the existence of a relationship between the hysteresis loop shift (exchange bias field) and the interaction between NPs. The first part of this thesis is a methodological study to figure out the optimal conditions to simulate hysteresis loops correctly by MC. The results reveal that the coercive field Hc is linearly related to the effective anisotropy constant for non-biased conditions (free algorithm, cone algorithm, mixed algorithm). The second part is dedicated to the study of exchange-biased nanostructures at the atomic scale. We have been able to reproduce both characteristics of EB (hysteresis loop shift, significant increase in Hc). A method allowing the evaluation of the effective anisotropy has been proposed. Considering an assembly of nanoparticles, several models are studied. The experimental results are interpreted according to the degree of aggregation of NPs. It was shown that the aggregation (exchange interactions between NPs) has a direct effect on the exchange bias field, but the role of the ID on the exchange field requires complimentary calculations to be clarified.

Made available in DSpace on 2016-06-02T19:02:43Z (GMT). No. of bitstreams: 1 5706.pdf: 7869131 bytes, checksum: 3a35e736b3418ca357ef4fc2e657c0af (MD5) Previous issue date: 2013-10-16 / Universidade Federal de Minas Gerais / Enzymes are potent catalysts, but operationally fragile, expensive and soluble. Industrial applications of enzymes, often, are possible only using immobilized enzyme. Nowadays, various studies have been performed aiming to immobilize enzymes onto magnetic carriers, which allow the selective recovery of the derivative by applying an external magnetic field even in complex reaction media containing other suspended solids. There are many studies using magnetic carriers in enzymes immobilization procedures, however there are no commercially available enzymes immobilized onto magnetic materials. In these studies usually are used carriers with not ideal characteristics for applications in industrial processes. The present study aimed to develop new magnetic carriers and methods for immobilization of enzymes in these carriers, penicillin G acylase (PGA) and cellulases have been used as model enzymes. The thesis was divided into five parts, in the first part (Chapter 1) the state-of-art is presented. The second part (Chapter 2) describes the synthesis of magnetic carriers robust, cheap and with good characteristics for applications in bioprocesses. For this purpose were tested the synthesis of silica magnetic microparticles (SMMps) in water-in-oil micro-emulsion using sodium silicate as silica source and superparamagnetic iron oxide nanoparticles as magnetic core. Materials with good magnetic properties, high surface area and mesoporous structure were obtained. SMMps structure was characterized, it was possible to control the final structure of the material according to the synthesis conditions. In the third part of this study (Chapter 3) was evaluated a new concept in enzymes immobilization using magnetic materials. Magnetic tags were co-aggregated with PGA and cross-linked with glutaraldehyde, producing magnetic cross-linked enzymes aggregates (M-CLEAs). Several reaction conditions were tested producing M-CLEAs with different characteristics and strong response to external magnetic fields. Derivatives with good recovered activity and increased thermal and methanol 50% (v/v) stabilities were obtained. M-CLEAs presented superior performance, in comparison with the free enzyme, in penicillin G hydrolysis experiments, being reused for three reaction cycles without loss of activity. In the fourth part of this study (Chapter 4) the immobilization of the Trichoderma reesei cellulolytic complex onto 17 carriers using 60 different immobilization conditions was evaluated. Covalent methods to cellulases immobilization resulted in total loss of the enzymatic activity. The immobilization by adsorption allowed preserving a portion of the enzymatic activity, however, the enzyme was desorbed from the carrier with the increase in the ionic strength. The best results were achieved for adsorption in MANAE-agarose followed by cross-linking with glutaraldehyde. Hydrolysis experiments using insoluble substrates showed that it is possible to hydrolyze such substrates even using immobilized enzyme onto porous carriers. The derivative was reused for ten reaction cycles (hydrolysis of filter paper) saving more than 90% of its activity. Finally, in Chapter 5, the T. reesei cellulolytic complex was immobilized by adsorption onto SMMp activated with amino groups followed by glutaraldehyde cross-linking achieving good results in terms of recovered activity. / Enzimas são potentes catalisadores, porém frágeis operacionalmente, caras e solúveis. Aplicações industriais desses catalisadores, muitas vezes, são possíveis apenas com o uso de enzima imobilizada. Estudos indicam que o uso de suportes magnéticos para imobilizar enzimas pode permitir a recuperação seletiva do derivado através da aplicação de um campo magnético externo mesmo em meios complexos contendo outros sólidos em suspensão. Apesar de existirem muitos estudos empregando suportes magnéticos para imobilização de enzimas, não existem enzimas imobilizadas em materiais magnéticos disponíveis comercialmente. Nestes estudos geralmente são utilizados suportes magnéticos com características não ideais para aplicações em bioprocessos. O presente estudo teve como principal objetivo o desenvolvimento de novos suportes magnéticos e métodos para imobilização de enzimas nestes suportes, a enzima penicilina G acilase (PGA) e celulases foram utilizadas como modelo. O estudo foi dividido em cinco partes, no Capítulo 1 é apresentada uma introdução indicando o estado da arte. O Capítulo 2 apresenta o preparo de novos suportes magnéticos robustos, baratos e com características ótimas para aplicações em bioprocessos. Nesta etapa foi testada a síntese de micro-partículas magnéticas de sílica (SMMps) em micro-emulsão água-em-óleo, empregando silicato de sódio como fonte de sílica e nanopartículas superparamagnéticas de óxido de ferro como núcleo magnético. Os materiais obtidos apresentaram excelentes propriedades magnéticas, alta área de superfície e estrutura mesoporosa. A partir da caracterização físico-química e morfológica das SMMps foi possível controlar a estrutura final do material de acordo com as condições de síntese. No Capítulo 3 foi avaliado um novo conceito em imobilização de enzimas empregando materiais magnéticos. Neste estudo etiquetas magnéticas foram co-agregadas com PGA e entrecruzadas com glutaraldeído, gerando agregados enzimáticos entrecruzados com propriedades magnéticas (M-CLEAs). Várias condições reacionais foram testadas rendendo M-CLEAs com diferentes características e com resposta robusta a campos magnéticos externos. Derivados imobilizados com boa atividade recuperada e incremento na estabilidade térmica e frente a metanol 50% (v/v) foram obtidos. M-CLEAs apresentaram desempenho superior ao observado para a enzima livre em experimentos de hidrólise de penicilina G, sendo reutilizados por três ciclos reacionais sem perda de atividade. No Capítulo 4 foi avaliada a imobilização do complexo celulolítico de Trichoderma reesei em 17 suportes, empregando 60 diferentes condições de imobilização. Os experimentos de imobilização realizados empregando técnicas de imobilização por união covalente ocasionaram perda total de atividade enquanto métodos de imobilização por adsorção permitiram conservar boa atividade enzimática, porém a enzima dessorveu do suporte com o aumento na força iônica do meio. Os melhores resultados foram alcançados para adsorção em MANAE-agarose seguido de entrecruzamento com glutaraldeído. Experimentos de hidrólise de substratos insolúveis mostraram que é possível hidrolisar este tipo de substrato mesmo com enzima imobilizada em suportes porosos. O derivado foi reutilizado por dez ciclos (hidrólise de papel filtro) conservando mais de 90% de sua atividade. Por fim, no Capítulo 5, o complexo celulolítico de T. reesei foi imobilizado por adsorção em SMMp ativado com grupos amino seguido de entrecruzamento com glutaraldeído apresentando bons resultados em termos de atividade recuperada.

Les particules des polymères avec des propriétés magnétiques sont utilisées dans des applications thérapeutiques in vivo, comme des agents de libération contrôlée de principes actifs, pour des applications ex vivo, dans l’extraction de cellules cancéreuses dans le corps, et finalement pour le diagnostic in vitro. La nécessité de matériaux biocompatibles et intelligents, comme agent d’encapsulation de nanoparticules magnétiques, conduit à l’utilisation de polymères hydrophiles, biodégradables, biocompatibles et dans certains cas stimulables. Dans les applications thérapeutiques, cette technologie est basée sur le déplacement des particules en appliquant un champ magnétique et sur la concentration du médicament dans la zone d’intérêt. Cette étape est suivie par la libération du médicament, en utilisant les propriétés des polymères stimulables. Ainsi, cette thèse est consacrée à l’étude de la préparation de nanoparticules composées d’une matrice polymère sensible aux stimuli et des particules d’oxyde de fer (γ-Fe2O3 et Fe3O4). Tout d’abord, nous avons étudié l’obtention de nanogels à base de poly(NVCL-co-AA) en utilisant la polymérisation par précipitation. La poly (N-vinylcaprolactama) (PNVCL)et un polymère qui possède une température critique inférieure de solubilité (LCST), proche de la température physiologique (35-38°C). Ce polymère est connu pour avoir une bonne biocompatibilité plus haute, par rapport à des autres polymères sensibles à la température. En plus, le poly (acide acrylique) (PAA) est un polymère qui présente la sensibilité au pH. Dans cette étape, on a étudié l’influence de quelques paramètres de synthèse sur les diamètres des particules, la distribution de la taille des particules et la sensibilité à la température des nanogels. La sensibilité au pH a été également évaluée en fonction de la concentration d’AA ajouté dans les synthèses. Ensuite, nous avons effectué l’étude de l’incorporation de nanoparticules magnétiques stabilisées par le dextran dans les nanogels de PNVCL réticulé en utilisant la technique de polymérisation en mini-émulsion inverse. Les nanogels magnétiques thermosensibles ont été caractérisés en termes de taille (DP), distribution de la taille des particules (DTP) en utilisant la diffusion de la lumière. Le caractère thermosensible des nanogels magnétiques a également été étudié en mesurant le diamètre hydrodynamique en fonction de la température. Les propriétés magnétiques (aimantation spécifique et la magnétisation) ont été examinées en utilisant un magnétomètre à échantillon vibrant (VSM). L’analyse par infrarouge (GTIR) et par diffraction des rayons X ont montré qualitativement l’incorporation des nanoparticules magnétiques dans la matrice polymère. L’efficacité d’encapsulation de nanoparticules d’oxyde de fer a été étudiée par l’analyse thermo-gravimétrique (TGA) et par les mesures d’aimantation. Les caractéristiques morphologique des nanogels magnétiques et stimulables ont été examinées par la microscopie électronique en transmission (TEM). / Polymeric particles with magnetic properties can be useful for in vivo therapeutic applications, as agents for controlled drug release, for ex vivo applications, as agents for the extraction of cancer cells, and finally, for the diagnosis in vitro. The search for biocompatible and smart materials as agents for the encapsulation of magnetic particles, leads to the use of stmuli-responsive polymers. In therapeutic applications, this technology is based on the localization and the concentration of the particles containing the drug in the area of interest by applying a magnetic field. This step is followed by the release of the drug, using the sensitive properties of the polymers. In this context, this thesis is devoted to the preparation of nanoparticles constituted by a stimuli-responsive polymer matrix and particles of iron oxide (γ-Fe2O3 e Fe3O4). First of all, we performed the synthesis of poly(NVCL-co-AA)-based nanogels using the precipitation polymerization method. Poly(N-vinilcaprolactama) (PNVCL) is a thermo-responsive polymer which presents the lower critical solution temperature (LCST) near the physiological temperature (35-38 °C) and it is well known by its greater biocompatibility, in comparison with other themallysensitive polymers. On the other hand, the poly(acrylic acid) (PAA) is known by its sensibility to changes in the enviromental pH. In this stage, the influence of some synthesis parameters on the particles diameter, polydispersity and themally-sensitive behavior of the nanogels was evaluated. The pH-sensibility behavior was also studied as a function of the AA concentration in the synthesis. As a second step, the study of the incorporation of dextran-coated magnetic nanoparticles in the PNVCL-based nanogels using the inverse miniemulsion polymerization was preformed. The thermo-responsive magnetic nanogels were characterized in terms of particles diameter (PD) and particles size distribution (PSD) using light scattering. The temperature sensitivity of the magnetic nanogels was also studied by light scattering, with measurements of particles diameter as a function of temperature. The magnetization measurements were obtained on a vibrating sample magnetometer (VSM). Analysis of infra-red (FTIR) and X-ray diffraction revealed qualitatively the incorporation of magnetic nanoparticles. The incorporation efficiency of iron oxide nanoparticles was studied by thermo-gravimetric analysis (TGA) and magnetic measurements. The morphological characteristics of the magnetic nanogels were observed by transmission electron microscopy (TEM). / Partículas poliméricas com propriedades magnéticas podem ser utilizadas tanto em aplicações terapêuticas in vivo, como agentes de liberação controlada de princípios ativos, ex vivo, na extração de células cancerígenas do organismo, ou ainda in vitro, em diagnósticos. A necessidade de materiais inteligentes e biocompatíveis, como agentes de encapsulação destas partículas magnéticas, leva ao uso de polímeros sensíveis a estímulos. Em aplicações terapêuticas, esta tecnologia é baseada na localização das partículas através da aplicação de um campo magnético e na concentração da droga na área de interesse. Esta etapa é seguida pela liberação da droga, utilizando-se as propriedades sensíveis dos polímeros. Dessa forma, este trabalho de tese se dedica ao estudo da obtenção de nanopartículas constituídas de uma matriz polimérica sensível a estímulos e de partículas de óxido de ferro (γ-Fe2O3 e Fe3O4). Inicialmente, nanogéis à base de poli(NVCL-co-AA) foram obtidos através do método de polimerização por precipitação. A Poli(Nvinilcaprolactama) (PNVCL) é um polímero termo-sensível, que possui temperatura crítica inferior de solubilização (LCST) próxima à temperatura fisiológica (35-38 ºC) e é conhecida, ainda, por possuir maior biocompatibilidade, em comparação a outros polímeros do gênero. O poli(ácido acrílico) (PAA), por sua vez, é um polímero que apresenta sensibilidade ao pH. Nesta etapa estudou-se a influência de alguns parâmetros de síntese nos diâmetros de partículas, na polidispersidade e na sensibilidade à temperatura dos nanogéis. A sensibilidade ao pH também foi estudada em função da concentração de ácido acrílico adicionado nas sínteses. Em seguida, realizou-se o estudo da encapsulação de nanopartículas magnéticas complexadas com dextrana em nanogéis de PNVCL, utilizando-se a técnica de polimerização em miniemulsão inversa. Os nanogéis magnéticos sensíveis à temperatura foram caracterizados quanto ao diâmetro de partículas (DP) e distribuição do diâmetro de partículas (DDP), pela técnica de espalhamento de luz. A sensibilidade à temperatura dos nanogéis magnéticos também foi estudada por espalhamento de luz, através de medidas de diâmetro de partículas em diferentes temperaturas. As medidas de magnetização foram obtidas em um magnetômetro de amostra vibrante (MAV). Análises de infravermelho (FTIR) e de difratometria de raios X revelaram qualitativamente a encapsulação das nanopartículas magnéticas. A eficiência de incorporação das nanopartículas de óxido de ferro foi estudada através de análi ses termogravimétricas (TGA) e medidas de magnetização. As características morfológicas dos nanogéis magnéticos foram observadas por microscopia eletrônica de transmissão (TEM).

Made available in DSpace on 2014-07-29T15:07:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-05-26 / In this work we investigated the encapsulation of magnetite nanoparticles into the nanopores of anodic alumina membranes using atomic force microscopy (AFM), vibrating sample magnetometer (VSM) and electron magnetic resonance (EMR). Three biocompatible magnetic fluids, with different nanoparticle diameters, stably dispersed in water at physiological conditions, were used. The nanoparticles were obtained through the coprecipitation method and characterized by X-ray diffraction, from which we obtained the nanoparticle size and confirmed the crystal structure. The Scherrer´s relation revealed a nanoparticle diameter of 10.1nm, 12.3nm and 13.8nm. The alumina membrane were prepared through anodization process. The nanopores were arranged on a hexagonal lattice with an alumina thickness of 4 μm, a distance between pores (center to center) of 105 nm, and samples containing nanopores with diameter of 35 nm or 80 nm. The method of encapsulation of nanoparticles consisted of depositing a drop of magnetic fluid into the surface of alumina. The fluid enters the nanopores through capillarity carrying the nanoparticles into it. AFM images prove that we had success in encapsulating nanoparticles only for the alumina samples with nanopores with a size of 80 nm. Magnetization data of the alumina sample containing nanoparticles with a diameter of 13.8nm encapsulated into nanopores of 80 nm, revealed an increase, with respect to the first procedure of encapsulation, of 48 % of the nanoparticles internalized into the nanopore after the second process of encapsulation. Further, different from all the samples investigated, EMR data for the alumina containing nanopores of 80 nm and nanoparticles of 13.8 nm, after the first procedure of encapsulation, had shown perpendicular magnetization with respect to the alumina surface. The EMR spetra were curve fitted using two Gaussian lines, one representing the nanoparticles with magnetization parallel to the surface and the other perpendicular. AFM images suggest, in our sample, that residues on the alumina surface are responsible for the parallel component. The magnetic resonance field data, for the perpendicular contribution, were analyzed taking into account in the energy density terms with uniaxial and cubic symmetry. The uniaxial energy contribution had a term due to magnetic dipolar interaction, between nanoparticles forming a linear chain, a magnetostatic term, due to the nanostructures self-organization, and also a magnetoelastic contribution, which came from the stress generated by the packing of nanoparticles, whose origin were related to the dipolar interaction between nanoparticles forming the linear chain. Indeed, the theoretical analysis allowed us to conclude that the mean size of the chain could vary from 4 to 9.5 nanoparticles. Finally, after heating the alumina, at 300°C for one hour, which contained nanoparticles with a size of 10.1 nm, and dissolving it in NaOH aqueous solution, AFM data were obtained. The AFM images confirmed the existence of nanowires. The diameter distribution, obtained from the AFM images, were curve fitted with a lognormal distribution revealing a modal diameter for the nanowires of 25,8 0, ± 4nm and diameter dispersity of 0,30 ± 0,02nm . / Neste trabalho investigamos o encapsulamento de nanopartículas de magnetita (Fe3O4) em nanoporos de alumina anódica utilizando as técnicas de Microscopia de Força Atômica (AFM), Magnetometria de Amostra Vibrante (VSM) e Ressonância Magnética Eletrônica (RME). Utilizamos três fluidos magnéticos com nanopartículas de diâmetros diferentes dispersas em solução fisiológica. As nanopartículas foram sintetizadas pelo método da coprecipitação e foram caracterizadas por difração de raios-x, de onde confirmamos sua estrutura cristalina e obtivemos o diâmetro. A relação de Scherrer forneceu os seguintes diâmetros: DRX=10,1nm, DRX=12,3nm e DRX=13,8nm. As membranas de alumina foram preparadas pelo método da anodização de um filme de alumínio puro, gerando nanoporos em um arranjo hexagonal, sendo a espessura da alumina de 4μm com distância entre poros centro a centro de 105nm e amostras contendo diâmetros de nanoporos de 35nm ou 80nm. O método de encapsulamento das nanopartículas consistiu em depositar uma gota do fluido magnético sobre a alumina, que penetra nos nanoporos por capilaridade, carreando as nanopartículas. Imagens de AFM mostraram que obtivemos sucesso no encapsulamento das nanopartículas em alumina somente nas amostras com nanoporos de 80nm. Uma comparação entre as curvas de magnetização da amostra com nanopartículas de DRX=13,8nm em nanoporos de 80nm, encapsuladas uma vez e duas vezes, mostrou um acréscimo de 48% no número de nanopartículas encapsuladas do primeiro para o segundo processo de encapsulamento. Além disso, diferentemente de todas as outras amostras estudadas, os dados de RME para alumina com nanoporos de 80 nm e nanopartículas com diâmetro de 13,8 nm, após o primeiro processo de encapsulamento, apresentaram magnetização perpendicular ao plano da membrana de alumina. O espectro de RME foi ajustado por duas gaussianas, uma representando uma componente com magnetização paralela e outra perpendicular. Imagens de AFM sugerem, na nossa amostra, que resíduos na superfície são responsáveis pela componente paralela. A análise dos dados do campo de ressonância para a componente perpendicular foram ajustados considerando termos de simetria uniaxial e cúbica para a densidade de energia. Na contribução uniaxial foi explicitado o termo devido à interação dipolar magnética, entre nanopartículas formando uma cadeia linear, o termo magnetostático, devido à autoorganização das nanoestruturas, e um magnetoelástico, proveniente do stress gerado pelo empacotamento das nanopartículas, cuja origem foi atribuída à interação dipolar entre as nanoestruturas formando a cadeia. A análise teórica permitiu, ainda, concluir que o tamanho médio das cadeias lineares formadas no interior dos nanoporos corresponde a 6,0 nanopartículas, podendo variar entre 4 e 11. Essas cadeias podem existir não somente em nanoporos diferentes, mas também no interior de um mesmo nanoporo. Por fim, após aquecermos a membrana de alumina, a 300°C por 1 hora, que continha nanopartículas com DRX=10,1nm e a dissolvermos em uma solução aquosa de NaOH, obtivemos imagens de AFM dos nanofios. Uma distribuição de tamanho construída a partir das imagens e ajustada por uma lognormal nos forneceu um diâmetro modal para os nanofios de 25,8 0, ± 4nm e uma dispersidade de 0,30 ± 0,02nm.