O transplante de ilhotas microencapsuladas constitui uma alternativa terapêutica interessante para o Diabetes Mellitus tipo 1, permitindo um melhor controle glicêmico e eliminando a necessidade de imunossupressão. Entretanto, a manutenção a longo prazo da viabilidade das células-&#946; ainda é um desafio. No isolamento, a perda da matriz extracelular e as condições hipóxicas subsequentes afetam decisivamente a sobrevivência e funcionalidade das ilhotas. Objetivo Para diminuir o estresse sobre o enxerto, levando a um sucesso prolongado do transplante, propôs-se a adição de perfluorocarbono (PFC) ou laminina (LN), moléculas associadas respectivamente à oxigenação e interações célula-célula, ao biomaterial baseado em alginato, Biodritina, adequado ao encapsulamento celular. Metodologia Para testar a estabilidade das formulações PFC-Biodritina e LN-Biodritina, microcápsulas foram submetidas a diferentes estresses (rotacional, osmótico, temperatura e cultura) por 7 e 30 dias. A pureza do biomaterial foi avaliada pela coincubação com macrófagos murinos RAW264.7, por 3, 9 e 24h, quando a ativação dos macrófagos foi observada pela expressão gênica de IL- 1&#946; e TNF&#945;. Microcápsulas implantadas i.p. em camundongos foram recuperadas após 7 ou 30 dias, para análises de biocompatibilidade. A expressão de níveis de mRNA (bax, bad, bcl-2, bcl-XL, xiap, caspase 3, mcp1/ccl2, hsp70, ldh, insulina 1 e 2), proteínas (Bax, Bcl-XL e Xiap) e a atividade de Caspase3 foram avaliadas em ilhotas microencapsuladas com PFC- e LN-Biodritina, após cultura de 48h em condições de normóxia e hipóxia (<2% O2). Camundongos diabéticos foram transplantados com ilhotas encapsuladas nas diferentes formulações e os animais foram monitorados pelas variações de massa corporal, glicêmicas e pela funcionalidade do enxerto (TOTGs). As ilhotas foram recuperadas de animais normo ou hiperglicêmicos e uma análise de biocompatibilidade das cápsulas foi realizada, assim como a avaliação funcional das células-&#946;. Após o explante, a glicemia dos animais normoglicêmicos foi monitorada para se atestar a eficiência das ilhotas transplantadas. Resultados Microcápsulas de PFC- e LN-Biodritina são tão estáveis e biocompatíveis quanto as de Biodritina. Para ilhotas encapsuladas em ambos os materiais, em normóxia ou hipóxia, observou-se uma modulação gênica que sugere proteção contra apoptose. Adicionalmente, encontrou-se uma diminuição na expressão de genes indicadores de estresse (mcp1, hsp70). Uma diminuição nos níveis de mRNA de ldh foi vista para PFC-Biodritina, mas o oposto foi encontrado para LN-Biodritina. As diferenças encontradas na expressão proteica sugerem o mesmo padrão anti-apoptótico. Caspase3 não foi modulada por nenhum biomaterial. Nos experimentos de transplante, apenas LN-Biodritina levou reversão prolongada do diabetes, com 60% dos animais normoglicêmicos, 198 dias pós-cirurgia, comparado a 9% do grupo Biodritina. O TOTG demonstrou que camundongos transplantados com ilhotas encapsuladas secretaram mais insulina do que controles, 60 (LN-Biodritina) ou 100 (PFC- e LN-Biodritina) dias pós-cirurgia. O explante restabeleceu a hiperglicemia nos camundongos. Microcápsulas recuperadas de animais hiperglicêmicos apresentavam uma extensa adesão celular. Testes de secreção de insulina in vitro demonstraram que somente ilhotas do grupo normoglicêmico responderam às variações da concentração de glicose. Conclusão A adição de moléculas bioativas à Biodritina é capaz de diminuir o estresse em ilhotas isoladas e tem o potencial de melhorar a terapia pelo transplante de ilhotas. / Transplantation of microencapsulated islets represents an attractive therapeutical approach to treat type 1 Diabetes Mellitus, accounting for an improved glycemic control and the abolishment of immunosuppressive therapies. However, maintenance of long-term &#946;-cell viability remains a major problem. During islet isolation, the loss of extracellular matrix interactions and the hypoxic conditions thereafter dramatically affect &#946;-cell survival and function. Objective To lessen the burden of islet stress and achieve a better outcome in islet transplantation we tested the addition of perfluorocarbon (PFC) or laminin (LN), molecules associated respectively with oxygenation and cell-cell interaction, to Biodritin, an alginate-based material suitable for cell microencapsulation. Methodology To test the stability of PFC-Biodritin and LN-Biodritin composites, microcapsules were subjected to different stresses (rotational, osmotic, temperature and culture) for 7 and 30 days. To assess biomaterial purity microcapsules were co-incubated with RAW264.7 murine macrophage cell line for 3, 9 and 24h and macrophage activation was detected through mRNA levels of IL-1&#946; and TNF&#945;. Microcapsules were implanted i.p. in mice and retrieved after 7 or 30 days, for biocompatibility analyses. Gene expression at mRNA (bax, bad, bcl-2, bcl-XL, xiap, caspase 3, mcp1/ccl2, hsp70, ldh, insulin 1 and 2) and protein (Bax, Bcl-XL and Xiap) levels, together with Caspase3 activity, were evaluated in islets microencapsulated in PFC- or LN-Biodritin, upon culturing for 48h in normoxic or hypoxic (<2% O2) conditions. Diabetic mice were transplanted with PFC- or LN-Biodritin microencapsulated islets, followed by assessments of body weight, glycemia and graft function by oral glucose tolerance tests (OGTTs). Microencapsulated islets were retrieved from normoglycemic or hyperglycemic mice and biocompatibility analyses of the beads together with a functional assessment of the graft followed. After graft removal, normoglycemic animals had their glycemias monitored to attest the efficacy of the transplanted islets. Results PFC- and LN-Biodritin microcapsules were as stable and biocompatible as Biodritin. For both biomaterials in normoxia and hypoxia a modulation in gene expression was observed in islets associated with a protection against apoptosis. Also, a decreased expression of stress-related genes (mcp1, hsp70) was evidenced. ldh mRNA levels were down-regulated in PFC-Biodritin microencapsulated islets but upregulated in the presence of LN. Increased levels of insulin mRNA were observed. The differences seen in protein expression indicated the same anti-apoptotic pattern. Caspase3 activity was not different between groups. Concerning diabetes reversal experiments, only mice transplanted with LN-Biodritin microencapsulated islets presented a better outcome, with 60% remaining euglycemic at 198 days post-surgery, compared with 9% for the Biodritin group. OGTT showed that mice transplanted with encapsulated islets secreted more insulin than normal mice, 60 (LN-Biodritin) or 100 days (PFC- and LN-Biodritina) posttransplant. Hyperglycemia was achieved after the retrieval of microcapsules showing graft efficacy. Retrieved microcapsules revealed an extensive overgrowth in most beads from hyperglycemic mice. A static glucose stimulated insulin secretion test revealed that only islets from normoglycemic subjects were able to secrete insulin according to glucose concentration. Conclusion- The addition of bioactive molecules to Biodritin may lessen the stress of isolated islets and have the potential to improve islet transplantation therapy.

Neste trabalho foi feito um estudo para obtenção de formulações multiparticuladas, formadas por micropartículas de polímeros naturais, solúveis em água, não tóxicos e biodegradáveis. Os polímeros utilizados foram: Caseína, Hidroxietilcelulose (HEC), Hidroxipropilmetilcelulose (HPMC), Alginato de sódio e Quitosana. As técnicas utilizadas para obtenção das micropartículas foram o spray drying e a atomização de alginato de sódio em uma solução de CaCl2, para gelificação das gotículas formadas, com uma complexação ou não com quitosana. O estudo de micropartículas secas de alginato de cálcio e alginato de cálcio recoberta com uma membrana de quitosana revelou que diferentes procedimentos e variáveis de processo influenciavam nas características das micropartículas obtidas. O diâmetro médio das micropartículas de alginato e alginato-quitosana variou de 60 a 553 &#181;m. A superfície de micropartículas com quitosana se mostrou mais rugosa, com uma grande quantidade de poros menores que 1 &#181;m. As micropartículas com diâmetro médio de 60 &#181;m apresentaram uma boa esfericidade e uma distribuição de tamanho de partícula uniforme. O teor de cálcio apresentou variações, diminuindo em processos com quitosana. A maioria das micropartículas de alginato e alginato-quitosana eram estáveis em água, mas instáveis em soro fisiológico e tampão fosfato 0,1M. Após a realização do estudo das características destas micropartículas foram incorporados a elas materiais com atividade bioinseticida (Bacillus thuringiensis var. krusfaki (Btk) e de Baculovirus (Bv)). As micropartículas obtidas com alginato de cálcio formaram pós contendo sistemas matriciais capazes de microencapsular e reter microrganismos entomopatogênicos, inclusive após redispersão em água. Os processos de microencapsulação de desenvolvidos e avaliados demonstraram ser adequados para a manutenção da viabilidade de Btk e da integridade dos poliedros de Bv, assim como, a conservação da capacidade bioinseticida destes microrganismos. As micropartículas obtidas com as misturas poliméricas Caseína/HEC e Caseína/HPMC pelo processo de spray drying revelaram-se inadequadas na medida que se dissolviam rapidamente depois de dispersas em água e não poderiam assim proteger o material bioinseticida no meio ambiente. / The aim of this work was to develop powder formulations containing microparticles, to be used as multiparticulate delivery systems. Two different methods were investigated : 1) the preparation of microparticles by spray drying using casein, hydroxyethylcellulose (HEC) and hydroxypropilmetilcellulose (HPMC) and 2) the preparation of calcium alginate and chitosan-alginate microparticles by using an atomizer device. Different experimental procedures to prepare calcium alginate and chitosanalginate microparticles were evaluated and variables beHeved to be important for the membrane formation were examined. The mean particles diarneter ranged from 60 to 553 &#181;m. When a comparison was made between the surface morphology of calcium alginate and alginate-chitosan microparticles, remarkable roughness and more porous structure was observed in the chitosan-alginate microparticles. Some properties of the microparticles depended on the method and the procedure conditions of forming the chitosan-alginate complex. Calcium alginate and chitosan-alginate microparticles containing two different bioinsecticides were also prepared: 1) a spore/&#948;-endotoxinscomplex of Bacillus thuringiensis var.kurstaki (Btk) and 2) polyhedra of Baculovirus anticarsia (Bv), a viral insecticide. The results shown that the encapsulation of suspensions of Btk containing spore/toxin complex or polyhedra of B. anticarcia in calcium alginate and chitosan-alginate microparticles did not decrease the larvicidal activity of these biopesticides against lepidopterous pests. The formulations developed in this study remained unchanged, did not swell, did not release the spores (Btk) or the polyhedra (Bv), when dispersed in water and could be useful to be applied by aqueous spray as bioinsecticides in agriculture. On the other hand, casein, HEC and HPMC microparticles prepared by spray drying were not suitable to encapsulate bioinsecticides because they dissolved fastly after dispersion in water.

Neste trabalho foi feito um estudo para obtenção de formulações multiparticuladas, formadas por micropartículas de polímeros naturais, solúveis em água, não tóxicos e biodegradáveis. Os polímeros utilizados foram: Caseína, Hidroxietilcelulose (HEC), Hidroxipropilmetilcelulose (HPMC), Alginato de sódio e Quitosana. As técnicas utilizadas para obtenção das micropartículas foram o spray drying e a atomização de alginato de sódio em uma solução de CaCl2, para gelificação das gotículas formadas, com uma complexação ou não com quitosana. O estudo de micropartículas secas de alginato de cálcio e alginato de cálcio recoberta com uma membrana de quitosana revelou que diferentes procedimentos e variáveis de processo influenciavam nas características das micropartículas obtidas. O diâmetro médio das micropartículas de alginato e alginato-quitosana variou de 60 a 553 &#181;m. A superfície de micropartículas com quitosana se mostrou mais rugosa, com uma grande quantidade de poros menores que 1 &#181;m. As micropartículas com diâmetro médio de 60 &#181;m apresentaram uma boa esfericidade e uma distribuição de tamanho de partícula uniforme. O teor de cálcio apresentou variações, diminuindo em processos com quitosana. A maioria das micropartículas de alginato e alginato-quitosana eram estáveis em água, mas instáveis em soro fisiológico e tampão fosfato 0,1M. Após a realização do estudo das características destas micropartículas foram incorporados a elas materiais com atividade bioinseticida (Bacillus thuringiensis var. krusfaki (Btk) e de Baculovirus (Bv)). As micropartículas obtidas com alginato de cálcio formaram pós contendo sistemas matriciais capazes de microencapsular e reter microrganismos entomopatogênicos, inclusive após redispersão em água. Os processos de microencapsulação de desenvolvidos e avaliados demonstraram ser adequados para a manutenção da viabilidade de Btk e da integridade dos poliedros de Bv, assim como, a conservação da capacidade bioinseticida destes microrganismos. As micropartículas obtidas com as misturas poliméricas Caseína/HEC e Caseína/HPMC pelo processo de spray drying revelaram-se inadequadas na medida que se dissolviam rapidamente depois de dispersas em água e não poderiam assim proteger o material bioinseticida no meio ambiente. / The aim of this work was to develop powder formulations containing microparticles, to be used as multiparticulate delivery systems. Two different methods were investigated : 1) the preparation of microparticles by spray drying using casein, hydroxyethylcellulose (HEC) and hydroxypropilmetilcellulose (HPMC) and 2) the preparation of calcium alginate and chitosan-alginate microparticles by using an atomizer device. Different experimental procedures to prepare calcium alginate and chitosanalginate microparticles were evaluated and variables beHeved to be important for the membrane formation were examined. The mean particles diarneter ranged from 60 to 553 &#181;m. When a comparison was made between the surface morphology of calcium alginate and alginate-chitosan microparticles, remarkable roughness and more porous structure was observed in the chitosan-alginate microparticles. Some properties of the microparticles depended on the method and the procedure conditions of forming the chitosan-alginate complex. Calcium alginate and chitosan-alginate microparticles containing two different bioinsecticides were also prepared: 1) a spore/&#948;-endotoxinscomplex of Bacillus thuringiensis var.kurstaki (Btk) and 2) polyhedra of Baculovirus anticarsia (Bv), a viral insecticide. The results shown that the encapsulation of suspensions of Btk containing spore/toxin complex or polyhedra of B. anticarcia in calcium alginate and chitosan-alginate microparticles did not decrease the larvicidal activity of these biopesticides against lepidopterous pests. The formulations developed in this study remained unchanged, did not swell, did not release the spores (Btk) or the polyhedra (Bv), when dispersed in water and could be useful to be applied by aqueous spray as bioinsecticides in agriculture. On the other hand, casein, HEC and HPMC microparticles prepared by spray drying were not suitable to encapsulate bioinsecticides because they dissolved fastly after dispersion in water.

A quitosana é um biopolímero produzido a partir da quitina, presente na casca de crustáceos. O grande interesse nesta substância é atribuído à sua diversidade de aplicações industriais. A quitosana utilizada neste estudo foi quimicamente modificada para tornar-se solúvel em água. Estudou-se a estabilidade de emulsões de d-limoneno com esta quitosana, a retenção e liberação do aroma contido nas microcápsulas liofilizadas e o comportamento higroscópico destas microcápsulas. Os mesmos estudos foram realizados com maltodextrina, para comparação, já que é um produto muito utilizado como agente de encapsulação de aromas. A estabilidade das emulsões foi analisada por espectrofotometria e por cromatografia gasosa associada à técnica da análise do espaço livre. As emulsões foram caracterizadas por microscopia óptica. A análise da liberação do aroma contido nas microcápsulas foi realizada em função da umidade relativa do ambiente e da temperatura. Observou-se boa estabilidade de emulsões de d-limoneno com quitosana succinilada ao longo do tempo e características muito distintas em relação às observadas nas outras emulsões. Verificou-se 72% de retenção de d-limoneno em maltodextrina (40% p/p) e 62% de retenção de d-limoneno em quitosana modificada (1% p/p) após o processo de liofilização, sendo que durante todas as etapas do processo de encapsulação a perda de aroma em maltodextrina foi de 72%, enquanto em quitosana modificada foi de 50%. As microcápsulas de maltodextrina sofreram colapso estrutural em ambiente de 90% de umidade relativa. Os valores de umidades de monocamada das microcápsulas decaíram com o aumento da temperatura. O calor total de sorção de microcápsulas de quitosana modificada apresentou-se superior, assim como seu coeficiente de difusividade. A área superficial dos poros de microcápsulas de quitosana modificada apresentou-se superior, absorvendo água por capilares. Conclui-se que microcápsulas de quitosana modificada podem apresentar-se como uma nova opção na escolha de agentes de encapsulação de aromas. / Chitosan is a biopolymer derived from chitin, a component of the shells of crustaceans. It has been of interest due to their broad range of industrial applications. In this study, the chitosan used was chemically modified in order to become water soluble. It was studied the stability of d-limonene emulsion with modified chitosan, flavour retention and release and the hygroscopic behaviour of the microcapsules. The same tests were made with maltodextrin that is a product widely used as a flavour encapsulating agent. The stability of the emulsion was analyzed by spectrofotometry and by headspace/gas chromatography technique. The emulsion characterization was obtained by optical microscopy. Flavour release of microcapsules was measured under different relatives humidities and temperatures. It was observed an improvement on the stability of d-limonene emulsion with modified chitosan along the time and different characteristics in relation to the other. Retention of d-limonene after lyophilization in maltodextrin (40% w/w) was 72% and 62% in modified chitosan (1% w/w). Although, flavour released after encapsulation process in maltodextrin was 72% while in modified chitosan was 50%. Maltodextrin microcapsules presented structural collapse in 90% of relative humidity. The humidity monolayers decreased with temperature increasing for both microcapsules. The heat of sorption for the chitosan microcapsules was greater as well as its diffusivity coefficient. Its pore superficial area was greater. It was observed that chitosan microcapsules absorb water by capilariy absorption. The results suggest that chitosan microcapsules could be a new option of flavour encapsulating agent.

Fish oil is an excellent source of long chain polyunsaturated fatty acids (LC-PUFA), which can reduce the risk of cardiovascular disease in addition to other health benefits. However, the average intake of LC-PUFA in the Western diet is much lower than the recommended levels. Fish oil is prone to oxidative deterioration when exposed to oxygen and thus must be protected in order to be used in food products. Microencapsulation is one possibility that is already applied by the industry to protect fish oil. However, most of the conventional microencapsulation techniques suffer from shortcomings such as harsh processing conditions or the use of numerous chemicals. The main objective of this thesis was to develop a novel spray process to microencapsulate fish oil based on supercritical fluid (SCF) technology using supercritical carbon dioxide (SC-CO2) and CO2-expanded ethanol (CX EtOH). Fundamental physicochemical properties essential for optimal process design were lacking in the literature; therefore, density, interfacial tension (IFT) and viscosity of fish oil in the form of triglycerides and fatty acid ethyl esters were determined at different temperatures and pressures. Fish oil when equilibrated with SC-CO2 at elevated pressure expanded by up to about 40% in volume and increased in density by up to about 5%. Furthermore, IFT of fish oil in contact with SC-CO2 decreased substantially by an order of magnitude with an increase in CO2 pressure. When fish oil was in contact with CX EtOH, IFT decreased to ultra low levels at pressures of less than 10 MPa. Viscosity of fish oil equilibrated with SC-CO2 decreased substantially with pressure but increased with shear rate. Based on the physicochemical properties determined in this research, a novel process to produce micro- and nano-sized particles containing fish oil was developed based on a SCF spray-drying method. Key processing parameters have been evaluated and can be further optimized to improve encapsulation efficiency. Determination of physicochemical properties contributed to the fundamental understanding of the behavior of the fish oil+CO2 system with and without ethanol under high pressure conditions. The new microencapsulation process shows great potential for the delivery of bioactives in various product applications. / Bioresource and Food Engineering

Des microparticules de rifampicine pour l'administration intraoculaire ont été développées en vue du traitement prophylactique de l'endophtalmie post-chirurgicale. Ces microparticules à base d'un polymère biodégradable, le PLGA, sont produites par émulsion-diffusion. Tout d'abord l'influence des paramètres de formulation et de procédé sur les caractéristiques des microparticules (taille, charge, taux d'encapsulation...) a été étudiée. La formulation a par la suite pu être optimisée par la mise en place d'un plan d'expérience, ce qui a permis de mettre en valeur les paramètres clefs influençant les propriétés des microparticules (concentrations en PLGA et PVA, présence ou non d'un coeur huileux...) et donc d'optimiser la formulation pour obtenir des microparticules adéquates pour l'administration intraoculaire de rifampicine (1μm

This work deals with the fabrication process of low density foam shells and the sharp control of their shape (diameter, thickness, density, sphericity, non-concentricity). During this PhD we focused on the non-concentricity criterion which has to be lower than 1%. The shells are synthesized using a microencapsulation process leading to a double emulsion and followed by a thermal polymerization at 60°C. According to the literature, three major parameters, the density of the three phases, the deformations of the shells along the process and the kinetics of the polymerization have a direct influence on the shells non-concentricity. The results obtained showed that when the density gap between the internal water phase and the organic phase increases, the TMPTMA shells non-concentricity improves. A density gap of 0.078 g.cm-3 at 60°C, leads to an average non-concentricity of 2.4% with a yield of shells of 58%. It was also shown that the synthesis process can be considered as reproducible. While using the same internal water phase, equivalent non-concentricity results are obtained using either a straight tube, a tube with areas of constriction or a short wound tube. The time required to fix the shell's shape is at least 20 minutes with thermal polymerization. So, it seems that the time spent by the shells inside the rotating flask allows the centering of the internal water phase inside the organic phase, whatever the circulation process used. In order to get higher polymerization rates and to avoid destabilization phenomena, we then focused our study on photopolymerization. When the synthesis is performed using a UV lamp with an efficient light intensity, the shells have a slightly higher thickness than the shells synthesized by thermal polymerization. Moreover, a really higher yield, around 80%, is achieved with UV polymerization. However, the average non-concentricity of the shells synthesized lays around 20%, which is really high compared to the 2.4% average non-concentricity obtained with thermal polymerization. It would be interesting to expose the shells to UV light at different times after collection in order to study the influence of the agitation time on the shells non-concentricity.

No description available.

L’objectif principal de cette recherche est de contribuer au développement de biocapteurs commerciaux utilisant des surfaces de papier comme matrices d’immobilisation, capables de produire un signal colorimétrique perceptible dans les limites sensorielles humaines. Ce type de biocapteur, appelé papier bioactif, pourrait servir par exemple à la détection de substances toxiques ou d’organismes pathogènes. Pour atteindre l’objectif énoncé, ce travail propose l’utilisation de systèmes enzymatiques microencapsulés couchés sur papier. Les enzymes sont des catalyseurs biologiques dotés d’une haute sélectivité, et capables d'accélérer la vitesse de certaines réactions chimiques spécifiques jusqu’à des millions des fois. Les enzymes sont toutefois des substances très sensibles qui perdent facilement leur fonctionnalité, raison pour laquelle il faut les protéger des conditions qui peuvent les endommager. La microencapsulation est une technique qui permet de protéger les enzymes sans les isoler totalement de leur environnement. Elle consiste à emprisonner les enzymes dans une sphère poreuse de taille micrométrique, faite de polymère, qui empêche l’enzyme de s’echapper, mais qui permet la diffusion de substrats à l'intérieur. La microencapsulation utilisée est réalisée à partir d’une émulsion contenant un polymère dissous dans une phase aqueuse avec l’enzyme désirée. Un agent réticulant est ensuite ajouté pour provoquer la formation d'un réseau polymérique à la paroi des gouttelettes d'eau dans l'émulsion. Le polymère ainsi réticulé se solidifie en enfermant l’enzyme à l'intérieur de la capsule. Par la suite, les capsules enzymatiques sont utilisées pour donner au papier les propriétés de biocapteur. Afin d'immobiliser les capsules et l'enzyme sur le papier, une méthode courante dans l’industrie du papier connu sous le nom de couchage à lame est utilisée. Pour ce faire, les microcapsules sont mélangées avec une sauce de couchage qui sera appliquée sur des feuilles de papier. Les paramètres de viscosité i de la sauce et ceux du couchage ont été optimisés afin d'obtenir un couchage uniforme répondant aux normes de l'industrie. Les papiers bioactifs obtenus seront d'abord étudiés pour évaluer si les enzymes sont toujours actives après les traitements appliqués; en effet, tel que mentionné ci-dessus, les enzymes sont des substances très sensibles. Une enzyme très étudiée et qui permet une évaluation facile de son activité, connue sous le nom de laccase, a été utilisée. L'activité enzymatique de la laccase a été évaluée à l’aide des techniques analytiques existantes ou en proposant de nouvelles techniques d’analyse développées dans le laboratoire du groupe Rochefort. Les résultats obtenus démontrent la possibilité d’inclure des systèmes enzymatiques microencapsulés sur papier par couchage à lame, et ce, en utilisant des paramètres à grande échelle, c’est à dire des surfaces de papier de 0.75 x 3 m2 modifiées à des vitesses qui vont jusqu’à 800 m/min. Les biocapteurs ont retenu leur activité malgré un séchage par évaporation de l’eau à l’aide d’une lampe IR de 36 kW. La microencapsulation s’avère une technique efficace pour accroître la stabilité d’entreposage du biocapteur et sa résistance à l’exposition au NaN3, qui est un inhibiteur connu de ce biocapteur. Ce projet de recherche fait partie d'un effort national visant à développer et à mettre sur le marché des papiers bioactifs; il est soutenu par Sentinel, un réseau de recherche du CRSNG. / The main objective of this research is the development of a commercial biosensor immobilized on paper surfaces, able to produce a colorimetric signal detected by human sensorial limits. This kind of biosensor could be used, for example, in the detection of toxic substances or pathogens. To achieve this objective, microencapsulated enzymes fixed on paper are proposed. Enzymes are biological catalysts with a high selectivity that can accelerate the speed of some chemical reactions up to a million times. However, the enzymes are very sensitive substances that lose their functionality easily; it is therefore necessary to protect them from conditions that could damage them. Microencapsulation is a technique that protects the enzymes without totally isolating them from their environment. In fact, microencapsulation entraps the enzymes into a micron size sphere, made of a porous polymer which prevents the enzyme to be released but allows the diffusion of its substrate inside. The microencapsulation process consists in making an emulsion containing a polymer dissolved in an aqueous phase with the desired enzyme, and the wall of the microcapsule is formed by adding a crosslinking agent that forms a polymer network at the interface of the emulsion. The crosslinked polymer solidifies and it encloses the enzyme in the interior of the capsule. Thereafter, this kind of microcapsules are used to give biosensor properties to the paper. Blade coating technique is used in order to immobilize the enzyme capsules on paper because it is the most widely used method in the paper industry. The microcapsules are mixed with a coating suspension and applied on sheets of paper. The viscosity parameters of the suspension and those of the coating are optimized to obtain a uniform coating in order to meet the industry standards. Bioactive paper obtained is first studied to assess whether the enzymes are still active or not after all the treatments because, as described above, enzymes are iii very sensitive substances. An enzyme known as laccase is used, which allows an easy evaluation of its activity. Enzymatic activity was evaluated through existing analytical techniques or new analysis techniques developed in the Rochefort lab. The results demonstrate the possibility to transfer microencapsulated enzyme systems onto paper by blade coating, by using large scale settings, with paper surfaces of 0.75 x 3 m2 modified at speeds ranging up to 800 m/min. Biosensors retained their activity, despite a drying process by evaporation of water using an IR lamp of 36 kW. The microencapsulation technique proposed here is an effective technique to increase the storage stability of the biosensor and its resistance to exposure to NaN3, which is a known inhibitor of this biosensor. This research is part of a national effort in order to develop a commercial device called bioactive paper; it is supported by the NSERC research network Sentinel.

This dissertation presents a new class of cancer diagnostic agents composed of quantum dots, magnetic nanoparticles, and amphiphilic polymers. The central hypothesis is that biocompatible, amphiphilic block copolymers can be used to create multinanoparticle micellar probes with imaging capabilities and surface properties optimized for applications in cancer diagnostics. To test this hypothesis, we investigated a number of different block copolymer structures and synthetic procedures. We found that use of a poly(methyl methacrylate)-poly(ethylene oxide) polymer in conjunction with a dialysis-based procedure produced uniform probes with excellent imaging properties. We also found that the probes formed using these materials and methods were surprisingly stable, even after incubation in whole human blood for 24 hrs at 37oC. As a corollary, we hypothesized that modified polymer structures could be used to introduce functional groups for use in linking the micellar probes to biological molecules. To test this hypothesis, we used a modified version of our synthetic procedure and utilized a novel method for studying nanoparticle binding to biological molecules in real time. We found that active amine groups could be added to the polymer shell using these methods, and that surface plasmon resonance could be used for studying nanoparticle binding. In sum, this dissertation makes several contributions to the field of cancer nanotechnology. First, we provide a new encapsulation procedure and nanostructure that has promising physical and biological properties. Secondly, we provide general strategies that can be used for future nanoprobe development. Finally, we demonstrate the capability of a new method for quantitative study of probe binding characteristics. Together, these contributions drive the field of cancer nanotechnology forward by providing a deeper understanding of the relationship between surface design and behavior in biological systems.