This experiment aimed to evaluate the replacement of soybean meal by non-protein nitrogen sources (NPN) in combination with ground or whole corn in the diet concentrate fraction of confined cattle on the dietary intake, blood protein metabolite analyzes, and performance and ingestive behavior. The experiment was developed at the Laboratório de Bovinocultura de Corte of the Department of Zootechnics of UFSM. We used 53 steers, with Charolais or Nellore predominance, with initial mean age and weight of 22 months and 250 kg. Experimental diets contained voluminous:concentrate ratio of 50:50 based on dry matter. The treatments were: soybean meal + whole grain corn; soybean meal + milled corn; common urea + whole grain corn; common urea + milled corn; protected urea + whole grain corn; protected urea + milled corn. The experimental design was completely randomized in a 3x2 factorial scheme. There was no significant interaction between nitrogen source and corn grain form for any of the variables studied (P>0.05). Dry matter and crude protein intakes were higher for animals fed with soybean meal both in kg and in kg/100 live weight. Daily weight gain was higher for steers fed with soybean meal (1,600 kg/day) vs common urea (1,238 kg/day) and protected urea (1.217 kg/day). Corn presentation did not reflect any differences in the parameters of consumption, performance and behavior. The concentration of circulating albumin showed a significant interaction between nitrogen source and date of collection (P<0.05). Animals fed with soybean meal presented increasing albumin over the experimental period, while animals fed with NPN had growth in albumin levels from the first to the second collection, stabilizing up to the end of the study. The values of globulin presented interaction of the dates of collection with nitrogen sources and also with forms of corn grain, having an inverse behavior to the value of albumin. The albumin/total protein ratio increased throughout the experimental period, except for steers that consumed protected urea, which had similar values between the first and last collection. Feeding time of soybean meal was higher than protected urea, while common urea had intermediate behavior. Idling time differed among all treatments, with superiority to soybean meal, while common urea was higher than protected urea. NPN sources promoted higher rumination time and number of chewing per ruminal bolus and lower ruminating efficiencies of DM and NDF than soybean meal (P<0.05). Higher daily chewing times were observed for protected urea, followed by common urea, which was higher than soybean meal. In the number of chews similar behavior was observed, only with common urea presenting intermediate values. The use of true protein source in the diet of confined steers provides better results in aspects of performance, blood metabolites and ingestive behavior in relation to NPN in the two evaluated forms. / O objetivo deste experimento foi avaliar a substituição do farelo de soja por fontes de nitrogênio não proteico (NNP) em combinação com milho moído ou inteiro na fração concentrado da dieta de bovinos confinados através do desempenho, comportamento ingestivo e análises dos metabólitos proteicos sanguíneos. O experimento foi desenvolvido no Laboratório de Bovinocultura de Corte do Departamento de Zootecnia da UFSM. Foram utilizados 53 novilhos, mestiço Charolês Nelore com idade e peso médios iniciais de 22 meses e 250 kg. As dietas experimentais continham relação volumoso: concentrado de 50:50, com base na matéria seca. Os tratamentos foram: farelo de soja + milho grão inteiro; farelo de soja + milho moído; ureia comum + milho grão inteiro; ureia comum + milho moído; ureia protegida + milho grão inteiro; ureia protegida + milho moído. O delineamento experimental foi o inteiramente casualizado, em esquema fatorial 3x2. Não houve interação significativa entre fonte nitrogenada e forma do grão de milho para nenhuma das variáveis estudadas (P>0,05). Os consumos de matéria seca e proteína bruta foram superiores para animais alimentados com farelo de soja tanto em kg quanto em kg/100 de peso vivo. O ganho de peso diário foi superior para novilhos alimentados com farelo de soja (1,600 kg/dia) vs ureia comum (1,238 kg/dia) e ureia protegida (1,217 kg/dia). A forma de apresentação do milho não refletiu nenhuma diferença nos parâmetros de consumo, desempenho e comportamento. A concentração de albumina circulante apresentou interação significativa entre fonte nitrogenada e data de coleta (P<0,05). Animais alimentados com farelo de soja apresentaram albumina crescente ao longo do período experimental, enquanto os animais alimentados com NNP tiveram crescimento nos níveis de albumina da primeira para a segunda coleta, estabilizando até o final do estudo. Os valores de globulina apresentaram interação das datas de coleta com fontes nitrogenadas e também com formas do grão de milho, tendo comportamento inverso ao valor de albumina. A relação albumina/proteínas totais aumentou ao longo do período experimental, exceto para os novilhos que consumiram ureia protegida, os quais tiveram valores semelhantes entre a primeira e última coleta. O tempo de alimentação da ureia protegida foi superior ao farelo de soja, enquanto a ureia comum teve comportamento intermediário. O tempo de ócio diferiu entre todos os tratamentos, com superioridade para o farelo de soja, enquanto a ureia comum foi superior à ureia protegida. As fontes de NNP promoveram maior tempo de ruminação e número de mastigadas por bolo e menores eficiências de ruminação de MS e FDN do que o farelo de soja (P<0,05). Maiores tempos de mastigação diária foram observados para a ureia protegida, seguida da ureia comum, que foi superior ao farelo de soja; no número de mastigadas mericíclicas se observou comportamento semelhante, apenas com a ureia comum apresentando valores intermediários. A utilização de fonte de proteína verdadeira na dieta de novilhos confinados proporciona melhores resultados nos aspectos de desempenho, metabólitos sanguíneos e comportamento ingestivo em relação ao NNP nas duas formas avaliadas.

Le but de notre travail était l’étude des dysfonctions artérielle et musculaire dans des modèles d’agression aiguë et chronique puis l’évaluation de l’effet de la modulation pharmacologique du stress oxydant sur les anomalies vasculaires induites. Dans le modèle aigu de choc endotoxinique chez la souris, l’administration d’albumine humaine a permis d’augmenter leur survie, d’améliorer l’hyporéactivité vasculaire et la dysfonction endothéliale grâce à des propriétés ,concentration-dépendante, anti-oxydantes, modulatrices de l’inflammation, aboutissant à une diminution de l’expression de la NOS inductible et à une amélioration fonctionnelle et quantitative de la NOS endothéliale. L’administration chronique chez le rat de ciclosporine ou d’évérolimus induit une dysfonction endothéliale caractérisée par une altération de la voie de signalisation du NO et de l’EDHF et associée à une augmentation locale du stress oxydant. Au niveau musculaire cardiaque, seul l’évérolimus induit une augmentation de radicaux libres et une diminution de la respiration mitochondriale ventriculaire droite. L’administration préventive de polyphénols a permis, une amélioration de la composante EDHF de la dysfonction endothéliale, ciclosporine-induite, via une diminution de la surexpression de la NADPH oxydase et du stress oxydant vasculaire. Ainsi, le stress oxydant est une entité commune, impliquée dans la genèse de dysfonctions vasculaire et musculaire. Sa modulation par l’administration d’albumine ou de polyphénols a permis respectivement l’amélioration de l’hyporéactivité vasculaire du choc endotoxinique et de la dysfonction endothéliale dans nos deux modèles. / The aim of this work was to study the arterial and muscular dysfunctions in two rodent models of acute and chronic injuries, then the evaluation of pharmacological evaluation of oxidative stress on vascular abnormalities induced in these models.In the acute model, a mouse endotoxic shock, human serum albumin administration improved survival, endothelial dysfunction and reduced vascular hyporeactivity thanks to its concentration-dependant , anti-oxidative and anti-inflammatory properties, leading to a decrease of inductible NOS expression and an qualitative and quantitative increase of endothelial NOS.In the chronic model, rat’s administration of ciclopsorine and everolimus induced an endothelial dysfunction characterized by alterations of both NO and EDHF pathways and associated with an in situ oxidative stress. In the cardiac muscle, an increase of reactive oxygen species and a decrease of right ventricle mitochondrial respiratory chain complexes activities was observed only with everolimus. The preventive administration of polyphenols markedly increased the blunted EDHF component, but not the NO component, of the cyclosporine-induced endothelial dysfunction by reducing NADPH oxidase over expression and oxidative vascular stress. Finally, oxidative stress is a common entity, implicated in genesis of vascular and muscular dysfunctions. Its modulation by human albumin or polyphenols administration improves respectively arterial hyporeactivity of endotoxinic shock and endothelial dysfunction in our two models.

Au laboratoire, une méthode originale d'encapsulation par transacylation entre l'alginate de propylène-glycol (PGA) et une protéine a été mise au point. Cette méthode est basée sur la création de liaisons amides entre les fonctions amines libres de la protéine et les groupes esters du PGA dans une phase aqueuse émulsionnée (E/H) après alcalinisation. Les microparticules obtenues, stables, biocompatibles et biodégradables, sont potentiellement intéressantes pour la délivrance de substances actives en thérapeutique ou en cosmétique.Le premier objectif de ce travail est d'étudier l'influence des propriétés physicochimiques des deux biopolymères (protéine et PGA) et de leurs solutions, ainsi que l'effet des paramètres de préparation sur la réaction de transacylation et sur les propriétés des microparticules obtenues. Pour cela, la sérumalbumine humaine (HSA) a servi de protéine modèle et les microparticules ont été préparées dans différentes conditions physicochimiques puis caractérisées. Différents liens ont été établis entre les propriétés physicochimiques des solutions initiales des deux polymères et les propriétés fonctionnelles des microparticules obtenues.Le deuxième objectif est de remplacer le PGA, seul polysaccharide utilisable jusqu'à présent pour la microencapsulation par transacylation, par d'autres polysaccharides naturels, dans la préparation de microparticules. Etant donné ses propriétés intrinsèques limitantes, le remplacement du PGA par d'autres esters polysaccharidiques parait avantageux dans le domaine d'application des microparticules.Dans ce travail, le PGA a été remplacé par une série d'esters semi-synthétiques d'alginate puis par d'autres polysaccharides estérifiés naturels (pectines) ou semi-synthétiques (esters polypectiques et esters de l'acide hyaluronique). Les conditions optimales pour l'utilisation de chaque ester ont été alors déterminées. / In our laboratory, an original method of microencapsulation was developed, based on the use of a transacylation reaction, creating covalent bonds between proteins and propylene glycol alginate (PGA). The covalent bonds are created after alkalization of the aqueous phase of a W/O emulsion, without using bifunctional crosslinking reagent.The resulting microparticles, which are stable, biocompatible and biodegradable, have potential applications for the delivery of active compounds for therapeutics or cosmetics.The first aim of this work is to study the influence of the physicochemical properties of the two polymers (protein and PGA) and of their solutions, as well as the effect of the preparation parameters on the transacylation reaction and on microparticle characteristics. For this purpose, human serum albumin (HSA) was picked as a model protein and microparticles were prepared using several physicochemical conditions then characterized. Several relationships were established between the physicochemical properties of the initial solutions of the two polymers and the functional properties of the resulting microparticles.The second purpose is to replace the PGA, only polysaccharide used for microencapsulation by transacylation so far, by other natural polysaccharides in the preparation of microparticles. Given its limiting intrinsic properties, the replacement of PGA by other polysaccharidic esters seems advantageous in the field of microparticle applications.In this work, the PGA was successfully replaced by a series of semisynthetic alginate esters, and then by other polysaccharidic esters, either natural esters (pectin) or semisynthetic esters (polypectate esters and hyaluronate esters). The optimal conditions for the use of each ester were then determined.

Interaction between bovine serum albumin (BSA) and thimerosal (TM), organic mercury compound, was investigated by spectroscopic methods. The results, by molecular fluorescence, show that the interaction takes place by static quenching with electrostatic forces spontaneously (ΔG = - 4.40 kJ mol-1 at 30°C). The binding constant (Kb) was 3.24 ± 0.01x103 L mol-1 (30°C) is considered a moderate interaction. Fluorescence in three dimensions revealed that TM causes structural involving the the polypeptide chain BSA changes in the polarity of the tryptophan and tyrosine residues confirmed by circular dichroism (CD) showed an increase in α-helix content after interaction with TM. In addition, the TM decreases the surface hydrophobicity of the protein. Bilirubin was used as a marker for the subdomain IB, confirming that TM interacts in this region of the protein. The study of the interaction mechanism proposed that TM is reacted with BSA through the free cysteine residue, forming the adduct BSA-HgEt release of thiosalicylic acid (ATS), which interacts with amino acids with side chain positive. Besides, it was seen that TM accelerates the protein fibrillation kinetics by 42%, with a possible indication of the toxicity of this compound in biological systems. / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A interação entre albumina do soro bovino (BSA) e timerosal (TM), composto orgânico de mercúrio, foi investigada utilizando métodos espectroscópicos. Os resultados, por fluorescência molecular, evidenciam que a interação acontece por quenching estático através de forças eletrostáticas de forma espontânea (ΔG = - 4,40 kJ mol-1 a 30ºC). A constante de ligação (Kb) foi de 3,24 ± 0,01x103 L mol-1 (30ºC) sendo considerada uma interação moderada. A fluorescência em três dimensões revelou que TM causa mudanças estruturais envolvendo a cadeia polipeptídica da BSA assim como altera a polaridade dos resíduos de triptofano e tirosina, confirmada por dicroísmo circular (DC) que evidenciou aumento no conteúdo de α-hélice após interação com TM. Além disto, TM diminui a hidrofobicidade superficial da proteína. Bilirrubina foi utilizada como marcador para o subdomínio IB, confirmando que TM interage nesta região da proteína. O estudo do mecanismo de interação propôs que TM reage com BSA através do resíduo de cisteína livre, formando o aduto BSA-HgEt com liberação de ácido tiosalicílico (ATS), que interage com os aminoácidos com cadeia lateral positiva. Por fim, foi visto que TM acelera a cinética de fibrilação proteica em 42%, sendo um possível indício da toxicidade deste composto em sistemas biológicos.

A importância de toda a cadeia produtiva de aves no Brasil é fortemente impulsionada pela produção, exportação de carne de frango e pelo seu consumo per capita, que em 2009 foi de 11 milhões de toneladas, 3.3 milhões de toneladas e 40,1 Kg/hab/ano respectivamente. Porém a exportação de cortes e a preferência dos consumidores por cortes de frangos e filés ao invés dos frangos inteiros despertaram a necessidade de encontrar meios para o aproveitamento de dorsos, pescoços e ossos resultantes da desossa. A produção de carne mecanicamente separada com essas partes tornou-se ao longo dos anos uma alternativa viável, produzindo uma matéria-prima de baixo custo. O desenvolvimento de um produto reestruturado cozido congelado com carne de ave, com a utilização de pedaços desossados de peito e sobrecoxa, adicionados de carne mecanicamente separada do dorso da carcaça das aves tornou-se interessante para a indústria e consumidor. Porém as características desejadas pelo consumidor deste tipo de produto são consistência, força de adesão e suculência, ou seja, características de um filé de frango íntegro. Para conseguir essas características desejadas objetivou-se utilizar ingredientes não cárneos como a transglutaminase e albumina de ovo em pó, sendo esses ingredientes adicionais somados a adição de carne mecanicamente separada o estudo deste projeto. Para determinar a qualidade do produto foi realizada análise microbiológica no produto cru e após o cozimento e posteriormente para avaliar sua qualidade durante o armazenamento e sua vida útil a -18°C, foram realizadas análises como pH, cor instrumental, valor de TBARS e análise sensorial, sendo esta última para determinar aceitação do produto por consumidores. / The importance poultry production chain in Brazil is strongly stimulated by meat poultry exportation 11 million of tons, 3,3 million of tons and by per capita consumption 40,1 kg/hab/year. Although the preference of exportation cuts and the consumer preference by cuts instead of whole chicken so theres a search for the use of backs, necks, and bones from the boning. The mechanically separated meat production with that parts over the years one viable alternative, resulting in low cost raw material. Then the develop of restructured product frozen and cooked with poultry meat, the utilization of boning meat from chest and thighs , added to most from back bones is extremity interesting for industry and consumers. But the features desired by consumers for this product are the cohesiveness, richness and strength of accession, that means whole chicken characteristics. To achieve these desired features aimed to use non meat ingredients as transglutaminase and egg albumin powder were used, and these non meat ingredients added to the minced meat in the objective of this project. To determine the quality of the product, microbiological analyses in the raw product and after cooking were performed, and later, to evaluate its quality during storage and service life at -18°C, analyses such as pH, instrumental color, TBARS values and analysis sensory were performed, the latter being performed to determine the acceptance of the product by consumers.

This thesis is focused on synthesis of quantum dots (QDs) for protein detection. It comprises three parts. The first part summaries the theory of QDs, their synthesis, functionalization, interactions and applications in medicine. In the second part synthesis of CdTe/ZnS core/shell QDs modified by glutathione (GSH) is described, followed by the conjugation with biomolecules BSA and IgG. Several coupling agents such as EDC with NHS and CDI were used. In the last part, the final products were characterized by fluorescence spectroscopy and capillary electrophoresis. The results show the dependence of the fluorescence intensity of the QDs on pH range, concentration of BSA and IgG concentrations using different crosslinkers.

Dizertační práce se zabývá mechanismy mazání v náhradách kyčelního kloubu. Byla provedena systematická studie formování proteinového filmu při zahrnutí různých materiálů a provozních podmínek. Hlavní pozornost je přitom věnována roli jednotlivých proteinů obsažených v synoviální kapalině při současné přítomnosti dalších proteinů. Jelikož metody aplikované v předchozích studiích neumožňovaly separovat jednotlivé složky maziva, byla vyvinuta optická měřící metoda na principu fluorescenční mikroskopie. Z důvodu verifikace metody byly provedeny dvě nezávislé studie zaměřené na měření tloušťky mazacího filmu a dělení maziva na výstupu mazaného kontaktu. Z důvodu určitých limitací fluorescenční mikroskopie byla dále využita i metoda optické interferometrie, jejíž využití je ilustrováno ve studii zabývající se formováním mazacího filmu v náhradách kyčelního klubu při uvažování reálné konformity třecích povrchů. Závěrečná část práce představuje nový metodologický přístup založený na in situ pozorování kontaktní oblasti umožňující popsat roli jednotlivých proteinů ve vztahu k vývoji tloušťky mazacího filmu. Práce obsahuje originální výsledky, které přináší nové poznání v oblasti biotribologie náhrad kyčelního kloubu vedoucí k dalšímu vývoji implantátů při snaze zabránit jejich selhání v důsledku omezené životnosti.

Des biomolécules telles que les protéines et les acides nucléiques sont d'un grand intérêt dans l'immunothérapie, la thérapie génique tels que des cancers, le traitement de SIDA, les troubles auto-immuns liés à la déficience immunitaire sévère et combinés à autres maladies. Afin de cibler ces objectifs, l'encapsulation et le ciblage sont incontestablement nécessaires pour protéger les biomolécules contre la dégradation rapide et aussi pour éviter les effets secondaires. Ainsi, dans le cadre de cette thèse, l'encapsulation de protéines et de l'ADN en utilisant le procédé basé sur la double émulsion évaporation de solvant a été étudiée. Mais avant de conduire l’étude expérimentale, l'état de l'art des procédés d'encapsulation a été effectué et une attention particulière a été consacrée à l’encapsulation de l'ADN et les protéines et aux procédés d’encapsulations basés sur la formulation des polymères biodégradables. En ce qui concerne l’étude expérimentale, une étude systématique des paramètres pertinents sur les propriétés physico-chimique et colloïdales des particules a été examinée. Cette étude a permis de mettre en évidence les facteurs clés capables d’affecter non seulement le procédé mais aussi les propriétés colloïdales des particules élaborées (taille, distribution en taille, propriétés électrocinétiques et stabilité colloïdale…). En second lieu, la formulation optimisée permettant l’obtention de tailles submicroniques a été utilisée pour l’encapsulation de l'ADN et les protéines. L'encapsulation de l'ADN a été étudiée en fonction de la concentration de l'ADN (sperme de saumon) et une encapsulation totale d'ADN a été observée sans modification ou altération des propriétés colloïdales des particules finales. L'encapsulation de protéines a été également examinée en utilisant HAS et une autre protéine fluorescente comme modèles. L'efficacité de l'encapsulation a été déterminée et trouvée de l’ordre de 100%. L'utilisation de la protéine fluorescente indique l'encapsulation totale avec une bonne répartition dans les cavités de la matrice polymère. En conclusion, le procédé de la double émulsion – évaporation de solvant est efficace pour l’encapsulation de l'ADN et des protéines sans modification des propriétés colloïdales des particules finales qui restent bien submicroniques / Biomolecules such as proteins and nucleic acids are of great interest in immunotherapy and gene therapy such as cancers, AIDS, autoimmune disorders, X-linked severe combined immune deficiency and many other diseases. In order to target such objectives, the encapsulation and the targeting are incontestably necessary principally to protect biomolecules against rapid degradation and also to avoid side effects. Then, in this thesis, the encapsulation of proteins and DNA using double emulsion solvent evaporation technique was performed. But before the encapsulation, the state of the art of the encapsulation processes was performed and special attention has been dedicated to DNA and proteins encapsulation. Firstly, systematic study of double emulsion solvent evaporation process was investigated in order to point out the key parameters controlling the particles size and the colloidal stability. Secondly, the optimized formulation conditions leading to submicron colloidal was used for DNA and Proteins encapsulation. The DNA encapsulation was investigated as a function of DNA concentration and it was found total encapsulation of DNA without any modification of the colloidal properties. The protein encapsulation was examined using HAS and fluorescent proteins as model. The encapsulation efficiency was found to be high and reaches 100%. The use of fluorescent protein shows the total encapsulation and good distribution of proteins in polymer matrix. As a general conclusion, the high encapsulation efficiency of DNA and protein, reveled the compatibility of the used process to encapsulated hydrophilic biomolecules even in submicron size biodegradable colloidal submicron particles

The impact was assessed of clinical and nutritional factors on prognosis of 107 oesophageal cancer patients treated with neoadjuvant chemoradiotherapy (CHRT) and surgery. Individualised nutritional support, according to grade of dysphagia was carried out in all the patients. Serum leptin, soluble leptin receptors (SLR), TNF, IGF and fatty acid (FA) profiles in plasma phosphatidylcholine (PC) were studied as well. Addition of paclitaxel to carboplatin and continual fluorouracil significantly increased toxicity without influencing efficacy of the treatment. Post-operative node negativity, grade of dysphagia, weight loss and serum albumin were proved to be prognostic factors of survival and time to progression. CHRT led to decrease of SLR, palmitoleic and oleic acid and increase of n-3 polyunsaturated FA in PC. Lower concentrations of SLR were associated with improved survival of the patients. Key words: oesophageal cancer, neoadjuvant chemoradiotherapy, weight loss, paclitaxel, albumin, soluble leptin receptor, fatty acids

Development of methods for studying drug delivery systems is of great significance for the improvement of topical formulations. Active compounds for topical drug delivery are often formulated into gels and creams, that can be applied onto skin surfaces. It is important to know the extent of the permeability of the active compounds, in order to determine the medical effect. This study examines the possibilities of using an animal-free skin equivalent for penetration and permeation experiments, i.e. a silk scaffold integrated with viable human dermaland epidermal cells. Mammalian cell culturing together with silkconstruct formation, constituted the upstream bioprocess and acquisition of the skin equivalents. Permeability of fluorescently labelled Bovine Serum Albumin and Sodium Fluorescein salt was assessed, using a Franz- cell setup incorporated with the skin equivalents. Furthermore, fluorescence analysis and SDS-PAGE was performed on the collected samples, along with cryosectioning and image analysis of the skin equivalents. The results indicate variations in tissue integrity, leading to both high and low permeability. Fluorescence intensity can be correlated with the amount of sample liquid passing through. The model is still under development, hence more research is needed to draw a conclusion regarding the cellular composition of the skin equivalents, and how it influences permeability. / NextBioForm