The development of bio-resorbable implant materials is rapidly going on. Sterilization of those materials is inevitable to assure the hygienic requirements for critical medical devices according to the medical device directive (MDD, 93/42/EG). Biopolymer-containing biomaterials are often highly sensitive towards classical sterilization procedures like steam, ethylene oxide treatment or gamma irradiation. Supercritical CO2 (scCO2) treatment is a promising strategy for the terminal sterilization of sensitive biomaterials at low temperature. In combination with low amounts of additives scCO2 treatment effectively inactivates microorganisms including bacterial spores. We established a scCO2 sterilization procedure under addition of 0.25% water, 0.15% hydrogen peroxide and 0.5% acetic anhydride. The procedure was successfully tested for the inactivation of a wide panel of microorganisms including endospores of different bacterial species, vegetative cells of gram positive and negative bacteria including mycobacteria, fungi including yeast, and bacteriophages. For robust testing of the sterilization effect with regard to later application of implant materials sterilization all microorganisms were embedded in alginate/agarose cylinders that were used as Process Challenge Devices (PCD). These PCD served as surrogate models for bioresorbable 3D scaffolds. Furthermore, the impact of scCO2 sterilization on mechanical properties of polysaccharide-based hydrogels and collagen-based scaffolds was analyzed. The procedure was shown to be less compromising on mechanical and rheological properties compared to established low-temperature sterilization methods like gamma irradiation and ethylene oxide exposure as well as conventional steam sterilization. Cytocompatibility of alginate gels and scaffolds from mineralized collagen was compared after sterilization with ethylene oxide, gamma irradiation, steam sterilization and scCO2 treatment. Human mesenchymal stem cell viability and proliferation were not compromised by scCO2 treatment of these materials and scaffolds. We conclude that scCO2 sterilization under addition of water, hydrogen peroxide and acetic anhydride is a very effective, gentle, non-cytotoxic and thus a promising alternative sterilization method especially for biomaterials.

CAPES; CNPq; Fundação Araucária / Este trabalho apresenta um estudo sobre o efeito do agente compatibilizante de anidrido maleico (PE-g-MAH) nas propriedades da blenda polimérica híbrida contendo o composto low smoke zero halogen e polietileno de alta densidade (LSZH/HDPE), visando a obtenção de um material adequado para uso em cruzetas separadoras de cabos de telecomunicações. Os requisitos exigidos a esta função são capacidade de isolamento elétrico, para evitar que os campos magnéticos gerados pelos pares condutores causem interferência na transmissão de dados dos pares adjacentes, e resistência à flamabilidade, devido à crescente preocupação sobre os riscos à saúde e ambientais no caso de incêndios. Os compostos LSZH são utilizados na fabricação de cabos por serem materiais resistentes à chama e livres de halogênios, porém não detém capacidade de isolamento elétrico. O HDPE é um material dielétrico, mas não possui a resistência à flamabilidade. O PE-g-MAH é um agente compatibilizante que melhora a resistência a flamabilidade de blendas poliméricas híbridas pelo aumento da dispersão das cargas antichama. A blenda LSZH/HDPE foi analisada com a adição de diferentes proporções de compatibilizante com o objetivo de se obter a proporção que apresenta propriedades mais adequadas à aplicação em cruzetas separadoras. Foram fabricadas amostras contendo 75% de LSZH-A (índice de oxigênio 33 %) com aplicação de 8%, 10% e 12% de PE-g-MAH, e amostras contendo 75% do LSZH-B (índice de oxigênio 37 %) com a aplicação de 10% e 12 % de PE-g-MAH em blendas. Observou-se que a adição do compatibilizante aumenta a dispersão das cargas antichama, melhora a resistência a flamabilidade e diminui a capacidade de isolamento elétrico da blenda. A proporção 75/13/12 LSZH-B/HDPE/PE-g-MAH apresentou as propriedades mais apropriada para emprego em cruzetas separadoras, com índice de oxigênio de 33,6%, constante dielétrica em 1 MHz de 3,06, resistividade volumétrica de 3,05.10 15 Ohm.cm, resistência a tração de 14,47 MPa e alongamento a ruptura de 412 %. / This paper presents a study on the effect of the compatibilizing PE-g-MAH on the properties of the hybrid polymer blends of low smoke zero halogen and high density polyethylene LSZH / HDPE, aiming at obtaining a suitable material for use in cross web of telecommunication cables. The requirements for this function are electrical insulation capability, to avoid electromagnetic interference between the conductor pair’s constituent of the cable, and resistance to flammability, due to the growing concern about the health and environmental risks in the event of fires. LSZH compounds are used in the manufacture of cables because they are flame-resistant and halogen-free materials, but have no electrical insulation capacity. HDPE is a dielectric material, but it lacks the flammability resistance. PE-g-MAH is a compatibilizing that improves the flammability resistance of hybrid polymer blends by increasing the dispersion of the flame retardant loads. The LSZH / HDPE blend was analyzed with the addition of different proportions of PE-g-MAH in order to obtain the proportion that presents properties more suitable for the application in cross web. Samples containing 8%, 10% and 12% PE-g-MAH were prepared in blends containing 75% LSZH-A (LOI 33 %) and 10% and 12% PE-g-MAH samples in blends containing 75% of LSZH- B (LOI 37 %). It was observed that the addition of PE-g-MAH increases the dispersion of the flame-retardants, improves the flammability resistance and decreases the electrical insulation capacity of the blender. The ratio 75/13/12 LSZH-B / HDPE / PE-g-MAH presented the most suitable properties for use in cross web, with oxygen index of 33.6%, dielectric constant at 1 MHz of 3.06, resistivity 3.05.1015 Ohm.cm, tensile strength of 14.47 MPa and elongation at break of 412%.

CAPES; CNPq; Fundação Araucária / Este trabalho apresenta um estudo sobre o efeito do agente compatibilizante de anidrido maleico (PE-g-MAH) nas propriedades da blenda polimérica híbrida contendo o composto low smoke zero halogen e polietileno de alta densidade (LSZH/HDPE), visando a obtenção de um material adequado para uso em cruzetas separadoras de cabos de telecomunicações. Os requisitos exigidos a esta função são capacidade de isolamento elétrico, para evitar que os campos magnéticos gerados pelos pares condutores causem interferência na transmissão de dados dos pares adjacentes, e resistência à flamabilidade, devido à crescente preocupação sobre os riscos à saúde e ambientais no caso de incêndios. Os compostos LSZH são utilizados na fabricação de cabos por serem materiais resistentes à chama e livres de halogênios, porém não detém capacidade de isolamento elétrico. O HDPE é um material dielétrico, mas não possui a resistência à flamabilidade. O PE-g-MAH é um agente compatibilizante que melhora a resistência a flamabilidade de blendas poliméricas híbridas pelo aumento da dispersão das cargas antichama. A blenda LSZH/HDPE foi analisada com a adição de diferentes proporções de compatibilizante com o objetivo de se obter a proporção que apresenta propriedades mais adequadas à aplicação em cruzetas separadoras. Foram fabricadas amostras contendo 75% de LSZH-A (índice de oxigênio 33 %) com aplicação de 8%, 10% e 12% de PE-g-MAH, e amostras contendo 75% do LSZH-B (índice de oxigênio 37 %) com a aplicação de 10% e 12 % de PE-g-MAH em blendas. Observou-se que a adição do compatibilizante aumenta a dispersão das cargas antichama, melhora a resistência a flamabilidade e diminui a capacidade de isolamento elétrico da blenda. A proporção 75/13/12 LSZH-B/HDPE/PE-g-MAH apresentou as propriedades mais apropriada para emprego em cruzetas separadoras, com índice de oxigênio de 33,6%, constante dielétrica em 1 MHz de 3,06, resistividade volumétrica de 3,05.10 15 Ohm.cm, resistência a tração de 14,47 MPa e alongamento a ruptura de 412 %. / This paper presents a study on the effect of the compatibilizing PE-g-MAH on the properties of the hybrid polymer blends of low smoke zero halogen and high density polyethylene LSZH / HDPE, aiming at obtaining a suitable material for use in cross web of telecommunication cables. The requirements for this function are electrical insulation capability, to avoid electromagnetic interference between the conductor pair’s constituent of the cable, and resistance to flammability, due to the growing concern about the health and environmental risks in the event of fires. LSZH compounds are used in the manufacture of cables because they are flame-resistant and halogen-free materials, but have no electrical insulation capacity. HDPE is a dielectric material, but it lacks the flammability resistance. PE-g-MAH is a compatibilizing that improves the flammability resistance of hybrid polymer blends by increasing the dispersion of the flame retardant loads. The LSZH / HDPE blend was analyzed with the addition of different proportions of PE-g-MAH in order to obtain the proportion that presents properties more suitable for the application in cross web. Samples containing 8%, 10% and 12% PE-g-MAH were prepared in blends containing 75% LSZH-A (LOI 33 %) and 10% and 12% PE-g-MAH samples in blends containing 75% of LSZH- B (LOI 37 %). It was observed that the addition of PE-g-MAH increases the dispersion of the flame-retardants, improves the flammability resistance and decreases the electrical insulation capacity of the blender. The ratio 75/13/12 LSZH-B / HDPE / PE-g-MAH presented the most suitable properties for use in cross web, with oxygen index of 33.6%, dielectric constant at 1 MHz of 3.06, resistivity 3.05.1015 Ohm.cm, tensile strength of 14.47 MPa and elongation at break of 412%.

L’étude de la structure de l’ARN est reconnue un sujet d’actualité qui permet de connaître sa fonction. Il existe différentes techniques enzymatiques ou chimiques classiques et récentes couplées au séquençage à haut débit pour étudier la structure de l’ARN. Le projet de thèse vise l’optimisation de deux techniques de cartographie déjà existantes et se divise en deux parties. La première partie concerne l’optimisation de la technique de hSHAPE et ceci en synthétisant de nouvelles sondes fluorescentes à transfert d’énergie capables d’étudier de faibles quantités d’ARN et adaptées au séquenceur d’ADN. Mon travail a permis la synthèse de 5 molécules fluorescentes intermédiaires et l’obtention d’une sonde finale. L’étude de cette sonde a montrée que l’interaction entre les deux fluorophores est majoritairement due à un mécanisme de couplage excitonique causé par la flexibilité et la distance entre les deux entités fluorescentes. De plus, les propriétés de la molécule sont fortement dépendantes du solvant. Dans la deuxième partie, nous cherchons à optimiser la cartographie en solution afin d’étudier le génome entier du VIH-1 in virio. Pour cela, nous avons synthétisé 4 molécules dérivées de l’anhydride isatoïque et modifiées sur la fonction amine N1 par un groupement propyn-2-yle. Nous avons aussi préparé 8 molécules de référence. De plus, des études préliminaires réalisées sur l’ARN 1-311 du VIH-1 ont montré que nos composés modifient l’ARN et qu’il est possible de fixer la biotine-PEG3-azoture par click chemistry sur les ARN ainsi modifiés. / Understanding the function of RNA involved in biological processes requires a thorough knowledge of RNA structure. Traditional chemical and enzymatic reagents and backbonebased cleavage are useful for mapping RNA secondary structure, and on going advances in nucleotide resolution RNA structure probing have made possible increasingly rigorous and quantitative analysis. Although chemical and enzymatic probes are recently coupled to high throughput sequencing, these techniques still suffer from some disadvantages. This project aims the optimization of two existing techniques and is divided into two major topics. The first part aims the optimization of “hSHAPE chemistry”. A practical method is the synthesis of energy transfer dyes from the “BigDyes” family that are useful to study small amount of RNA and are compatible with DNA sequencing. We have done the synthesis of 5 modified dyes and one BigDye. The characterization of this BigDye shows that it exists excitoncoupling mechanism due to the strong interaction between the two transition dipoles. In addition, the solvent influences strongly the photophysical properties of this BigDye. The purpose of the second part is to develop and synthesize new class of isatoic anhydride derivatives useful to map the entire genome of HIV-1 in virio. We have done the synthesis of 4 N-substituted molecules with a propyn-2-yl function and 8 reference molecules. In addition, preliminary results on HIV-1 1-311 RNA showed that these compounds are able to modify RNA and that it is possible to fix a biotin-azide by click chemistry.

Bisacetylenic cabamates/carbonates are most useful compounds in finger mark development, for the synthesis of polymeric gels and other material applications. Organochalogen derivatives are the organic compounds containing chalcogen (S, Se) atoms. They have been used as chiral ligands for enatioselective catalysis, glycosyl donors and in the synthesis of heterocyclic compounds etc. This thesis describes our efforts towards synthesis of bisacetylenic cabamates/carbonates and development new synthetic strategies using rongalite (Na+HOCH2SO2-) and benzyltriethyl ammonium tetrathiomolybate [BnEt3N]2MoS4 as a reducing agents led to obtain various organochalcogen derivatives. We developed a new reagent, hexa-2,4-diyne-1,6-bisoxycarbonyl chloride [Hbc Cl] for the synthesis of symmetrical diacetylenic biscarbamates/biscarbonates and further studied the solid state structures using X-ray crystallography. Later we described a stereoselective method for the hydrothiolation of buta-1,3-diynes derivatives using diaryldichalcogenides in the presence of rongalite and K2CO3. The buta-1,3-diynes underwent stereoselective addition reaction with in situ generated chalocgenate anion from diaryl dichalcogenides which afforded the corresponding (Z)-chalcogenynes. The reactivity of buta-1,3-diynes with diaryl dichalcogenides was further studied at higher temperature led to a mixture of mono chalcogenated and bischalcogenated products. Then an efficient method was developed for the synthesis of enatiopure β-amino sulfides/selenides via ring opening of sulfamidates using diarylchalcogenides with rongalite as reducing agent. Further we synthesized chalcogeno derivatives of sugars from glycosyl halides and diaryl dichalcogenides in the presence rongalite. In addition, the synthesis of mixed glycosyl dichalcogenides has been demonstrated using [BnEt3N]2MoS4 as sulfur transfer agent as well as reducing agent. Finally the reactivity of [BnEt3N]2MoS4 was studied in detail with various isatioc anhydrides which led to the formation of S-benzyl 2-aminobenzothioate derivatives. Further we synthesized S-alkyl/aryl 2-aminobenzothioate derivatives via ring opening of isatoic anhydrides and diaryl/dialkyl chalcogenides by mean of [BnEt3N]2MoS4 as a reducing agent. We extended this method in a one-pot, tandem fashion with various alkyl halides. In this thesis, details of all of the above studies have been described.

The development of bio-resorbable implant materials is rapidly going on. Sterilization of those materials is inevitable to assure the hygienic requirements for critical medical devices according to the medical device directive (MDD, 93/42/EG). Biopolymer-containing biomaterials are often highly sensitive towards classical sterilization procedures like steam, ethylene oxide treatment or gamma irradiation. Supercritical CO2 (scCO2) treatment is a promising strategy for the terminal sterilization of sensitive biomaterials at low temperature. In combination with low amounts of additives scCO2 treatment effectively inactivates microorganisms including bacterial spores. We established a scCO2 sterilization procedure under addition of 0.25% water, 0.15% hydrogen peroxide and 0.5% acetic anhydride. The procedure was successfully tested for the inactivation of a wide panel of microorganisms including endospores of different bacterial species, vegetative cells of gram positive and negative bacteria including mycobacteria, fungi including yeast, and bacteriophages. For robust testing of the sterilization effect with regard to later application of implant materials sterilization all microorganisms were embedded in alginate/agarose cylinders that were used as Process Challenge Devices (PCD). These PCD served as surrogate models for bioresorbable 3D scaffolds. Furthermore, the impact of scCO2 sterilization on mechanical properties of polysaccharide-based hydrogels and collagen-based scaffolds was analyzed. The procedure was shown to be less compromising on mechanical and rheological properties compared to established low-temperature sterilization methods like gamma irradiation and ethylene oxide exposure as well as conventional steam sterilization. Cytocompatibility of alginate gels and scaffolds from mineralized collagen was compared after sterilization with ethylene oxide, gamma irradiation, steam sterilization and scCO2 treatment. Human mesenchymal stem cell viability and proliferation were not compromised by scCO2 treatment of these materials and scaffolds. We conclude that scCO2 sterilization under addition of water, hydrogen peroxide and acetic anhydride is a very effective, gentle, non-cytotoxic and thus a promising alternative sterilization method especially for biomaterials.

In this work the organic markers that serve for identification of sources of aerosols are monitored. Theoretic part deals with detailed survey of organic markers emitted from the most significant sources of atmospheric aerosols (biomass combustion, combustion of fossil fuels and traffic) as well as with sampling of aerosols and analysis of organic markers by analytical techniques. Monosaccharide anhydrides (emissions from biomass combustion) and polyaromatic hydrocarbons (emissions from traffic and incomplete combustion) were observed especially. Hopanes and steranes (traffic, coal combustion) and fatty acids (cooking) were next groups of monitored organic markers. Markers were studied mainly in size fraction PM1 because this fraction of aerosols is the most harmful to human health. Aerosols were sampled in two seasons (winter and summer) in two towns in 2009. Higher concentrations of aerosols and organic compounds were found in winter season, which resulted from increased combustion of biomass, coal and other organic material while traffic was the most significant source of aerosols in summer.

Diploma thesis deals with the analysis of saccharides and markers of biomass and wood combustion in atmospheric aerosol, PM2,5. Theoretic part is focused on general characterisation of atmospheric aerosols and on the properties, sources and analysis methods of selected analysed organic compounds. The experimental part is focused on the optimization of method for simultaneous analysis of saccharides and tracers from biomass and wood combustion. Optimization of method includes selection of solvent for the extraction of compounds and optimization of derivatization process and GC/MS analysis. The optimized method was then applied for the analysis of selected compounds in real aerosol samples in the size fraction PM2.5. The concentrations of analysed compounds were compared in term of sampling seasons.

L'étude de la structure de l'ARN est reconnue un sujet d'actualité qui permet de connaître sa fonction. Il existe différentes techniques enzymatiques ou chimiques classiques et récentes couplées au séquençage à haut débit pour étudier la structure de l'ARN. Le projet de thèse vise l'optimisation de deux techniques de cartographie déjà existantes et se divise en deux parties. La première partie concerne l'optimisation de la technique de hSHAPE et ceci en synthétisant de nouvelles sondes fluorescentes à transfert d'énergie capables d'étudier de faibles quantités d'ARN et adaptées au séquenceur d'ADN. Mon travail a permis la synthèse de 5 molécules fluorescentes intermédiaires et l'obtention d'une sonde finale. L'étude de cette sonde a montrée que l'interaction entre les deux fluorophores est majoritairement due à un mécanisme de couplage excitonique causé par la flexibilité et la distance entre les deux entités fluorescentes. De plus, les propriétés de la molécule sont fortement dépendantes du solvant. Dans la deuxième partie, nous cherchons à optimiser la cartographie en solution afin d'étudier le génome entier du VIH-1 in virio. Pour cela, nous avons synthétisé 4 molécules dérivées de l'anhydride isatoïque et modifiées sur la fonction amine N1 par un groupement propyn-2-yle. Nous avons aussi préparé 8 molécules de référence. De plus, des études préliminaires réalisées sur l'ARN 1-311 du VIH-1 ont montré que nos composés modifient l'ARN et qu'il est possible de fixer la biotine-PEG3-azoture par click chemistry sur les ARN ainsi modifiés.

No / A systematic identification of the degradation products of hydroxypropyl methylcellulose phthalate (HPMCP) during hot melt extrusion (HME) has been performed. A reverse phase HPLC method was developed for the extrudates of both hydroxypropyl methylcellulose acetate succinate (HPMCAS) and HPMCP polymers to quantify their thermal hydrolytic products: acetic acid (AA), succinic acid (SA) for HPMCAS and phthalic acid (PA) for HPMCP, without hydrolysing the polymers in strong alkaline solutions. The polymers were extruded in the temperature range of 160-190 degrees C at different screw rotation speeds and hydrolytic impurities were analysed. Investigation of extruded HPMCP showed an additional thermal degradation product, who is structural elucidation revealed to be phthalic anhydride (PAH). Moreover, two environmental analytical impurities, dimethyl phthalate and methyl benzoate formed in situ were recorded on GC-MS and their origin was found to be associated with PAH derivatization. Using the experimental data gathered during this study, a degradation mechanism for HPMCP is proposed.