This thesis contains investigations in two different areas, described under six chapters. Chapter 1 contains a broad introduction to the area of foldamers, while Chapters 2, 3, 4, and 5 deal with various novel classes of synthetic polymers which can form folded structures in solution utilizing different non-covalent interactions. Chapter 6 deals with a distinctly different topic, where the objective was to study the effect of phenyl ring location on the micellization properties of a series of isomeric cationic surfactants. Synthetic polymers typically adopt a random coil conformation in solution, which is primarily an entropy driven process. So the generation of well-defined secondary structures in synthetic polymers requires specific intra-chain inter-segment interactions that will give adequate enthalpic contribution to overcome the entropic penalty associated with the formation of well-ordered conformations. During the past decade, various research groups have made significant effort to understand the essential design elements that could enable secondary structure formation in synthetic macromolecules through intra-chain inter-segment interactions, such as hydrogen bonding, solvophobic and solvophilic interaction, acid-base interaction, bond angle constraint, steric interaction, charge-transfer interaction, metal-ion complexation etc.1 Gellman2 first used the term “foldamer” to describe “any polymer with a strong tendency to adopt a specific compact conformation” which was more precisely defined by Moore and coworkers3 as “any oligomer that folds into a conformationally ordered state in solution, the structures of which are stabilized by a collection of non-covalent interactions between nonadjacent monomer units” and where the folded conformation is one of the various possible conformations. Several classes of foldamers have been studied during the past decade; a majority of them are well-defined oligomers that possess relatively restricted conformational degrees of freedom. Relatively fewer studies have explored conformational control in flexible high molecular weight polymers that possess greater conformational freedom.4 A few years ago, Ghosh et al. designed a polymeric system wherein charge-transfer interactions between alternatively placed electron-rich and electron-deficient aromatic units, aided by metal-ion complexation and solvophobic interactions, causes the polymer chain to adopt a specific folded conformation.5 Such charge-transfer induced folding was first studied by Iverson and co-workers6 in well-defined oligomers and was later elaborated by Zhao et al.7 to generate alternate designs to fold oligomeric systems. In all these studies, the C-T interactions served not only to assist the folding process but it also served as a valuable spectroscopic signature to study the folding process. The objectives of the present study are to develop simple synthetic strategies to generate different types of polymers that could be fold in solution using various noncovalent interactions. We have developed a simple synthetic strategy to design a new type of donor (1,5-dialkoxynaphthalene-DAN) containing polymer that carries a tertiary amine unit in the spacer segment, which could interact strongly with a suitably designed acceptor (pyromellitic diimide-PDI) bearing folding agent that carries a carboxylic acid group, as shown in Scheme 1.8 This acid-base interaction, brings the acceptor unit in a suitable position so as to form a C-T complex with the adjacent donors, resulting in the folding of the polymer chain. The folded conformation was studied using UV-vis and NMR spectroscopy and the folding propensities were rationalized using DFT studies. The highest association constant between the folding agent and the polymer was estimated to be around 1200 M-1. Scheme 1. Schematic representation of folding aided by two-point interactions with a folding agent. This value of association constant was not adequate to realize some potentially interesting properties in solid state. In an attempt to develop alternate systems, that could exhibit stronger propensity to fold, we designed a new type of cationic ionene,9 wherein electron-rich (DAN) and electron-deficient (PDI) aromatic units were included within the alkylene segments in an alternating fashion, as shown in Scheme 2.10 The charge-transfer (C-T) interaction between the donor and acceptor units in neighbouring segments of the ionene not only reinforced the transition to the collapsed nano-bundle form but also provides a useful spectroscopic handle to monitor the conformational change. The UV-visible spectra of these novel D-A ionene solutions at a fixed concentration in four different solvents, namely water, methanol, acetonitrile and DMSO, show different extents of charge-transfer interaction. The colour of the solution in water was deep-red, whereas in acetonitrile, it was light-yellow. The conformational transition could also be induced by titrating an acetonitrile solution of the ionene with increasing amounts of water causing a dramatic increase in the intensity of the charge-transfer band, which reflects the extent of collapse to the zig-zag state that brings the donor and acceptor units together. AFM studies confirmed the presence of flat pancake-like aggregates having nearly constant height of about 3-5 nm, which was in accordance with the estimated thickness of the postulated zig-zag structure. Scheme 2. Schematic depiction of folding of D-A ionene (left), AFM micrograph showing pancake-like aggregates of D-A ionenes (right-top), a line scan depicting the heights and diameters of the aggregates along with a schematic depiction of the aggregate (right-bottom). Scheme 3. Schematic representation of folding aided by interactions with a folding agent. In order to explore this concept further, we designed a two component system wherein the solvophobically-driven collapse of a DAN-containing ionene chain in a polar solvent is reinforced by intercalation with a suitably designed electron-deficient acceptor-containing external folding agent. DAN containing ionene polymer chains in polar solvent form an accordion-type zig-zag structure that brings adjacent donor units in close proximity; this provided an ideal hydrophobic pocket for intercalation of suitably designed electron-deficient acceptor molecules, the additional driving motivation for the intercalation being the formation of a C-T complex as shown in Scheme 3.11 Several acceptor-bearing molecules were prepared by the derivatization of pyromellitic dianhydride and naphthalene tetracarboxylic dianhydride with two different oligoethylene glycol monomethyl ether monoamines. UV-vis spectroscopic studies were carried out by using a 1:1 mixture of the DAN-ionenes and different acceptor molecules in water/DMSO solvent mixtures. The intensity of the charge-transfer (C-T) band was seen to increase with the water content in the solvent mixture, thereby suggesting that the intercalation is indeed aided by solvophobic effects. The naphthalene diimide (NDI) bearing acceptor molecules consistently formed significantly stronger C-T complexes when compared to the pyromellitic diimide (PDI) bearing acceptor molecules, which is a reflection of the stronger π-stacking tendency of the former. The highest association constant between the folding agent and the polymer was estimated to be around 4519 M-1, which was a substantial improvement over the earlier reported values.9 With a slight modification in the pendant group, we designed a water-soluble DAN-containing ionene, which can intercalate hydrophobic electron-deficient molecules, like TNT (2,4,6-trinitrotoluene), within the hydrophobic interstices between DAN units (as shown in Scheme 4), causing a depletion in fluorescence from the DAN units; TNT at concentration as low as 30 nM could be detected in this manner. Scheme 4. Schematic representation of folding of water soluble ionene and interactions with an electron-deficient hydrophobic moiety TNT. Scheme 5. Schematic representation of folded D-A allyl ionene. In light of the growing interest in single-chain polymeric nanoparticles, the fully collapsed D-A ionenes in water could be viewed as polymeric nanoparticles that are stitched together by reversible weak noncovalent interactions. In an attempt to transform the folded structure into a polymeric nanoparticle using covalent bonding, we designed D-A ionene that carries potentially polymerizable allyl units on the cationic head group instead of the dimethyl amine head group that was used in previous examples (as shown in Scheme 5). Preliminary studies showed that polymerization does not proceed readily; however, thiol-ene based clicking strategy enabled partial stitching of the folded segments, by the use of a suitably designed dithiol. In the last section of this thesis, we examined the effect of phenyl ring location on the micellization properties of a series of isomeric cationic surfactants, wherein the phenyl ring location was varied from head to tail region (as shown in Scheme 6).12 Thus, cationic surfactants (S1-S5) bearing a long alkyl chain that carries a 1,4phenylene unit and a trimethyl ammonium headgroup was synthesized and their solution properties were examined. Micellization behavior was studied using conductivity, ITC (Isothermal Titration Calorimetry), SANS (Small-Angle Neutron Scattering) and NMR. These present studies demonstrated that the presence of a large rigid ring near the hydrocarbon tail-end of the surfactant leads to a dramatic change in the micelle structure; the driving motivation to form micelles in such systems is greatly reduced and the micelles that are formed are relatively smaller and contain significantly fewer surfactants. NMR studies of micellar solutions of these surfactants indicate that the variation in the phenyl ring location may also help to probe the microenvironment at various depths within the micellar aggregates. Scheme 6. Structures of the various surfactant molecules carrying the 1,4-dioxyphenylene unit at different locations within hydrophobic segment (left), variation of CMC values (right). References (1) Foldamers - structure, properties, and applications, edited by Stefan Hecht and Ivan Huc, Wiley-VCH, 2007. (2) Gellman, S. H. Acc. Chem. Res. 1998, 31, 173. (3) Hill, D. J.; Mio, M. J.; Prince, R. B.; Huges, T. S.; Moore, J. S. Chem. Rev. 2001, 101, 3893. (4) (a) Wang, W.; Li, L. S.; Helms, G.; Zhou, H. H.; Li, A. D. Q. J. Am. Chem. Soc. 2003, 125, 1120. (b) Li, A. D. Q.; Wang, W.; Wang, L. Q. Chem. Eur. J. 2003, 9, 4594. (c) Neuteboom, E. E.; Meskers, S. C. J.; Meijer, E. W.; Janssen, R. A. J. Macromol. Chem. Phys. 2004, 205, 217. (d) Balbo Block, M. A.; Hecht, S. Macromolecules 2004, 37, 4761. (5) (a) Ghosh, S.; Ramakrishnan, S. Angew. Chem. Int. Ed. 2004, 43, 3264. (b) Ghosh, S.; Ramakrishnan, S. Angew. Chem. Int. Ed. 2005, 44, 5441. (6) Lokey, R. S.; Iverson, B. L. Nature 1995, 375, 303. (7) Zhao, X.; Jia, M. X. Jiang, X. K.; Wu, L. Z.; Li, Z. T.; Chen. G. J. J. Org. Chem. 2004, 69, 270. (8) De, S.; Koley, D.; Ramakrishnan, S. Macromolecules 2010, 43, 3183. (9) Williams, S. R.; Long, T. E. Prog. Polym. Sci. 2009, 34, 762. (10) De, S.; Ramakrishnan, S. Macromolecules 2009, 42, 8599. (11) De, S.; Ramakrishnan, S. Chem. Asian J. 2011, 6, 149. (12) De, S.; Aswal, V. K.; Ramakrishnan, S. Langmuir 2010, 26, 17882. (For structural formula pl see the abstract file.

Tendo em vista que os amidos (catiônicos ou não) não são nocivos a saúde do ser humano, uma vez que são utilizados na indústria de alimentos, e observando-se a potencialidade do seu uso como auxiliares de floculação, foi proposto o presente trabalho. Consiste na realização de ensaios de coagulação (com sulfato alumínio), floculação e sedimentação em equipamento de reatores estáticos, objetivando verificar a influência do gradiente de velocidade e do tempo de agitação na floculação com polímero sintético catiônico e amidos de milho e mandioca catiônicos. Concluiu-se que as condições de aplicação exercem influência na eficiência de remoção de turbidez e cor aparente, sendo que cada polímero estudado apresentou uma condição ótima específica. Em todos os ensaios realizados, o amido de mandioca catiônico foi o mais eficiente, indicando que os amidos catiônicos podem ser substitutos em potencial dos polímeros sintéticos no tratamento de águas de abastecimento, quando utilizados como auxiliares de floculação. / This present work was based on the fact that native and cationic starches are not harmful to man\'s health, since they have been largely used in food processing industries, and that they may be potentially used as flocculation aids in water treatment. Jar Test assays were performed, including coagulation with aluminum sulphate, flocculation and sedimentation, aiming to study the influence of the velocity gradient and mixing time on flocculation, using a cationic synthetic polymer and cationic com and manioc starches as aids. In the main, it was concluded that the conditions under which the polymers were applied, affect the efficiency of turbidity and color removals and that each polymer studied presented an specific optimum condition. It was also observed that manioc cationic starch resulted more effective than the others polymers studied, when used as flocculation aids.

<p>Denna avhandling presenterar experimentella studier av polyelektrolytadsorption på motsatt laddade ytor, där substrat av både kiseloxid och blekt barrsulfatmassa har använts. Ett huvudsakligt syfte med denna forskning var att karaktärisera konformationen hos adsorberade skikt av katjonisk polyakrylamid (CPAM) i jämförelse med katjonisk dextran (Cdextran) och relatera denna information till inbindningskapaciteten av kolloidal kiselsyra. Ett andra syfte i denna avhandling var att studera kinetiken för sekventiell adsorption av polyamidamin epiklorhydrin (PAE) och karboxymetyl cellulosa (CMC) på massafibrer och att bestämma adsorptionsisotermer för deponering av polyelektrolyter skikt för skikt på massafibrer.</p><p>Adsorptionen av CPAM på kiseloxidytor studeras med stagnationspunkts-reflektometri och kvartskristalls-mikrogravimetri för att bestämma adsorptionskinetiken och mättnadsadsorptionens beroende av polyelektrolytens laddningstäthet, pH och NaCl koncentration. Konformationen hos adsorberade skikt av CPAM och Cdextran bestämdes både före och efter sekundär tillsats av kolloidal kiselsyra (CS) och adsorptionen av CS kvantifierades också som funktion av yttäckningen av polyelektrolyt.</p><p>Resultaten indikerar att laddningstätheten hos CPAM kontrollerar den adsorberade mängden på kiseloxidytor vid låga NaCl koncentrationer. Både adsorptionen av CPAM och Cdextran på kiseloxid visades vara effektiv i NaCl koncentrationer upp till 1 M, vilket indikerar ett signifikant bidrag av icke-jonisk interaktion mellan polyelektrolyterna och kiseloxid. Adsorptionen av CS var högre på föradsorberad CPAM än Cdextran. Konformationen hos de adsorberade skikten efter tillsats av CS sågs expandera signifikant för skikt baserade på CPAM medan skikt av Cdextran vid låga salthalter verkade återta sin konformation efter en temporär expansion.</p><p>I den andra delen av avhandlingen studerades sekventiell adsorption av PAE och CMC på massafibrer. Adsorptionsisotermer skikt för skikt på avkryllad massa visade att PAE adsorberade i större mängd än CMC, både i hänseende av massa och laddning. Adsorptionen av PAE var signifikant långsammare än CMC och adsorptionstiden till 90% av mättnadsadsorptionen bestämdes till 3 respektive 1 minut. Zetapotentialen för kryll bestämdes för adsorption av de två första polyelektrolytskikten och resultaten tydde på att kryllmaterialet omladdade inom en minut efter tillsatserna av polyelektrolyt. Reflektometriförsök inom sekventiell adsorption av PAE och CMC på kiseloxid antydde att den låga molekylviktsfraktionen av PAE störde uppbyggnaden av polyelektrolyt-multiskikten.</p> / <p>This thesis presents experimental studies of polyelectrolyte adsorption on oppositely charged surfaces, where substrates of both silica and bleached softwood kraft pulp were used. A major aim of this research was to characterise the conformation of adsorbed layers of cationic polyacrylamide (CPAM), in comparison to cationic dextran (Cdextran), and relate this information to the binding capacity of colloidal silica. A second aim in this thesis was to study the kinetics of the sequential adsorption of polyamide epichlorohydrine (PAE) and carboxymethyl cellulose (CMC) on pulp fibres, and to determine the adsorption isotherms for the layer-by-layer deposition of polyelectrolytes on pulp fibres.</p><p>The adsorption of CPAM on silica surfaces was studied using stagnation point adsorption reflectometry and quartz crystal microgravimetry to determine its adsorption kinetics as well as the dependencies of polyelectrolyte charge densities, pH, and NaCl concentration on saturation adsorption. The conformation of adsorbed layers of CPAM and Cdextran, analysed in terms of amount of water and layer thickness, was determined both before and after the secondary adsorption of colloidal silica (CS), and the adsorption of CS was also quantified as a function of the surface coverage of the polyelectrolyte.</p><p>Results indicate that the charge density of CPAM controlled the amount of the polyelectrolyte adsorbed on silica surfaces at low NaCl concentrations. The adsorption of both CPAM and Cdextran on silica was shown to be effective at up to 1 M NaCl concentrations, which indicates that non-ionic interactions between the polyelectrolytes and silica contribute significantly. CS adsorption was higher on pre-adsorbed CPAM than on Cdextran. The conformation of the adsorbed layer after CS addition was seen to expand significantly in CPAM-based layers, while the Cdextran layer appeared to restore its conformation after a temporary expansion at low salt concentrations.</p><p>In the second part of the thesis, the sequential adsorption of PAE and CMC on pulp fibres was determined using the polyelectrolyte titration technique. Layer-by-layer adsorption isotherms derived on fractionated pulp showed that PAE adsorbed in higher amounts than CMC did, both in terms of adsorbed mass and adsorbed charge. The adsorption of PAE was significantly slower compared to CMC, and the adsorption times required to reach 90% of the saturation adsorption were 3 and 1 min, respectively. The zeta potential of pulp fines was determined for the adsorption of the two first polyelectrolyte layers, and data indicated that the fines recharge within one minute after the polyelectrolyte additions. Reflectometry experiments regarding the sequential adsorption of PAE and CMC on silica indicated that the low-molecular-weight fraction of PAE disturbed the formation of polyelectrolyte multilayers.</p>

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2005. / Härtill 4 uppsatser.

Made available in DSpace on 2014-12-17T15:18:13Z (GMT). No. of bitstreams: 1 LourenaMV.pdf: 713655 bytes, checksum: d2c61219ecefc4633620bb6eebd45325 (MD5) Previous issue date: 2007-02-26 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Broadly speaking, the concept of gene therapy involves the transfer of a genetic material into a cell, tissue, or organ in order to cure a disease or at least improve the clinical status of a patient. Making it simple, gene therapy consists in the insertion of functional genes into cells containing defective genes by substituting, complementing or inhibiting them. The achievement of a foreigner DNA expression into a population of cells requires its transfer to the target. Therefore, it is a key issue to create systems able to transfer and protect the DNA until it reaches the target, the vectors. The disadvantages related to the use of viral vectors have encouraged efforts to develop emulsions as non-viral vectors. In fact, they are easily produced, present controllable stability and enable transfection. The aim of this work was to develop an emulsion for gene therapy and evaluate its ability to compact nucleic acids by the development of a complex with the plasmid pIRES2-EGFP. The first step was to determine the Hydrophilic Lipophilic Balance (HLB) of the Captex? 355 (oily internal phase of the emulsion) through long and short term stability assays. Based on the results, emulsions composed of Captex? 355, Tween 20? and Span 60? with 10.7 HLB were produced by three different methods: phase inversion, spontaneous emulsification and sonication. The results showed that the lowest diameter and best stability of the emulsions were achieved by the sonication method. The cationic emulsions were made by adding DOTAP to the basic emulsion. Its association with pIRES2-EGFP was evaluated by electrophoresis. Several rates of emulsion and DNA were evaluated and the results showed that 100% of the complex was formed when the rate DOTAP/DNA(nmol/?g) was 130. In conclusion, the overall results show the ability of the proposed emulsion to compact pIRES2-EGFP, which is a requirement to a successful transfection. Therefore, such formulation may be considered a promising candidate for gene therapy / Terapia g?nica, em uma ampla defini??o, ? o tratamento de doen?as baseado na transfer?ncia de material gen?tico a uma c?lula, tecido ou ?rg?o com o intuito de curar ou melhorar o estado cl?nico do paciente. Em sua forma mais simples, a terapia g?nica consiste na inser??o de genes funcionais em c?lulas com genes defeituosos objetivando substituir, complementar ou inibir esses genes causadores de doen?as. Para que o DNA ex?geno seja expresso em uma popula??o celular faz-se necess?ria a sua transfer?ncia at? o local. Assim, ? necess?rio criar ve?culos, os vetores, que transportem e protejam o DNA at? que este chegue a uma popula??o celular alvo. Os obst?culos encontrados com a utiliza??o de vetores virais t?m proporcionado o interesse no desenvolvimento de emuls?es cati?nicas como vetores n?o-virais, por serem f?ceis de produzir, apresentarem estabilidade control?vel e facilitarem a transfec??o g?nica. O objetivo deste trabalho foi desenvolver um sistema emulsionado para terapia g?nica e avaliar sua capacidade de compacta??o de ?cidos nucl?icos atrav?s da sua associa??o com o plasm?deo pIRES2-EGFP. Primeiramente o EHLc do TCM utilizado, o Captex? 355, foi determinado atrav?s de ensaios de estabilidade acelerada e a longo termo. Com base nos resultados obtidos, emuls?es de EHL 10,7 compostas de Captex? 355, Tween 20? e Span 60? foram preparadas pelos m?todos de invers?o de fases, emulsifica??o espont?nea e sonica??o e elegeu-se o melhor m?todo para o preparo das emuls?es cati?nicas. As emuls?es de menor granulometria e maior estabilidade foram obtidas atrav?s do m?todo de sonica??o. As emuls?es cati?nicas foram preparadas acrescendo-se ? emuls?o base o DOTAP e a sua associa??o com o pIRES2-EGFP foi avaliada atrav?s da t?cnica de eletroforese em gel de agarose. V?rias propor??es de emuls?o e DNA foram testadas e os resultados demonstraram que houve forma??o de 100% dos complexos quando a propor??o DOTAP/DNA(nmol/?g) foi igual a 130. Em conclus?o, o conjunto dos resultados obtidos demonstra a capacidade da emuls?o proposta neste trabalho de compactar o DNA, requisito necess?rio para uma boa transfec??o, tornando a formula??o uma forte candidata ? utiliza??o em terapia g?nica

Tendo em vista que os amidos (catiônicos ou não) não são nocivos a saúde do ser humano, uma vez que são utilizados na indústria de alimentos, e observando-se a potencialidade do seu uso como auxiliares de floculação, foi proposto o presente trabalho. Consiste na realização de ensaios de coagulação (com sulfato alumínio), floculação e sedimentação em equipamento de reatores estáticos, objetivando verificar a influência do gradiente de velocidade e do tempo de agitação na floculação com polímero sintético catiônico e amidos de milho e mandioca catiônicos. Concluiu-se que as condições de aplicação exercem influência na eficiência de remoção de turbidez e cor aparente, sendo que cada polímero estudado apresentou uma condição ótima específica. Em todos os ensaios realizados, o amido de mandioca catiônico foi o mais eficiente, indicando que os amidos catiônicos podem ser substitutos em potencial dos polímeros sintéticos no tratamento de águas de abastecimento, quando utilizados como auxiliares de floculação. / This present work was based on the fact that native and cationic starches are not harmful to man\'s health, since they have been largely used in food processing industries, and that they may be potentially used as flocculation aids in water treatment. Jar Test assays were performed, including coagulation with aluminum sulphate, flocculation and sedimentation, aiming to study the influence of the velocity gradient and mixing time on flocculation, using a cationic synthetic polymer and cationic com and manioc starches as aids. In the main, it was concluded that the conditions under which the polymers were applied, affect the efficiency of turbidity and color removals and that each polymer studied presented an specific optimum condition. It was also observed that manioc cationic starch resulted more effective than the others polymers studied, when used as flocculation aids.

Dans les procédés classiques d'extraction de la cellulose pour la fabrication de la pâte à papier à partir de bois, les hémicelluloses sont dégradées et sont brûlées pour alimenter l'usine en énergie. En incluant une étape préalable d'extraction de ces polysaccharides, ils pourraient alors être réintroduits en aval du procédé, après cationisation, en tant qu'agent de renfort dans la pâte à papier. Dans ce travail, la cationisation de deux hémicelluloses modèles commerciales de type galactomannane et xylane a été effectuée par éthérification avec un époxyde cationique en solution aqueuse basique, avec des degrés de substitution de 0,1 à 1,3. Les paramètres prépondérants de la réaction sont la concentration des espèces et la quantité d'agent éthérifiant, tandis que la température et le temps de réaction influent principalement sur le rendement massique. Un plan d'expérience a permis de modéliser l'interaction entre les quantités de soude et d'époxyde et d'optimiser les conditions en fonction du DS avec une efficacité de réaction de 50%. La cationisation a également été appliquée à deux hémicelluloses extraites au FCBA. La mise en oeuvre de la réaction en absence de solvant a permis d'atteindre une efficacité de réaction de 70 à 90% et de réduire considérablement les quantités de réactif utilisées et d'effluents à traiter. Parmi la gamme de DS, les dérivés cationiques de DS 0,3 ont donné les meilleures propriétés après ajout dans la pâte à papier, le galactomannane étant plus efficace que le xylane. Enfin, la réticulation des chaînes du xylane extrait a permis d'augmenter les propriétés physiques du papier de 15% supplémentaires / With current paper pulp processes, hemicelluloses are decomposed during the extraction of cellulose fibres. By including a previous step of hemicellulose extraction, these compounds could be reused as additives in the paper pulp, instead of cationic starch. In this work, two model galactomannan and xylan hemicelluloses were cationized in aqueous solution under basic conditions by etherification with a cationic epoxide. Degrees of substitution from 0.1 to 1.3 were obtained. Species concentration and epoxide amount appeared as preponderant parameters of the reaction. With an experimental design, the interaction between sodium hydroxide and epoxide amounts was modelized, and reactional conditions were optimized depending on the desired DS with a reaction efficiency of 50%. Then, cationisation was applied to two hemicelluloses extracted in FCBA. The reaction was also carried out in absence of solvent with a reaction efficiency of 70-90%, by minimising reagents consumption and effluents. Cationic derivatives with DS 0.3 gave the best performance to increase the paper resistance when they were added in paper pulp, galactomannan being more efficient than xylan. Chain reticulation allowed increasing mechanical properties of paper of 15% for the extracted xylan

Cathelicidins are cationic host defence peptides (CHDP) with essential roles in the innate defence system. These peptides have antimicrobial potential and the capacity to modulate innate immunity and inflammatory processes. Neutrophils (PMN) are the main reservoir of cathelicidins and play key roles in first line defence against infection. The appropriate regulation of PMN function, death, and clearance is critical to innate immunity, and the efferocytosis of apoptotic PMN, in contrast to necrotic cells, is proposed to promote the resolution of inflammation. In this thesis I demonstrate that the human cathelicidin LL-37 rapidly induced secondary necrosis of apoptotic human PMN and identify the essential C-terminal region of LL-37 required for this activity. In addition to the induction of secondary necrosis, higher concentrations of LL-37 also promoted PMN granule contents release. LL-37-induced secondary necrosis did not affect PMN ingestion by human monocyte-derived macrophages and, in contrast to expectation, was not proinflammatory. Interestingly, the anti-inflammatory effects of apoptotic PMN on activated macrophages were retained and even potentiated where LL-37-mediated secondary necrosis induced anti-inflammatory granule content release. Consistent with the results of in vitro studies, in vivo murine sterile peritonitis model revealed the same phenomenon: LL-37-induced secondary necrosis diminished inflammatory responses with decreased PMN influx. I also present data on LL-37- mediated modulation of innate immune effector cell cytokines responses to inflammatory signals. I propose that during acute inflammation LL-37 can modulate innate immune responses through its activity on cytokine production, and that LL-37-mediated secondary necrosis of apoptotic PMN has anti-inflammatory effects, but may also mediate host damage by promoting the release of potentially harmful intracellular contents under chronic or dysregulated conditions.

Cationic lipids are efficient tools to introduce nucleic acids and proteins into cells. Elucidation of the mechanism and cellular pathways associated to such a transport has been relatively slow, even though significant progress has been made in the characterization of the intracellular trafficking of cationic lipid/DNA complexes. Surprisingly, little is known about the effects of these delivery vectors on cell functioning. In the present thesis, we show that cationic lipids and cationic lipid/DNA complexes strongly increase the intracellular Ca2+ concentration. The end point of the Ca2+ increase was ~400 nM from a basal level of ~100 nM. The [Ca2+]i increase was studied using K562 and Jurkat cells cultured in vitro. This effect is weakened following addition of DNA to cationic liposomes, although remaining very large at cationic lipid/DNA ratios commonly used for cell transfection experiments. Removal of extracellular Ca2+ did not abolish this effect significantly and preincubating K562 cells with the Ca2+-ATPase inhibitor thapsigargin strongly abolished intracellular Ca2+ concentration increase, indicating that Ca2+ was released mainly from internal Ca2+ stores sensitive to thapsigargin. Pretreatment of the cells with the phospholipase C inhibitor U73122 blocked the intracellular Ca2+ concentration rise, suggesting an inositol pathway-dependent mechanism. LDH release assay indicates that in the conditions used for fluorescence measurement and in those used to transfer DNA into cells, cationic liposomes diC14-amidine and DOTAP had no massive cytotoxic effects. Cationic liposomes showed more toxicity than their corresponding complexes; this toxicity decreases in the presence of serum. The effect of cationic lipids on phosphatidylinositol-specific phospholipase C (PI-PLC) was quantitatively assessed using phosphatidylinositol (PI) and radiolabeled phosphatidylinositol ([3H]-PI). Incorporation of diC14-amidine into PC/PI vesicle activated PI-PLC and was shown to activate the hydrolysis of PI and [3H]-PI. Our data may suggest that mobilization of intracellular Ca2+ by complex could have an effect on the transfection process itself. These results indicate for the first time that cationic lipids and cationic lipid/DNA complexes are not inert and can affect the functioning of the cells by increasing their intracellular Ca2+.

No description available.