CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Neste trabalho, estudou-se o efeito da temperatura nos espectros Raman de cristais de L-alanina + Ãcido oxÃlico, C3H8NO2+.C2HO4-. Foram realizadas medidas de espectroscopia Raman em policristais a diferentes temperaturas no intervalo compreendido entre a temperatura ambiente e a temperatura de 20 K, sendo fornecida uma identificaÃÃo tentativa para todos os modos normais de vibraÃÃo observados. Para auxiliar o entendimento do comportamento do cristal tambÃm foram obtidos os difratogramas de raios-X bem como estudada a dependÃncia dos parÃmetros de rede em funÃÃo da temperatura atravÃs de dilatometria no intervalo entre 290 K e 93 K. As trÃs tÃcnicas utilizadas em conjunto permitiram mostrar o comportamento estrutural do material em baixas temperaturas. Deste quadro foi possÃvel inferir que os cristais de L-alanina + Ãcido oxÃlico apresentam trÃs diferentes transiÃÃes de fase durante o resfriamento. Em 250 K o aparecimento de um dubleto em 90 cm-1 e a anomalia num dos parÃmetros de rede apontam para a ocorrÃncia da primeira transiÃÃo de fase. Em 150 K surgem pelo menos duas novas bandas no espectro Raman, ao mesmo tempo em que ocorrem bruscas mudanÃas de inclinaÃÃo nas curvas que representam as dimensÃes dos eixos a e c do cristal. TambÃm se verifica que, de forma semelhante ao que ocorre com os espectros Raman, aparecem novos picos no difratograma de raios-X em torno desta temperatura, caracterizando assim a segunda transiÃÃo de fase. A uma temperatura ainda mais baixa, em torno de 43 K, foi verificada a ocorrÃncia da terceira transiÃÃo de fase, que tem como principal caracterÃstica a separaÃÃo de dois modos Raman associados a modos da rede. MudanÃas nos ambientes dos grupos CH3 e do NH3+ durante o resfriamento sÃo discutidas. Um importante aspecto apresentado pelos espectros Raman com o resfriamento da amostra foi o deslocamento da banda de mais baixa energia para menores valores de frequÃncias, semelhantemente ao que ocorre com vibraÃÃes do tipo soft-mode em materiais ferroelÃtricos, embora a frequÃncia do modo no cristal de L-alanina + Ãcido oxÃlico nÃo tenha ido à zero. Baseado nos resultados acima e nos possÃveis sÃtios de simetria ocupados pelas molÃculas atravÃs do grupo O=CC nas diversas fases, sugere-se a seguinte sequÃncia de transiÃÃes de fase D24 para C2h5 para Cs3 para C23, que aconteceriam, respectivamente, nas temperaturas de 250 K, 150 K e 43 K. / In the present word we have studied the effect of temperature on the Raman spectra of crystals of L-alanine + oxalic acid, C3H8NO2+.C2HO4-. Raman spectroscopy measurements were performed on polycrystalline samples at different temperatures varying in the range from room temperature to T = 20 K; a tentative assignment of all normal modes was furnished. In order to help the understanding of the crystal behavior we have also obtained X-ray diffractograms and studied the dependence of lattice parameters through dilatometry as a function of temperature in the 290 K â 93 K range. The three different techniques allowed us to construct a picture of the material under low temperature conditions. As a consequence we have realized that L-alanine + oxalic acid crystal undergoes three phase transitions at low temperatures. The splitting of a band at 90 cm-1 and an anomaly in one of the lattice parameters are the signature for the first phase transition that is observed at 250 K. At 150 K it was observed the appearance of two new bands in the Raman spectrum and, simultaneously, it was observed change in the curves of a and c lattice parameters. Additionally, it was verified the appearance of new peaks in the X-ray diffractogram at the same temperature, characterizing the second phase transition. At a temperature even lower, at about 43 K, it was verified the occurrence of the third phase transition that has as main characteristic the splitting of two bands that are associated to the lattice modes. Changes in the modes associated with CH3 and NH3+ during the cooling is discussed. An important behavior of the crystal with the cooling process was the red shift of the band of lower frequency, similar to the soft-mode vibration of ferroelectric materials, although the frequency of the mode in L-alanine + oxalic acid does not goes to zero. Based on the results on Raman spectroscopy, dilatometry and X-ray diffraction, and on the possible symmetry sites occupied by the molecules through the O=CC group in the various phases, it is suggested the following sequence of phase transitions D24  C2h5  Cs3  C23, which should occur at 250 K, 150 K and 43 K.

O metabolismo da glutamina (Gln) é alvo de atenções recentes para a compreensão da reprogramação metabólica para o suprimento energético das células tumorais em proliferação e para o desenvolvimento de novas estratégias terapêuticas em câncer. Tanto a absorção de glutamina quanto a taxa de glutaminólise, o catabolismo da Gln para gerar adenosina trifosfato (ATP) e lactato na mitocôndria estão aumentados em diferentes tumores. A Gln e glicose participam do processo da proliferação de células tumorais tanto na produção de (ATP) como no fornecimento de produtos intermediários utilizados na síntese de macromoléculas e Gln é utilizado para anaplerose do ciclo do ácido tricarboxílico. Nesse estudo, nosso objetivo foi analisar a expressão dos genes envolvidos na glutaminólise: ASCT2, LAT1, GLS, GLSISO1, GLSISO2, GLS2, GOT1, GOT2, GLUD1 e GPT2 em astrocitomas de diferentes graus de malignidade (AGI-AGIV), classificados de acordo com a Organização Mundial de Saúde (OMS), em relação às expressões em tecidos cerebrais não neoplásicos e correlacionar os níveis de expressão destes genes aos dados clínicos. PCR quantitativo em tempo real (qRT-PCR) foi realizado em 175 amostras, sendo 22 dentre estes de tecidos não neoplásicos. Observou-se tempo de sobrevida menor entre os pacientes com hiperexpressão de LAT1, na presença de hipoexpressão de ASCT2. A expressão de GLS foi comparativamente maior que a expressão de GLS2 entre os astrocitomas de diferentes graus de malignidade, corroborando descrições prévias de que GLS relaciona-se à proliferação tumoral e GLS2 à supressão do crescimento tumoral. Observou-se, adicionalmente, o aumento da associação das expressões destes genes conforme o aumento do grau de malignidade, culminando em GBM, onde estas correlações foram estatisticamente significativas. Apesar da demonstração da ativação gradativa desta via da glutaminólise com o aumento da malignidade, a hiperexpressão dos genes relacionados a esta via mostrou-se hiperexpressa em apenas um subgrupo de pacientes com GBM. Esta observação ressalta a heterogeneidade observada em GBM e a elegibilidade restrita deste subgrupo a eventuais estratégias terapêuticas que forem desenvolvidas com alvos nesta via / The metabolism of glutamine (Gln) is the target of recent attentions to understand the metabolic reprogramming of cancer cell for the energetic needs for cell proliferation, and to develop new cancer therapeutic strategies. Glutamine absortion and glutamine conversion to ATP and lactate in the mitochondria through glutaminolysis are both increased in different cancer types. Gln and glucose participate in metabolic pathways which provide ATP and intermediate substrats for synthesis of macromolecules, and Gln is used for anaplesoris of tricyclic acid cycle. The aims of the present study were to analyze the differential mRNA expressions of genes involved in the glutaminolysis pathways: ASCT2, LAT1, GLS, GLSISO1, GLSISO2, GLS2, GOT1, GOT2, GLUD1 e GPT2 in astrocytomas of different grades of malignancy (WHO grades I to IV) compared to non-neoplastic brain tissues, and to correlate these expression data to clinical outcome. Shorter overall survival time was observed among a subset of GBM patients presenting hyperexpression of LAT1 while ASCT2 was hypoexpressed. GLS expression was comparatively higher than GLS2 expression among astrocytomas of different grades of malignancy, which corrobates previous reports relating GLS to tumor proliferation and GLS2 to suppression of tumor growth. Additionally, increased gene expression correlation was observed in parallel to the increase of malignancy, and these associated expressions were significant among GBM. Although a stepwise increase of the glutaminolysis pathway was demonstrated with the increase of malignancy in astrocytomas, the hyperexpression of genes involved in this pathway were detected only in a subset of GBM patients. This finding confirm the heterogeneity observed among GBM, and highlights the fact that any therapeutic strategy aiming this pathway will be restricted to a subset of GBM patients

Chapter -I Introduction Salmonella is a pathogen well-known for its ability to infect a wide variety of hosts and causes disease ranging from mild gastroenteritis to typhoid fever. During infection, it is exposed to a myriad of conditions; from the aquatic environment, the gut lumen to the phagolysosome. The success of Salmonella as a pathogen lies in its ability to sense each of these environments and adapt itself for survival and proliferation accordingly. This is done mainly via the action of specific two-component systems (TCSs) which sense cues specific to each of these niches and trigger the appropriate transcriptional reprogramming. This reprogramming is best studied for the genes directly known to be involved in virulence. In the case of Salmonella, most of these genes are a part of specific clusters, acquired through horizontal gene transfer, known as Salmonella Pathogenicity Islands (SPIs). Of the various SPIs, the two most important are SPI-1 and SPI-2. SPI-1 is classically involved in orchestrating bacterial invasion of non-phagocytic cells in the gut, allowing the pathogen to invade the host. Furthermore, its role is well characterized in the classic inflammation associated with gastroenteritis. On the other hand, SPI-2 is specialized for survival within the harsh intracellular environment of host cells such as macrophages and epithelial cells. Other important virulence determinants include motility, chemotaxis as well as adhesins. The transcription of these virulence genes is under tight regulation and responsive to environmental conditions. Many small molecules such as short chain fatty acids, pp(p)Gpp, bile and acyl homoserine lactones among others are known to be potent regulators of virulence in Salmonella. Furthermore, the metabolic products of the normal flora in the gut also affect its virulence. Thus the metabolic status, of both the host as well as the pathogen, plays an important role in determining the outcome of the infection. Many metabolic enzymes and their products are now known to directly or indirectly affect virulence gene expression. In this study, we explore one such class of metabolic enzymes viz amino acid racemases. They catalyze the chiral conversion of L-amino acids to D-amino acids and vice versa. We have studied the biochemical properties of two such non-canonical racemases as well as their role in bacterial survival and pathogenesis. Chapter-II Identification and characterization of putative aspartate racemases in Salmonella Amino acid racemases, such as alanine and glutamate racemases, are ubiquitously found in all bacteria and they play an essential role in cell wall biosynthesis. Recently it has been found, that bacteria possess other amino acid racemases which produce non-canonical D-amino acids. These D-amino acids, upon secretion, further orchestrate various phenotypes such as cell wall remodeling and biofilm dispersal. In this study, we have explored the ability of Salmonella to produce such non-canonical D-amino acids. The genome of S. Typhimurium possesses genes encoding two putative aspartate racemases; ygeA and aspR. These genes were maximally expressed in mid-log phase of bacterial growth and their corresponding proteins ar localized in the outer membrane of the bacterium. The biochemical characterization of the proteins YgeA and AspR revealed that only the latter is catalytically active under in vitro conditions. AspR could catalyze the conversion of L-Aspartate to D-Aspartate and vice versa, however was unable to use any other amino acid as its substrate. With atleast one of the racemases showing catalytic activity, the profiling of the secreted D-amino acids in Salmonella conditioned medium was undertaken using LC-MS. It was observed that the bacterium actively secreted specific D-amino acids such as D-Ala and D-Met into the culture medium in a growth-phase dependent manner. Furthermore, analysis of the secreted D-amino acid profile of the strains lacking either one or both the racemases revealed that atleast a subset of the secreted D-amino acids were dependent on the activity of YgeA and AspR. Thus, D-amino acids secreted by S. Typhimurium might represent a novel class of signaling molecules. Chapter – III Role of aspartate racemases in growth and survival of S. Typhimurium In order to understand the role of ygeA and aspR in vivo, we created knockouts of these genes (both single as well as double knockout) in S. Typhimurium using λ Red recombinase strategy. These knockouts were then assessed for their growth and morphology. The aspartate racemase knockouts behave similar to the wild type during growth in LB as well as M9 minimal medium. While their gross morphology remained the same as the wild type, the size distribution of the racemase knockouts was slightly different in the stationary phase. Unlike the wild type bacteria, the mutants did not exhibit the characteristic reduction in cell size upon entry into stationary phase. In addition, the survival of the mutants in the presence of cell wall damaging agents such as bile and Triton-X 100 was compromised as compared to the wild type. This can be ascribed to changes in the cell wall of the bacterium, wherein the mutants accumulated peptidoglycan in the stationary phase of growth. This suggests that aspartate racemases might have an effect on cell wall biosynthesis in Salmonella in the stationary phase. Another important strategy employed by bacteria to survive in stress conditions is biofilm formation. It was seen that the mutants were compromised in their ability to form a biofilm at the liquid-air interface in vitro. This defect is due to a transcriptional downregulation of the genes required for biofilm formation. These results demonstrate that, contrary to the established inhibitory effects of D-amino acids on biofilms of various bacteria, the aspartate racemases appear to act as positive regulators of biofilm formation in Salmonella. Chapter – IV Involvement of aspartate racemases in the regulation of Salmonella pathogenesis Salmonella’s success as a pathogen can be broadly assessed by its ability to invade and replicate within two major cell types: epithelial cells and macrophage-like cells. We have studied the fate of the aspartate racemase knockout strains in both these cell types. While the mutants replicate as well as the wild type in macrophage cell lines, their ability to invade epithelial cell lines is highly compromised. This defect can be ascribed to the downregulation of the Salmonella Pathogenicity Island-1 (SPI-1) in the racemase knockouts at the transcriptional level. One of the major pathways that regulate SPI-1 activation is the flagellar pathway. It was observed that in addition to SPI-1, the motility of the racemase mutants was also highly compromised. The mutants did not possess any flagella and showed a high transcriptional downregulation of all the three classes of flagellar genes. Transcriptome analysis revealed a global reprogramming in the aspartate racemase mutants, resulting in the differential regulation of motility, adhesion, amino acid transport, cell wall biosynthesis and other pathways. Of the genes upregulated in the knockouts, FimZ is known for its negative effect on motility and might be responsible for the observed downregulation of the flagellar regulon. This suggests that ygeA and aspR might be repressors of fimbrial gene expression. In totality, the racemases affected the pathogenesis of Salmonella, where the double knockout was severely compromised in the colitis model of infection. Overall the study is the first to identify secretion of non-canonical D-amino acids by Salmonella and suggests that YgeA and AspR might be the source of the same. This is supported in part by in vitro studies with the purified proteins. Studies in vivo further highlight the possible substrates that might be utilized by these enzymes. Physiologically, the aspartate racemases appear to regulate cell wall remodeling and biofilm formation. In contrast to the established literature, aspartate racemases (and their possible D-amino acid products) seem to be essential for formation of biofilms and regulate this phenotype at the transcriptional level. Furthermore, our studies put forth aspartate racemases as novel positive regulators of Flagella and SPI-1, affecting the success of Salmonella in the colitis model of infection in mice. Transcriptome analysis hints at the pleiotropic effects of aspartate racemases in Salmonella, bringing forth hitherto unexplored roles for this class of enzymes in the biology of this pathogen.

Chirality is the property of objects, including molecules, which are not superimposable on their materialized mirror image. Chiral molecules are omnipresent in living organisms and the constituents of biological macromolecules (proteins and nucleic acids) are chiral. Amino-acids (constituting proteins), ribose and 2-deoxy-ribose (the only chiral constituent of RNA and DNA nucleotides respectively) are furthermore generally present in living organisms only under one of their enantiomeric forms. This is referred to as the homochirality of the living world. The origin of this homochirality is still unexplained, even if many partial scenarios have been proposed in the literature. All scenarios involve the creation of a small enantiomeric excess for certain molecules, amplification of this excess and chirality transfer to other chiral molecules. The origin of homochirality on Earth is closely related to the origin of life, and is currently supposed to have preceded life. As no-one will ever be able to directly observe the phenomena which lead to homochirality, and life, on our planet, the only scientific approach to try and help explain how this occurred is to build scenarios, and test them taking into account all available information on the physical and chemical conditions on the primitive Earth (Earth before life appeared). In our work, we investigated three scenarios related to the origin of homochirality on Earth. One of these scenarios also relates to a very precise step of the origin of life: the selection of beta-d-ribofuranose as component of RNA nucleotides.<p><p>Enantiomeric excesses (up to 15 %) of alpha-methylated alpha-amino-acids have been detected in meteorites which fell on Earth during the 20th century. No enantiomeric excess is detected for the corresponding alpha-hydroxy-acids in the same meteoritic samples and small (2% at most) or no enantiomeric excesses have been measured for non-methylated alpha-amino-acids. In the first part of our work, we investigated if photolysis by circularly polarized light (CPL) in space could be at the origin of the presence (or absence) of an enantiomeric excess for these compounds. Experiments to reproduce UV-CPL photolysis are difficult to undertake: they require high-energy circularly polarized photons, hence the use of a synchrotron. In our work, we used quantum mechanical calculations to obtain the electronic circular dichroïsm (ECD) spectra of two -methylated -amino-acids, their corresponding alpha-hydroxy-acids and one non-methylated alpha-amino-acid. Differences are observed between these spectra, and we propose a scenario to explain the experimental measurements reported here above: the enantioselective photolysis, in the gas phase at low temperatures (20K at most), of the alpha-amino-acids by UV-CPL with lambda>210 nm. Under these conditions no photolysis of the alpha-hydroxy-acids would occur. This scenario concerns the first step in the origin of homochirality on Earth: the creation of a small enantiomeric excess for some chiral molecules.<p><p>The second scenario that we investigated relates to the enantiomeric amplification step of the origin of homochirality on Earth, for which the role of the alpha-amino-acid serine has been suggested in the literature. Serine clusters have been observed in the gas phase by mass spectrometry. Among these clusters the octamer has been shown to be a magic number cluster and to have a preference for homochirality. An enantiomeric amplification via cycles of formation and dissociation of the octamer has been suggested. No complete scenario has however been proposed in the literature to explain how this could have occurred on the primitive Earth, but any scenario would most probably include an aqueous phase. We aimed at determining if the homochiral preference of serine octamers also exists in solution and therefore we first investigated if serine octamers exist in solution. For this study, we used nuclear magnetic resonance and infrared spectroscopies, which are well-adapted to the study of molecular assemblies in solution. We were able to demonstrate that most probably serine clusters are not present in solution, and if they are it could only be in extremely low concentration. The scenario suggested in the literature is discussed in the light of our results and of literature data on serine clusters.<p><p>As last hypothesis, we investigated a possible scenario for the selection of beta-d-ribofuranose as component of RNA nucleotides. The currently known prebiotic synthesis pathways to ribose also lead to the formation of many other carbohydrates, and ribose is only a minor product of these syntheses. Our hypothesis is that beta-d-ribofuranose could have been selected through favorable interactions with alpha-amino-acids already present on the primitive Earth under one enantiomeric form. Indeed, it is plausible that a peptidic world emerged before the presence of RNA and that homochiral alpha-amino-acids were present on Earth when RNA was synthesized. Under this hypothesis, we investigated the role that alpha-l-amino-acids could have played in the selection of alpha-d-ribofuranose as component of RNA nucleotides. This work is related to the last step of the origin of homochirality: chirality transfer. Our scenario was investigated via nuclear magnetic resonance studies of the interaction between alpha-amino-acids and carbohydrates. We were able to show that, in the systems that we studied, when an interaction occurs it is very weak (affinity constant less than 1M−1) and non enantioselective. Our results most probably discard the role that alpha-amino-acids alone could have played in the selection of beta-d-ribofuranose as component of RNA nucleotides, but does not discard the role that peptides could have played in this selection. / Doctorat en Sciences de l'ingénieur / info:eu-repo/semantics/nonPublished

Meiosis is functionally conserved across eukaryotes, thus not expected to vary considerably among different species, and even less so among lineages within a species. However, recent studies showed that this is not necessarily the case in Arabidopsis arenosa. Genome scanning identified an excess differentiation in meiosis genes between A. arenosa diploids and tetraploids, interpreted as meiosis adaptation to the whole genome duplication in tetraploids and differentiation was also found between two diploid lineages. Thus, I present a population-based analysis of positive selection acting on meiosis proteins across multiple lineages of A. arenosa. I showed that meiosis proteins were under positive selection in all diploid lineages, mainly in the Pannonian and South-eastern Carpathian lineage. The evidence for positive selection in diploid lineages suggested differential pathways of meiosis adaptations in the species, probably reflecting the necessity to adapt to local environments, among all to temperature. The highest enrichment of amino acid substitutions (AASs) under positive selection was identified in tetraploids, in consistence with previous genome-scan results. As several interacting meiosis proteins were under positive selection in the same A. arenosa lineage, I hypothesize that the close...

La chimie des foldamères est un domaine de recherche en pleine expansion où les chimistes explorent la construction d’architectures artificielles variées mimant les structures repliées des biopolymères naturels. Les foldamères d’oligoamides quinoline, constituent une branche importante des foldamères montrant de nombreuses caractéristiques attractives, incluant la stabilité et la prédictibilité de leurs conformations repliées, qui en font de bons candidats pour des applications biologiques. Jusqu’à présent, la plupart des études sur les foldamères d’oligoamides quinolines ont été menées dans des solvants organiques. Cette thèse a pour objectif d’étendre leur portée au milieu aqueux et présente plusieurs méthodologies pour parvenir à leur solubilité, leur repliement, la variation de leurs chaines latérales, leur agrégation et leur capacité à former des cristaux dans l’eau.Tout d’abord, une méthode de synthèse en phase solide a été développée permettant l’accès rapide aux foldamères hybrides α-amino acide/quinoline (X/Q). Leur étude dans l’eau montre que contrairement aux foldamères hybrides de type (XQ)n, ceux de type (XQ2)n sont capables d’adopter une conformation hélicoïdale présentant un alignement des chaines α-amino acides dans l’espace. Ensuite, plusieurs chaines latérales courtes ont été identifiées pour doter les foldamères aromatiques d’une solubilité et d’une capacité à cristalliser dans l’eau. Six oligoamides quinoline ont ainsi été synthétisés pour une étude modèle. Des cristaux ont été obtenus pour toutes les séquences sauf une, présentant une excessive solubilité dans l’eau. Enfin, des efforts ont été faits pour construire des faisceaux d’hélices auto-assemblés dans l’eau à base d’effets hydrophobes et d’interactions électrostatiques. Les études RMN et cristallographiques ont indiqué que les effets hydrophobes étaient plus faibles qu’attendu et ne provoquaient pas d’agrégation forte. / Foldamer chemistry is a rapidly expanding research field where chemists explore the construction of various artificial architectures that mimic the folded structures of biopolymers found in nature. Quinoline oligoamide foldamers, as an important branch of foldamers, have been shown to possess many desirable features, including stability and predictability of their folded conformations, and are promising candidates to achieve biological applications. Up to now, most investigations of quinoline oligoamide foldamers have been carried out in organic solvents. This thesis is aimed to expand their scope in aqueous medium and presents several methodologies to achieve solubility, folding, side-chain variation, aggregation and crystal growth ability in water.First, a solid phase synthesis method was developed to enable the fast access to α-amino acid/quinoline (X/Q) hybrid oligoamide foldamers. The study of these hybrid foldamers in water showed that contrary to (XQ)n-type foldamers the (XQ2)n-type foldamers could adopt aromatic helical conformations with α-amino acid side chains aligned in space. Then, several short side chains were identified to endow aromatic foldamers with both solubility in, and crystal growth ability from water. Six quinoline oligoamides displaying these side chains were synthesized as a case study. Crystals were obtained from aqueous medium in all cases but one, exceedingly soluble in water. At last, efforts were made to construct self-assembled aromatic helix bundles in water based on hydrophobic effects and electrostatic interactions. NMR and crystallographic studies indicated that hydrophobic effects are weaker than expected and not strongly conducive of aggregation.

Chapter 1. Supramolecular Gels and their Applications Supramolecular gels are viscoelastic materials composed of a solid like three dimensional fibrillary network that is embedded in a liquid. Supramolecular gels are derived from low molecular weight compounds (typically MW < 3000). In the 1990s, the investigations on gels were mainly focused on designing new gelator molecules. However, during the last decade, research focus shifted towards designing functional gels and their applications. As a result of extensive work in this area, gels have been found to have varied applications in the templated synthesis of inorganic nanomaterials, hybrid materials, light harvesting systems, as responsive system and sensors, and also in drug delivery, tissue engineering etc. This chapter gives an introduction to supramolecular hydrogels/organogels and relevant bile acid chemistry touching upon the gelation properties of the bile acid derivatives. Diverse applications of the supramolecular gels are also illustrated with several examples. Scheme 1. Various applications of functional supramolecular gels Chapter 2. Bile Acid derived novel Hydrogelators Part 1. Hydrogelation of Bile acid protected Amino acids and Hybrid Materials Hydrogels from low molecular weight molecules have significant importance in biomedical applications. In this chapter, we report injectable hydrogel formation from bile acid conjugates of various amino acids. Hydrogel formation was found to be dependent on multiple factors such as bile acid backbone structure, linkage between the bile acid and the amino acid, pH etc. Single crystal structures of lithocholyl phenylalanine, lithocholyl-glycine, lithocholyl-L valine and lithocholyl-L alanine were also determined. Finally, the hydrogel frameworks were utilized to produce hybrid materials with Gold and ZnO nanoparticles. Scheme 2. (a) Crystal structure of LC-LF-OH gelator molecule, (b) photograph of gel, (c) SEM and (d) AFM image of LC-LF-OH xerogel Part 2. Hydrogelation of bile acid-dipeptide conjugates and in situ synthesis of silver and gold nanoparticles in the hydrogel matrix Fabricating supramolecular hydrogels with embedded metal nanostructures are important for the design of novel hybrid nanocomposite materials for diverse applications such as bio sensing and chemo sensing platforms, catalytic and antibacterial functional materials etc. Supramolecular self-assembly of bile acid-dipeptide conjugates have led to the formation of new supramolecular hydrogels. Gelation of these molecules depends strongly on the hydrophobic character of the bile acids. Ag+ and Au3+ salts were incorporated in the hydrogels, and photo reduction and chemical reduction led to the in situ generation of Ag and Au NPs in these supramolecular hydrogels without the addition of any external stabilizing agent. The color, size and shape of silver nanoparticles formed by photo reduction depended on the amino acid residue on the side chain. Furthermore, the hydrogel-Ag nanocomposite was tested for its antimicrobial activity. Scheme 3. Bile acid based dipeptide hydrogelators and soft hybrid materials Chapter 3. Sonogels of bile salts of In(III): use in the formation of self-templated indium sulfide nanostructures In this chapter, facile hydrogel formation by Indium(III) cholate and deoxy cholate are reported. When In(III) solution was added to aqueous solutions of sodium cholate and sodium deoxy cholate and sonicated, the mixtures formed gels. The gels thus obtained were translucent/turbid and thermos irreversible. Rheological measurements showed that all of them could be classified as viscoelastic soft solids. Scanning electron microscopy and atomic force microscopy showed typical entangled three dimensional fibrous networks. The In-Ch hydrogel were further used to prepare nanostructured In2S3 in which the cholate units possibly acted as a surfactant to confine the growth of the Nano flakes. Scheme 4. In-Ch hydrogel (Photograph and SEM image of In-Ch gel) Chapter 4. Palladium-Hydrogel Nanocomposite for C-C Coupling Reactions Supported metallic nanoparticles are important composite materials owing to their enormous potential for applications in various fields. This chapter describes the in situ formation of palladium nanoparticles in a calcium-cholate (Ca-Ch) hydrogel by reduction with sodium cyan borohydride. The hydrogel matrix appeared to assist the controlled growth as well as stabilization of palladium nanoparticles. The palladium nanoparticle/Ca-Ch hydrogel hybrid was characterized by scanning and transmission electron microscopy, atomic force microscopy, X-ray diffraction, and energy-dispersive X-ray spectroscopy. Furthermore, PdNP/Ca-Ch hybrid xerogel was shown to act as an active catalyst for Suzuki reaction under aqueous aerobic conditions, up to 4 cycles. This PdNP/Ca-Ch xerogel retained its catalytic activities on storage for several months. Scheme 5. Palladium-hydrogel nanocomposite for C-C coupling reactions in water Chapter 5. Sensitization of Terbium/Europium in self-assembled cholate hydrogel: An approach towards the detection of amine vapours "Luminescent" lanthanides have intrinsic low molar absorptivity, although this problem can be addressed by complexing the lanthanide ion with suitable chelating ligands which improve the luminescence properties drastically. However the design of such systems often involves careful planning and laborious synthetic steps. It is therefore desirable to have a simpler way to sensitize lanthanides with high efficiency. It was observed in our group that trivalent lanthanides formed hydrogels on the addition of sodium cholate. This chapter describes the discovery of the several biphenyl derivatives (such as 4-biphenylcarbaxaldehyde, 4-acetylbiphenyl) for sensitization of Tb(III) and Eu(III) in lanthanide hydrogels. Sensitization of Tb(III) and Eu(III) were observed by doping was characterized by scanning and transmission electron microscopy, atomic force microscopy, X-ray diffraction, and energy-dispersive X-ray spectroscopy. Furthermore, PdNP/Ca-Ch hybrid xerogel was shown to act as an active catalyst for Suzuki reaction under aqueous aerobic conditions, up to 4 cycles. This PdNP/Ca-Ch xerogel retained its catalytic activities on storage for several months. Scheme 6. Schematic representation of the sensitization process (the arrangement of themolecules in the gel fiber is arbitrary)(For figures pl refer the abstract pdf file)

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Ce travail décrit la synthèse et les propriétés gélifiantes de nouveaux dérivés d’acides aminés de faible poids moléculaire dans des solvants organiques ainsi que l’élaboration d’aérogels correspondants par séchage au CO2 supercritique. Nous avons pu montrer, dans notre cas, que seuls les dérivés de la leucine et de la phénylalanine étaient nécessaires au phénomène de gélation. L’étude des paramètres des solvants a permis de montrer que les paramètres de Hansen h des solvants gélifiés s’inscrivaient dans un domaine étroit de valeurs faibles ; il inclut des solvants aromatiques et des solvants chlorés. L’usage de spectroscopies diverses (IR, RMN, dichroïsme circulaire et fluorescence) a permis de mettre en évidence les interactions responsables du phénomène de gélation. Les liaisons hydrogène permettent l’empilement unidimensionnel des molécules gélatrices, ces empilements s’associent ensuite grâce à des interactions de - stacking intercolonnaires. Des aérogels monolithiques ont pu être obtenus. Ils présentent des propriétés remarquables parmi lesquelles une conductivité thermique sous vide extrêmement faible / This work describes the synthesis and the gelation properties of new amino-acid-based low-molecular-weight derivatives in organic solvents as well as the development of the corresponding aerogels by supercritical CO2 drying. We have proved that in our case the presence of phenylalanine or leucine lateral chains were necessary for gelation. A solvent parameters study led us to define a favourable narrow h Hansen parameter domain for gelation including aromatic and chlorinated solvents. The use of several spectroscopy methods (IR, NMR, circular dihroism and fluorescence) allowed to settle the interactions accountable for gelation phenomenon. Hydrogen bonds enable the unidimensional stacking-up of gelator molecules; next, the stacking-up are associated through intercolumnar - stacking interactions. Monolithic aerogels were obtained. They display noteworthy properties among them an extremely low thermal conductivity under vacuum

É sabido que: (i) as sínteses individual e múltipla (manual e automática), bem como a construção de bibliotecas e micro-arranjos de peptídeos sintéticos empregam a metodologia da fase sólida (SPFS); (ii) apesar do desenvolvimento atual desta metodologia sintética, os químicos de peptídeos continuam se deparando com problemas e limitações inerentes a ela; (iii) muitos trabalhos relatam a sua agilização pelo uso de altas temperaturas, mas poucos revelam preocupação com a integridade quiral dos peptídeos-alvo. Portanto, o presente trabalho objetivou dar continuidade à nossa investigação pioneira das: 1) incidência da enantiomerização dos aminoácidos e/ou de outras reações secundárias nas SPFS passo a passo de peptídeos a 60 °C; 2) viabilidade de realização de todas as etapas da síntese convergente em fase sólida (SCPFS) a 60 °C. Em relação ao tópico 1, os peptídeos-alvo escolhidos tinham tamanho e sequência variáveis que incluiam os aminoácidos trifuncionais problemáticos Cys, Ser, His, Met e Trp. Todos eles foram obtidos por SPFS passo a passo convencional e a 60 °C usando aquecimento convencional e micro-ondas. A identificação e a quantificação dos isômeros contaminantes foram feitas com a ajuda de padrões resultantes da SPFS passo a passo convencional e de métodos analíticos (RP-HPLC, LC-ESI/MS, CE e análise quiral de aminoácidos) em condições estabelecidas por nós. Foi constatado que: (i) as nossas condições de acoplamento são mais econômicas que as usuais, pois empregam menor concentração e excesso molar de N-acil-aminoácidos; (ii) nelas, a SPFS a 60 °C usando o aquecimento convencional é simples, prática, de custo relativamente baixo, demanda ½ do tempo da SPFS convenciona e não compromete significativamente a integridade quiral dos aminoácidos; (iii) nas condições similares, a SPFS passo a passo a 60 °C assistida por micro-ondas é mais rápida (realizada em ¼ do tempo gasto na SPFS convencional), porém mais cara e acompanhada de aumento significativo da enantiomerização da Cys; (iv) a mistura 25% DMSO/tolueno, nunca antes utilizada na SPFS assistida por micro-ondas, favoreceu a formação de contaminantes contendo Met oxidadas a sulfóxidos durante as sínteses do fragmento CCK24-33NS, mas o mesmo ocorreu quando DMF foi usado nas sínteses da CCK-33 NS; (v) outras reações secundárias típicas da SPFS passo a passo não foram intensificadas significativamente nas nossas condições sintéticas a 60 °C. Quanto ao tópico 2, foi escolhida a CCK-33 NS como modelo peptídico. Foi constatado que: (i) a etapa de obtenção dos fragmentos peptídicos protegidos que atuariam como doadores de acila (D.A.) e aceptor de acila (A.A.) de partida pode ser ágil e bem sucedida pela SPFS passo a passo a 60 °C nas nossas condições experimentais usando o aquecimento convencional; (ii) DMF, NMP e 25% DMSO/Tolueno foram adequados à solubilização dos fragmentos D.A. e dos reagentes necessários à sua ativação a 60 °C; (iii) a 60 °C, tais solventes também intumesceram satisfatoriamente a CCK24-33NS-resina Rink amida, A.A. de partica; (iv) o aquecimento convencional permitiu que algumas reações de acoplamento entre os D.A. e A.A. escolhidos fossem realizadas com sucesso; na maioria dos casos em que isso não ocorreu, o uso combinado das micro-ondas e agitação sob atmosfera inerte mediaram a formação do produto desejado; (v) a natureza dos fragmentos D.A. e A.A. é fator limitante na SCPFS, mesmo a 60 °C e usando o aquecimento convencional ou as micro-ondas, e, portanto, ele precisa ser melhor estudado. / It is well known that: (i) individual and multiple peptide syntheses (manual and automatic) as well as construction of synthetic peptide libraries and micro-arrays are all based on solid phase chemistry (SPPS); (ii) despite the current development of such synthetic methodology, peptide chemists are still facing its problems and inherent limitations; (iii) many previous works employed high temperatures to accelerate stepwise SPPS, but only a few showed concern about the preservation of the chiral integrity of the target peptide. Therefore, the main goal of the present work was to continue our pioneering investigation of: 1) the incidence of amino acids enantiomerization and/or other side-reactions in the stepwise SPPS at 60°C, 2) the viability of performing all steps of the convergent synthesis on a solid support (CSPPS) at 60°C. With regard to the topic 1, the peptides chosen as targets had variable size and sequence, which included the tricky trifunctional amino acids Cys, Ser, His, Met and Trp. The peptides were synthesized by conventional stepwise SPFS and at 60 °C using conventional or microwave heating. Identification and quantification of the contaminant isomers was done with the aid of standards resultant from conventional stepwise SPPS and of analytical methods (RP-HPLC, LC-ESI/MS, CE and chiral amino acids analysis) in conditions established in our laboratory. It was shown that: (i) our coupling conditions are cheaper than the usual ones as they employ lower concentration and excess of N-acyl-amino acids; (ii) under them, stepwise SPPS at 60 °C using the conventional heating is simple, practical, relatively low-cost, demands half of the time required by conventional stepwise SPPS and does not cause the enhancement of amino acids enantiomerization; (iii) under similar conditions, microwave-assisted stepwise SPPS at 60 °C is faster (it demands only one-fourth of the time spent in the conventional stepwise SPPS), but it is more expensive and causes significant damage specially to the chiral integrity of Cys; (iv) the binary mixture 25% DMSO/toluene, never used earlier in microwave-assisted stepwise SPPS, led to the formation of contaminants with Met oxidized to its sulfoxides during the synthesis of CCK24-33NS; however, it also occurred when DMF was used in the synthesis of CCK-33 NS; (v) other side reactions typical of stepwise SPPS were not significantly intensified under our conditions at 60 °C. Concerning to the topic 2, CCK-33 NS was chosen as the model peptide. It was shown that: (i) the synthesis of the protected peptides that would act as acyl donor (A.D.) or as the starting acyl aceptor (A.A.) can be fast and successfully achieved at 60 °C under our experimental conditions using conventional heating; (ii) DMF, NMP and 25% DMSO/toluene dissolved all A.D. and the reagents required for their activation at 60 °C; (iii) at this temperature, such solvents were also able to properly swell CCK24-33NS-Rink amide resin, the starting A.A.; (iv) the conventional heating allowed for some coupling reactions between A.D. and A.A., but in most cases in which it did not occur, the combined use of microwaves and stirring under inert atmosphere mediated the formation of the desired products; (v) the nature of fragments A.D. and A.A. is a limiting factor in the CSPPS even at 60 °C and using the conventional or microwave heating; therefore, it should be further studied.