A series of pyrene end-labeled monodisperse poly(ethylene oxide)s (PEO(X)-Py2 where X represents the number average molecular weight (Mn) of the PEOs and equals 2, 5, 10 and 16.5 K) and one pyrene mono-labeled PEO (PEO(2K)-Py1) were synthesized and characterized in solution using fluorescence. First, the end-to-end cyclization (EEC) of PEO(X)-Py2 was investigated in seven organic solvents with viscosities (η) ranging from 0.32 to 1.92 mPa•s. The classical Birks scheme was used to globally fit the pyrene monomer and excimer fluorescence decays. The fraction of pyrenes that did not form excimer (ffree) was found to increase with increasing η and Mn. This result was contrary to the assumptions made by Birks’ scheme. To account for this, ffree was assumed to represent the fraction of PEO chains other than the monolabeled polymer impurities that cannot accomplish EEC. A fluorescence blob model (FBM) was applied to handle this assumption in the process of excimer formation for the PEO(X)-Py2 samples in solution. The radius of a blob, Rblob, in organic solvents was determined according to the results retrieved from the FBM. To quantitatively account for the existence of pyrene impurity in pyrene-labeled macromolecules, known amounts of PEO(2K)-Py1 were added into a PEO(2K)-Py2 solution and the fluorescence decays were fitted globally according to the Birks scheme and “model free” (MF) analysis to verify the validation of the MF analysis. The MF analysis was then applied to determine the amounts of 1-pyrenebutyric acid (PyBA) that had been added to a solution of pyrene end-labeled fourth generation dendritic hybrid (Py16-G4-PS). The results demonstrated that the contribution from unwanted fluorescent species could be isolated and quantitatively accounted for by fitting the fluorescence decays of the pyrene monomer and excimer globally with the MF analysis. Since the PEO(X)-Py2 samples form hydrophobic pyrene aggregates in aqueous solution, a sequential model (SM) was proposed to characterize the pyrene excimer formation of PEO(X)-Py2 in water at different polymer concentration (CP). The capture distance over which the pyrenyl end-groups experience hydrophobic forces in water was determined by assuming that the end-to-end distances of the PEO(X)-Py2 samples adopt a Gaussian distribution and that the fraction of pyrenes that are aggregated (fE0) determined by the sequential model corresponds to the fraction of PEO(X)-Py2 chains whose end-to-end distance is smaller than the hydrophobic capture distance. Since a surfactant can interact with a hydrophobically modified water-soluble polymer in aqueous solution, the interactions taking place between PEO(X)-Py2 and sodium dodecyl sulfate (SDS) were investigated at a low PEO(X)-Py2 concentration. The pyrene monomer and excimer fluorescence decays of the PEO(X)-Py2 and SDS solutions were acquired at various SDS concentrations and globally fitted according to the MF analysis to retrieve the parameters that described the kinetics of pyrene excimer formation. At high SDS concentrations above the critical micelle concentration (CMC), the pyrene end-groups of the short-chain samples (PEO(2K)-Py2 and PEO(5K)-Py2) were incorporated inside the same micelle and excimer was formed intramolecularly, while most pyrene groups of the long-chain samples (PEO(10K)-Py2 and PEO(16.5K)-Py2) were isolated into different micelles. Lastly, both the rheological properties and fluorescence behavior of a pyrene-labeled hydrophobically-modified alkali-swellable emulsion (Py-HASE) polymer in basic aqueous solution with SDS were studied. Furthermore, a joint experimental setup that combined a rheometer and a steady-state fluorometer was applied to investigate at the molecular level the effect that a shearing force had on the polymeric network. However, despite the dramatic decrease in solution viscosity with increasing shear rate, no change in the fluorescence spectra was detected, suggesting that changes in the polymeric network that affected the balance of intra- versus intermolecular pyrene associations did not impact the process of excimer formation. Together the experiments described in this thesis represent the broadest set of examples found in the scientific literature where information on the dynamics and level of association of pyrene-labeled polymers has been retrieved through the quantitative analysis of the fluorescence decays acquired with pyrene-labeled polymers in solution.

The biological function of a protein is determined by its amino acid sequence, structure, and internal dynamics. In turn the prediction of a protein structure from its folding pathway involves the characterization of the dynamics of the polypeptide backbone. This study addresses how the internal dynamics of arborescent polypeptides are affected by increased crowding of the interior of these branched polymer molecules. Linear, comb-branched, and arborescent poly(L-glutamic acid) (PGA) samples were analyzed by 1H NMR spectroscopy to determine their chain conformation. The PGA chains of these constructs were shown to adopt α-helical and random coil conformations in N,N-dimethylformamide (DMF) and in dimethyl sulfoxide (DMSO), respectively. The hydrodynamic diameter (Dh) of the arborescent PGAs, determined using dynamic light scattering measurements, increased with increasing generation number and when the side-chains adopted random coil instead of α-helical conformations. The PGA samples were labelled with 1-pyrenemethylamine to determine how their structure affected the internal dynamics of the arborescent polymers in solution, from the analysis of their fluorescence spectra and decays. For each pyrene-labelled polymeric construct excimer formation increased with increasing pyrene content, and the efficiency of excimer formation increased with the generation number due to the increased density of the macromolecules. Comparison of the time-resolved fluorescence results acquired in DMF and in DMSO demonstrated that the helical conformation led to slower chain dynamics in DMF and that despite the higher viscosity of DMSO, the polypeptide side-chains were more mobile as a consequence of the random coil conformation of the linear PGA segments. These results suggest that the formation of structural motives inside a polypeptide slows down its internal dynamics.

Thesis (Ph. D.)--University of Oregon, 2007. / Typescript. Includes vita and abstract. Includes bibliographical references (leaves 143-151). Also available for download via the World Wide Web; free to University of Oregon users.

Thesis (Ph. D.)--University of Texas at Austin, 2007. / Vita. Includes bibliographical references.

Les nanoparticules inorganiques luminescentes ont suscité un intérêt croissant au cours des dernières décennies, notamment pour leur application en imagerie biologique. Un certain nombre d’entre elles présentent toutefois des limitations telles que toxicité, absence de biodégradabilité, faible brillance, clignotements…. Dans cette optique, les nanoparticules fluorescentes à base de petites molécules organiques (FONs) offrent une solution alternative prometteuse aux nanoparticules inorganiques pour l'imagerie biologique. Le principal défi réside dans l'élaboration des nanoparticules organiques possédant une brillance élevée, une bonne stabilité dans l'eau (y compris en milieu biologique), une bonne biocompatibilité ainsi qu'une émission accordable dans le visible et au-delà dans le proche infrarouge (pour une détection plus aisée en milieu diffusant). Dans cette optique, nous avons utilisé une stratégie basée sur l’utilisation de chromophores dipolaires de type "push pull" « adaptés ». Au cours du travail, la synthèse de séries de chromophores homologues bâtis sur le même système conjugué et ayant en commun un groupe donneur de type triphénylamine (destiné à préserver les propriétés de luminescence) présentant ou non des motifs encombrants positionnés a été réalisée. Les nanoparticules correspondantes ont été préparées selon un protocole classique, simple et rapide à mettre en oeuvre (précipitation). L’étude des propriétés photophysiques des nanoparticules organiques fluorescentes ainsi obtenues a été réalisée et mise en perspective avec celles des chromophores en solution dans des solvants organiques de polarité variable. Une étude systématique de l’évolution dans le temps des propriétés optiques des nanoparticules organiques a été réalisée permettant de mettre en lumière des relations entre la structure des sous-unités chromophoriques et la stabilité colloïdale et « optique » des nanoparticules. Ces études ont permis d’identifier des nanoparticules émettant dans le proche infrarouge extrêmement brillantes et présentant une stabilité colloïdale remarquable dans l’eau, une photostabilité accrue et une très bonne biocompatibilité. De ce fait, ces nanoparticules ont pu être utilisées avec succès dans l'imagerie biologique des cellules et le suivi (tracking) à l'échelle de la particule unique, démontrant l'intérêt de la démarche d'ingénierie mise en oeuvre. / During the last decades, luminescent inorganic nanoparticles have attracted a large interest in different fields including biological imaging. However, a number of them have drawbacks such as toxicity and absence of biodegradability. Recently, molecular-based fluorescent organic nanoparticles (FONs) have emerged as a promising alternative to inorganic nanoparticles for bioimaging. The main challenge lies in the elaboration of organic nanoparticles that combine large brightness, good colloidal stability in biological environments) and biocompatibility as well as NIR emission (to allow improved detection in thick tissues). To achieve this objective, we have implemented a molecular engineering strategy based on dedicated polar and polarizable "push pull" chromophore built from a triphenylamine donor moiety and a specific pi-conjugated system. The corresponding nanoparticles were readily prepared by the reprecipitation method. In the present manuscript, the synthesis of the chromophores and the preparation and characterization of the organic fluorescent nanoparticles is described. A comprehensive investigation of their photophysical properties and study of their colloidal stability is presented allowing to derive structure-property relationships. The implemented study led to innovative NIR-emitting nanoparticles combining large brightness (superior to those of QDs and NIR-emitting organic dyes), remarkable colloid stability and suitable photostability. These nanoparticles have been successfully used for single particle tracking and imaging in cells, while no toxic effect was observed.

Conçues à partir d’une approche rationnelle, nous avons créé de nouvelles sondes membranaires permettant l’imagerie de l’organisation de la membrane plasmique cellulaire. Dans ce travail, nous avons d’abord développé un groupe d’outils, à partir du fluorophore solvatochrome Nile Red et de Black Hole Quencher-2, capable de marquer spécifiquement les domaines ordonnés et désordonnés (radeaux) en les identifiant par leur couleur d’émission. Les études cellulaires, à l’aide de ces sondes, suggèrent que la membrane plasmique est composée de deux phases distinctes. Puis dans le but de créer de nouvelles sondes basées sur Nile Red compatibles avec le sérum et fixables par formaldéhyde/glutaraldéhyde, nous avons modifié la sonde, préalablement développée, NR12S avec un groupement PEG ou amino, respectivement. Etonnamment, la sonde PEGylée est rapidement internalisée dans la cellule et le dérivé animo agrège avec l’agent fixant. D’un autre côté,basée sur Nile Red, nous avons conçu une sonde capable de détecter un récepteur donné et de visualiser son environnement lipidique. Initialement, nous avons obtenu des sondes capables d’allumer leur fluorescence en se liant sur le RCPG à l’ocytocine. Puis, nous avons conjugué NR12Spar l’intermédiaire d’un espaceur PEG(12) au ligand de l’intégrine, RGD. Les résultats préliminaires montrent que la molécule peut se lier à la membrane et détecter l’ordre lipidique, cependant les études cellulaires nécessitent un achèvement. Nous avons aussi travaillé sur des sondes membranaires fluorogéniques (turn-on) pour de l’imagerie multi-couleurs. Basées sur le fluorophore3-méthoxychromone, nous avons obtenu des sondes plus brillantes et plus photostables que la sonde développée originellement à partir de 3-hydroxychromone (F2N12S). Grâce à l’important déplacement de Stokes, elles permettent une imagerie de la membrane cellulaire avec une autofluorescence minimale dans la région spectrale bleue, compatible avec les marqueurs communs verts et rouges. Pour finir, basées sur le fluorophore squaraine, nous avons développé trois nouvelles sondes opérant dans la région rouge lointain, qui est particulièrement intéressante pour l’imagerie in vitro et in vivo. Ces sondes montrent une orientation parallèle avec la membrane lipidique, alors que les expériences cellulaires indiquent que seule la sonde avec deux ancres lipidiques est capable de marquer de façon stable la membrane plasmique. Ces sondes développées ici sont prévues pour être utilisées dans la recherche des radeaux lipidiques aussi bien que pour l’imagerie super-résolution et multi-couleurs de cellules vivantes. / Based on rational molecular design, we design new membrane probes that enable fluorescence imaging of cell plasma membrane organization. In this work, we first synthesized a toolkit, based on solvatochromic Nile Red dye and Black Hole Quencher-2, that can stain specifically ordered and disordered lipid domains (rafts) and identify them by the emission color. Cellular studies with these probes suggested that the plasma membrane is composed of two distinct phases. Then,with the idea to make Nile Red-based probes compatible with serum medium and fixable by formaldehyde/glutaraldehyde, we modified previously developed probe NR12S with PEG and aminogroups, respectively. Surprisingly, the PEGylated probe is quickly internalized inside the cell and the amino-derivative aggregates with the fixing agent. On the other hand, based on Nile Red we designed probes capable to detect a given receptor and visualize its lipid environment. Initially, we obtained probes that can turn-on fluorescence on binding to the oxytocin GPCR receptor. Then, we conjugated NR12S through a PEG(12) spacer to the ligand of intergrin, RGD. The first data show that the molecule can bind to the membrane and detect the lipid order, though cellular studies have to be completed. We also worked on fluorogenic (turn-on) membrane probes for multi-color imaging. Based on blue 3-methoxychromone dyes, we obtained probes that are brighter and more photostable than the originally developed probe based on 3-hydroxychromone (F2N12S). Due to large Stocks shift, they enabled cell membrane imaging with minimal auto-fluorescence in the blue spectral region, compatible with common green and red probes. At the end, based on squaraine fluorophore, we developed three new probes operating in the far red region, which is also very interesting for in vitro and in vivo imaging. These dyes show a parallel orientation with the lipid membrane, while the cellular experiments point out that only the probe with two anchor groups is able to stain stably the plasma membrane. The probes developed here are expected to be used for lipid rafts research as well as for super-resolution and multi-color imaging of living cells.

La production et l’utilisation croissante des nanomatériaux et nanoparticules (NP) dans de nombreux secteurs d’activité conduisent inévitablement à un relargage de NP dans l'environnement et notamment dans les eaux, devenant ainsi une pollution émergente dans le schéma de la production d’eau potable. Les membranes d’ultrafiltration (UF) semblent présenter un réel potentiel de rétention envers les NP du fait de leur taille de pores proche de 20 nm. La filtration de NP fluorescentes de diamètre 100, 10 et 1,5 nm en suspensions, seules ou en mélange, a été étudiée. Les tailles des NP ainsi sélectionnées permettent de travailler avec des dimensions plus grande, plus petite et du même ordre de grandeur que la taille des pores. La considération de la concentration des flux de la filtration en nombre de NP a permis d’estimer le nombre de NP bloquées sur et/ou dans la membrane. Une méthodologie précise et fiable permettant la localisation de ces NP bloquées a été consolidée par une précision de mesure plus importante grâce à une caractérisation multi-échelle. Des profils de pénétration des NP fluorescentes dans la membrane ont été réalisés grâce à une imagerie au Microscope Confocal à Balayage Laser (MCBL). L’application des modèles de colmatage aux données expérimentales a montré une bonne adéquation avec la localisation microscopique des NP et les résultats expérimentaux obtenus. L’influence des conditions opératoires, de la présence de sel et/ou de la polydispersité de la suspension d’alimentation sur l’établissement et la localisation du colmatage a pu être déterminée, notamment grâce à la mise en place de plan d'expérience. / The increasing use of nanotechnologies and nanoparticles (NPs) in many sectors of activities leads to their inevitable discharge in the environment and thus in water. The properties and the toxicity of these NP are still unclear, so that this new pollution have to be considered in drinking water treatment plant. Ultrafiltration (UF) membranes, which have nanometric pore size, present a great potential for NP retention. The mass transport through the UF membrane of fluorescent NP, with size of 100, 10 and 1.5 nm, has been studied. The influence of operating conditions (transmembrane pressure, volumetric concentration factor) have been treated by experimental design. Consideration of the NP concentration in number in each flux thanks to a Nanosight NS300, allowed to estimate the number of NPs blocked on and/or into the membrane. An accurate and reliable methodology allowed the localization of the fouling by Confocal Laser Scanning Microscopy (CLSM). Application of fouling models to experimental data showed good agreement with microscopic NP location. The influence of the operating conditions, the salinity and/or the polydispersity of the feed suspension on the fouling establishment and fouling profile has been studied.

La perfusion intestinale est un facteur clé dans la guérison des anastomoses digestives. L'évaluation clinique de la perfusion demeure subjective et s’avère insuffisante pour prédire les complications anastomotiques, quelle que soit l'expérience du chirurgien. La vidéographie par fluorescence peut fournir une évaluation en temps réel de la perfusion intestinale. Suite à l'administration systémique d'une substance fluorescente, il est possible d’évaluer les unités de fluorescence, lorsque la zone d'intérêt est illuminée par une source de lumière spécifique. La présence de fluorescence est un marqueur de perfusion tissulaire mais cela reste une donnée subjective. Pour obtenir des données quantitatives, nous avons mis au point une solution d'analyse assistée par ordinateur permettant l'évaluation intraopératoire du site optimal de résection. Le logiciel peut générer une cartographie virtuelle de perfusion et celle-ci peut être superposée aux images laparoscopiques en pour obtenir l'effet de réalité augmentée. Cette thèse décrit le développement progressif de ce concept. / Pre-anastomotic bowel perfusion is a key factor for a successful healing process. Clinical evaluation of bowel perfusion is subjective and not accurate at predicting anastomotic complications, irrespective of the surgeon’s experience. Fluorescence videography analysis can provide real-time evaluation of bowel perfusion. Following the systemic administration of a fluorescent substance, fluorescence videography can compute the units of fluorescence, when the area of interest is illuminated by a specific light source. Fluorescence intensity is a marker of blood supply to the tissue. However, fluorescence intensity alone remains a subjective approach. We have developed a software solution enabling quantitative estimation of bowel perfusion, for intra-operative evaluation of the optimal resection site. The software can generate a “virtual perfusion cartography”, which can be overlapped onto real-time laparoscopic images to obtain the Enhanced Reality effect. This thesis describes the stepwise development of such concept.

The aim of this thesis was to determine the response of trees to air pollution in the city. For this purpose, APTI index (Air pollution tolerance index), the measurement of chlorophyll fluorescence and the analysis of the content of photosynthetic pigments were used. The examined trees were Platanus acerifolia and Prunus subhirtella v. autumnalis rosea growing in Lannova třída in České Budějovice. For evaluation of APTI index, four parameters are required. These are the quantity of ascorbic acid and the photosynthetic pigments in the leaves, the relative water content in the leaves and the leaf extract pH. The samples of leaves were taken in August 2015. From Lannova třída, three trees of each species were chosen. This research has showed that in terms of evaluation APTI index of Platanus acerifolia is more tolerant and less sensitive to air pollution than Prunus subhirtella. The value of the APTI index at the Platanus acerifolia was 9,33, at the Prunus subhirtella APTI index was 7,84. During the evaluation of the maximum photochemical efficiency of photosystem II, statistically significant differences between the trees were not found. The values obtained via the method of chlorophyll fluorescence at both tree species were low, at the Platanus acerifolia it was 0,65, at the Prunus subhirtella it was 0,68. In terms of content of photosynthetic pigments, Prunus subhirtella has come out better. In almost all, it had greater content of pigments than the Platanus acerifolia. The results of the research were used in the draft for excursions for students of 7th grades of elementary schools to this area.

Ce travail de recherche consiste à proposer une fonctionnalisation des surfaces textiles par le procédé sol-gel à partir d’alcoxysilanes. L‘objectif est de conférer aux textiles des propriétés hydrofuges, ignifuges et fluorescentes. La première partie de notre étude traite dans un premier temps les paramètres de synthèse des sols et leur application sur différentes étoffes textiles. Ensuite, nous nous sommes intéressés à la synthèse des sols avec des alcoxydes de silicium permettant d’avoir des propriétés hydrofuges. Par la suite, les propriétés d’adsorption et de non mouillabilité des textiles traités ont été déterminées selon le standard AATCC- 22 et celles mécaniques selon la norme ISO 13934. Nous avons également étudié la stabilité thermique par des analyses ATG et les propriétés de non mouillabilité de ces textiles en fonction de leur photovieillissement accéléré. Dans un deuxième temps et dans l’optique de développer des textiles ignifuges, nous avons montré que l’immobilisation des LIs à la surface des textiles peut être réalisée par le procédé sol-gel. Pour ce faire nous avons tout d’abord synthétisé des liquides ioniques à partir d’un alcoxyde de silicium halogéné. Le sel obtenu est ensuite greffé sur des matrices textiles par le procédé sol- gel.Enfin, la dernière partie de l’étude consiste en la synthèse de fluorophores organiques par un procédé utilisant le couplage de type Heck. Leurs propriétés de photoluminescence ont été étudiées. Nous avons ainsi pu les greffer sur des surfaces pour développer des traceurs textiles fluorescents et des sondes pH fluorescentes. / This work consists to provide a functionalization of textile surfaces by the sol-gel process using alcoxysilanes. The final aim is to impart water repellent, flame retardant and fluorescent features to textile surfaces. Firstly, the effects of synthesis parameters such as the concentration of the alcoxysilanes and the impregnation time of the fabrics in the solution (sol) prepared have been thoroughly investigated. Then, we have prepared a silica modified sol which is expected to induce a water repellent feature to textile. To investigate the water repellency under practical conditions, the common “rain test” was used according to AATCC test method 22-2005 also mechanical properties of treated un untreated textile was studied according to ISO 13934. Furthermore, thermal properties of sol-gel modified textiles were investigated and their water repellency abilities were assessed and compared after photo ageing upon UV irradiations.Secondly, we have showed that the immobilization of ionic liquids onto the surface of textile fabrics can be achieved by the sol-gel process, which is of a growing interest due to its simplicity and flexibility. For this aim, two ionic liquids were synthesized using halogenated alkoxysilane, and then grafted onto the cotton fabrics by sol- gel process .The functionalized cotton fabrics exhibit good water repellency and flame stability. Finally, a fluorophores bearing phenolic hydroxyl and pyridinium end groups have been synthesized and grafted into cellulosic fiber by sol -gel process. This fluorophores have been firstly synthesized by double Heck couplings, and then we have studied their photo physical behavior as function of their concentration both in ethanol and in the textile. We have also grafted the fluorophores in cellulosic textile by sol gel method to have fluorescent and PH-sensitive properties.