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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
31

Methods for protein analysis by capillary electrophoresis and mass spectrometry

Romson, Joakim January 2018 (has links)
Protein analysis is important to understanding biological systems, but sample diversity necessitates a multitude of analysis techniques and methods. Challenges that are addressed include analysis of low abundance samples, fractionation to reduce sample complexity, and automation to reduce time and cost. Matrix assisted laser desorption/ionization mass spectrometry (MALDI-MS) is an important technique for protein characterization. In Paper I, the sensitivity of MALDI-MS was enhanced through the fabrication of a hydrophobic coating for the MALDI target plate, yielding analyte concentration. The plate outperformed a commercial concentration plate. Capillary electrophoresis (CE) separation offers low sample consumption and high efficiency, and in Paper II, offline CE-MALDI-MS fractionation was employed. A robot system for automation was constructed and used in analysis of spermatophore proteins from the butterfly Pieris napi. The robot was also used in automated on-target trypsin digestion under a lid of liquid fluorocarbons, a simpler and cheaper alternative to controlled humidity chambers. An indication of indigenous proteolysis of the sample was seen. Electrospray ionization (ESI) is the other technique for protein analysis in MS. In Paper III, the biomarker protein osteopontin (OPN) was analyzed by ESI-MS in order to find suitable conditions for its detection. A preliminary optimization of solvents and ionization conditions was done, and tandem MS (MSn) performed to increase the reliability of identification. / Proteinanalys är viktigt för att förstå biologiska system, men mångfalden av prov kräver en mängd olika analystekniker och metoder. Utmaningar som tas upp inkluderar analys av små provmängder, fraktionering för att minska provkomplexiteten, och automatisering för att minska tidsåtgång och kostnad. Matris-assisterad laserjoniserings-masspektrometri (MALDI-MS) är en viktig teknik för proteinkarakterisering. I Artikel I förbättrades känsligheten i MALDI-MS genom tillverkning av en hydrofob beläggning på MALDI-provplattan, vilket gav en koncentration av provet. Provplattan gav bättre resultat än en kommersiell koncentrationsprovplatta. Kapillärelektroforesseparation (CE) har låg provåtgång och hög separationseffektivitet och i Artikel II användes offline CE-MALDI-MS-fraktionering. Ett robotsystem för automatisering konstruerades och användes för analys av spermatoforproteiner från fjärilen Pieris napi. Roboten användes även i automatiserad trypsinklyvning under en yta av en flytande fluorkolförening, ett billigare alternativ tilli nkubationskammare med kontrollerad luftfuktighet. En indikation på naturlig enzymatisk proteinklyvning i provet hittades. Elektrospray jonisering (ESI) är den andra tekniken för proteinanalys i MS. I Artikel III analyserades biomarkören osteopontin (OPN) med ESI-MS för att hitta lämpliga förhållanden för dess detektion. En preliminär optimering av lösningsmedel och jonisationsförhållanden gjordes, och tandem-MS (MSn) utfördes för att öka identifikationens tillförlitlighet. / <p>Full text will not be uploaded due to unpublished results. QC 20181121</p>
32

Limites et potentiels de la polymérisation radicalaire par ouverture de cycle pour la synthèse de polyesters / Limits and Potential of the Radical Ring-Opening Polymerization for the Synthesis of Polyesters

Tardy, Antoine 18 April 2014 (has links)
La Polymérisation Radicalaire par Ouverture de Cycle (R-ROP) est une méthode de synthèse de polymères contenant des fonctions chimiques de choix dans le squelette carboné grâce à un mécanisme d'addition-fragmentation. L'utilisation de monomères spécifiques, les Acétals Cétènes Cycliques (CKA), permet dans certaines conditions l'obtention de polyesters aliphatiques dont la propriété de (bio)dégradation présente de nombreuses applications. Cette méthode relativement peu étudiée depuis les années 1980 présente un fort potentiel mais également de nombreuses limites. Ce travail de thèse a consisté à comprendre l'origine de ces limites pour tenter d'y apporter des solutions, grâce à une approche combinée expérience-théorie.Nous avons montré que l'obtention exclusive de polyesters découle d'une compétition cinétique et que le comportement des différents monomères s'explique par des interactions orbitalaires dépendant de la géométrie, la flexibilité et la substitution des cycles. D'autre part, nous avons mis en évidence l'extrême difficulté de propagation des monomères propageant via des radicaux stabilisés par des cycles aromatiques. Cette faible réactivité inhérente à la double liaison riche en électrons des CKA est également la cause de l'incorporation restreinte des monomères cycliques en copolymérisation avec des monomères vinyliques usuels. La rationalisation de la copolymérisation a été mise à profit pour réaliser des copolymérisations de type statistique et alternée. Enfin, l'étude du contrôle de la R-ROP par les nitroxydes a montré la présence de réactions secondaires propres à ce système et permettant actuellement un contrôle partiel de la polymérisation. / The Radical Ring-Opening Polymerization (R-ROP) is a synthetic pathway to introduce chemical functions into a polymer backbone due to an addition-fragmentation mechanism. Using specific monomers like Cyclic Ketene Acetals (CKA) in the right conditions allows preparing aliphatic polyesters which have numerous applications thanks to their (bio)degradability. This method has been quite faintly investigated since the 1980s and even if it has a great potential, it suffers of numerous limitations. This PhD work consisted in the understanding of those limitations to try bringing solutions to them, with a combined approach of experiments and theory.We first demonstrated that the exclusive preparation of polyesters comes from a kinetic competition. The behavior of the distinct monomers is explained by orbital interactions depending on the geometry, flexibility and substitution of the cycles. Then, we highlighted the extremely difficult propagation of the monomers propagating with stabilized aromatic radicals. This low reactivity inherent to the electron-rich double link of the CKAs is also the cause of low polyester introduction during the copolymerization with usual vinyl monomers. We took advantage of the CKA copolymerization rationalization to realize statistical and alternate copolymerizations. At last, the study of the nitroxide mediated R-ROP demonstrated the occurrence of side reactions characteristic of this system that allow at present a partial control of the polymerization.

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