Les travaux de recherche présentés dans cette thèse sont orientés vers la compréhension des mécanismes réactionnels propres aux produits pétroliers tels que les lubrifiants et les graisses par analyse thermique. La technique d’analyse thermique principalement utilisée était la calorimétrie de réaction. Dans un premier temps, une étude approfondie de la réponse instrumentale a été conduite. Une nouvelle méthode d’étalonnage a été proposée en se basant sur l’étalonnage par effet Joule. Avec ce type de correction, une étude cinétique plus précise peut alors être conduite, elle permet en outre d’étudier la cinétique de mécanisme complexes comme celles rencontrées avec les produits pétroliers. Dans un second temps, l’étude de la neutralisation de l’acide sulfurique par un lubrifiant marin a été effectuée. Cette étude a permis de développer une méthode d’analyse de routine permettant de comparer l’efficacité d’un lubrifiant à neutraliser une quantité précise d’acide sulfurique. Il devient alors possible d’estimer le temps nécessaire pour neutraliser l’acide afin d’éviter qu’il n’endommage l’intégrité des parties métalliques sensées être protégées par le lubrifiant marin. Une étude cinétique du phénomène de neutralisation, par l’intermédiaire des méthodes isoconversionnelles, a permis d’avoir une indication sur le modèle réactionnel pilotant ce mécanisme. Pour finir, l’influence du choix des paramètres tels que la température, les vitesses de chauffe où les temps de cuisson de graisse, a été étudié par en mêlant des techniques d’analyses thermique et rhéologiques. / The research work presented in this thesis are oriented on the understanding of reactional mechanisms occurring in the field of oil products, such as lubricants and greases, by thermal analysis. The main thermal analysis technique used was the reaction calorimetry. Firstly, the instrumental response study of the calorimeter was led. A new calibration method have been proposed, this method is based on the calibration by Joule effect. From this correction, a more accurate kinetic evaluation can be done in order to study the complex mechanisms as those met with oil products. In a second time, the study of the neutralization of sulfuric acid by a marine cylinder lubricant was carried out. This work allowed to develop a routine analysis method to compare the lubricant ability to neutralize a precise amount of sulfuric acid. The time estimation is important to determine the frequency of lubricant replacement and to avoid the damage on the metallic parts. A kinetic study of the neutralization phenomena using isoconversional kinetic method, allowed to obtain the reactional model. Finally, the influence of parameters choice such as temperature, heating rate or cooking time have been evaluated by thermal and rheological analysis. Greases have been produced in a reaction calorimeter using a stirring system specifically developed. Information obtained allow to choose accurately the parameters used during the manufacturing processes in order to save time and decrease production costs.

Esta tese apresenta resultados da caracterização estrutural de três sistemas de classes diferentes por meio, principalmente, da modelagem de dados de espalhamento de raios X a baixos ângulos (SAXS). No sistema surfactante-surfactante as micelas mistas são formadas por dodecil sulfato de sódio (SDS) e um dos surfactantes da série Tween (Tween 20, 40, 60 e 80). A modelagem adotada impôs vínculos moleculares uma vez que os dados de SAXS estavam em escala absoluta. Isso reduziu a ambiguidade nos valores dos parâmetros ajustáveis e permitiu verificar que os dados de SAXS são satisfatoriamente descritos considerando-se que as micelas são elipsoides de revolução core-shell, podendo ser prolatas ou oblatas dependendo do tipo de Tween empregado. Para o sistema proteína-surfactante, a metodologia experimental utilizada permitiu um estudo estrutural e termodinâmico dos complexos formados por meio do acompanhamento do processo de ligação de SDS às proteínas lisozima e alfa-lactalbumina. A técnica de calorimetria de titulação isotérmica (ITC) forneceu um panorama geral sobre a desnaturação proteica e norteou os experimentos seguintes de SAXS e dicroísmo circular (CD). Por meio da análise dos dados de CD concluiu-se que as proteínas perdem quase totalmente sua estrutura terciária, mas não a secundária (esse estado é conhecido como molten globule). Já a modelagem de SAXS em escala absoluta com imposição de vínculos permitiu concluir que os complexos proteína-surfactante podem ser entendidos micelas decoradas, isto é, a proteína está distribuída sobre a superfície de uma micela de SDS. Esse modelo, aliado à abordagem experimental empregada, permitiu a caracterização sistemática dos complexos durante a desnaturação proteica. Em relação ao sistema lipossomas-(bio)ativos, a análise dos dados de SAXS por meio do Método da Deconvolução Gaussiana usando bicamadas simétricas, para os sistemas Phospholipon 90H curcumina/vitamina D3 e dipalmitoilfosfatidilcolina de soja (DPPC) ácido láurico (LA), e assimétricas, para o caso fosfatidilcolina de ovo (EPC) sumatriptano (SMT), permitiu acompanhar mudanças na estrutura das mesmas ocasionadas pela presença dos (bio)ativos. Verificou-se em todos os casos que a espessura da bicamada se mantém praticamente constante. A flexibilidade membranar aumenta, seja em função da temperatura, para o sistema Phospholipon 90H curcumina/ vitamina D3, seja em função da concentração de (bio)ativos, como nos outros dois casos. Para estes, concluiu-se pela análise dos perfis de contraste de densidade eletrônica que os (bio)ativos interagem preferencialmente com as cabeças polares dos fosfolipídeos que constituem os lipossomas, possivelmente causando defeitos topológicos nessa região e ocasionando o aumento da flexibilidade membranar mencionada antes. LA, diferentemente de SMT, induz uma transição de lipossomas multilamelares para unilamelares, e esse fenômeno é grandemente influenciado pelo pH do meio. / This thesis presents a structural characterization of three systems of different classes through, mainly, the small angle X-ray scattering (SAXS) technique. In the surfactant-surfactant system the mixed micelles are composed by sodium dodecyl sulfate (SDS) and one of the Tween surfactants (Tween 20, 40, 60 and 80). The adopted modeling imposed molecular constraints since the SAXS data was in absolute scale. This procedure reduced the ambiguity in the values of the adjustable parameters and allowed to verify that SAXS data is satisfactorily described considering that the micelles are core-shell revolution ellipsoids which can be prolate or oblate depending on the type of Tween used in the micelle. For the protein-surfactant system, the applied experimental methodology allowed a structural and thermodynamic study of the complexes formed through monitoring the binding of SDS to the proteins lysozyme and alpha-lactalbumin. Isothermal titration calorimetry (ITC) technique provided an overview of proteic denaturation and guided the following experiments of SAXS and circular dichroism (CD). From CD data analysis it was concluded that the proteins lose almost totally their tertiary structure, but not the secondary one (this state is known as \"molten globule\"). On the other hand, SAXS data modeling in absolute scale with molecular constraints leaded to the conclusion that protein-surfactant complexes can be considered decorated micelles in which the protein is distributed over a SDS micelle surface. This model, combined to the adopted experimental procedure, allowed the systematic characterization of the complexes along the protein denaturation. In the liposome-(bio)actives system, the SAXS data analysis using the Gaussian Deconvolution Method assuming symmetric bilayers, for the systems Phospholipon 90H curcumin /vitamin D3 and soybean dipalmitoyl phosphatidylcholine (DPPC) lauric acid (LA), and asymmetric bilayers, in the case of egg phosphatidylcholine (EPC) sumatriptan (SMT) system, allowed to follow changes in the lipid bilayer structure induced by the presence of the (bio)actives. It has been found, in all cases, that the bilayer thickness remains approximately. Membrane flexibility increases, depending on the temperature, for the Phospholipon 90H curcumin /vitamin D3 system, or as a function of the (bio)actives concentrations, as in the other two cases. For those, it was concluded, by the analysis of electron density contrast profiles, that the (bio)actives preferentially interact with the polar heads of the phospholipids forming the liposomes, possibly causing topological defects in that region and leading the membrane flexibility increase. LA, unlike SMT, induces a transition from multilamellar to unilamellar liposomes, and this phenomenon is greatly influenced by the pH of the medium.

Small quantities of amorphous content can have a profound influence on the properties of a material, however their instability means that quantifying amorphous content over time is important for proving the stability of a drug. Quantifying amorphous content in α-lactose monohydrate by solid state 13C CP MAS NMR, has been carried out by use of proton saturation recovery relaxation and differentiating between spectra by partial least squares (PLS), however these techniques have not proved sensitive on their own, this work investigates their sensitivity in combination. Crystalline α-lactose monohydrate and a rapidly quenched melt were combined to create a set of calibration mixes, whose spectra were recorded using proton saturation recovery relaxations ranging from 2 to 60 seconds. This technique showed a limit of detection of 0.17% (LOD = intercept + 3xSy/x), with a relaxation delay of 15 s and was able to recognise amorphous materials generated by spray and freeze drying. The atmospheric effects on the proton saturation recovery relaxation times of different amorphous lactose preparations were investigated. This found that an oxygen atmosphere reduced the relaxation times, of amorphous lactose that was prepared from a rapidly quenched melt. The loss of moisture from spray dried and freeze dried samples to less than 1% removed the significance of this effect. Lactose is an important excipient in pharmaceuticals and a key ingredient of confectionary, very little research has been carried out in to the quantification of the isomers of different preparations of amorphous lactose. This work quantifies the isomer content by Gas Chromatography with Flame Ionisation Detection (GC-FID) using a DB-17 15m 0.53mm 1.00 μm column and derivatisation with N- (trimethylsilyl)imidazole.

La première partie rapporte la caractérisation de prototypes de chambre MICROMEGAS en tests sous faisceaux et en laboratoire. L'étude détaille notamment l'efficacité de détection, la multiplicité de la réponse et dépendance du gain du détecteur vis à vis des conditions environnementales. Dans la seconde partie, La détection d'un boson de jauge Z', prédit par différends modèles de types Randal-Sundrum est étudiées dans le cadre de l'expérience CLIC. Le canal considéré implique l'émission du Z' par une paire de quarks top. Le Z' se désintègre ensuite en particules de matière noire. / The first part of the thesis reports on the characterisation of MICROMEGAS prototypes in beam and laboratory tests. Detection efficiency, response multiplicity and detector gain dependency against environmental conditions are detailed. In the second part, the detection of a Z' gauge boson predicted by a Randal & Sundrum inspired model together with a dark matter candidate (nu') is investigated in the framework of the CLIC experiment. The considered channel involves a top quark pair emmiting the Z' which, in turn, decays into nu', anti-nu'.

The research programme studied the cure reaction of a phenolic novolak resin and the effects of various additives and fillers on the reaction. The programme utilised the recently developed thermal analysis technique of temperature-modulated differential scanning calorimetry (TMDSC) performed in conjunction with other available thermal analysis techniques. TMDSC enables the signal for the heat of reaction to be separated from the underlying specific heat change in the resin. This meant that the reaction could be studied without interference from any physical changes in the resin. The manufacture of composite brake materials required the use of numerous additives and fillers to produce the desired properties. The influence of such additives on the cure rate and final properties of the resin was known to occur but had not previously been measured due to the difficulties presented by the presence of opaque additives. Some additives also underwent thermally induced physical changes in the temperature range of the cure. The final properties and the processing of new brake materials undergoing development often required trial and error adjustments to compensate for changes in cure rate. An understanding of the influence of additives would enable more rapid commercial development of brake materials through an improvement in the ability to predict both the properties of the product and the optimal processing parameters. Processing efficiency could also be improved through detailed knowledge of the kinetics. Moulding cycle times and post-baking times and temperatures were longer than necessary in order to ensure adequate cure at the end of each stage because of the lack of kinetic data. The cure of phenolic resin has been shown to be highly complicated with numerous alternate and competing reactions. For the manufacture of composite materials, knowledge of the kinetic parameters of individual reactions is not considered to be important; rather the overall kinetic parameters are required for prediction. Therefore the kinetic model parameters that best described the observed behaviour were chosen even though the model had no basis in the molecular interaction theory of reaction. Rather it served as a convenient tool for predictions. Characterisation of the resin proved to be difficult due to the presence of overlapping peaks, and volatile reaction products. TMDSC was successfully used to determine the reaction kinetics of the pure resin and the influence of certain additives on the reaction kinetics. The determination of the kinetic parameters using TMDSC agreed well with the traditional Differential Scanning Calorimetry isothermal and non-isothermal techniques. Both the Perkin-Elmer and TA Instruments were utilised for the research and were found to provide reasonably good agreement with each other. The capabilities and limitations of the individual instruments were critically examined, frequently beyond the manufacturers' specifications. TMDSC suffers from a limitation in the heating rate of the sample compared to DSC. However, it was observed that valuable information could still be obtained from TMDSC despite using heating rates that were higher than specified by manufacturers. Hot Stage Microscopy and thermogravimetry were additional experimental techniques used to aid in the characterisation of the resin. Some inhomogeneity of the resin was identified as well as differences in the behaviour of the cure between open (constant pressure) and closed (constant volume) environments were observed. A novel method of determining the orders of the cure reactions and their kinetic parameters was utilised. Reaction models for the overall cure reactions were postulated and tested by fitment to sections of experimental data in temperature regions which appeared to be free of interference from overlapping peaks. Once an individual peak was reasonably well modelled, adjacent overlapping peaks were able to be modelled both individually and in combinations by fitment to experimental data. The Solver function in Microsoft Excel was utilised to find the best fitting model parameters for the experimental data. The model parameters were able to be refined as overlapping peaks were progressively incorporated into the calculations. This method produced results that agreed well with the traditional method of analysing reaction peak temperatures at multiple scanning rates. Model fitment was shown to be of benefit where overlapping reactions occur. Various model scenarios could be tested and optimised to particular sections of experimental data. This enabled the researcher to easily identify areas of possible anomalies and postulate alternative scenarios. The accuracy of the postulated model was able to be determined by its successful fitment to experimental data from experiments run under different conditions.

<p>In this thesis, the structure and stability of globular proteins adsorbed onto nanometer-sized hydrophilic silica particles were investigated using differential scanning calorimetry (DSC), hydrogen/deuterium exchange (HDX), and mass spectrometry (MS). The adsorption process itself was characterized with fluorescence and absorption spectroscopy and surface plasmon resonance (SPR). The combination of these methods offered a unique insight into adsorption-induced changes within proteins related to their adsorption characteristics. DSC contributed with thermodynamic information on the overall structural stability within the protein population. HDX in combination with MS contributed information on the structure and stability of adsorbed proteins with focus on changes within the secondary structure elements. In order to increase the structural resolution in this part of the investigation, proteolysis was performed prior to the MS analyzing step. Knowledge on the protein adsorption process was utilized in a practical approach called ligand fishing. In this approach, SPR was used to monitor the chip-based affinity purification of a protein with MS used for protein identification.</p><p>Adsorption isotherms revealed that electrostatic interactions play an important role in the adsorption of proteins to hydrophilic surfaces. DSC investigation revealed that the thermal stability of proteins reduces with increasing electrostatic attraction between the protein and the surface and that this effect diminishes at higher surface coverage. The mass-increase due to exchange between protein hydrogen atoms and deuterium atoms in solution was investigated as a function of time. This gave insight into adsorption-induced changes in the structural stability of proteins. By combining DSC and HDX-MS, it was possible to differentiate between adsorption-induced changes in the secondary and tertiary structure. Additionally, if limited proteolysis was performed, the investigations gave insight into the orientation and protein segment specific changes in the stability of proteins adsorbed to silica surfaces. The adsorption of proteins to silica particles also provided the basis for a new experimental design that allows handling of minute amounts of proteins in a ligand fishing application, as used in the field of functional proteomics.</p>

In this thesis, the structure and stability of globular proteins adsorbed onto nanometer-sized hydrophilic silica particles were investigated using differential scanning calorimetry (DSC), hydrogen/deuterium exchange (HDX), and mass spectrometry (MS). The adsorption process itself was characterized with fluorescence and absorption spectroscopy and surface plasmon resonance (SPR). The combination of these methods offered a unique insight into adsorption-induced changes within proteins related to their adsorption characteristics. DSC contributed with thermodynamic information on the overall structural stability within the protein population. HDX in combination with MS contributed information on the structure and stability of adsorbed proteins with focus on changes within the secondary structure elements. In order to increase the structural resolution in this part of the investigation, proteolysis was performed prior to the MS analyzing step. Knowledge on the protein adsorption process was utilized in a practical approach called ligand fishing. In this approach, SPR was used to monitor the chip-based affinity purification of a protein with MS used for protein identification. Adsorption isotherms revealed that electrostatic interactions play an important role in the adsorption of proteins to hydrophilic surfaces. DSC investigation revealed that the thermal stability of proteins reduces with increasing electrostatic attraction between the protein and the surface and that this effect diminishes at higher surface coverage. The mass-increase due to exchange between protein hydrogen atoms and deuterium atoms in solution was investigated as a function of time. This gave insight into adsorption-induced changes in the structural stability of proteins. By combining DSC and HDX-MS, it was possible to differentiate between adsorption-induced changes in the secondary and tertiary structure. Additionally, if limited proteolysis was performed, the investigations gave insight into the orientation and protein segment specific changes in the stability of proteins adsorbed to silica surfaces. The adsorption of proteins to silica particles also provided the basis for a new experimental design that allows handling of minute amounts of proteins in a ligand fishing application, as used in the field of functional proteomics.

Background: Cerebral Palsy is usually divided into three subgroups: ataxic, spastic and dyskinetic, where children and adolescents can, because of misestimation in nutrition and energy intake, suffer from weight problems. Aim: To compare calculations with equations with the individual's measured RMR in the subgroups, to see if any equation is more suitable. This could be used as a tool to calculate the resting metabolism at times when it is not possible to perform clinical measurements. Method: The RMR has been measured in 37 children and adolescents aged 3-15 years through indirect respiratory calorimetry. Those values have been compared with calculations from five equations. The results were then analyzed in order to find if any equation is better to apply for calculating resting metabolism for each subgroup. Results: The ataxic group was overestimated by 56.5% of the calculated values. WHO/FAO/UNU’s equation indicates a significantly strong correlation between the measured and calculated values (r=0.85, p&lt;0.05). The spastic group was underestimated by 53.3%. There is significance for all equations, however it is a relatively low correlation (r=0.63–0.66, p=0.02–0.03). The dyskinetic group was underestimated by 95%. There was no significant association between clinically measured values and the calculated resting metabolism (r=0.21–0.45, p=0.26–0.61). Conclusion: The equations are not reliable for calculation of energy need for all subgroups. There is a substantial risk of over- and underestimation, therefore awareness of the equations insufficiency is needed. Studies should be performed with additional basic data, and with more equations. Especially equations including further variables that involves the body composition. / Bakgrund: Cerebral Pares brukar delas in i undergrupper: ataktisk, spastisk och dyskinetisk, där barn och ungdomar kan, beroende på felskattningar av näringsintag och energiberäkning, lida av över- eller undervikt. Syfte: Att jämföra uträkningar med ekvationer med individernas uppmätta vilometabolism i de tre undergrupperna, för att se om någon ekvation lämpar sig bättre. Detta skulle kunna ge ett verktyg för att kunna räkna ut vilometabolismen vid de tillfällen då det inte är möjligt att utföra kliniska mätningar. Metod: Mätningar av vilometabolismen har utförts på 37 barn och ungdomar i åldern 3-15 år genom indirekt respiratorisk kalorimetri. Värdena har jämförts med uträkningar från fem ekvationer. Resultatet har sedan analyserats, för att hitta någon ekvation som är bättre lämpad för att räkna ut vilometabolismen, för respektive undergrupp. Resultat: Den ataktiska gruppen överskattades i 56,5% av uträkningarna av vilometabolism. WHO/FAO/UNUs ekvation påvisar ett signifikant starkt samband mellan det uppmätta och uträknade värdena (r=0,85, p&lt;0,05). Den spastiska gruppen underskattades 53,3% av de uträknade värdena. Det föreligger signifikans för alla ekvationer, dock ett relativt lågt samband (r=0,63–0,66, p=0,02–0,03). Den dyskinetiska gruppen underskattades i 95 %. Det förelåg inget signifikant samband mellan kliniskt uppmätt och uträknad vilometabolism (r=0,21–0,45, p=0,26–0,61). Slutsats: Ekvationerna är inte tillförlitliga för uträkning av energibehov för alla undergrupper. Risk för över- och underskattning är överhängande, och en ökad medvetenhet om ekvationernas ofullständighet måste belysas. Studier bör göras med större underlag och med fler ekvationer. Framförallt ekvationer med fler variabler som tar hänsyn till kroppssammansättning.

The aim of this study was to investigate the molecular interactions of the anti-cancer drug Doxorubicin (DOX) with poly(methacrylic acid) grafted starch nanoparticles (PMAA-g-St). In order to fully understand the DOX/PMAA-g-St system, we conducted in-depth studies on DOX dimer dissociation and DOX/PMAA-g-St binding interactions using various techniques such as isothermal titration calorimetry (ITC), dynamic light scattering (DLS), and fluorescence and absorption spectroscopy. Based on our experimental results, we developed a quantitative thermodynamic model with relevant parameters such as dissociation constant, Kd, as well as enthalpy of binding, ΔH, in order to explain DOX/PMAA-g-St interactions. In addition, we also studied the effect of environmental factors such as pH and NaCl on DOX self-association and DOX/PMAA-g-St complex formation. In conclusion, the combination of results obtained from various techniques as well as the multispecies equilibrium model, enables us to interpret quantitatively the data of drug loading onto and release from polymeric nanoparticles.

The persistence and propagation of bacterial antibiotic resistance presents significant challenges to the treatment of drug resistant bacteria with current antimicrobial chemotherapies, while a dearth in replacements for these drugs persists. The thiopeptide family of antibiotics may represent a potential source for new drugs and thiostrepton, the prototypical member of this antibiotic class, is the primary subject under study in this thesis. Using a facile semi-synthetic approach novel, regioselectively-modified thiostrepton derivatives with improved aqueous solubility were prepared. In vivo assessments found these derivatives to retain significant antibacterial ability which was determined by cell free assays to be due to the inhibition of protein synthesis. Moreover, structure-function studies for these derivatives highlighted structural elements of the thiostrepton molecule that are important for antibacterial activity. Organisms that produce thiostrepton become insensitive to the antibiotic by producing a resistance enzyme that transfers a methyl group from the co-factor S-adenosyl-L-methionine (AdoMet) to an adenosine residue at the thiostrepton binding site on 23S rRNA, thus preventing binding of the antibiotic. Extensive site-directed mutagenesis was performed on this enzyme to generate point mutations at key active site residues. Ensuing biochemical assays and co-factor binding studies on these variants identified amino acid residues in the active site that are essential to the formation of the AdoMet binding pocket and provided direct evidence for the involvement of an active site arginine in the catalytic mechanism of the enzyme. Certain bacteria that produce neither thiostrepton nor the resistance methyltransferase express the thiostrepton binding proteins TIP-AL and TIP-AS, that irreversibly bind to the antibiotic, thereby conferring resistance by sequestration. Here, it was found that the point mutation of the previously identified reactive amino acid in TIP-AS did not affect covalent binding to the antibiotic, which was immediately suggestive of a specific, high affinity non-covalent interaction. This was confirmed in binding studies using chemically synthesized thiostrepton derivatives. These studies further revealed structural features from thiostrepton important in this non-covalent interaction. Together, these results indicate that thiostrepton binding by TIP-AS begins with a specific non-covalent interaction, which is necessary to properly orient the thiostrepton molecule for covalent binding to the protein. Finally, the synthesis of a novel AdoMet analogue is reported. The methyl group of AdoMet was successfully replaced with a trifluoromethyl ketone moiety, however, the hydrated form (germinal diol) of this compound was found to predominate in solution. Nevertheless, the transfer of this trifluoroketone/ trifluoropropane diol group was demonstrated with the thiopurine methyltransferase.