La résonance magnétique nucléaire est une technique de choix pour étudier la dynamique de l'échange dans des composés à portée thérapeutique. Cependant, les spectres acquis avec les méthodes 1D et 2D classiques présentent souvent des recouvrements importants, ce qui rend l'attribution de chaque espèce en échange difficile. Le développement méthodologique d'une approche pure shift permettrait donc d'améliorer la résolution de ces données. Au cours de cette thèse, nous avons effectué une série de développements méthodologiques des expériences de type EXSY dans lesquelles nous avons mis en œuvre différentes méthodes de découplage homonucléaire. En particulier, deux approches ont été testées: la méthode PSYCHE appliqué aux dimensions F1 et F2 des cartes EXSY, et la méthode Zangger-Sterk en F2. Ces approches ont toutes mené à une amélioration significative de la résolution qui nous a permis de caractériser les deux conformères s-cis/s-trans du Trandolapril. Nous avons également étudié l’intérêt de la technique d'échantillonnage non-uniforme (NUS) du signal pour réduire le temps d'analyse. Cette approche nous a permis de gagner en temps d'analyse, mais la présence d'artefacts à certaines températures suggère que d'autres développements seront encore nécessaires. Ces résultats ouvrent la voie vers une analyse plus fine du processus d’échange dans des composés présentant un spectre RMN complexe. / Nuclear magnetic resonance is a technique of choice for studying chemical exchange in therapeutic compounds. However, spectra acquired with standard 1D and 2D methods often show spectra with overlapping signals, which makes the assignment of each species difficult. Methodological development of a pure shift approach would therefore make it possible to improve the resolution of these data. During this thesis, we carried out a series of developments of the EXSY type experiments in which we implemented different homonuclear decoupling methods. Two approaches were tested: the PSYCHE method applied to the F1 and F2 dimensions of EXSY maps, and the Zangger-Sterk method in F2. These approaches all led to a significant resolution improvements allowing us to characterize the s-cis/ s-trans conformers in Trandolapril. We have also studied the interest of the non-uniform sampling (NUS) technique to reduce the analysis time. This approach allowed us to accelerate the experiment, but the presence of artifacts at various temperatures suggests that further developments will still be needed. These results pave the way for more analysis of the exchange process in compounds with complex NMR spectrum.

Final publication is available at http://link.springer.com/article/10.1007/s11060-014-1715-8 / 京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19605号 / 医博第4112号 / 新制||医||1014(附属図書館) / 32641 / 京都大学大学院医学研究科医学専攻 / (主査)教授 村井 俊哉, 教授 平岡 眞寛, 教授 山田 泰広 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

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Understanding transport of water molecules and salt ions from a molecular level up to macroscopic length scales is critical to the design of novel materials for many applications, including separations membranes for fuel cell and desalination applications, as well as rechargeable battery technology. This work aims to investigate and develop new models correlating the dynamics and structure of polymeric materials, to the transport of small molecules within them, using a variety of Nuclear Magnetic Resonance (NMR) techniques. We present three studies through which we utilize two chemically similar membranes: hydroxyethyl acrylate-co-ethyl acrylate (HEA-co-EA) and hydroxymethyl methacrylate-co-methyl methacrylate (HEMA-co-MMA), which greatly differ in glass transition temperature, in order to understand the fundamental relationships from polymer chain dynamics and small molecule diffusion. From observations of the micron scale diffusion of these materials we find that the more dynamic, rubbery HEA-co-EA exhibits lower water to salt selectivity than HEMA-co-MMA, and that this difference arises from nanoscale morphology of the materials. From this, we propose a new model for hydrophilic pathways inside polymeric materials consisting of nanometer scale interconnected pathways are interrupted by micron scale arrangements of so-called "dead ends". We also for the first time show the separation of material tortuosity into two regimes, ranging from the nanometer-bulk and micron-bulk length scales. We further separate the contributions of structure from chemical interactions in the chemically similar desalination materials by investigating the local activation energy of diffusion in both materials, as well as aqueous solutions of the hydrophilic monomers analogous to the internal membrane environment. We find that the effects of local geometric confinement are very similar between the two materials, however the intermolecular interactions between water and the hydrophilic monomers, with respect to water transport, are significantly different between the two hydrophilic species. Geometric confinement accounts for a 5 ± 1 kJ/mol increase in diffusive activation energy from solution to membrane for both chemistries, and a 4 ± 1 kJ/mol difference in activation energy is seen between the two chemistries in both solution and membrane form. We propose that the entropic contributions to transport, are strongly impacted by the rigid environment of the HEMA material, and is related to the increased water-salt selectivity, as well as the increasing selectivity with increased ionic size observed compared to the HEA system. Using Dynamic NMR spectroscopy, we further investigate the differences seen in water-monomer intermolecular proton exchange by NMR. We utilize an iterative least-squares solving method to fit our exchange lineshape to a model of an uncoupled, two-site exchange lineshape in order to obtain rate and equilibrium population data from -50 to 70 °C. We find that, similar to the diffusive activation energy, the HEA-water system shows reduced enthalpy and entropy of the transition state compared to HEMA-water, such that there is faster exchange between HEMA and water at all temperatures measured, in addition to more biased populations in the HEA-water system. / Doctor of Philosophy / Understanding transport of water molecules and salt ions from a molecular level up to macroscopic length scales is critical to the design of novel materials for many applications, including separations membranes for fuel cell and desalination applications, as well as rechargeable battery technology. This work aims to investigate and develop new models correlating the dynamics and structure of polymeric materials, to the transport of small molecules within them, using a variety of Nuclear Magnetic Resonance (NMR) techniques. We will present three studies in which we seek to further understand the relationships between a material's physical and chemical properties, with the behavior of small molecules like water absorbed within the material. NMR spectroscopy, while not the standard method for characterizing desalination membranes, allows us to specifically probe direct effects on molecular motion of polymer structure from the microscopic level to the bulk, a feat not easily achieved by any other single technique. The first study presented within focuses on the differences in micrometer scale structure in two near identical sets of materials; differing only in that one is rubbery with flexible polymer chains, and the other is rigid with relatively immobile polymer chains. The second study takes these two materials and investigates them through a different lens, probing the molecular scale differences in water motions imparted by the flexible versus rigid polymer chains. The third and final study looks into the fundamental differences seen in how the two chemistries used to create the polymers in the first two studies interact with water molecules through a different NMR technique. These three studies together represent a series of methods and techniques that can be applied to many other classes of polymer materials, such as those destined for use in fuel cells and rechargeable batteries, in order to better understand the fundamental forces at work in those systems to aid in the design of the next generation's materials.

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Stroke is one of the leading causes of death and adult disability worldwide. The major therapeutic intervention for acute ischemic stroke is the administration of recombinant tissue plasminogen activator (rtPA) to help to restore blood flow to the brain. This has been shown to increase the survival rate and to reduce the disability of ischemic stroke patients. However, rtPA is associated with intracranial haemorrhage and thus its administration is currently limited to only about 5% of ischemic stroke patients. More advanced imaging techniques can be used to better stratify patients for rtPA treatment. One new imaging technique, chemical exchange saturation transfer (CEST) magnetic resonance imaging, can potentially image intracellular pH and since tissue acidification happens prior to cerebral infarction, CEST has the potential to predict ischemic injury and hence to improve patient selection. Despite this potential, most studies have generated pH-weighted rather than quantitative pH maps; the most widely used metric to quantify the CEST effect is only able to generate qualitative contrast measurements and suffers from many confounds. The greatest clinical benefit of CEST imaging lies in its ability to non-invasively measure quantitative pH values which may be useful to identify salvageable tissue. The quantitative techniques and work presented in this thesis thus provide the necessary analysis to determine whether a threshold for the quantified CEST effect or for pH exists to help to define tissue outcome following stroke; to investigate the potential of CEST for clinical stroke imaging; and subsequently to facilitate clinical translation of CEST for acute stroke management.

Magnetic Resonance Imaging (MRI) is one of noninvasive imaging and diagnostic methods in today's medicine. The most commonly measured nucleus is H1 of the water molecules present in the human body. There are also methods that use signal saturation transfer between the contrast agent and water molecules via exchangeable H1 nuclei, or use a different nucleus like F19 . Compounds that show a high potential in this area are complexes of paramagnetic ions such as Gd3+ , Eu3+ , Co2+ , Cu2+ and Ni2+ , which can affect relaxation times and chemical shifts of other atoms due to their magnetic properties. This Master's thesis focuses on ligands L1 and L2 which were prepared in the Bachelor thesis. The main focus is on preparation of complexes with selected paramagnetic ions and subsequent study of their properties relevant for a potential use as contrast agents for F19 -MRI and CEST methods. Keywords: F19 -MRI, CEST, macrocyclic ligands, coordination chemistry

ABSTRACT This work is focused on the synthesis of a family of new macrocyclic ligands with exchangeable protons on coordinating groups that could potentially serve (after complexation with suitable paramagnetic lanthanide(III) ions) as responsive contrast agents (CAs) for magnetic resonance imaging (MRI). It is expected that measurement of extracellular pH should bring information for tumorous disease diagnoses and/or for suggesting the most efficient treatment. Therefore, our attention was focused on pH-dependent CAs based on a PARAmagnetic Chemical Exchange Saturation Transfer (PARACEST) mechanism capable of reporting pH changes in tissue. The PARACEST-related properties of a series of Ln(III) complexes with the CEST effect caused by amino groups coordinated to the central Ln(III) metal ions were investigated. Such a kind of PARACEST CA is new and has had no precedent in the literature. It was shown that these Ln(III) complexes produce a pH-sensitive PARACEST effect in the pH region relevant for living systems. The study brings proof-of-principle for utilization of complexes with a linear diamine pendant arm, i.e. complexes with two exchanging proton pools, for ratiometric pH determination by MRI independently on the probe concentration. In addition, to ensure a higher kinetic inertness of the...

In ischaemic stroke a disruption of cerebral blood flow leads to impaired metabolism and the formation of an ischaemic penumbra in which tissue at risk of infarction is sought for clinical intervention. In stroke trials, therapeutic intervention has largely been based on perfusion-weighted measures, but these have not been shown to be good predictors of tissue outcome. The aim of this thesis was to develop analysis techniques for magnetic resonance imaging (MRI) of chemical exchange saturation transfer (CEST) in order to quantify metabolic signals associated with tissue fate in patients with acute ischaemic stroke. This included addressing robustness for clinical application, and developing quantitative tools that allow exploration of the in-vivo complexity. Tissue-level analyses were performed on a dataset of 12 patients who had been admitted to the John Radcliffe Hospital in Oxford with acute ischaemic stroke and recruited into a clinical imaging study. Further characterisation of signals was performed on stroke models and tissue phantoms. A comparative study of CEST analysis techniques established a model-based approach, Bloch-McConnell model analysis, as the most robust for measuring pH-weighted signals in a clinical setting. Repeatability was improved by isolating non-CEST effects which attenuate signals of interest. The Bloch-McConnell model was developed further to explore whether more biologically-precise quantification of CEST effects was both possible and necessary. The additional model complexity, whilst more reflective of tissue biology, diminished contrast that distinguishes tissue fate, implying the biology is more complex than pH alone. The same model complexity could be used reveal signal patterns associated with tissue outcome that were otherwise obscured by competing CEST processes when observed through simpler models. Improved quantification techniques were demonstrated which were sufficiently robust to be used on clinical data, but also provided insight into the different biological processes at work in ischaemic tissue in the early stages of the disease. The complex array of competing processes in pathological tissue has underscored a need for analysis tools adequate for investigating these effects in the context of human imaging. The trends herein identified at the tissue level support the use of quantitative CEST MRI analysis as a clinical metabolic imaging tool in the investigation of ischaemic stroke.