Nesta DissertaÃÃo sÃo apresentados os resultados oriundos do estudo do efeito da aplicaÃÃo de pressÃo hidrostÃtica sobre cristais de L-alanina deuterada. A L-alanina à um dos vinte aminoÃcidos formadores das proteÃnas, sendo entre os quirais o mais simples deles. A L-alanina cristaliza-se numa estrutura ortorrÃmbica (D24) com quatro molÃculas por cÃlula unitÃria formando uma complexa rede de sete diferentes ligaÃÃes de hidrogÃnio. Quando pressÃo à aplicada a um cristal de aminoÃcidos ocorre a aproximaÃÃo das molÃculas com a consequente variaÃÃo nas distÃncias e Ãngulos das ligaÃÃes de hidrogÃnio, eventualmente obrigando a estrutura a ir para uma nova simetria. Investigou-se nesta DissertaÃÃo o comportamento dos espectros Raman da L-alanina deuterada na regiÃo espectral entre 50 e 2500 cm-1 para pressÃes variando entre 0 e 6 GPa. AlÃm do usual comportamento do desvio da freqÃÃncia de quase todos os modos para altos valores, observou-se tambÃm descontinuidades nas freqÃÃncias de modos internos e modos externos para valores de pressÃo em 1,5 e 4,5 GPa. Estas descontinuidades foram interpretadas como sendo consequÃncias de transiÃÃes de fase estruturais de uma estrutura ortorrÃmbica para uma estrutura tetragonal e, a seguir, para uma estrutura monoclÃnica em analogia ao que ocorre com a L-alanina nÃo deuterada. O comportamento do modo de torÃÃo da estrutura ND3 tambÃm foi estudado e uma analogia do seu comportamento com aqueles observados na L-alanina, na L-treonina e na taurina em termos das dimensÃes das ligaÃÃes de hidrogÃnio tambÃm à fornecida. / In this dissertation we present of Raman spectroscopy on deuterated L-alanine crystals under hydrostatic pressure. L-alanine is one of twenty proteic amino acids, and among them it is the simplest chiral one. L-alanine crystallizes in an orthorhombic (D24) structure with four molecules per unit cell, forming a complex network of seven hydrogen bonds. When one applies pressure on an amino acid crystal the distances between molecules are changed and, eventually, the crystal goes to a new structure with different symmetry. In this work we present the Raman spectra of deuterated L-alanine in the 50 - 2500 cm-1 spectral region with pressures changing from 0 to 6 GPa. Beyond the conventional frequency blue shift of almost all modes, we observed frequency discontinuities of both internal and external modes at 1,5 and 4,5 GPa. These discontinuities were interpreted as structural phase transitions undergone by deuterated L-alanine crystal from the orthorhombic to the tetragonal, and from the tetragonal to a monoclinic symmetry, respectively. This suggestion is based on previous results on non - deuterated L-alanine. The behavion of torcional vibration of ND3 was also studied and a comparison with studies on L-alanine, L-threonine and taurine crystals (related with dimensions of hydrogen bonds) is also given.

Using ion-trapping techniques, selected laboratory experiments on ion-molecule reactions of astrophysical interest have been performed. For the first time a carbon beam source has been integrated into an ion trapping machine for studying collisions between ions and neutral carbon atoms and molecules. Results are presented for the interaction of D3+ ions stored in a ring-electrode trap (RET), with a beam of hot neutral carbon molecules, Cn (n = 1, 2, 3). The measured reaction rate coefficients are up to a factor two smaller than values presently used in astrophysical models. In order to complete our knowledge about the ion chemistry involving three carbon atoms, detailed investigations of reactions of C3+, C3H+ and C3H3+ with H2 and HD have been performed between 15 K and room temperature. These studies have been performed in a second apparatus, a variable-temperature 22-pole trap machine (VT-22PT). Results include reactive collisions, deuteration and radiative association. It is discussed in connection with the increase in lifetime of the C3+ + H2 collision complexes with falling temperature, what could be responsible for producing more C3H+ at 15 K. Tunneling is excluded. In C3+ + HD collisions an isotope effect has been detected, the C3D+ product ions being slightly more abundant than C3H+. Comparison of the reaction of C3H+ primary ions with HD and H2 gas revealed that the deuterated molecules are significantly more reactive. The process of radiative association of C3H+ and for the first time of C3+ with hydrogen molecules has been observed. An analysis of the data shows that radiative association becomes slower, if the neutral reactant is deuterated. Finally, the theoretical prediction from ab initio calculations that C3H3+ does not exchange an H for a D in collisions with HD, has been proven in an ion trap experiment. Careful analysis of all competing processes allows the conclusion that the rate coefficient is smaller than 4x10-16 cm3s-1 at 15 K. / Unter Verwendung von zwei Speicherapparaturen wurden ausgewählte, astrophysikalische wichtige Ionen-Molekülreaktionen untersucht. Durch die Kombination einer Kohlenstoffquelle mit einem Ionenspeicher, in dem so Reaktionen zwischen Ionen und Kohlenstoffmolekülen oder -atomen untersucht werden können, wurde Neuland betreten. Es werden Ergebnisse vorgestellt für die Reaktion von D3+ Ionen, die in einem Ringelektrodenspeicher gefangen sind, mit einem Strahl von heißen Cn (n = 1, 2, 3). Die gemessenen Ratenkoeffizienten sind nur halb so groß wie die Werte, die in astrophysikalischen Modellen verwendet werden. Um die Kenntnis über alle möglichen Reaktionen, bei denen drei C-Atome beteiligt sind, abzurunden, wurden zwischen 15 K und Zimmertemperatur die Reaktionen zwischen C3+, C3H+ und C3H3+ Ionen mit H2 und HD in vielen Details untersucht. Diese Experimente wurden in einer zweiten Apparatur durchgeführt, in der ein temperaturvariabler 22-Polspeicher das zentrale Element ist (VT-22PT). Berichtet werden Ergebnisse zu reaktiven Stößen, zur Deuterierung von Kohlenwasserstoffen und zur Strahlungsassoziation. In der Diskussion bleibt offen, was - in Verbindung mit der von 300 K zu 15 K zunehmenden Lebensdauer - der Grund dafür sein kann, daß die Bildung des exothermen Produkts C3H+ anwächst. Der Tunneleffekt scheidet aus. Bei der Reaktion C3+ + HD wurde ein Isotopeneffekt beobachtet, das C3D+ Produkt wird etwas häufiger gebildet als C3H+. Ein Vergleich der Reaktion zwischen C3H+ Ionen mit HD bzw. H2 zeigt, daß das deuterierte Molekül wesentlich reaktiver ist. Es wurden Ratenkoeffizienten für die Strahlungsassoziation von H2 Molekülen mit C3H+ und erstmals mit C3+ Ionen gemessen. Die Auswertung der Daten zeigt, dass der Prozeß langsamer abläuft, wenn der neutrale Stoßpartner deuteriert ist. Schließlich wurde experimentell die theoretische Vorhersage überprüft, dass C3H3+ keinen H-D Austausch mit HD eingeht. Eine sorgfältige Analyse aller konkurrierenden Prozesse ergab, dass bei 15 K der Raten koeffizient kleiner als 4x10-16 cm3s-1 ist.

Líquidos iônicos imidazólios (LIIs) associados a ânions com caráter básico foram sintetizados, caracterizados e seu comportamento estudado em solução. Inicialmente a troca isotópica H/D preferencial no grupo C2-CH3 de sais de 1-nbutil-2,3-dimetilimidazólio (BMMI) foi avaliada. Ficou evidente que o contato entre os pares de íons e a atuação do ânion como uma base neutra influenciaram fortemente a deuteração. Após a análise dos resultados de RMN, cálculos de DFT (Teoria do Funcional de Densidade) e Difração de Raios X, um complexo entre uma molécula de água e alguns LI foi sugerido, modificando fortemente a estrutura organizacional e afetando também a troca isotópica nestes sais. Através do mecanismo reacional proposto, surgiu a possibilidade de deuteração de novos substratos contendo H ácidos, usando os LIIs como catalisadores. A reação de troca isotópica em alcinos e cetonas foi estudada e bons resultados foram obtidos. O efeito da variação de cátions e ânions na atividade catalítica foi analisado, assim como a variação dos substituintes dos substratos. Além disso, um estudo cinético foi realizado através de análises de RMN de 1H e o mecanismo reacional foi proposto. Por fim, a capacidade de formação de pares iônicos em soluções aquosas de LIIs foi avaliada na captura de CO2. Ótimos resultados foram obtidos e atribuídos à ocorrência de sorção física e química devido à ativação da água pelo LII. / Imidazolium based ionic liquids (ImIL) associated with basic anions were synthesized, characterized and studied. Initially the preferred H/D isotopic exchange to C2-CH3 group of 1-nbutyl-2,3-dimethylimidazolium salts (BMMI) was studied. In this context, it became evident that the ion-pairing formation and the anion action as a neutral base strongly influence the deuteration reaction. NMR analysis, theoretical calculation (Density Functional Theory) and X-ray Diffraction have been performed and a complex between a water molecule and the IL have been suggested, strongly modifying the IL structure and characteristics. The isotope exchange reaction in alkynes and ketones was studied using imidazolium based ionic liquids as catalysts and good results obtained. The effect of cation and anion variation on the catalyst activity were analyzed, also a variation of the substrate substituent’s. Also, a kinetic study was performed by 1H NMR analyzes and the reaction mechanism were proposed. Finally, the ability to form ion pair contact of ILs in aqueous solutions was evaluated for CO2 capture. Great results were obtained and this success can be attributed to the occurrence of physical and chemical sorption due to water activation by the IL.

Líquidos iônicos imidazólios (LIIs) associados a ânions com caráter básico foram sintetizados, caracterizados e seu comportamento estudado em solução. Inicialmente a troca isotópica H/D preferencial no grupo C2-CH3 de sais de 1-nbutil-2,3-dimetilimidazólio (BMMI) foi avaliada. Ficou evidente que o contato entre os pares de íons e a atuação do ânion como uma base neutra influenciaram fortemente a deuteração. Após a análise dos resultados de RMN, cálculos de DFT (Teoria do Funcional de Densidade) e Difração de Raios X, um complexo entre uma molécula de água e alguns LI foi sugerido, modificando fortemente a estrutura organizacional e afetando também a troca isotópica nestes sais. Através do mecanismo reacional proposto, surgiu a possibilidade de deuteração de novos substratos contendo H ácidos, usando os LIIs como catalisadores. A reação de troca isotópica em alcinos e cetonas foi estudada e bons resultados foram obtidos. O efeito da variação de cátions e ânions na atividade catalítica foi analisado, assim como a variação dos substituintes dos substratos. Além disso, um estudo cinético foi realizado através de análises de RMN de 1H e o mecanismo reacional foi proposto. Por fim, a capacidade de formação de pares iônicos em soluções aquosas de LIIs foi avaliada na captura de CO2. Ótimos resultados foram obtidos e atribuídos à ocorrência de sorção física e química devido à ativação da água pelo LII. / Imidazolium based ionic liquids (ImIL) associated with basic anions were synthesized, characterized and studied. Initially the preferred H/D isotopic exchange to C2-CH3 group of 1-nbutyl-2,3-dimethylimidazolium salts (BMMI) was studied. In this context, it became evident that the ion-pairing formation and the anion action as a neutral base strongly influence the deuteration reaction. NMR analysis, theoretical calculation (Density Functional Theory) and X-ray Diffraction have been performed and a complex between a water molecule and the IL have been suggested, strongly modifying the IL structure and characteristics. The isotope exchange reaction in alkynes and ketones was studied using imidazolium based ionic liquids as catalysts and good results obtained. The effect of cation and anion variation on the catalyst activity were analyzed, also a variation of the substrate substituent’s. Also, a kinetic study was performed by 1H NMR analyzes and the reaction mechanism were proposed. Finally, the ability to form ion pair contact of ILs in aqueous solutions was evaluated for CO2 capture. Great results were obtained and this success can be attributed to the occurrence of physical and chemical sorption due to water activation by the IL.

Using ion-trapping techniques, selected laboratory experiments on ion-molecule reactions of astrophysical interest have been performed. For the first time a carbon beam source has been integrated into an ion trapping machine for studying collisions between ions and neutral carbon atoms and molecules. Results are presented for the interaction of D3+ ions stored in a ring-electrode trap (RET), with a beam of hot neutral carbon molecules, Cn (n = 1, 2, 3). The measured reaction rate coefficients are up to a factor two smaller than values presently used in astrophysical models. In order to complete our knowledge about the ion chemistry involving three carbon atoms, detailed investigations of reactions of C3+, C3H+ and C3H3+ with H2 and HD have been performed between 15 K and room temperature. These studies have been performed in a second apparatus, a variable-temperature 22-pole trap machine (VT-22PT). Results include reactive collisions, deuteration and radiative association. It is discussed in connection with the increase in lifetime of the C3+ + H2 collision complexes with falling temperature, what could be responsible for producing more C3H+ at 15 K. Tunneling is excluded. In C3+ + HD collisions an isotope effect has been detected, the C3D+ product ions being slightly more abundant than C3H+. Comparison of the reaction of C3H+ primary ions with HD and H2 gas revealed that the deuterated molecules are significantly more reactive. The process of radiative association of C3H+ and for the first time of C3+ with hydrogen molecules has been observed. An analysis of the data shows that radiative association becomes slower, if the neutral reactant is deuterated. Finally, the theoretical prediction from ab initio calculations that C3H3+ does not exchange an H for a D in collisions with HD, has been proven in an ion trap experiment. Careful analysis of all competing processes allows the conclusion that the rate coefficient is smaller than 4x10-16 cm3s-1 at 15 K. / Unter Verwendung von zwei Speicherapparaturen wurden ausgewählte, astrophysikalische wichtige Ionen-Molekülreaktionen untersucht. Durch die Kombination einer Kohlenstoffquelle mit einem Ionenspeicher, in dem so Reaktionen zwischen Ionen und Kohlenstoffmolekülen oder -atomen untersucht werden können, wurde Neuland betreten. Es werden Ergebnisse vorgestellt für die Reaktion von D3+ Ionen, die in einem Ringelektrodenspeicher gefangen sind, mit einem Strahl von heißen Cn (n = 1, 2, 3). Die gemessenen Ratenkoeffizienten sind nur halb so groß wie die Werte, die in astrophysikalischen Modellen verwendet werden. Um die Kenntnis über alle möglichen Reaktionen, bei denen drei C-Atome beteiligt sind, abzurunden, wurden zwischen 15 K und Zimmertemperatur die Reaktionen zwischen C3+, C3H+ und C3H3+ Ionen mit H2 und HD in vielen Details untersucht. Diese Experimente wurden in einer zweiten Apparatur durchgeführt, in der ein temperaturvariabler 22-Polspeicher das zentrale Element ist (VT-22PT). Berichtet werden Ergebnisse zu reaktiven Stößen, zur Deuterierung von Kohlenwasserstoffen und zur Strahlungsassoziation. In der Diskussion bleibt offen, was - in Verbindung mit der von 300 K zu 15 K zunehmenden Lebensdauer - der Grund dafür sein kann, daß die Bildung des exothermen Produkts C3H+ anwächst. Der Tunneleffekt scheidet aus. Bei der Reaktion C3+ + HD wurde ein Isotopeneffekt beobachtet, das C3D+ Produkt wird etwas häufiger gebildet als C3H+. Ein Vergleich der Reaktion zwischen C3H+ Ionen mit HD bzw. H2 zeigt, daß das deuterierte Molekül wesentlich reaktiver ist. Es wurden Ratenkoeffizienten für die Strahlungsassoziation von H2 Molekülen mit C3H+ und erstmals mit C3+ Ionen gemessen. Die Auswertung der Daten zeigt, dass der Prozeß langsamer abläuft, wenn der neutrale Stoßpartner deuteriert ist. Schließlich wurde experimentell die theoretische Vorhersage überprüft, dass C3H3+ keinen H-D Austausch mit HD eingeht. Eine sorgfältige Analyse aller konkurrierenden Prozesse ergab, dass bei 15 K der Raten koeffizient kleiner als 4x10-16 cm3s-1 ist.

The ¹HNMR spectra of all possible structural isomers of mono and dihydroxy substituted naphthalenes are measured in trifluoracetic acid and trifluormethanesulfonic acid as well as in their deuterated derivatives. These spectra indicate different protonation/deuteration positions in the compounds studied. Whereas with the relative weak TFA the OH group as most basic group of the substrates is protonated, with the more acidic TFS also aromatic positions with lower basicity are protonated. However, some of these position were indicated only by using deutertated acids. To quantify the degree of proton/deuterium exchange at the aromatic rings, dioxane as proton standard was used. For the OH-protonation a complex between the appropriate naphthol and the acid used, in which a quick proton exchange can be occur, is assumed. Furthermore, the formation of corresponding trifluoroacetates or triflates by reaction of the naphthols with the corresponding acids has been indicated. In some cases, in which the ring positions of protonation are not definitely clear, 2D NOESYNMR experiments have been performed.

In analogy with the reaction of phenols with very strong super acids, these compounds can be protonated by the weaker super acid trifluoroacetic acid (TFA) and trifluoromethanesulphonic acid (TFS) also giving rise to the formation of OH and/or ring protonated products, which can be identified unambiguously by means of ¹HNMR spectroscopy. The same reactions are possible with the deuterated acids TFA-d and TFS-d. However, from these reactions, additional information on the proton and deuterium attack at different positions in the phenolic compounds is possible not detectable by the hitherto used non-deuterated acids. Moreover, the reaction of phenols with these deuterated acids allows the preparation of partially or fully deuterated phenols that are also important as markers in biological application and material chemistry.

Deuteration is the replacement of a hydrogen atom by deuterium atom in a molecule. The replacement begins at the most acidic hydrogen in the molecule. In ATP, the deshielded hydrogen is C8-H which is the first replaced during deuteration. During ATP deuteration some of the ATP is hydrolysed to ADP concurrently. Using kinetic analysis, it was confirmed that the ATP hydrolysis that occurs is 1st order in ATP concentration, while the hydrogen replacement is 2nd order. The ATP and its C8 deuterated analogue were tested against three enzymes shikimate kinase (SK), acetate kinase (AK) and glutamine synthetase (GS) to determine if a kinetic isotope effect (KIE) exists in these systems. With AK and GS, the KIED increased as the KIEH decreased, while with SK the KIED decreased as the KIEH increased as the concentration of the ATP or deuterated analogue increased. Deuteration of imidazole and purine compounds reduced the specific activity of AK or SK at low concentrations in an enzyme-catalysed reaction. From a library of imidazole-containing compounds that inhibited SK, three compounds were selected and their IC50 values were determined on the SK-catalysed reaction. These compounds show a differential potency and efficiency between their protonated and deuterated analogues when compared in a 1:1 mixture. Synthesized purines incorporating three different substituents at N-9 were tested against AK or SK for their ability to lower the specific activity of the enzymes used / Physics / M. Sc. (Physics)

In der hier vorgestellten Arbeit werden neuartige Festkörper-NMR (Nuclear Magnetic Resonance) Experimente vorgestellt, die eine direkte Detektion von Protonen einbeziehen. Die Technik basiert auf einer weitgehenden Verdünnung der Protonen durch Deuteronen in vollständig isotopenmarkierten, rekombinant exprimierten Proteinen und ermöglicht Festkörper-NMR mit sehr schmalen Linienbreiten aller standardmäßig erfassbaren Kerne (Protonen, Stickstoff, Kohlenstoff) ohne Hochleistungsentkopplung. Zusätzlich werden Methoden für ein besonders hohes Signal-zu-Rausch durch Paramagnetic Relaxation Enhancement (PRE) entwickelt. Die so präparierten Proteine erweisen sich tauglich für eine stark verbesserte NMR-Charakterisierung durch eine Vielzahl neuer Struktur- und Zuordnungsexperimente, in denen Techniken aus Festkörper- und Lösungs-NMR vereint werden. Dabei können hier erstmals auch Bereiche im Protein mit einbezogen werden, die langsame Dynamik vollziehen. Die Experimente finden Anwendung auf die SH3-Domäne von alpha-Spektrin, das Alzheimer-Peptid Abeta1-40 und das Membranprotein Omp G. / In this work, novel solid-state NMR (Nuclear Magnetic Resonance) experiments are presented that imply direct detection of protons. The technique is based on extensive dilution of protons with deuterons in uniformly labelled, recombinantly expressed proteins and allows for solid-state NMR providing very narrow lines of all commonly accessible nuclei (protons, nitrogen, carbon) without high-power decoupling. In addition, methods are developed that yield a particularly high signal-to-noise through Paramagnetic Relaxation Enhancement (PRE). The accordingly prepared proteins are shown to be applicable for a significantly improved NMR-characterization by manifold new experiments for assignment and structure elucidation, in which techniques from solid-state and solution NMR are united. For the first time, also those regions in a protein can be accessed that undergo slow dynamics. The experiments are employed on the SH3-domain of alpha-spectrin, Alzheimer’s peptide Abeta1‑40, and the membrane protein Omp G.

Deuteration is the replacement of a hydrogen atom by deuterium atom in a molecule. The replacement begins at the most acidic hydrogen in the molecule. In ATP, the deshielded hydrogen is C8-H which is the first replaced during deuteration. During ATP deuteration some of the ATP is hydrolysed to ADP concurrently. Using kinetic analysis, it was confirmed that the ATP hydrolysis that occurs is 1st order in ATP concentration, while the hydrogen replacement is 2nd order. The ATP and its C8 deuterated analogue were tested against three enzymes shikimate kinase (SK), acetate kinase (AK) and glutamine synthetase (GS) to determine if a kinetic isotope effect (KIE) exists in these systems. With AK and GS, the KIED increased as the KIEH decreased, while with SK the KIED decreased as the KIEH increased as the concentration of the ATP or deuterated analogue increased. Deuteration of imidazole and purine compounds reduced the specific activity of AK or SK at low concentrations in an enzyme-catalysed reaction. From a library of imidazole-containing compounds that inhibited SK, three compounds were selected and their IC50 values were determined on the SK-catalysed reaction. These compounds show a differential potency and efficiency between their protonated and deuterated analogues when compared in a 1:1 mixture. Synthesized purines incorporating three different substituents at N-9 were tested against AK or SK for their ability to lower the specific activity of the enzymes used / Physics / M. Sc. (Physics)