A series of 31 sulfur-nitrogen donor ligands and 64 metal(II) complexes have been investigated. The thiomethylated aniline ligands 2–(methylthiomethyl)aniline 2MT and 2–(methylthio)aniline 2MA were synthesized with their substituted derivatives (-Me, -MeO, -Cl, -Br, -NO2) to serve as chelating agents. These ligands behave as bidentate ligands with SN donor group with Co(II), Ni(II) and Cu(II). The Co(II) and Ni(II) complexes have the ML2Cl2 molecular formula while the Cu(II) complexes formed with MLCl2 stoichiometry where L is the bidentate ligand. The ligands and their metal(II) complexes have been characterized by elemental analysis and with spectroscopic techniques. The trend observed in the NMR spectra and IR frequencies of the thiomethylated compounds shows there is a significant difference between the 2MT and 2MA series as a result of sulfur lone pairs extending the conjugation of the aromatic ring in the case of the latter. The effect of the position and electronic nature of ring substituent on the NMR shifts of the amine protons is discussed. The 6- and 5-membered chelate complexes formed by the 2MT and 2MA ligands respectively do not show significant diversity in their spectroscopic properties. From the elemental analysis for the Co(II) and Ni(II) complexes, their compositions reveal 1:2 M:L stoichiometry with 2 chlorine atoms from the respective metal salts. In addition, the spectroscopic data are largely indicative of tetragonally distorted structures for these solid complexes. The X-ray crystallography data reveal the Cu(II) complexes exist as square pyramidal dimers and with long Cu–Cl equitorial bonds fit into the tetragonally distorted octahedral structure. The electrolytic nature of Co(II) and Cu(II) complexes in DMF were found to be similar, they behave as non electrolytes in contrast to Ni(II) complexes which are 1:1 electrolytes. The electronic spectra of these metal(II) complexes were found to be different for both their solid forms and in solutions of DMF and DMSO and this has been discussed. The thiomethylated aniline ligands possess the amine and thioether groups which are present in many known biologically active compounds, hence the biological activity of the ligands and their metal complexes were tested against three strains of bacteria and one fungus. The methoxy-substituted derivatives were found to possess better inhibitory activity and this was similarly reflected in the metal(II) complexes. The activity of the complexes can be said to be in the order, Cu(II) > Co(II) > Ni(II). The Schiff-base derivatives were prepared from the ligands and para-methoxysalicylaldehyde and their Cu(II) complexes were synthesized in order to determine their biological activity. The Schiff-base ligands were found to be less active than their parent ligands. The Cu(II) complexes are not soluble in water, DMSO or DMF, as a result and could not be evaluated for their biological activity. Based on the good results from the antimicrobial evaluation, the antiplasmodial activity of some of the Co(II), Ni(II) and Cu(II) complexes of the thiomethylated ligands against Plasmodium falciparum (FCR-3) was determined. At 50 μM concentration level, the Cu(II) complexes show activity equal or better than the prophylactic chloroquine. The Cu(II) complexes with the methoxy-substituted demonstrated exceptional activity but their Co(II) and Ni(II) analogues did not show any activity. The cytotoxicity of the active Cu(II) complexes at 50 μM concentration was determined against the breast cancer cell line (MDA-MB-231). The compounds destroyed the cancer cell in the range of 28–40%, thus showing their preferred activity against the parasitic cell instead of the cancer cell. The selectivity demonstrated by these compounds have shown them to be potential antimalarial agents and this could be further investigated.

There is currently no definitive biomarker for Alzheimer's Disease (AD), confirmation of diagnosis is only possible post-mortem. Magnetic resonance spectroscopy (MRS) has potential in aiding diagnosis, an MRS scan can be performed during an MRI scan, only adding around 10 minutes to scan time. Use of data from the two scans may allow more accurate diagnosis of AD. This thesis investigates a transgenic mouse model of AD, the TASTPM mouse, using in vitro and in vivo MRS as well as matrix assisted laser desorption ionisation mass spectrometry imaging (MALDI MS Imaging). The first aim of the study was to search for a biomarker of AD that may allow better diagnosis or further our understanding of the pathology of the disease. The second aim was to evaluate the TASTPM mouse as a model of AD for use in preclinical testing of amyloid lowering agents. The third aim was to investigate a thalamic pathology in the TASTPM mice using MALDI MS Imaging. Metabolically, we found differences between the brains of TASTPM mice and their wild type base strain in both in vitro and in vivo scans. These differences may be exploited in the preclinical testing of novel amyloid lowering therapies. We also found similarities with human AD and other mouse models, lower N-acetylaspartate, lower glutamate and higher myo-inositol are all observed in human AD, as well as the TASTPM mice in vivo. We also found further evidence of impaired neuronal energy metabolism in TASTPM mice, such as lower succinate. Cerebral hypometabolism is a symptom of human AD. The TASTPM mouse seems to be a fairly good approximation of the human disease, sharing several traits. In our investigation of the thalamic pathology, we discovered a peptide which was strongly localised to the regions of the pathology and isolated it, but were unable to identify it, the work in this area will continue.

There is a general lack of atomic resolution data of mobile regions of membrane proteins embedded in lipid bilayers. As an inherently complex system, few techniques can capture information about the mobile portions of an otherwise immobilized protein. The nature of crystallography and solid-state NMR relies on structural rigidity. Solution-state NMR relies on overall mobility of a protein for resolution. In the middle regime, there are few solutions to study these systems. The inward-rectifying, pH-gated potassium channel KcsA from Streptomyces lividans makes an excellent model for the development of methods to study mobile regions of membrane proteins. Of its 160 residues, more than a third are in extracellular do- mains and are not typically captured by solid-state NMR or crystallographic techniques. These pages present evidence that KcsA’s C-terminus is highly mobile and becomes increasingly dynamic when the protein is at low pH and high K+ concen- tration, where the channel is known to be active. By applying proton-detected, high-resolution magic angle spinning NMR (HR-MAS) to fractionally deuterated KcsA, previously unattainable correlations are collected and new resonance assignments are made, demonstrating the utility of the technique. The lipid environment is well known to regulate the function of KcsA in particular and membrane proteins in general. It is generally assumed that reconstituting KcsA into a synthetic phospholipid membranes provides the protein a well-defined environment. Data is presented here which shows that KcsA co-purifies with phosphoglycerol lipids from the E. coli membrane and that these molecules are 13C enriched in the course of isotopically labeling KcsA. Further, significant hydrolysis of both co- purifying and synthetic lipids occurs under ordinary experimental conditions. These findings demand that routine analysis of samples must include verification of the chemical integrity of lipids. Finally, the feasibility of applying dynamic nuclear polarization-enhanced NMR (DNP) to KcsA is investigated as a means of elucidating information about its termini. Although KcsA is known to enhance poorly by DNP, data presented here show that this is not an intrinsic property of the protein but rather an effect of the matrix in which KcsA is investigated. The use of a 15N-enriched free amino acid dissolved into buffers used for DNP is shown to be a powerful diagnostic internal standard.

No description available.

A lo largo de la última década hemos asistido a un desarrollo sin precedentes de las tecnologías de la salud. Los avances en la informatización, la creación de redes, las técnicas de imagen, la robótica, las micro/nano tecnologías, y la genómica, han contribuido a aumentar significativamente la cantidad y diversidad de información al alcance del personal clínico para el diagnóstico, pronóstico, tratamiento y seguimiento de los pacientes. Este aumento en la cantidad y diversidad de datos clínicos requiere del continuo desarrollo de técnicas y metodologías capaces de integrar estos datos, procesarlos, y dar soporte en su interpretación de una forma robusta y eficiente. En este contexto, esta Tesis se focaliza en el análisis y procesado de señales biomédicas y su uso en problemas de clasificación automática. Es decir, se focaliza en: el diseño e integración de algoritmos para el procesado automático de señales biomédicas, el desarrollo de nuevos métodos de extracción de características para señales, la evaluación de compatibilidad entre señales biomédicas, y el diseño de modelos de clasificación para problemas clínicos específicos. En la mayoría de casos contenidos en esta Tesis, estos problemas se sitúan en el ámbito de los sistemas de apoyo a la decisión clínica, es decir, de sistemas computacionales que proporcionan conocimiento experto para la decisión en el diagnóstico, pronóstico y tratamiento de los pacientes. Una de las principales contribuciones de esta tesis consiste en la evaluación de la compatibilidad entre espectros de resonancia magnética (ERM) obtenidos mediante dos tecnologías de escáneres de resonancia magnética coexistentes en la actualidad (escáneres de 1.5T y de 3T). Esta compatibilidad se evalúa en el contexto de clasificación automática de tumores cerebrales. Los resultados obtenidos en este trabajo sugieren que los clasificadores existentes basados en datos de ERM de 1.5T pueden ser aplicables a casos obtenidos con la nueva tecnolog / Fuster García, E. (2012). Biomedical signal analysis in automatic classification problems [Tesis doctoral]. Editorial Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/17176 / Palancia

Arctic soils contain a large fraction of our planets terrestrial carbon (C) pool. When tundra soils become warmer and permafrost thaws, non-native geoengineering earthworms can enter these soils and ingest organic matter accumulated over long timescales. Previous studies have found that earthworms increase mineralization rates of soil organic matter into carbon dioxide (CO2) when introduced. Yet, this initial mineralization boost seems transient with time and it has been hypothesized that earthworms stimulate the formation of persistent C forms. In this study, I investigated how non-native, geoengineering earthworms affected the relative proportions of seven carbon forms in the O and A1 horizon of tundra soil and if their effect induced a change in pH. I used Nuclear Magnetic Resonance (NMR) spectroscopy to understand what happens to soil carbon compounds in two different tundra vegetation types (heath and meadow), that had been subjected to earthworm treatment for three summers. I found that O-aromatic C increased from 7.22% ± 0.24 (mean ± stderr) in the meadow soil lacking earthworms to 8.98% ± 0.30 in the meadow exposed to earthworms, and that aromatic C increased from 8.71% ± 0.23 to 9.93% ± 0.25. In similar, the result suggested that alkyl C decreased in this vegetation type from 20.43% ± 0.38 to 18.70% ± 0.25 due to earthworm activities. I found no effect on the chemical properties in the heath. I conclude that geoengineering earthworms affect the two vegetation types differently and that earthworms seem to enhance the accumulation of recalcitrant aromatic C forms.

Ribonuclease HI (RNase HI), a ubiquitous, non-sequence-specific endonuclease, cleaves the RNA strand in RNA/DNA hybrids. The enzyme has roles in replication, genome maintenance, and is the C-terminal domain of retroviral multi-domain reverse transcriptase (RT) proteins. Murine Leukemia Virus (MLV) and Human Immunodeficiency Virus (HIV) are two such retroviruses and their RNase HI (RNHI) domains are necessary for viral replication, making it an attractive drug target. RNase HI has a “handle region”, an extended loop with a large cluster of positive residues, that is critical for substrate recognition. MLV-RNHI is active in isolation and contains a handle region, but, HIV-RNHI is inactive in isolation and does not contain a handle region. HIV-RT, however, has a region in its polymerase domain (positive charge cluster and aromatic cluster) that makes contact with the RNHI domain that may be serving as a “pseudo” handle region; additionally, insertion of a handle region into isolated HIVRNHI restores its activity. Overall, a breadth of information exists on this region’s dynamics, but important gaps remain unfilled; gaps that may potentially lead to creating effective drugs to treat the above-mentioned viruses. Solution-state nuclear magnetic resonance (NMR) spectroscopy combined with Molecular Dynamic (MD) simulations suggest a model in which the extended handle region domain of the mesophilic Escherichia coli RNHI (EcRNHI) populates "open" (substrate-bindingcompetent) and "closed" (substrate-binding incompetent) states, while the thermophilic Thermus thermophilus RNHI (TtRNHI) mainly populates the closed state at 300 K. In addition, an in silico designed mutant Val98Ala (V98A) EcRNHI was predicted to populate primarily the closed state. Understanding the structural features and internal motions that lead RNase HI to adopt these various conformers is of central importance to better understanding RNase HI’s role in retroviral infection. To formulate a comprehensive model on handle region dynamics, an integrative approach of NMR spectroscopy, X-ray crystallography, and MD simulations is employed. The sensitivity to internal conformational dynamics at multiple time scales of NMR spectroscopy, molecular range and resolution of X-ray crystallography, and structural interpretations of dynamic processes by MD simulations create a synergistic trio capable of tackling this issue. First, the in silico 2-state Kinetic model is validated through NMR observables that correlate with the respective conformers, thus serving as experimental analogs. The NMR parameters also correlate with the Michaelis constants (KM) for RNHI homologs and help to confirm the in silico predictions of V98A EcRNHI. This study shows the important role of the handle region in modulation of substrate recognition. It also illustrates the power of NMR spectroscopy in dissecting the conformational preferences underlying enzyme function. Next, a deeper dive is taken into handle region dynamics, specifically focusing on residue 88 and the impact its identity has on this region. Its sidechain interactions are shown to directly correlate with handle region conformations and helps to amend the originally proposed in silico 2-state Kinetic model. Lastly, looking at RNHI handle region dynamics through an evolutionary lens opens the door to uncovering novel mutations that have been previously overlooked or not identified. Through a phylogenetic analysis, researchers have reconstructed seven ancestral RNHI mutants and three of them have been expressed here. The sequence identity of these three ancestral mutants range from 60-87% to extant homologs and this is reflected by similar peak positions in their 15N HSQC spectra. Requisite experiments to assign the NMR backbone have been completed.

Includes bibliographical references (leaves 73-79). / In vivo magnetic resonance spectroscopy (MRS) is an imaging technique that allows the chemical study of human tissue non-invasively. The method holds great promise as a diagnostic tool once its reliability has been established. Inter-scanner variability has, however, hampered this from happening as results cannot easily be compared if acquired on different scanners. In this study a phantom was constructed to determine the localisation efficiency of the 3 T Siemens Allegra MRI scanner located at the Cape Universities Brain Imaging Centre (CUBIC). Sufficient localisation is the key to acquiring useful spectroscopic data as only the signal from a small volume of interest (VOI) is typically acquired. The phantom consisted of a Perspex cube located inside a larger Perspex sphere. Solutions of the cerebral metabolites N-acetyl aspartate (NAA) and choline (Cho) were placed in the inner cube and outer sphere respectively. The phantom was scanned at a range of voxel sizes and echo times in order to determine parameters that typically indicate the performance of the scanner in question. The resultant full width at half maximum (FWHM) and signal to noise ratio (SNR) values indicated that optimal results were obtained for a voxel with dimensions 20 x 20 x 20 mm3. The selection efficiency could not be measured due to limitations in the scanner, but two other performance parameters ' extra volume suppression (EVS) and contamination ' could be determined. The EVS showed that the scanner was able to eliminate the entire background signal from the out-of-voxel region when voxel sizes with dimensions (20 mm)3 and (30 mm)3 were used. This performance decreased to 96.2% for a voxel size of (50 mm)3. The contamination indicated that the unwanted signal, weighted by the respective proton densities of the chemicals, ranged from 12% in the (20 mm)3 voxel to 24% in the (50 mm)3 voxel. These ranges are well within acceptable limits for proton MRS. Analysis of the water suppression achieved in the scanner showed an efficiency of 98.84%, which is acceptable for proton spectroscopy. It was also found that manual iv shimming of the scanner improved the spectra obtained, as compared to the automated shimming performed by the scanner. The second objective of the study was to quantify absolute metabolite concentrations in the familiar SI units of mM as results were previously mostly expressed as metabolite ratios. The LCModel software was used to assess two methods of determining absolute metabolite concentrations and the procedure using water scaling consistently showed superior performance to a method using a calibration factor. The method employing water scaling was then applied to a study of fetal alcohol spectrum disorder (FASD) where the deep cerebellar nuclei of children with FASD and a control group were scanned. The cerebellar nuclei were of interest as children with FASD show a remarkably consistent deficit in eye blink conditioning (EBC). The cerebellar deep nuclei is known to play a critical role in the EBC response. The results show significant decreases in the myo-inositol (mI) and total choline (tCho) concentrations of children with FASD in the deep cerebellar nuclei compared to control children. The FAS/PFAS subjects have a mean mI concentration of 4.6 mM as compared to a mean of 5.3 mM in the controls. A Pearson correlation showed that there was a significant relationship between decreasing mI concentrations with increasing prenatal alcohol exposure. The mean tCho concentrations are 1.3 mM for FAS/PFAS and 1.5 mM for the controls. There was no significant differences between the heavily exposed group and either the FAS/PFAS or the control subjects for either metabolite. The decreased mI and tCho concentrations may indicate deficient calcium signalling or decreased cell membrane integrity ' both of which can explain the compromised cerebellar learning in FASD subjects.

The IR spectra of twenty-one transition metal complexes of 8-hydroxyquinoline over the range 700 - 50 cm⁻¹ are discussed in relation to their known or inferred structures. The complexes are of three types: (a) the bis(aquo) complexes of the first row transition metal(II) ions, [M(ox)₂(H₂O)₂] (M =Mn, Fe, Co, Ni, Cu, Zn); (b) the corresponding anhydrous complexes, [M(ox)₂]n (M=Mn, Co, Ni, Cu, Zn) and (c) the complexes of the metal(III) ions, [M(ox)₃] (M = Sc, V, Cr, Mn, Fe, Co, Ga, Rh and In). Deuterated 8-hydroxyquinoline was. synthesized by the Skraup synthesis and has been used to assist in the assignment of the metal-ligand modes. The assignment of these bands was further based on ⁶⁴,⁶⁸Zn labellihg of the bis(aquo) zinc chelate and on the effects of metal ion substitution in relation to structural considerations based on crystal field theory. An investigation of the IR spectra of a series of -tris, bis and mono(8-aminoquinoline) complexes of the first transition row metal(II) perchlorates and halides is reported.

The time-dependent visible spectra and the crystal structure of [Fe2(C20H24N8O2)(CH3CN)4]·PF6 (diketo-dimer) were studied. The spectra showed that the most significant chemistry occurred during the initial 1.5 hours of the synthetic reaction. The starting materials 343 nm peak shifted to a lower energy, at 360 nm, and a new shoulder appeared at 490 nm. This change suggests the formation of a new intermediate whose spectrum has an exceptional resemblance to the starting materials mixed valent species, [Fe2(TIED)(Cl)4]+1 (TIED = tetraiminethylene dimacrocycles). Two isosbestic points were found at 538 and 371 nm. The diketo-dimer's crystals appear to have individual colors, a physical characteristic called pleochroism. Pleochroism is a topic in the study of optical crystallography which is discussed and applied to the diketo-dimer. The extinction angle was estimated to be 14°, a value consistent for triclinic crystals. X-ray crystallography found that the diketo-dimer is triclinic, and has a space group of P-1. A noteworthy feature is the bond length, 1.406 Å, between the two linking bridgehead carbons. This bond length matches the value for partial double bonds of aromatic compounds. This argues for a delocalized electron circulating within the macrocycle. The NMR spectra of a diiron macrocycle, [Fe2(TIED)(CH3CN)4]4+, were examined. Temperature dependent, pH dependent, D+ substitution, selectively decoupled, and COSY 1H NMR experiments were performed. Two sets of structural equilibria were found. One set is temperature dependent, and the other is pH dependent. Of particular interest are the peaks centered at 9.7 ppm and assigned to the imine carbon protons H2. Its resonance indicates an imine proton in an extensively conjugated aromatic environment with an electron deficient metal.