Dissertations / Theses on the topic 'Linear dichroism'

Circular dichroism (CD) is an important technique in the structural characterization of proteins, and especially for secondary structure determination. The CD of proteins can be calculated from first principles using the matrix method, with an accuracy that is almost quantitative for helical proteins. Thus, for proteins of unknown structure, CD calculations and experimental data can be used in conjunction to aid structure analysis. The vacuum-UV region (below 190 nm), where charge-transfer transitions have an influence on the CD spectra, can be accessed using synchrotron radiation circular dichroism (SRCD) spectroscopy. Calculations of the vacuum-UV CD spectra have been performed for 71 proteins, for which experimental SRCD spectra and X-ray crystal structures are available. The theoretical spectra are calculated considering charge-transfer and side chain transitions, which significantly improves the agreement with experiment, raising the Spearman correlation coefficient between the calculated and experimental intensity at 175 nm from 0.12 to 0.79. The influence of the different conformations used for the calculation of charge-transfer transitions is discussed in detail, focussing on the effect in the vacuum-UV. Linear dichroism (LD) provides information on the orientation of molecules but is more challenging to analyze than CD. To aid the interpretation of LD spectra, the calculation of protein LD using the matrix method is established and the results compared to experimental data. The orientations of five prototypical proteins are correctly reproduced by the calculations. Using a simplified approach, matrix method parameter sets for the nucleic bases and naphthalenediimide (NDI) have been created and are used to determine DNA/RNA conformations and to study NDI nanotubes. Finally, to make CD and LD calculations available for the scientific community in an easy-to-use fashion, the web interface DichroCalc is introduced.

Existing methodologies for biomolecular detection are limited in several key areas. Heterogeneous assays struggle with various wash steps which can prolong assay time, while other assays require costly reagents, lack mobility and can be highly complex in nature. This project demonstrates how a bio-nano particle in the form of M13 bacteriophage (M13) can be used for the basis of a novel homogeneous immunoassay which incorporates the use of linear dichroism spectroscopy (LD). M13 has a high aspect ratio which allows it to align easily in shear flow, this in turn generates a large LD signal. This property of M13 has been manipulated for use in a new in-vitro diagnostic technique. Existing M13 production yields are much lower than those required for this assay. A new method was developed which increased the yield 10 fold. Chemical modifications were made by covalently attaching chromophores, this enabled the M13 LD signal to be visualised in the visible region and develops the potential for multiplexing. By chemically modifying M13 with chromophores and antibodies it was possible to create an assay capable of detecting 10⁵ cells/mL of Escherichia coli O157. This is 100 times more sensitive than the M13 based assay developed by Pacheco-Gomez et al. (2012). The assay was reassembled to detect small molecules and was found to have a sensitivity of 0.01 mM. The assays presented form a sensitive, specific, fast diagnostic tool capable of detecting pathogens and small molecules. It offers significant improvements over existing methods, and could act as a platform in developing a multimodal detection system.

Though Parkinson’s Disease is known to be caused by cell death in one region of the brain, and though the protein α-synuclein is known to be associated with it, the causes are still poorly understood. It has not yet been shown how α-synuclein may cause cells to die, with research focusing on the range of structures the protein is able to adopt. The classic amyloid fibrils are currently believed to be non-toxic, but smaller, soluble oligomers appear to be the toxic species. Key to toxicity and to the normal function of α-synucein (also unknown to-date) appear to be the ability of the protein to bind to lipids, as toxicity may be due to oligomers forming pores in cells, and the normal function of the protein may be in vesicle transport. The work presented in this thesis represents a collection of studies, across several disciplines, that test aspects of the behaviour of α-synuclein. Circular dichroism and fluorescence data presented here show that the protein interacts with the lipid POPS in a concentration-dependent manner. Linear dichroism was an important and complementary technique to these, but required some method refinement and sample preparation improvement. This work is presented in this thesis, alongside experimental and theoretical studies into the behaviour of lipid vesicles in Couette flow. It was shown that certain lipids perform better in Couette flow, particularly the mixture of POPC and POPS. This informed the linear dichroism studies, and enabled experiments suggesting that oligomers of α-synuclein may insert across the membrane of vesicles. If confirmed, this would support the amyloid pore theory of α-synuclein toxicity.

La microscopie de fluorescence a récemment été complétée par une technique appelée dichroïsme linéaire résolu angulairement, basé sur le fait que l'absorption de la lumière est un processus sensible à l'orientation moléculaire. En analysant la réponse d'émission de fluorescence en fonction de l'orientation de la polarisation de la lumière excitatrice, cette technique permet de remonter à l'information d'orientation sur un ensemble de molécules fluorescentes, plus précisément son angle d'orientation moyenne et l'amplitude de ses fluctuations angulaires autour de cette moyenne. Dans cette thèse, nous mettons en oeuvre de nouvelle méthodes et instrumentations capables d'améliorer la robustesse et la rapidité de l'analyse de données de réponses résolues en polarisation, la vitesse de l'acquisition de données, et d'explorer la possibilité de mesurer l'orientation 3D de molécules. Nous proposons une méthode capable de mesurer les propriétés d'orientation de sondes lipidiques fluorescentes par l'utilisation d'un disque de Nipkow couplé à une imagerie par caméra, et combiné avec la modulation rapide de la polarisation par modulateur électro-optique. Une nouvelle méthode de traitement de données est développée pour considérablement améliorer la rapidité et la précision de l'information par une étude des sources de bruit et d'incertitude, dues au bruit et aux facteurs instrumentaux. Cette technique a été testée avec succés sur des vésicules géantes uni-lamellaires et sur cellules vivantes, marquées par les sondes lipidiques DiIC18 et di-8-ANEPPQ. Cette méthode est capable d'acquérir une information précise sur l'orientation moléculaire à une cadence d'une image par seconde. Enfin, afin de sonder de manière non ambiguë l'orientation 3D d'un ensemble de molécules, une nouvelle méthode est proposée, supportée par des simulations numériques, basée sur la variation hors plan de la polarisation d'excitation dans le volume focal par une somme cohérente de champs polarisés linéairement et radialement
Based on the fact that the absorption of light is a molecular-orientation sensitive process, fluorescence microscopy has been recently completed by a technique called angle-resolved linear dichroism. By analyzing the fluorescence emission response with respect to the polarization orientation of the exciting light, this technique allows retrieving orientation information of an ensemble of fluorescent molecules, namely the average orientation angle and the amplitude of the angular fluctuations around this average. In this PhD thesis, we implement new methods and instrumentation tools able to improve the robustness and speed of the polarization resolved data analysis, the rate of the data acquisition, and at last to explore the possibility to record molecular 3D orientation information. A scheme able to monitor the real-time orientation properties of fluorescent lipid probes is proposed using a high-speed spinning disk coupled to camera imaging, combined with fast switching of the polarization state by an electro optical modulator. A new data processing method is developed which considerably improves the speed and the precision of the retrieved information by investigating the sources of bias and uncertainty due to noise and instrumentation factors. The technique has been successfully tested on giant unilamellar vesicles and on living cells labeled with different fluorescent lipid probes, DiIC18 and di-8-ANEPPQ. It was able to acquire precise molecular orientation images at full frame rates in the range of one frame per second. At last in order to probe unambiguously the 3D orientation information of an ensemble of molecules, a new method is proposed and supported by simulations, based on the out-of-plane tuning of the excitation polarization realized in the focusing volume by coherently summing linearly and radially polarized fields

Within this thesis the use of linear dichroism (LD) spectroscopy was utilised as a new platform for the development of diagnostic sensors. To develop a novel diagnostic sensor in combination with LD spectroscopy a particle with a high aspect ratio is need. Such a particle is M13 bacteriophage, this micron long molecular scaffold is easily alignable in shear flow generating a large LD signal. Using LD spectroscopy, it was possible to demonstrate the successful detection of multiple DNA targets, and that the sensor can also discriminate between DNA sequences differing in length, with a limit of detection (LOD) of 2 M, competitive with the current non amplification methods of DNA detection. The reversibility and regeneration of the sensor were also investigated. Finally, the development of an M13 bacteriophage sensor for the detection of proteins was designed in an unprecedented plug-and-play format. The assay was designed to detect thrombin, which is important in the monitoring of blood clotting disorders. LD spectroscopy enabled the detection of thrombin with a LOD of 10 pM and with a dynamic range from 10 pM to 47 nM.

Linear dichroism (LD) spectroscopy is a technique which uses polarised light to make inferences on the relative orientations of chemical groups within a molecule. A requirement for experiments using this technique is the alignment of molecules in the analyte. This thesis considers the use of fluid flows to align molecules. The use of bead-spring and bead-rod models to simulate the behaviour of polymers and particles in fluid flows has been extensively discussed in the literature. In this work, the FENE-Fraenkel spring force law is implemented in bead-spring chain simulations, to assess the alignment behaviour of molecules in flow. Analytical properties of FENE-Fraenkel springs are derived and used to inform the coding of the simulation programme and to assess its outputs. The results of these simulations were comparable to both analytical results in zero-shear conditions and to data previously published by Hsieh et al. [48] on bead-spring chains in extensional flow, albeit with a reduced signal-to-noise ratio. Simulations of the alignment of FENE-Fraenkel spring dumbbells in shear flows show increased alignment with high shear rates, similar to modelling data published by McLachlan et al. [89]. This work also considers the use of microfluidic devices manufactured from polydimethylsiloxane to perform linear dichroism studies on DNA and vesicles in solution. A wide channel and cross-slot device are developed to provide conditions for pressure-driven and extensional flow to align molecules. Both devices are shown to align DNA molecules to measure an LD signal, with the cross-slot performing better than the wide channel device. With solutions of vesicles containing 1,6-Diphenylhexa-1,3,5-triene, a membrane probe, the wide channel device is shown to provide weak vesicle alignment to measure an LD signal. However, both microfluidic devices are unable to align molecules as well as Couette flow, the method normally used to align molecules for LD experiments.

All biological processes are fundamentally inter-molecular interactions. In order to understand, and hence control, biomolecular structure and function, methods are required that probe biological systems at the molecular level, ideally with those molecules being in their native environment. The research summarized herein has at its core the development and application of ultra violet (UV)-visible spectrophotometric techniquies for this prupose, in particular circular dichrosim (CD) and linear dichrosim (LD) but also absorbance, fluorescence and resonance light scattering. The spectroscopy is complemented by fundamental theoretical work on molecular structure and reactivity that forms the basis for designing molecules to bind to biomolecules for a particular structural or functional effect. A brief summary of the contributions of the listed publications to our understanding of 'Molecular aspects of biololecule structure and function' is given below under five headings: Circular dichroism theory Molecular geometry and reactivity Small molecule-macromolecule interactions: spectroscopic probes of inter-molecular geometries Molecular design for nucleic acid structure and control Spectroscopic probes of biomolecule structure: instrumentation and application In general terms these correspond to successive phases of the research programme, however, all areas have been present since the first publications in 1983 and can be traced weaving through all subsequent activity.

All biological processes are fundamentally inter-molecular interactions. In order to understand, and hence control, biomolecular structure and function, methods are required that probe biological systems at the molecular level, ideally with those molecules being in their native environment. The research summarized herein has at its core the development and application of ultra violet (UV)-visible spectrophotometric techniquies for this prupose, in particular circular dichrosim (CD) and linear dichrosim (LD) but also absorbance, fluorescence and resonance light scattering. The spectroscopy is complemented by fundamental theoretical work on molecular structure and reactivity that forms the basis for designing molecules to bind to biomolecules for a particular structural or functional effect. A brief summary of the contributions of the listed publications to our understanding of 'Molecular aspects of biololecule structure and function' is given below under five headings: Circular dichroism theory Molecular geometry and reactivity Small molecule-macromolecule interactions: spectroscopic probes of inter-molecular geometries Molecular design for nucleic acid structure and control Spectroscopic probes of biomolecule structure: instrumentation and application In general terms these correspond to successive phases of the research programme, however, all areas have been present since the first publications in 1983 and can be traced weaving through all subsequent activity.

The magnetization profile of iron oxide magnetic nanoparticles (MNPs) and the binding ability of MNPs with metal ions in aqueous solutions were investigated by means of the magnetic linear dichroism (MLD) and the magneto-optical Kerr effect (MOKE) measurements. The ultra-visible MLD spectra of the non-modified and surface modified MNPs with -COOH showed a typical magnetization curve for a superparamagnetic particles. Also, the MLD spectra were changed drastically by the addition of dysprosium(III) ion and showed that MLD spectra can be used for the evaluation of metal-MNPs binding ability. Furthermore, the magnetization of MNPs solution was measured sensitively by the transversal MOKE measurements, where a polarized beam was reflected at the interface of prism/solution of MNPs.

This study will focus on strain induced by thermal history and thickness on antiferromagnetic epitaxial LaFeO3(110) thin films grown on Nb:SrTiO3(001) substrates. Lattice mismatching between film and substrate induce strain in the film lattice during growth of the film. This strain can be relaxed by thermal treatment after growth and may show tendencies in the domain size, shape and distribution related to strain. Thin films are grown in different thicknesses by rf magnetron sputtering and selected films are relaxed by thermal anneal treatment. The samples are investigated in room temperature and in heating experiments to obtain images of antiferromagnetic domain contrast along the L2 and L3 absorbtion egde of Fe by XMLD spectromicroscopy. Size of domains areas and contrast level are measured and related to the strain in the thin films. Averaged spectra of L2 absorbtion edge antiferromagnetic contrast showed a clearly observable domain contrast with consistently shaped energy spectrums. No difference in antiferromagnetic contrast due to lattice strain caused by lattice mismatching was observed. All as-grown samples showed comparable size, distribution and shape of antiferromagnetic domains on both polished and etched substrates. Relaxation of prepared samples in 1000±C for 12 hours in a 1 atm atmosphere of oxygen gave a distinct increase in size for the domains. Area calculation show a increase from typically 0.2 ¹m2 for as-grown films to 2 ¹m2 for relaxed films. Heating experiments estimate a Néel temperature of 625oK for as-grown films and 740oK for relaxed samples. The thermal anneal thus has a significant effect on LaFeO3 thin films as the domains increase in size and the Néel temperature in which the film is no longer antiferromagnetic increases to the Neel temperature of bulk LaFeO3. This preparation approach may be useful for further investigations of the exchange-bias effect.

This thesis focuses on peptides derived from the Prion, Doppel and Influenza haemagglutinin proteins in the context of bilayer interactions with model membranes and live cells. The studies involve spectroscopic techniques like fluorescence, fluorescence correlation spectroscopy (FCS), circular and linear dichroism (CD and LD), confocal fluorescence microscopy and NMR.

The peptides derived from the Prion and Doppel proteins combined with their subsequent nuclear localization-like sequences, makes them resemble cell-penetrating peptides (CPPs). mPrPp(1-28), corresponding to the first 28 amino acids of the mouse PrP, was shown to translocate across cell membranes, concomitantly causing cell toxicity. Its bovine counterpart bPrPp(1-30) was demonstrated to enter live cells, with and without cargo, mainly via macropinocytosis. The mPrPp(23-50) peptide sequence overlaps with mPrPp(1-28) sharing the KKRPKP sequence believed to encompass the driving force behind translocation. mPrPp(23-50) was however found unable to cross over cell membranes and had virtually no perturbing effects on membranes.

mDplp(1-30), corresponding of the first 30 N-terminal amino acids of the Doppel protein, was demonstrated to be almost as membrane perturbing as melittin. NMR experiments in bicelles implied a transmembrane configuration of its alpha-helix, which was corroborated by LD in vesicle bilayers. The positioning of the induced alpha-helix in transportan was found to be more parallel to the bilayer surface in the same model system.

Positioning of the native Influenza derived fusion peptide in bilayers showed no pH dependence. The glutamic acid enriched variant however, changed its insertion angle from 70 deg to a magic angle alignment relative the membrane normal upon a pH drop from 7.4 to 5.0. Concomitantly, the alpha-helical content dramatically rose from 18% to 52% in partly anionic membranes, while the native peptide’s helicity increased only from 39% to 44% in the same conditions.

This thesis explores and expands upon theoretical means of quantifying a number of nonlinear spectroscopies, including two-photon absorption, resonant-inelastic x-ray scattering, Jones birefringence, and magnetic circular dichroism. On top of that, detailed information is given for the derivation and program implementation of damped cubic response functions. Complex-valued cubic response functions have been implemented in the quantum chemistry package DALTON, based on working equations formulated for an approximate-state wave function. An assessment of the implementation, such that for small frequencies the second-order hyperpolarizability should behave according to an analytic function that depends quadratically on the optical frequencies. It is demonstrated how two-photon absorption (TPA) can be described either through second-order transition moments or via the damped cubic response function. A few calculated TPA profiles are produced for a set of smaller molecules, in order to display the capability of the cubic response function in the x-ray frequency region. Resonance-inelastic x-ray scattering (RIXS) is explored in a similar manner as two-photon absorption. It is shown how the second-order hyperpolarizability can represent RIXS in the limit of intermediate-state and final-state resonances. Complications emerging from the complex dispersion of the hyperpolarizability are discussed. Moreover, linear birefringences, with focus on the Jones birefringence, are investigated for noble gases, monosubstituted benzenes, furan homologues, and a pure acetonitrile liquid. A linear relation between the Jones birefringence and the empirical para-Hammett constant as well as the permanent electric dipole moment is presented. Estimations of three linear birefringences --- Kerr, Cotton--Mouton, and Jones ---are obtained by averaging over a set of liquid snapshots. The Jones effect for acetonitrile turns out to be unusually large inmparison to the other two investigated linear birefringences. The final chapter of the thesis investigates magnetically induced circular dichroism (MCD). A question regarding relative stability of the first set of excited states for DNA-related molecular systems is resolved through MCD by exploiting the signed nature of circular dichroisms. Furthermore, to what extent solvent contributions affect MCD spectra is explored. The effect on uracil MCD spectrum due to thionation is studied, for which the degree of redshifting for systems 2-thiouracil and 4-thiouracil can be seen to be addative as compared to the 2,4-dithiouracil system.

QC 20171129

Dans cette thèse, les propriétés optiques de biomolécules importantes ont été étudiées en utilisant une approche théorique et, dans un cas, aussi expérimentale. La Théorie de la fonctionnelle de la densité (DFT) et la timedependent-DFT (TD-DFT) sont les principales méthodes de chimie quantique utilisées dans cette thèse. Plusieurs spectroscopies ont été étudiées (au niveau théorique et, dans certains cas, également au niveau expérimental) : absorption électronique linéaire (absorption à un photon, OPA) et non-linéaire (absorption à deux ou trois photons, TPA et 3PA), dichroïsme circulaire électronique (DCE) et spectroscopie de fluorescence. Les effets de l’environnement, particulièrement importants dans des systèmes biologiques, ont été pris en compte, pour les propriétés de l’état fondamental et des états excités en utilisant une méthode multi-échelles QM/MM appelée Polarizable Embedding (PE). L’échantillonnage des conformations a été pris en compte avec des simulations de dynamique moléculaire (MD) qui sont basées sur la mécanique classique. Deux thématiques ont été étudiées dans cette thèse : le cholestérol et le design in silico de ses analogues fluorescents ainsi que la caractérisation des coudes de type β dans différentes conformations grâce à la simulation des spectres DCE. La simulation de plus d’une spectroscopie a été importante dans l’étude des états excités du cholestérol dans des solutions organiques. Le design in silico a suggéré un nouveau stérol-polyénique (P-stérol) qui montre despropriétés optiques améliorées pour le mécanisme d’excitation à trois photons par rapport au déhydroergostérol (DHE), une sonde du cholestérol déjà très utilisée. Ce nouveau P-stérol a été suggéré pour la synthèse. L’étude des spectres de DCE des coudes β en différentes conformations a mené une double conclusion : même si deux allures de DCE pour les conformations des coudes β étudiées (4) ont été trouvées (dans la majorité des cas), la spectroscopie de DCE doit toujours être associée à d’autres techniques spectroscopiques dans la caractérisation en solution des coudes β
In this thesis, the optical properties of important biomolecules were studied using a theoretical approach and, in one case, also an experimental one. Density Functional Theory (DFT) and time-dependent-DFT (TD-DFT), were the principal quantum chemical methods adopted in this thesis.Various spectroscopies were studied (theoretically and, in some cases, also experimentally) : linear (one-photon, OPA) and non-linear (two- and three-photon, TPA and 3PA) electronic absorption, electronic circular dichroism (ECD) and fluorescence spectroscopy. The environment effects, which are particularly important in biological systems, were taken into account, for both ground and excited states properties, using a multiscale QM/MM method called Polarizable Embedding (PE). The sampling of conformations was addressed by Molecular Dynamic (MD) simulations based on classical mechanics. Two topics were studied in this thesis: cholesterol and the in-silico design of its fluorescent analogues, and the characterization of β-turns in different conformations by simulations of their ECD spectra in aqueous solutions. The simulation of more than one spectroscopy resulted to be important when studying the electronic excited states of cholesterol in organic solutions. The in-silico design study suggested a novel polyene-sterol (P-sterol) which shows improved optical properties for the three-photon excitation mechanism with respect to dehydroergosterol (DHE), an already widely used cholesterol probe. This new P-sterol was thus suggested for synthesis. The achievement from the study of ECD spectra for different β-turn conformations is two-fold: even if two ECD patterns for the β-turn conformations studied (4) were found (in most of cases), ECD spectroscopy should be always associated with other spectroscopic techniques when trying to characterize the β-turn conformations in solutions

L'objet de ce travail est l'etude theorique et experimentale des proprietes d'optique non lineaire (activite optique non lineaire) d'une solution de molecules chirales, en considerant principalement les effets resonnants du troisieme ordre analogues a l'effet kerr, et en particulier le dichroisme circulaire non lineaire. La resolution de l'equation de propagation non lineaire est effectuee pour un seul faisceau puis pour des configurations de type excitation-sondage, en incluant la non localite dans la reponse non lineaire. Des choix judicieux de polarisation permettent de decoupler dans une large mesure les effets non locaux des effets de dichroisme induits par l'effet kerr. On considere ensuite l'etude microscopique de l'activite optique non lineaire en developpant un formalisme classique puis quantique base sur l'extension des hyperpolarisabilites aux termes dipolaires magnetiques et quadrupolaires electriques. Ce formalisme est applique au cas de systemes simples, en particulier a trois niveaux. Des mesures du dichroisme circulaire en fonction de l'intensite incidente sont realisees a un puis deux faisceaux laser dans une solution d'ions complexes metalliques ruthenium(ii) tris-bipyridil chiraux. Les impulsions laser femtosecondes utilisees, accordables en frequence autour de 470 nm, sont generees a partir d'un amplificateur parametrique optique. Les experiences pour les deux enantiomeres et le melange racemique mettent en evidence une reponse non lineaire (saturation) du dichroisme circulaire. Une configuration d'excitation-sondage ou seule la polarisation du faisceau pompe est modulee permet en particulier de s'affranchir des effets de polarisation lineaires. Ces resultats sont en tres bon accord quantitatif avec une modelisation microscopique de l'ion etudie par un systeme d'oscillateurs couples a deux niveaux. Ils ouvrent des perspectives d'etude de modifications conformationnelles resolues temporellement a une echelle subpicoseconde.

Les systemes bidimensionnels metalliques (superreseaux et multicouches) sont probablement les futurs supports d'enregistrement a haute densite de stockage. Le dichroisme lineaire et circulaire de l'absorption x permet de caracteriser la structure atomique locale (exafs et xanes) et etudier le magnetisme d'interface. Le mxd est une technique nouvelle introduite a lure en 1989 juste apres les premieres etudes faites par une equipe allemande en 1987 le signal mxd est proportionnel a l'aimantation de l'element sonde, et permet de connaitre le sens du couplage dans un compose magnetique binaire. L'exafs et le xanes est une sonde atomique locale qui a permis de mettre en evidence des empilements nouveaux des metaux induits par la basse dimensionnalite dans les superreseaux et les multicouches

Dans certains semiconducteurs uniaxes, la birefringence naturelle s'annule en changeant de signe pour une longueur d'onde voisine du seuil d'absorption le plus bas. On peut alors realiser un transfert d'energie entre les modes ordinaire et extraordinaire en appliquant une perturbation convenable au cristal. Cette propriete a ete utilisee pour realiser, a partir de ces composes, des filtres a couplage de modes polarises. Toutefois, les proprietes dichroiques des echantillons n'ont jamais ete prises en consideration pour interpreter ce couplage. La partie theorique du travail presente ici, a consiste a calculer le champ electromagnetique d'une onde se propageant a travers un milieu birefringent, dichroique et soumis a une perturbation. Dans ces conditions, les modes propres de propagation ne verifient pas la condition d'orthogonalite. Pour resoudre ce probleme, nous avons generalise aux milieux perturbes, la methode decrite par jones pour les milieux non perturbes. Les expressions theoriques obtenues finalement mettent en evidence le couplage non negligeable entre le dichroisme lineaire et la perturbation au voisinage de la longueur d'onde annulant la birefringence. L'etude experimentale a ete effectuee sur deux types de cristaux, le thiogallate de cadmium et le sulfure de cadmium. Nous avons determine experimentalement d'une part, le pouvoir rotatoire du thiogallate de cadmium et d'autre part le coefficient elastooptique du sulfure de cadmium, a temperature ambiante et au voisinage de la longueur d'onde de birefringence nulle en tenant compte du couplage avec le dichroisme lineaire de ces echantillons et en determinant l'erreur effectuee en negligeant ce dernier

碩士
國立交通大學
物理研究所
104
We utilized synchrotron radiation linear polarized light to explore Sr3Cr2O8, CoO, Co (NO3)2 and SrCuO2 for its orbital ordering and electronic structure. In this thesis, Sr3Cr2O8 and CoO projects are supervised by Dr. Chun-fu Chang of Max Planck Institute, Dresden, Germany. For Sr3Cr2O8, we used linear polarized x-ray to measure the single-crystal sample for Cr L-edge absorption spectra to explore the chromium electronic structure and spin dimers formation, and Jahn - Teller Distortion. Along with the multiplet theory, we find the ground state wave functions, orbital ordering, as well as the electronic structure for Sr3Cr2O8. We also study cobalt oxide and cobalt nitrate using linear polarized light of Co L-edge x-ray absorption spectroscopy and deploy theoretical calculations in order to analyze the spectra. In the case of cobalt oxide, we use extendable substrate MICA in order to investigate the effect of the strain influence on the thin films. However, from the results, it seems that the mechanism for CoO growth on MICA is only related to Van deel war force. In the Co(NO3)2, cobalt ions have two different sites, which is divided by the way they are distorted. Assisted by the theoretical calculations, we find out that even the cobalt ions are in different local structure, they still show a total spin of 3 / 2 as the purely high-spin state. In addition, we also focus on SrCuO2. We want to investigate how the thickness of SrCuO2 influences the crystal field. It is an essential element for shaping superconductivity. Our result indicates that if the thickness is thinner than 10 unit cells, it grows in chain-like structure; while above 10 unit cells, it become plane-like structure leading to superconductivity with appropriate doping.

碩士
國立交通大學
光電工程系所
93
The Photoelastic Modulation Ellipsometer is used to measure an artificial circular polarizer (CP), which is a combination of a linear polarizer and a retarder. The multi-wavelength measurement technique is built up by using the Kr-Ar tunable laser. In the beginning, the azimuth direction of optical axis of this CP is roughly determined by comparing the theoretical model with its measurement for a single wavelength. The parameters: linear dichroism (LD), phase retardation and the azimuth angle between the linear polarizer and retarder (γ) are measured by fitting its 1st and 2nd harmonics signals of various azimuth positions of analyzer for the Kr-Ar tunable laser. Although the measured �� is independent of wavelength but it is not at 45o, and its phase retardation and LD are almost the same for every wavelength. Finally by comparing the theoretical transmission, which is derived by substituting its measured parameters, to its measured transmission under various azimuth positions of analyzer, we observed a well fitted result for every wavelength.

In this work we study thin epilayers of new antiferromagnetic semimetal CuMnAs by time- resolved magneto-optical experiments. In 10 nm layers of CuMnAs, we observed a harmonic dependence of the dynamical magneto-optical signal on the orientation of probe pulse linear polarization. This shows that in this 10 nm layer there is an in-plane uniaxial magnetic anisotropy which can be detected due to a quadratic magneto-optical effect - magnetic linear dichroism. From the measured data we also estimated the Néel temperature and the spectral variation of the magneto-optical coefficient describing the magnitude of the magnetic linear dichroism in this sample.

Probing orientations of fluorescent molecules embedded in or attached to cell membranes has a great potential to reveal information on membrane structure and processes occurring in living cells. In this thesis, we first describe one- and two-photon linear dichroism measurements on a fluorescent probe embedded in a phospholipid membrane with a well- defined lipid composition. On the basis of experimental data, we determine the distribution of the angle between the one-photon transition dipole moment of the probe and the membrane normal. At the same time, we perform molecular dynamics simulations of the fluorescent probe and quantum calculations of its one-photon and two-photon absorption properties. By comparing the orientational distribution gained from experiments with that predicted by simulations, we test the ability of linear dichroism measurements to report on the orientation of a fluorescent molecule in a lipid membrane. We also examine the applicability of molecular simulations as a basis for the interpretation of experimental data.

In these diploma thesis magnetically ordered materials are studied with the prospect of their application in spintronics. Specifically, we investigated metallic alloy FeRh, which undergoes a magnetic phase transition from antife- romagnetic phase to feromagnetic one around 100◦ C. This phenomenon can be readily used in memory devices. Laser spectroscopy is used as a nondestructive method without need of any electrical contacts. Magnetic properties of FeRh are studied by magnetooptical effects including quadratic magnetic linear dichroism. The measured polarization rotations are of the order of miliradians, therefore, the detection is realized by an optical bridge. At first, we concentrated on discrimina- ting of various magnetooptical effects from each other. The second part is focused on the phase transition induced by different means. Firstly, by heating the whole sample, secondly by illuminating the sample locally by continuous laser.

Spintronics is a new branch of electronics, which uses not only the charge of electron but also its spin for transfer and processing of information. For re- al applications it is necessary to understand, how the magnetic state of matter changes not only in time but also in space. This diploma thesis is therefore de- voted to the imaging of magnetic domains in ferromagnetic semiconductors by purely optical methods, which are based on the usage of the magneto-optical ef- fects. In first part of the thesis we concentrated on the optimalization of existing polarizing microscope, whose function is based solely on the polar Kerr effect. After the optimalization we try to use this setup to observe magnetic domains in ferromagnetic semiconductor GaMnAs by the means of the magnetic linear dichroism. 1

Ever since the discovery of exotic phenomena like high temperature (Tc) superconductivity in the cuprates and colossal magnetoresistance in the manganites, strongly correlated electron systems have become the center of attention in the field of condensed matter physics research. This renewed interest has been further kindled by the rapid development of sophisticated experimental techniques and tremendous computational power. Computation plays a pivotal role in the theoretical investigation of these systems, because one cannot explain their complicated phase diagrams by simple, exactly solvable models. Among the plethora of experimental techniques, various kinds of high energy electron spectroscopies are fast gaining importance due to the multitude of physical properties and phenomena which they can access. However the physical processes involved and the interpretation of the spectra obtained from these spectroscopies are extremely complex and require extensive theoretical modelling. This thesis is concerned with the theoretical modelling of a certain class of high energy electron spectroscopies, viz. the core-level electron spectroscopies, for strongly correlated systems of various kinds. The spectroscopies covered are Auger electron spectroscopy (AES), core-level photoemission spectroscopy (core-level PES) and X-ray absorption spec- troscopy (XAS), which provide non-magnetic information, and also X-ray magnetic circular and linear dichroism (XMCD and XMLD), which provide magnetic information. .

Ever since the discovery of exotic phenomena like high temperature (Tc) superconductivity in the cuprates and colossal magnetoresistance in the manganites, strongly correlated electron systems have become the center of attention in the field of condensed matter physics research. This renewed interest has been further kindled by the rapid development of sophisticated experimental techniques and tremendous computational power. Computation plays a pivotal role in the theoretical investigation of these systems, because one cannot explain their complicated phase diagrams by simple, exactly solvable models. Among the plethora of experimental techniques, various kinds of high energy electron spectroscopies are fast gaining importance due to the multitude of physical properties and phenomena which they can access. However the physical processes involved and the interpretation of the spectra obtained from these spectroscopies are extremely complex and require extensive theoretical modelling. This thesis is concerned with the theoretical modelling of a certain class of high energy electron spectroscopies, viz. the core-level electron spectroscopies, for strongly correlated systems of various kinds. The spectroscopies covered are Auger electron spectroscopy (AES), core-level photoemission spectroscopy (core-level PES) and X-ray absorption spec- troscopy (XAS), which provide non-magnetic information, and also X-ray magnetic circular and linear dichroism (XMCD and XMLD), which provide magnetic information. .

"Laser flash spectroscopy of photosystem II" Photosystem II (PS II) of plants and cyanobacteria oxidizes water in a light-powered reaction. Thereby, this protein is the ultimate source of the atmospheric oxygen. The capacity to oxidize water is owed to two properties of PS II: (i) The midpoint potential of the oxidizing chlorophyll moiety is increased by 0.6 V compared to photosystem I or photochemical reaction centers of anoxygenic bacteria, and (ii) the energy requirements of the four steps needed for the tetravalent oxidation of water are adapted to the energy of red light quanta. This thesis deals with two particular aspects, namely: 1. The coupling of the electron transfer from tyrosine Z (YZ) to the primary donor (P680+) to proton transfer, and an inquiry on the role of a positive charge on YZox (plus base cluster) in increasing the oxidizing potential at the catalytic site. 2. The localization of the electron hole, P680+, among the excitonically coupled four inner chlorophyll a molecules, and an estimation of the midpoint potential differences between them. Electron-proton-coupling by YZ This study was carried out with PS II core complexes from spinach or pea with a deactivated (removed) manganese cluster. The reduction of P680+ was investigated as a function of pH by detecting the laser flash induced absorption changes with nanosecond resolution. Two kinetic components were found with different pH-dependence and activation energies. The alteration of kinetic parameters by H/D isotope substitutions or by addition of divalent cations implied two different types of YZ-oxidation: At acidic pH the electron transfer was coupled with proton transfer, whereas in the alkaline region it was more rapid and no longer controlled by proton transfer. The conversion between both mechanisms occured at pH 7.4. This value corresponds either to the apparent pK of YZ itself (i.e. of the hydroxy group of the phenol ring) or to the pK of an acid-base-cluster, which includes YZ. Independent measurements of pH-transients by following the absorption changes of hydrophilic proton indicators corroborated this notion. The data were interpreted as indicating that the phenolic proton of YZ was released into the medium at acidic, but not at alkaline pH. The electron transfer and proton release characteristics of intact, oxygen-evolving PS II resembled those in deactivated samples kept at alkaline pH. We concluded that the electron transfer from YZ to P680+ in the native system was not coupled with proton transfer into the bulk. This has shed doubt on a popular hypothesis on the role of YZ as 'hydrogen abstractor' from bound water. On the other hand, the energetic constraints of water oxidation could be eased by the positive upcharging during oxidation of YZox plus its base cluster. On the localization of the electron hole of P680+ Photooxidation of PS II oxidizes the set of four innermost chlorophyll a molecules giving rise to the only spectroscopically defined species P680+. The deconvolution of difference spectra into bands of pigments is ambiguous. By using photoselective excitation of antennae, i.e. chl a molecules with site specific energies at the long wavelength border of the mean Qy-band, and by polarized detection, it was possible to tag P680+QA-/P680QA and 3P680/P680 difference spectra with a further parameter, the (wavelength-dependent) anisotropy r. Results obtained at liquid nitrogen temperature (77 K) can be clearly interpreted in terms of two chl a monomer bands. The two main components of the P680+QA-/P680QA difference spectrum were marked by two distinct values of the anisotropy and could be interpreted in a straightforward manner: the bleaching of a band at 675 nm belonging to the charged species (chl a+) and an electrochromic blue-shift of a nearby chl a from 684 to 682 nm. The main bleaching band of the 3P680/P680 spectrum (at 77 K) can be apparently attributed to a third (or several) chl a component(s). The analysis of the P680+QA-/P680QA spectrum at cryogenic temperature is compatible with monomeric chl a bands. On the other hand, one could assume a system of excitonically coupled core pigments, as it was recently introduced in the literature on the basis of energy transfer studies ('multimer model'). However, in view of the clear indications for an electrochromic band shift and the location of the bleaching band, which absorbs in a wavelength region of monomeric chl a, one assumption of the 'multimer model' should be questioned. Presumably, the excitonic couplings are rather weak, in particular between each of the two central chl a-molecules (PA/PB) and its respective accessory chl a (BA/BB), because of (i) the distances and (ii) different site energies of the monomeric chromophores. At room temperature, the absorption difference and anisotropy spectra of P680+QA-/P680QA were clearly altered. The anisotropy data indicated that the changes could no longer exclusively be ascribed to thermal broadening of individual bands. The localization of the positive charge on one pigment, analogous to the situation at 77 K, was now unlikely. Hence, the midpoint potential differences between the inner four chlorophyll a molecules were small and were estimated as approximately 15 meV.

Extremely important initial experiments on the interactions of DNA with various substances such as: anticancer, antibacterial and antifungal compounds started with creating a DNA-film electrode. The DNA film was formed on a typical Glassy Carbon electrode using extremely low DNA concentrations in solution, pg/mL instead of usually used µg/mL, but long time of DNA accumulation, up to a few hours. Such long and tedious procedure proved to be beneficial for obtaining a uniform and reproducible DNA layer. As an example of the anticancer drugs, the derivatives (-Cl, -Et, -Me) of the compound abbreviated as IPBD were studied. In every case, the complexation of plasmids was detected and was accompanied by the condensation within the DNA-compound layer. This result can be used as a proof of strong interactions between DNA and potential anticancer drugs, leading to the conclusion that such interactions might be crucial for the anticancer action of the IPBD compound and its derivatives. To prove the complexation of the DNA by these potential anticancer drugs, except for electrochemical, also the spectroscopic methods were used. From UV-Vis, the change in spectra were observed and the stability constants were calculated (in the range of 10^5 M-1). The CD spectra showed the change of the DNA structure with the addition of each potential anticancer drug. It was not clear, however, to which form of the plasmid DNA was transformed, initially probably form B-DNA into A-DNA. Then, with increasing concentration of the incorporated compound, it was anticipated that Z-DNA might be partially formed with increasing concentration of the incorporated compounds. In fact, a definite change from B-DNA to Z-DNA was indeed shown with the addition of pentamidine to accompany the potential anticancer drug Cl-IPBD as well as for the extract of Uncaria tomentosa. Pentamidine was basically studied to shed the light on the type of interactions between the potential anticancer drugs and DNA. Pentamidine incorporated into the oligonucleotide is one of a few examples of compounds which crystal structure is known and the accumulation in a specific part of the DNA helix called minor groove is proven. Therefore, I have used pentamidine to test whether it can be substituted by a potential anticancer drug (or vice versa). Indeed, such a substitution was proven, leading to the conclusion that IPBD and its derivatives might interact with DNA specifically in the minor groove. It should also be mentioned that pentamidine and the mixture of this compound with a potential anticancer drug Cl-IPBD was tested biologically on cancer cells. Both pentamidine and Cl-IPBD showed anticancer properties, but surprisingly, the combination of these drugs was more effective than each of them used separately. Not only Cl-IPBD, but also other potential anticancer drugs, namely the derivatives of IPBD were also tested biologically. Each compounds showed anticancer properties. Importantly, the differences detected between these derivatives correlated well with the differences in accumulation of the compounds within the plasmids, as detected simultaneously using electrochemical methods. Even though only two compounds were tested as typical antibacterial as well as antifungal substances, significant similarities were noticed within each group and differences between the groups, suggesting various modes of interactions with DNA. The antibacterial substances are difficult to oxidize on the GC electrode in contrast to antifungal substances which oxidize easily, similarly to the anticancer drugs. The differences are also observed in the degree of condensation within the DNA-compound films. In contrast to the interactions of anticancer, antibacterial and antifungal compounds, rutin and U. tomentosa, with plasmids, vitamin C, which oxidation can easily be studied electrochemically, does not cause the DNA condensation, proving again the importance of the condensation effect in other cases.
Przedstawiona praca doktorska dotyczy oddziaływania DNA z różnymi związkami chemicznymi, takimi jak: przeciwnowotworowe, przeciwbakteryjne, przeciwgrzybicze i przeciwutleniające. Pierwszym etapem każdego z eksperymentów było zaadsorbowanie DNA na powierzchni elektrody z węgla szklistego. Akumulację DNA prowadzono przez kilka godzin z roztworu DNA, którego stężenie było bardzo niskie, rzędu pg/ml, zamiast zwykle stosowanego μg/ml. Tak długa i żmudna procedura okazała się korzystna dla uzyskania jednolitej i powtarzalnej warstwy DNA. Jako przykład związków przeciwnowotworowych badano pochodne (-Cl, -Et, -Me) związku o skrócie IPBD. W każdym przypadku obserwowano kompleksowanie plazmidów, czemu towarzyszyła kondensacja warstwy DNA-związek. Wynik ten można wykorzystać jako dowód oddziaływania między DNA i potencjalnymi lekami przeciwnowotworowymi, co prowadzi do wniosku, że takie interakcje mogą być kluczowe dla działania przeciwnowotworowego związku IPBD i jego pochodnych. Aby udowodnić kompleksowanie DNA przez potencjalne związki przeciwnowotworowe, oprócz metod elektrochemicznych, zastosowano również metody spektroskopowe. Na podstawie danych UV-Vis obliczono stałe trwałości kompleksów DNA-związek, które wynosiły ok. 10^5 M-1. Widma CD wykazały zmianę struktury DNA z dodatkiem każdego potencjalnego leku przeciwnowotworowego. Nie było jednak jasne, do jakiej formy DNA zostało przekształcone, początkowo prawdopodobnie z B-DNA w A-DNA. Następnie przewidywano, że Z-DNA może być częściowo utworzone ze wzrostem stężenia badanych związków. W rzeczywistości wykazano wyraźną zmianę z B-DNA na Z-DNA dla mieszaniny potencjalnego związku przeciwnowotworowego Cl-IPBD i pentamidyny, jak również samej pentamidyny oraz dla roślinnego ekstraktu Uncaria tomentosa. Do badań zastosowano pentamidynę, czyli związek o znanym sposobie oddziaływania z DNA, który lokuje się w małym rowku helisy kwasu deoksyrybonukleinowego, tak, aby odpowiedzieć na pytanie w jaki sposób potencjalne związki przeciwnowotworowe oddziałują z DNA. Przeprowadzone eksperymenty polegały na sprawdzeniu czy pentamidyna zakumulowana w warstwie DNA może zostać zastąpiona przez potencjalny związek przeciwnowotworowy (lub odwrotnie). Rzeczywiście, takie podstawienie zostało udowodnione, co prowadzi do wniosku, że IPBD i jego pochodne mogą oddziaływać z DNA w małym rowku. Należy również podkreślić, że pentamidyna i mieszanina tej substancji z potencjalnym związkiem przeciwnowotworowym (Cl-IPBD) została przetestowana biologicznie na komórkach rakowych. Zarówno pentamidyna, jak i Cl-IPBD wykazywały właściwości cytotoksyczne, ale było zaskakujące, że połączenie tych leków okazało się bardziej skuteczne niż każdy z nich stosowany osobno. Nie tylko Cl-IPBD, ale także inne potencjalne związki przeciwnowotworowe, a mianowicie IPBD Me-IPBD i Et-IPBD, zostały również biologicznie przetestowane. Każdy związek wykazywał właściwości cytotoksyczne. Co ważne, różnice wykryte między tymi pochodnymi dobrze korelują z różnicami w akumulacji związków w plazmidach, wykrytymi jednocześnie za pomocą metod elektrochemicznych. Pomimo tego, w przedstawionej pracy testowano tylko dwa związki będące typowymi substancjami przeciwbakteryjnymi i przeciwgrzybiczymi, zauważono znaczące podobieństwa w obrębie każdej grupy i różnice między grupami, co sugeruje różne sposoby oddziaływania z DNA. Substancje przeciwbakteryjne są trudne do utlenienia na elektrodzie GC w przeciwieństwie do substancji przeciwgrzybiczych, które łatwo się utleniają, podobnie jak związki przeciwnowotworowe. Różnice obserwuje się także w stopniu kondensacji w warstwach DNA-związek. W przeciwieństwie do potencjalnych związków przeciwnowotworowych, przeciwbakteryjnych, przeciwgrzybiczych, rutyny i U. tomentosa, witamina C, której utlenianie można łatwo badać elektrochemicznie, nie powoduje kondensacji DNA, co ponownie potwierdza znaczenie efektu kondensacji w innych przypadkach.