In der vorliegenden Arbeit soll das weit gefächerte Thema der chemischen Nanotechnologie um neue Resultate bereichert werden. Im Einzelnen handelt es sich um neue Synthesemethoden für magnetische Nanorods (Nanoteilchen mit länglicher Form) und Nanoshells (oxidische Nanokristalle mit einer Hülle aus Edelmetall), ein Verfahren zur Modifizierung diverser Oberflächen mit heterogenen Polymerbürsten durch kontrollierte binäre radikalische Polymerisation, neuartige Copolymere zur Stabilisierung und Funktionalisierung von Nanopartikeln und Herstellung von amphiphilen Nanopartikeln durch Oberflächenbehandlung mit Niotensiden. Es sollen ferner die möglichen Anwendungen diskutiert werden. Die Abb. 1 illustriert die Zusammenhänge der bearbeiteten Thematik.

Mesenchymale Stammzellen (MSC) werden bereits in klinischen Studien zur Behandlung verschiedener Krankheiten eingesetzt. Über deren Wirkmechanismus und Verbleib nach Applikation ist jedoch noch wenig bekannt. Die in vivo-Nachverfolgung markierter MSC mittels Magnetresonanztomographie stellt eine mögliche Methode zur Erlangung weiterer Erkenntnisse dar. Zu diesem Zweck können die MSC mittels superparamagnetischen Eisenoxid (SPIO)-Partikeln markiert werden. In dieser Arbeit wurden 3 verschiedene SPIO-Produkte zur Markierung oviner und equiner MSC verwendet: Endorem™, Resovist® und Molday ION Rhodamine B™. Die Produkte wurden hinsichtlich ihrer Einflüsse auf die biologischen Eigenschaften der MSC, ihrer Markierungseffizienz und –selektivität verglichen. Desweiteren wurde die produktspezifische magnetresonanztomographische Nachverfolgbarkeit der SPIO-markierten MSC untersucht. Weiterführendes Ziel war die Selektion des am besten geeigneten SPIO-Produktes für die Verwendung in einem in vivo-Großtierversuch zur magnetresonanztomographischen Nachverfolgung SPIO-markierter MSC nach Applikation in arthrotische Gelenke. Die MSC wurden dazu aus dem Knochenmark von je 5 gesunden Schafen und Pferden isoliert, bis zur Passage 4 (P4) expandiert und schließlich mit den verschiedenen SPIO-Produkten markiert. Unmarkierte MSC der gleichen Tiere dienten zur Kontrolle. Proliferationsvermögen sowie tripotentes Differenzierungspotential wurden in vitro untersucht. Zur Evaluierung von Markierungsselektivität und -effizienz der SPIO-Produkte wurden die MSC ab der P4 bis zur P7 wöchentlich passagiert. Ein semiquantitatives histologisches Auswertungssystem basierend auf der Preußisch Blau-Färbung sowie T2*w-GRE-Sequenzen an einem 0,5T-MRT-System wurden zur Evaluierung genutzt. Markierungsselektivität bezeichnete die intra- oder extrazelluläre Lokalisation der SPIO-Partikel. Markierungseffizienz beschrieb die Menge intrazellulär vorhandener SPIO-Partikel. Es wurde gezeigt, dass sich ovine und equine MSC mit allen 3 untersuchten SPIO-Produkten erfolgreich markieren ließen. Die Ergebnisse der in vitro-Untersuchungen ergaben keine Unterschiede zwischen SPIO-markierten und unmarkierten MSC hinsichtlich des Proliferationsvermögens, der adipogenen oder osteogenen Differenzierungsfähigkeit. Jedoch wurde eine deutliche Verminderung des chondrogenen Differenzierungspotentials SPIO-markierter MSC beobachtet, welche von der Menge intrazellulär vorhandener SPIO-Partikel und somit von der Markierungseffizienz abhängig war. Zum Zeitpunkt der initialen Markierung konnte nur Molday ION Rhodamine B™ eine selektive und effiziente Zellmarkierung gewährleisten. Mit Endorem™ konnte eine selektive, jedoch keine ausreichend effiziente Zellmarkierung erreicht werden. Resovist® dagegen bewirkte zwar eine effiziente, aber sehr unselektive initiale Zellmarkierung: Mittels Preußisch Blau-Färbung wurde gezeigt, dass große Mengen von SPIO-Partikeln nur extrazellulär anhefteten. Die 3 verschiedenen SPIO-Produkte führten weiterhin zu unterschiedlich starken hypointensen MRT-Signalen der markierten MSC, welche im Verlauf der 3-wöchigen Versuchsdauer bei allen 3 Produkten stetig abnahmen. Unmarkierte MSC waren isointens, also mittels MRT nicht darstellbar und daher nicht nachverfolgbar. Stets verursachten Resovist®-markierte MSC das stärkste hypointense MRT-Signal, gefolgt von Molday ION Rhodamine B™ und Endorem™. Resovist®-markierte MSC konnten mittels MRT bei beiden Spezies über den längsten Zeitraum nachverfolgt werden (ovine MSC bis 16 Tage, equine MSC bis 23 Tage nach Markierung). Aufgrund der exzellenten initialen Markierungseigenschaften (hohe Markierungsselektivität und –effizienz sowie gute Nachverfolgbarkeit) eignet sich Molday ION Rhodamine B™ besonders gut für die SPIO-Markierung von MSC zur Nachverfolgung mittels MRT. Molday ION Rhodamine B™ verspricht somit eine erfolgreiche Anwendung in einem in vivo-Versuch zur magnetresonsztomographischen Nachverfolgung von MSC nach Applikation in arthrotische Gelenke. / Mesenchymal stem cells (MSC) are already used in clinical studies for treatment of different diseases. However, their mechanism of action and fate after application are still not fully understood. In vivo tracking of labeled MSC via magnetic resonance imaging (MRI) is a possible method to achive further knowledge. For this purpose MSC can be labelled with superparamagnetic iron oxide (SPIO) particles. For this study 3 different SPIO products were employed for labelling of ovine and equine MSC: Endorem™, Resovist®,, and Molday ION Rhodamine B™. The products were compared in terms of their influence on biologic behaviour of the MSC, their labelling efficiency, and selectivity. Furthermore, product specific magnetic resonance traceability of SPIO labelled MSC was evaluated. Final aim was the selection of the most suitable SPIO product to be used in an in vivo large animal study employing MRI tracking of SPIO labelled MSC after application into osteoarthritic joints. MSC therefore, were isolated from bone marrow of each 5 healthy sheep and horses, expanded up to passage 4 (p4), and labelled by the different SPIO products. Unlabelled MSC from the same animals served as control. Proliferation potential and tripotent differentiation capacities were assessed in vitro. For evaluation of labelling selectivity and efficiency of the SPIO products MSC were passaged weekly from p4 up to p7. Semiquantitative histological scoring based on Prussian blue staining and images using T2*w GRE sequences in a 0.5T MRI system were used. Labelling selectivity describes the intra- or extracellular localisation of the SPIO particles. Labelling efficiency describes the amount of intracellular SPIO particles. It was shown that ovine and equine MSC could be successfully labelled by all 3 evaluated SPIO products. The results of the in vitro experiments did not show differences between labelled and unlabelled MSC in terms of proliferation potential, adipogenic or osteogenic differentiation capacities. However, an inhibited chondrogenic differentiation capacity of SPIO labelled MSC was observed, which was dependend on the amount of intracellular SPIO particles and therefore, also on labelling efficiency. At the time of initial labelling, only Molday ION Rhodamine B™ showed selective and efficient cell labelling. With Endorem™ selective, but not efficient cell labelling was achieved. Resovist®, in contrast, caused efficient but very unselective initial cell labelling: By Prussian blue staining it was shown that large amounts of SPIO particles were attached extracellularly. These 3 different SPIO products led to variable hypointense MRI signals of the labelled MSC which decreased in all 3 products during the 3 week study period. Unlabelled MSC were isointense, thus not visible, and therefore, not traceable using MRI. At every point of time, Resovist® labelled MSC resulted in the most hypointense MR signals, followed by Molday ION Rhodamine B™ and Endorem™. Resovist® labelled MSC were traced over the longest time span (ovine MSC until 16 days, equine MSC until 23 days post labelling). Due to excellent initial labelling properties (high labelling efficiency and selectivity, good traceability) Molday ION Rhodamine B™ suits best for SPIO labelling of MSC to be tracked by MRI. Molday ION Rhodamine B™ therefore, promises a successful use in an in vivo study using MRI for MSC tracking after application into osteoarthritic joints.

Submitted by (ana.araujo@ufrpe.br) on 2016-08-02T14:20:48Z No. of bitstreams: 1 Leila Milfont Rameh.pdf: 1805419 bytes, checksum: 3c0a871188e0dc9ff8282000ec45fc1c (MD5) / Made available in DSpace on 2016-08-02T14:20:48Z (GMT). No. of bitstreams: 1 Leila Milfont Rameh.pdf: 1805419 bytes, checksum: 3c0a871188e0dc9ff8282000ec45fc1c (MD5) Previous issue date: 2010-08-26 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The method of unsupervised data classification proposed by Domany and coworkers is based on mapping the problem onto an inhomogeneous granular magnetic system whose properties can be investigated through some Monte Carlo Method. The array containing the data consists of n numeric attributes corresponding to points in an n-dimensional Euclidean space. Each data item is associated with a Potts spin. The interaction between such spins decays exponentially with the distance. This favors the alignment of the spins associated with similar objects. The physical system corresponds to a disordered ferromagnet which, in turn, is described by a Hamiltonian of a q-states Potts model. It is expected that the magnetic system exhibits three temperature-dependent regimes. For very low temperatures the system is completely ordered. At the other extreme, high temperatures, the system shows no magnetic order. In an intermediate range of temperatures, the spins within certain regions remain tightly coupled, forming grains. However, a grain does not influence the behavior of another grain. That is, the grains are non-correlated and this intermediate state is named a superparamagnetic phase. The transition from one regime to another can be identified by peaks in the specific heat versus temperature curve. We apply the method to several artificial and real-life data sets, such as classification of flowers, summary medical data and identification of images. We measure the spin-spin correlation at several temperatures to classify the data. In disagreement with the Domany and coworkers claims we found that the best classification of the data occurred outside the superparagnetic phase. / O método de agrupamento de dados não supervisionado proposto por Domany e colaboradores baseia-se no mapeamento do problema em um sistema magnético granular não homogêneo, cujas propriedades são investigadas através de algum método de Monte Carlo. A matriz que contém os dados é composta por n atributos de valor numérico e corresponde a um ponto em um espaço euclidiano n-dimensional. A cada item de dado é associado um spin de Potts. A interação entre tais spins decai exponencialmente com o aumento da distância entre eles. Isto favorece o alinhamento dos spins associados a objetos similares. O sistema físico corresponde a um ferromagneto desordenado que, por sua vez, é descrito por um hamiltoniano de Potts de q estados. Espera-se que o sistema magnético exiba três regimes quando sua temperatura seja variada. Para temperaturas muito baixas o sistema está completamente ordenado. No outro extremo, em altas temperaturas, o sistema não apresenta qualquer ordem magnética. Numa faixa intermediária de temperaturas, spins dentro de certas regiões permanecem fortemente acoplados, formando grãos. Porém, um grão não influencie o comportamento de outro grão. Ou seja, os grãos estão não correlacionados. Este estado intermediário caracteriza um estado superparamagnético. A transição de um regime para outro pode ser identificada por picos na curva de calor específico versus temperatura. Aplicamos o método aos conjuntos de dados reais da planta íris e de dados médicos, conhecido por BUPA, aos dados sintéticos conhecidos por Ruspini e a um conjunto de dados, gerado por nós, que consiste de duas figuras tridimensionais sobrepostas, um esfera e um toro. Procedemos a classificação dos dados através da correlação spin-spin em diversas temperaturas. O principal resultado foi a verificação que nem sempre o agrupamento realizado na fase superparamagnética é o ideal.

In this work is described a study of the magnetic anisotropy of a magnesioferrite nanoparticle system (MgFe2O4) obtained by precipitation in solid solution of magnesiowüstite [Mg, Fe]O. The anisotropy field was obtained from system’s resonance spectra as measured in the X band. Such spectra can be considered as composed by two spectral lines: A broad line assigned to blocked particles at the measuring temperature and a thin line, which can be attributed to unlocked (superparamagnetic) particles. The anisotropy field of four samples treated at 700 °C during different annealing times were obtained from the spectra measured with their easy magnetization axes oriented at 0 ° and 45 ° with the applied field. The use of a theoretical model to adjust the obtained anisotropy field data enabled the obtaining of the model’s parameter values and their posterior comparison with those reported by other authors. Beyond the dependence of the anisotropy field with the measurement temperature and the sample’s treatment time, it was observed the presence of low field resonance lines that may be related to transitions usually considered as forbidden, which can be enabled by the degenerescence breaking caused by dipolar magnetic interactions between particles. Such phenomenon, which is considerably rare to observe in magnetic nanoparticles systems, was clearly observed in the studied system not only for the half-field resonance line (2Q), but also for lines appearing at one third of field (3Q) and at one quarter of field (4Q). This feature shows that, at least in the investigated experimental system, the dipolar interactions between nearby particles cannot always be neglected as has been proposed by several other authors. / O presente trabalho aborda o estudo da anisotropia magnética do sistema de nanopartículas de magnesioferrita (MgFe2O4) precipitadas em solução sólida de magnesiowüstita [Mg;Fe]O a partir da análise de seus espectros de ressonância magnética medidos na banda X. Tais espectros podem ser aproximadamente decompostos em duas linhas, sendo uma atribuída às partículas bloqueadas na temperatura de medição (linha larga) e a outra às partículas desbloqueadas (linha estreita). Os campos de anisotropia de quatro amostras tratadas a 700 oC por diferentes tempos de tratamento foram obtidos a partir dos espectros medidos com seus eixos de magnetização fácil orientados a 0° e a 45° em relação ao campo aplicado. O emprego de um modelo teórico de anisotropia magnética para ajustar os dados obtidos possibilitou a obtenção dos valores dos parâmetros considerados no mesmo e a comparação destes valores com os que foram reportados por outros autores. Além da dependência do campo de anisotropia com a temperatura de medição e com o tempo de tratamento térmico das amostras, observou-se a presença de linhas de ressonância na região de baixos campos que podem estar relacionadas a transições normalmente consideradas como proibidas que ser viabilizadas com a quebra da degenerescência causada por interações magnéticas dipolares entre partículas. Tal fenômeno, que é consideravelmente raro de se observar em sistemas de nanopartículas magnéticas, foi claramente observado no sistema estudado não somente para a linha de ressonância a meio campo (2Q), mas também para as linhas a um terço (3Q) e um quarto (4Q) de campo. Isso evidencia que, ao menos no sistema experimental investigado, as possíveis interações dipolares entre partículas próximas não podem ser sempre desprezadas a priori como tem sido proposto por vários outros autores.

O carbono, em suas muitas formas distintas, tem sido largamente empregado como material adsorvente, do tratamento de água à tecnologia industrial em virtude de sua boa performance, baixo custo e compatibilidades biológica e ambiental. Nesta tese foi verificada a associação sinérgica entre carvão ativo e nanopartículas superparamagnéticas de óxido de ferro ensejando a exploração entre a capacidade adsortiva e o magnetismo para o desenvolvimento de processos de remoção de contaminantes de meios aquosos e melhorando sua análise por meio de concentração magnética. Para este propósito, nanopartículas magnéticas recobertas com ácidos oleico ou esteárico foram especialmente preparadas e combinadas com carvão ativo em proporções variadas (1, 5, 10, 20, 25, 50 % m/m), seguindo rigoroso controle químico. Os materiais de carbono magnetizáveis mantém suas afinidades química e característica porosa do material precursor com alguma diminuição de sua área superficial, embora exibindo uma forte atração por campos magnéticos externos, permitindo sua fácil concentração e remoção. Estes materiais foram extensivamente caracterizados por técnicas como BET, FTIR, XRD, DLS, SEM, TG/DTA, VSM e SQUID. Por conveniência, o compósito a 10 % de nanopartículas de magnetita foi escolhido como o principal a ser utilizado na maioria das aplicações exibindo ampla área superficial (700 m2.g-1), volume e diâmetro médio de poro (0,487 cc.g-1 e 8,5 nm, respectivamente) e uma magnetização de saturação de 5 emu.g-1, com histerese desprezível à temperatura ambiente. Suas características adsorventes foram exploradas com sucesso na captura de corantes orgânicos (pararosanilina e azul de metileno), nitrobenzeno, bisfenol-A e BTEX (benzeno, tolueno, etilbenzeno e o,m,p-xilenos), bem como para íons de metais pesados como Hg(II), Pb(II) e Ag(I) em solução aquosa. Estudos eletroanalíticos foram feitos explorando o efeito de pré-concentração magnética na superfície do eletrodo após realizar estudos de adsorção envolvendo o levantamento de isotermas afim de investigar parâmetros como capacidade adsortiva e constantes de adsorção. Em todos os casos, um elevado aumento de aumento na sensibilidade foi obtido através do confinamento magnético em relação aos métodos convencionais. Desta forma, o emprego do compósito de carbono com nanopartículas de óxido de ferro superparamagnéticas demonstrou ser uma ferramenta poderosa a ser explorada em Química Analítica e Ambiental. / Carbon in many distinct forms has been widely employed as absorbing material in industrial and water technology because of its good performance, low cost and bio/environmental compatibility. In this doctoral thesis, a synergistic association of activated carbon with superparamagnetic Fe3O4 nanoparticles was carried out, aiming the exploitation of their combined absorption capability and magnetism, for developing new processes of removing contaminants from water and improving their analysis by means of magnetic concentration effect. For this purpose, magnetite nanoparticles coated with stearic or oleic acids were specially prepared and combined with activated carbon, at several proportions (1, 5, 10, 20, 25, 50 % m/m), following a rigorous chemical and analytical control. The magnetized carbon materials kept the original porous characteristics and molecular affinity of the original absorber, with some decrease of the active surface area, but exhibiting a strong attraction by the magnetic field, allowing their easy concentration and removal. Such materials were extensively characterized by BET, FTIR, XRD, DLS, SEM, TG/DTA, VSM and SQUID techniques. For convenience, the carbon material containing 10 % of magnetite was elected as principal, exhibiting a surface area of 700 m2 g-1, average volume and porous diameter of 0,487 cm3 g-1 and 8,5 nm, respectively, and a saturation magnetization of 5 emu g-1, with null hysteresis at room temperature. Its absorbing characteristics were successfully explored for the capture of organic dyes (pararosaniline, and methylene blue), bisphenol-A, and BTEX (benzene, toluene, ethylbenzene, and o,m,p-xilene), as well as hazardous, heavy metal ions, such as Hg(II), Pb(II) and Ag(I) in aqueous solution. Electroanalytical studies were carried out by exploring the magnetic pre-concentration effect on the electrodes, after performing a detailed adsorption study involving the construction of isotherms, in order to evaluate the equilibrium and mass capacity parameters. In all the cases, a great increase of sensitivity was attained by the magnetic confined method, in relation to the conventional methods. In this way, the use of carbon with superparamagnetic nanoparticles demonstrated a powerful strategy to be explored in environmental and analytical chemistry.

Un des enjeux majeurs dans le domaine du magnétisme moléculaire est de mieux comprendre et prévoir, dans les composés à base moléculaire, les corrélations qui existent entre les propriétés structurales (modulables à partir de méthodes de synthèse de type « bottom-up ») et les propriétés magnétiques. En particulier, la compréhension et la maîtrise de l’anisotropie magnétique à l’échelle locale est primordiale, notamment en vue de concevoir des molécules-aimants avec de plus hautes températures de blocage. Dans ce contexte, ce travail de thèse s’organise autour de deux grands axes. La première partie se concentre sur la détermination et la caractérisation de l’anisotropie magnétique locale dans des complexes moléculaires d’ions de transition de faible nucléarité. La diffraction de neutrons polarisés (PND) nous a permis, pour la première fois, de mettre clairement en évidence le tenseur de susceptibilité magnétique locale dans un complexe moléculaire mononucléaire de Fe3+ Bas-Spin ainsi que dans deux complexes, mononucléaire et dinucléaire, de Co2+ Haut-Spin. Cette approche novatrice mène à l’établissement de relations magnéto-structurales claires et directes, en reliant les directions magnétiques locales propres à l’environnement de coordination des ions métalliques et en particulier aux axes locaux de distorsion. Nous avons également mené l’étude originale d’un complexe à transition de spin thermo-induite de Mn3+ par diffusion inélastique de neutrons (INS) dans les deux phases Haut-Spin (HS) et Bas-Spin (BS). Cette étude nous a conduits à la proposition d’un modèle d’hamiltonien de spin anisotrope dans les deux états HS et BS, en relation avec la structure du complexe. Dans une seconde partie plus exploratoire de la thèse, nous avons mené une étude complète des propriétés structurales et magnétiques de nanoparticules ferromagnétiques d’analogue du bleu de Prusse CsNiCr, par diffusion de neutrons aux petits angles (SANS). Les effets de taille, d’organisation et de concentration sur leurs propriétés superparamagnétiques ont ainsi été clairement mis en évidence. En particulier, nous avons mis en exergue, pour les particules de plus petite taille (5 nm de diamètre), une contribution magnétique qui résulte de la manifestation d’un phénomène collectif, tandis que celles de plus grande taille (28 nm de diamètre) apparaissent être dans un état complètement multidomaine. / One of the major issues in the field of molecular magnetism is to better understand and predict the correlations between the structural properties of molecule-based compounds and their magnetic properties, all of which may be tunable using “bottom-up” synthesis methods. In particular, the understanding and control of the magnetic anisotropy at the atomic scale is essential, especially with the aim to design Single-Molecule Magnets (SMM) with higher blocking temperatures. In this context, this thesis work is focused on two mains subjects. The first part deals with the determination and the characterization of the local magnetic anisotropy in low-nuclearity molecular complexes based on transition ions. Polarised neutron diffraction (PND) allows us, for the first time, to directly access the local susceptibility tensor in a Low-Spin Fe3+ mononuclear complex as well as in two, mononuclear and dinuclear, High-Spin Co2+ complexes. This innovative approach leads to the establishment of unique and direct magneto-structural correlations, by relating the local magnetic principal directions with the coordination environment of the metallic ions and, in particular, with the local distortion axes. We have also carried out an original investigation by inelastic neutron scattering (INS) of a Mn3+ thermo-induced spin-transition compound in both High-Spin (HS) and Low-Spin (LS) states. On the basis of this study, we were able to propose an anisotropic spin-Hamiltonian model in both HS and LS phases, and their relationships with the structure of the molecule are discussed. In a second more exploratory part of the thesis, we have carried out by small-angle neutron scattering (SANS) a complete study of the structural and magnetic properties of Prussian blue analogues (PBA) ferromagnetic nanoparticles CsNiCr. The effects of size, organization and concentration on their superparamagnetic properties have been clearly highlighted. In particular, a strong magnetic contribution has been observed for the smallest particles (5 nm diameter) which results from the manifestation of a collective process, while the biggest (28 nm diameter) appear to be in a multi-domain state.

In der vorliegenden Arbeit soll das weit gefächerte Thema der chemischen Nanotechnologie um neue Resultate bereichert werden. Im Einzelnen handelt es sich um neue Synthesemethoden für magnetische Nanorods (Nanoteilchen mit länglicher Form) und Nanoshells (oxidische Nanokristalle mit einer Hülle aus Edelmetall), ein Verfahren zur Modifizierung diverser Oberflächen mit heterogenen Polymerbürsten durch kontrollierte binäre radikalische Polymerisation, neuartige Copolymere zur Stabilisierung und Funktionalisierung von Nanopartikeln und Herstellung von amphiphilen Nanopartikeln durch Oberflächenbehandlung mit Niotensiden. Es sollen ferner die möglichen Anwendungen diskutiert werden. Die Abb. 1 illustriert die Zusammenhänge der bearbeiteten Thematik.

Mesenchymale Stammzellen (MSC) werden bereits in klinischen Studien zur Behandlung verschiedener Krankheiten eingesetzt. Über deren Wirkmechanismus und Verbleib nach Applikation ist jedoch noch wenig bekannt. Die in vivo-Nachverfolgung markierter MSC mittels Magnetresonanztomographie stellt eine mögliche Methode zur Erlangung weiterer Erkenntnisse dar. Zu diesem Zweck können die MSC mittels superparamagnetischen Eisenoxid (SPIO)-Partikeln markiert werden. In dieser Arbeit wurden 3 verschiedene SPIO-Produkte zur Markierung oviner und equiner MSC verwendet: Endorem™, Resovist® und Molday ION Rhodamine B™. Die Produkte wurden hinsichtlich ihrer Einflüsse auf die biologischen Eigenschaften der MSC, ihrer Markierungseffizienz und –selektivität verglichen. Desweiteren wurde die produktspezifische magnetresonanztomographische Nachverfolgbarkeit der SPIO-markierten MSC untersucht. Weiterführendes Ziel war die Selektion des am besten geeigneten SPIO-Produktes für die Verwendung in einem in vivo-Großtierversuch zur magnetresonanztomographischen Nachverfolgung SPIO-markierter MSC nach Applikation in arthrotische Gelenke. Die MSC wurden dazu aus dem Knochenmark von je 5 gesunden Schafen und Pferden isoliert, bis zur Passage 4 (P4) expandiert und schließlich mit den verschiedenen SPIO-Produkten markiert. Unmarkierte MSC der gleichen Tiere dienten zur Kontrolle. Proliferationsvermögen sowie tripotentes Differenzierungspotential wurden in vitro untersucht. Zur Evaluierung von Markierungsselektivität und -effizienz der SPIO-Produkte wurden die MSC ab der P4 bis zur P7 wöchentlich passagiert. Ein semiquantitatives histologisches Auswertungssystem basierend auf der Preußisch Blau-Färbung sowie T2*w-GRE-Sequenzen an einem 0,5T-MRT-System wurden zur Evaluierung genutzt. Markierungsselektivität bezeichnete die intra- oder extrazelluläre Lokalisation der SPIO-Partikel. Markierungseffizienz beschrieb die Menge intrazellulär vorhandener SPIO-Partikel. Es wurde gezeigt, dass sich ovine und equine MSC mit allen 3 untersuchten SPIO-Produkten erfolgreich markieren ließen. Die Ergebnisse der in vitro-Untersuchungen ergaben keine Unterschiede zwischen SPIO-markierten und unmarkierten MSC hinsichtlich des Proliferationsvermögens, der adipogenen oder osteogenen Differenzierungsfähigkeit. Jedoch wurde eine deutliche Verminderung des chondrogenen Differenzierungspotentials SPIO-markierter MSC beobachtet, welche von der Menge intrazellulär vorhandener SPIO-Partikel und somit von der Markierungseffizienz abhängig war. Zum Zeitpunkt der initialen Markierung konnte nur Molday ION Rhodamine B™ eine selektive und effiziente Zellmarkierung gewährleisten. Mit Endorem™ konnte eine selektive, jedoch keine ausreichend effiziente Zellmarkierung erreicht werden. Resovist® dagegen bewirkte zwar eine effiziente, aber sehr unselektive initiale Zellmarkierung: Mittels Preußisch Blau-Färbung wurde gezeigt, dass große Mengen von SPIO-Partikeln nur extrazellulär anhefteten. Die 3 verschiedenen SPIO-Produkte führten weiterhin zu unterschiedlich starken hypointensen MRT-Signalen der markierten MSC, welche im Verlauf der 3-wöchigen Versuchsdauer bei allen 3 Produkten stetig abnahmen. Unmarkierte MSC waren isointens, also mittels MRT nicht darstellbar und daher nicht nachverfolgbar. Stets verursachten Resovist®-markierte MSC das stärkste hypointense MRT-Signal, gefolgt von Molday ION Rhodamine B™ und Endorem™. Resovist®-markierte MSC konnten mittels MRT bei beiden Spezies über den längsten Zeitraum nachverfolgt werden (ovine MSC bis 16 Tage, equine MSC bis 23 Tage nach Markierung). Aufgrund der exzellenten initialen Markierungseigenschaften (hohe Markierungsselektivität und –effizienz sowie gute Nachverfolgbarkeit) eignet sich Molday ION Rhodamine B™ besonders gut für die SPIO-Markierung von MSC zur Nachverfolgung mittels MRT. Molday ION Rhodamine B™ verspricht somit eine erfolgreiche Anwendung in einem in vivo-Versuch zur magnetresonsztomographischen Nachverfolgung von MSC nach Applikation in arthrotische Gelenke. / Mesenchymal stem cells (MSC) are already used in clinical studies for treatment of different diseases. However, their mechanism of action and fate after application are still not fully understood. In vivo tracking of labeled MSC via magnetic resonance imaging (MRI) is a possible method to achive further knowledge. For this purpose MSC can be labelled with superparamagnetic iron oxide (SPIO) particles. For this study 3 different SPIO products were employed for labelling of ovine and equine MSC: Endorem™, Resovist®,, and Molday ION Rhodamine B™. The products were compared in terms of their influence on biologic behaviour of the MSC, their labelling efficiency, and selectivity. Furthermore, product specific magnetic resonance traceability of SPIO labelled MSC was evaluated. Final aim was the selection of the most suitable SPIO product to be used in an in vivo large animal study employing MRI tracking of SPIO labelled MSC after application into osteoarthritic joints. MSC therefore, were isolated from bone marrow of each 5 healthy sheep and horses, expanded up to passage 4 (p4), and labelled by the different SPIO products. Unlabelled MSC from the same animals served as control. Proliferation potential and tripotent differentiation capacities were assessed in vitro. For evaluation of labelling selectivity and efficiency of the SPIO products MSC were passaged weekly from p4 up to p7. Semiquantitative histological scoring based on Prussian blue staining and images using T2*w GRE sequences in a 0.5T MRI system were used. Labelling selectivity describes the intra- or extracellular localisation of the SPIO particles. Labelling efficiency describes the amount of intracellular SPIO particles. It was shown that ovine and equine MSC could be successfully labelled by all 3 evaluated SPIO products. The results of the in vitro experiments did not show differences between labelled and unlabelled MSC in terms of proliferation potential, adipogenic or osteogenic differentiation capacities. However, an inhibited chondrogenic differentiation capacity of SPIO labelled MSC was observed, which was dependend on the amount of intracellular SPIO particles and therefore, also on labelling efficiency. At the time of initial labelling, only Molday ION Rhodamine B™ showed selective and efficient cell labelling. With Endorem™ selective, but not efficient cell labelling was achieved. Resovist®, in contrast, caused efficient but very unselective initial cell labelling: By Prussian blue staining it was shown that large amounts of SPIO particles were attached extracellularly. These 3 different SPIO products led to variable hypointense MRI signals of the labelled MSC which decreased in all 3 products during the 3 week study period. Unlabelled MSC were isointense, thus not visible, and therefore, not traceable using MRI. At every point of time, Resovist® labelled MSC resulted in the most hypointense MR signals, followed by Molday ION Rhodamine B™ and Endorem™. Resovist® labelled MSC were traced over the longest time span (ovine MSC until 16 days, equine MSC until 23 days post labelling). Due to excellent initial labelling properties (high labelling efficiency and selectivity, good traceability) Molday ION Rhodamine B™ suits best for SPIO labelling of MSC to be tracked by MRI. Molday ION Rhodamine B™ therefore, promises a successful use in an in vivo study using MRI for MSC tracking after application into osteoarthritic joints.

Magnetic nanoparticles that display high saturation magnetization and high magnetic susceptibility are of great interest for medical applications. Nanomagnetite is particularly desirable because it displays strong ferrimagnetic behavior, and is less sensitive to oxidation than magnetic transition metals such as cobalt, iron, and nickel. Magnetite nanoparticles can be prepared by co-precipitating iron (II) and iron (III) chloride salts in the presence of ammonium hydroxide at pH 9-10. One goal of this work has been to develop a generalized methodology for stabilizing nanomagnetite dispersions using well-defined, non-toxic, block copolymers, so that the resultant magnetite-polymer complexes can be used in a range of biomedical materials. Hydrophilic triblock copolymers with controlled concentrations of pendent carboxylic acids were prepared. The triblock copolymers contain carboxylic acids in the central urethane segments and controlled molecular weight poly(ethylene oxide) tail blocks. They were utilized to prepare hydrophilic-coated iron oxide nanoparticles with biocompatible materials for utility in magnetic field guidable drug delivery vehicles. The triblock copolymers synthesized contain 3, 5, or 10 carboxylic acids in the central segments with Mn values of 2000, 5000 or 15000 g/mol poly(ethylene oxide) tail blocks. A method was developed for preparing ~10 nm diameter magnetite surfaces stabilized with the triblock polymers. The carboxylic acid is proposed to covalently bind to the surface of the magnetite and form stable dispersions at neutral pH. The polymer-nanomagnetite conjugates described in this thesis have a maximum of 35 wt. % magnetite and the nano-magnetite particles have an excellent saturation magnetization of ~66 - 78 emu/g Fe3O4. Powder X-ray diffraction (XRD) confirms the magnetite crystal structure, which appears to be approximately single crystalline structures via electron diffraction spectroscopy analysis (EDS). These materials form stable magnetic dispersions in both water and organic solvents. / Ph. D.

One of the most challenging goals in nanoparticle research is to develop successful protocols for the large-scale, simple and possibly low-cost preparation of morphologically pure nanoparticles with enhanced properties. The work presented in this thesis was focused on the synthesis, characterisation and testing of magnetic nanoparticles and their potential applications. There are a number of magnetic nano-materials prepared for specific applications such as metal oxide nanoparticles encapsulated with various porous materials including Fe₃O₄/Fe₂O₃ coated with soft bio-organic materials such as glycol chitosan and bovine serum albumin and hard materials such as silica (SiO₂) and zinc sulphide (ZnS). The preparation of these materials was achieved principally by bottom-up methods with different approaches including micro-emulsion, precipitation, electrostatic and thermolysis processes. The thesis also presents the uses of various analytical techniques for characterising different types of nano-materials including Attenuated Total Reflection Fourier Transformer Infrared Vibrational Spectroscopy (ATR-FTIR), Ultraviolet Visible- Near Infrared (UV-Vis-NIR) Spectroscopy, Zeta Potentiometric Surface Charge Analysis, Superconducting Quantum Interference Device (SQUID) and Vibration Sample Magnetometry (VSM) for magnetic analysis and powder X-Ray Diffraction (XRD) for crystallographic pattern analysis. There are many applications of magnetic nanoparticles, including nano-carriers for biological and catalytic reagents. The magnetic nanoparticles can facilitate separation in order to isolate the carriers from solution mixtures as compared to many inefficient and expensive classic methods, which include dialysis membrane, electrophoresis, ultracentrifugation, precipitation and column separation methods. There are six key chapters in this thesis: the first chapter introduces the up-to-date literature regarding magnetic nano-materials. The uses of magnetic nano-materials in drug binding and for protein separation are discussed in the second and third chapters. The fourth chapter presents the use of magnetic nanoparticle in conjunction with a photo-catalytic porous overlayer for the photo-catalytic reduction of organic molecules. The fifth chapter describes different analytical techniques used for the characterisation of nanoparticles and the underlying principles and the experimental details are also given. The sixth chapter summarises the results and provides an overview of the work in a wider context of future applications of magnetic nanoparticles.