@phdthesis{WaagHiersch2017, author = {Waag-Hiersch, Luisa}, title = {„iClick"-Reaktionen von Ru- und Rh-Azid-Komplexen mit elektronenarmen Alkinen: Regioselektivit{\"a}t, Stabilit{\"a}t und Kinetik}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-146286}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2017}, abstract = {Die regioselektive Funktionalisierung von Bio(makro)molek{\"u}len erfordert Reaktionen, die mit einem biologischen System weder interagieren noch interferieren. Bestimmte funktionelle Gruppen, wie Azide oder Alkine, sind unter physiologischen Bedingungen inert, kommen nicht in der Natur vor, lassen sich selektiv miteinander verkn{\"u}pfen und sind nicht-toxisch gegen{\"u}ber Zellen und Organismen. F{\"u}r die Einf{\"u}hrung metallbasierter Funktionalit{\"a}ten in solche Zielstrukturen stellen Click-Reaktionen daher einen schnellen Zugang dar, wobei Reaktionen, die ohne Zusatz von Katalysator und bei Raumtemperatur ablaufen von besonderem Interesse sind. Das Ziel der vorliegenden Arbeit war es daher die „iClick"-Reaktion von Ruthenium-Azid-Komplexen der allgemeinen Formel [Ru(N3)(aren)(N-N)]+ mit bidentaten Stickstoffliganden sowie Rhodium-Azid-Komplexen der allgemeinen Formel [Rh(Cp*)(N3)(bpyR,R)]+ mit unterschiedlich substituierten 2,2'-Bipyridin-Coliganden (R = OCH3, H, COOCH3) gegen{\"u}ber elektronenarmen Alkinen zu untersuchen. R{\"o}ntgenstrukturanalysen der resultierenden Triazolat-Komplexe sollten den Koordinationsmodus best{\"a}tigten, da die Produkte der Click-Reaktionen prinzipiell als zwei verschiedene Regioisomere auftreten k{\"o}nnen. Die [Rh(Cp*)(N3)(bpyR,R)]CF3SO3-Komplexe mit 2,2'-Bipyridin (bpy), dem elektronenziehenden Ligand 4,4'-Bis(methoxycarbonyl)-2,2′-bipyridin (bpyCOOCH3,COOCH3) sowie dem elektronenschiebenden Ligand 4,4'-Dimethoxy-2,2'-bipyridin (bpyOCH3,OCH3) wurden aus den entsprechenden Rhodium-Chlorido-Komplexen durch F{\"a}llung des Halogenids mit Silbertrifluormethansulfonat und anschließender Umsetzung mit Natriumazid hergestellt. In L{\"o}sung waren diese Verbindungen jedoch nur begrenzt stabil, wobei der Komplex mit bpyOCH3,OCH3 am wenigsten empfindlich war, w{\"a}hrend [Rh(Cp*)(N3)(bpyCOOCH3,COOCH3)]CF3SO3 aufgrund der sehr schnellen Zersetzung nicht isoliert werden konnte. Die „iClick"-Reaktion der Rhodium-Azid-Komplexe mit 4,4,4-Trifluorobut-2-ins{\"a}ureethylester ergab dann aber die stabilen Triazolat-Komplexe [Rh(Cp*)(triazolatCF3,COOEt)(bpyR,R)]CF3SO3 in sehr guter Ausbeute. Die Ruthenium-Azid-Komplexe [Ru(N3)(N-N)(p­cym)]PF6 mit N-N = bpy, bpyCOOCH3,COOCH3, bpyOCH3,OCH3, Bipyrimidin (bpym) sowie Dipyrido[3,2­a:2',3'­c]phenazin (dppz) wurden ausgehend von den jeweiligen Ruthenium-Chlorido-Komplexen durch F{\"a}llung des Halogenid-Liganden mit Silbertrifluormethansulfonat und anschließender Umsetzung mit Natriumazid in guter bis moderater Ausbeute hergestellt. Um den Einfluss des Aren-Liganden zu untersuchen wurde außerdem der entsprechende Hexamethylbenzol-Komplex [Ru(N3)(bpy)(hmb)]CF3SO3 in moderater Ausbeute hergestellt. Alle [Ru(N3)(aren)(N-N)]X-Komplexe mit X = PF6- oder CF3SO3- wurden mittels 1H, 13C NMR- und IR-Spektroskopie, CHN-Analyse sowie ESI-Massenspektrometrie charakterisiert. Die „iClick"-Reaktion dieser Komplexe erfolgte mit 4,4,4-Trifluorobut-2-ins{\"a}ureethylester und teilweise auch mit Dimethylacetylendicaboxylat (DMAD) in sehr guter bis guter Ausbeute. Außerdem konnten f{\"u}r die R{\"o}ntgenstrukturanalyse taugliche Einkristalle von [Ru(triazolatCF3,COOEt)(bpy)(hmb)]CF3SO3 und [Ru(triazolatCF3,COOEt)(bpyCOOCH3,COOCH3)(p­cym)]PF6 erhalten werden, die die N2-Koordination des Triazolat-Liganden an das Zentralatom best{\"a}tigten. Um diese als metallbasierte Marker einsetzen zu k{\"o}nnen, m{\"u}ssen die resultierenden Triazolat-Komplexe bei biologisch relevanten pH-Werten und gegen{\"u}ber Ligandenaustausch, zum Beispiel mit den Aminos{\"a}ureseitenketten von Proteinen, stabil sein. Durch HPLC-Untersuchungen an [Ru(triazolatCF3,COOEt)(bpy)(hmb)]CF3SO3 wurde gezeigt, dass dieser Komplex in w{\"a}ssriger L{\"o}sung {\"u}ber einen pH-Bereich von 1 bis 8 bei Raumtemperatur mindestens 24 h stabil ist. Außerdem konnte eine weitgehende Stabilit{\"a}t gegen{\"u}ber Ligandenaustausch mit den Seitenketten der Aminos{\"a}uren L­Cystein, L-Histidin, L­Methionin und L-Glutamins{\"a}ure bei 37 °C {\"u}ber mindestens 72 h festgestellt werden. Insbesondere die Geschwindigkeit der „iClick"-Reaktion ist in einem biologischen Kontext von Bedeutung, da die Konjugationsreaktionen schneller ablaufen m{\"u}ssen als interessierende biologische Prozesse. Mittels HPLC und IR-Spektroskopie wurde f{\"u}r die „iClick"-Reaktion der Rutheniumazid-Komplexe [Ru(N3)(bpyR,R)(p-cym)]PF6 mit R = OCH3, H oder COOCH3 sowie [Ru(N3)(bpy)(hmb)]CF3SO3 mit einem {\"U}berschuss an 4,4,4-Trifluorobut-2-ins{\"a}ureethylester Geschwindigkeitskonstanten pseudoerster Ordnung im Bereich von 1 ­ 3*10-3 s-1 bestimmt. Außerdem war es mittels IR-Spektroskopie in L{\"o}sung m{\"o}glich die Geschwindigkeits-konstante pseudoerster Ordnung f{\"u}r die „iClick"-Reaktion der Rhodiumazid-Verbindungen [Rh(Cp*)(N3)(bpyR,R)]CF3SO3 mit R = OCH3, H oder COOCH3 und 4,4,4-Trifluorobut-2-ins{\"a}ureethylester zu 2 ­ 4*10-3 s-1 zu ermitteln. Insgesamt zeigte sich, dass Komplexe mit elektronenreichen Coliganden schneller mit 4,4,4-Trifluorobut-2-ins{\"a}ureethylester reagieren als solche mit elektronen{\"a}rmeren Liganden. Auch war die Geschwindigkeitskonstante f{\"u}r die Reaktion der Rhodium-Komplexe h{\"o}her als f{\"u}r die Rutheniumverbindungen. Die Geschwindigkeitskonstanten zweiter Ordnung wurden aus der 19F NMR-spektroskopischen Untersuchung der Reaktion von 4,4,4-Trifluorobut-2-ins{\"a}ureethylester und [Ru(N3)(bpyR,R) (p-cym)]PF6 mit R = OCH3, H oder COOCH3 sowie [Ru(N3)(bpy)(hmb)]CF3SO3 bei 20 °C bestimmt. Bei ann{\"a}hernd gleichem Verh{\"a}ltnis von Alkin und Rutheniumazid-Komplexen wurden Geschwindigkeitskonstanten im Bereich von 1 - 2*10-2 L mol-1 s-1 erhalten. Diese sind gr{\"o}ßer als die der Staudinger-Ligation, aber kleiner als die der spannungsinduzierten Azid-Alkin Cycloaddition. Prinzipiell sollte damit also eine biologische Anwendung m{\"o}glich sein. Außerdem wurde die Aktivierungsenergie der Reaktion von [Ru(N3)(bpy)(p­cym)]PF6 mit 4,4,4-Trifluorobut-2-ins{\"a}ureethylester aus der Untersuchung der Temperaturabh{\"a}ngigkeit im Bereich von -20 °C bis +20 °C mit VT-NMR zu 46.1 kJ mol-1 bestimmt. In den 19F NMR-Spektren des Reaktionsgemisches zeigte sich bei -20 °C neben dem Signal des N2-koordinierten Triazolats außerdem ein weiteres, das dem N1-Isomer zuzuordnen ist, welches bei Erw{\"a}rmen jedoch wieder verschwand. In einer DFT-Rechnung wurde die Geometrie von [Ru(N3)(bpy)(hmb)]CF3SO3 optimiert. Dabei zeigte sich, dass nur etwa 25 - 30\% aller Trajektorien angreifender Alkinmolek{\"u}len einen Zugang zum Azid erm{\"o}glichen, sodass die Reaktionsgeschwindigkeit um etwa einen Faktor vier niedriger liegen sollte als f{\"u}r nicht oder nur wenig abgeschirmte Organoazid-Verbindungen. Die „iClick"-Reaktion der hier untersuchten Metall-Azid-Komplexe mit elektronenarmen Alkinen zeigt also bereits jetzt Reaktionsgeschwindigkeiten vergleichbar etablierter Biokonjugationsreaktionen. In Zukunft sollte daher das Potential anderer Metall-Azid-Bausteine untersucht und auch das Alkin variiert werden.}, subject = {Ruthenium}, language = {de} } @phdthesis{Schwenk2018, author = {Schwenk, Nicola}, title = {Seeing the Light: Synthesis of Luminescent Rhodacyclopentadienes and Investigations of their Optical Properties and Catalytic Activity}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-149550}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2018}, abstract = {Luminescent organotransition metal complexes are of much current interest. As the large spin-orbit coupling of 2nd and 3rd row transition metals usually leads to rapid intersystem crossing from S1 to T1, which enables phosphorescence, there is a special interest in using triplet-emitting materials in organic or organometallic light emitting diodes (OLEDs). Marder et al. have found that, reductive coupling of both para-R-substituted diarylbutadiynes and diaryldodecatetraynes on Rh(PMe3)4X leads to quantitative yields of bis(arylethynyl)-rhodacyclopentadienes with complete regiospecificity (R = BMes2, H, Me, OMe, SMe, CF3, CN, CO2Me, NMe2, NO2, C≡C-TMS and X = -C≡C-TMS, -C≡C-C6H4-4-NMe2, -C≡C-C≡C-C6H4-4-NPh2, Me, Cl).47,49 Unexpectedly, these compounds show intense fluorescence rather than phosphorescence (ɸf = 0.33-0.69, t = 1.2 3.0 ns). The substituent R has a significant influence on the photophysical properties, as absorption and emission are both bathochromically shifted compared to R = H, especially for R = π-acceptor. To clarify the mechanism of the formation of the rhodacyclopentadienes, and to investigate further their unique photophysical properties, a series of novel, luminescent rhodacyclopentadienes with dithiocarbamate as a bidentate ligand at the rhodium centre has been synthesised and characterised (R = NO2, CO2Me, Me, NMe2, SMe, Ar = C6F4-4-OMe). The rhodacyclopentadienes have been formed via reductive coupling of diaryl undecatetraynes with [Rh(k2-S,S`-S2CNEt2)(PMe3)2]. The structures of a series of such compounds were solved by single crystal X-ray diffraction and are discussed in this work. The compounds were fully characterised via NMR, UV/Vis and photoluminescence spectroscopy as well as by elemental analysis, high-resolution mass spectrometry (HRMS) and X-ray diffraction. When heating the reactions, another isomer is formed to a certain extent. The so-called dibenzorhodacyclopentadienes already appeared during earlier studies of Marder et al., when acetylacetonate (acac) was employed as the bidentate ligand at the Rh-centre. They are probably formed via a [4+2] cycloaddition reaction and C-H activation, followed by a β-H shift. Use of the perfluorinated phenyl moiety Ar = C6F4-4-OMe provided a total new insight into the mechanism of formation of the rhodacyclopentadiene isomers and other reactions. Besides the formation of the expected rhodacyclopentadiene, a bimetallic compound was generated, isolated and characterised via X-ray crystallography and NMR spectroscopy, elemental analysis and high resolution mass spectrometry. For further comparison, analogous reactions with [Rh(k2 S,S` S2CNEt2)(PPh3)2] and a variety of diaryl undecatetraynes (R = NO2 CO2Me, Me, NMe2, SMe, Ar = C6F4-4-OMe) were carried out. They also yield the expected rhodacyclopentadienes, but quickly react with a second or even third equivalent of the tetraynes to form, catalytically, alkyne cyclotrimerisation products, namely substituted benzene derivatives (dimers and trimers), which are highly luminescent. The rhodacyclopentadienes (R = NO2, CO2Me, Me, SMe, Ar = C6F4-4-OMe) are stable and were isolated. The structures of a series of these compounds were obtained via single crystal X-ray crystallography and the compounds were fully characterised via NMR, UV/Vis and photoluminescence spectroscopy as well as by elemental analysis and HRMS. Another attempt to clarify the mechanism of formation of the rhodacyclopentadienes involved reacting a variety of diaryl 1,3-butadiynes (R = CO2Me, Me, NMe2, naphthyl) with [Rh(k2 S,S` S2CNEt2)(PMe3)2]. The reactions stop at an intermediate step, yielding a 1:1 trans π-complex, confirmed by single crystal X-ray diffraction and NMR spectroscopy. Only after several weeks, or under forcing conditions (µw / 80 °C, 75 h), the formation of another major product occurs, having bound a second diaryl 1,3-butadiyne. Based on earlier results of Murata, the product is identified as an unusual [3+2] cycloaddition product, ϭ-bound to the rhodium centre.}, subject = {Rhodium}, language = {en} } @phdthesis{Sowik2014, author = {Sowik, Thomas}, title = {Assessment of the surface functionalization of SPION and DND nanomaterials for cellular uptake and fluorescence imaging}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-103709}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2014}, abstract = {The aim of this work was to synthesize and functionalize different bio-relevant nanomaterials like silica-coated superparamagnetic iron oxide nanoparticles (SPIONs) as contrast agents for T2 magnetic resonance imaging (MRI) and detonation nanodiamond (DND) with the neurohormone peptide allatostatin 1 (ALST1) and a fluorescent dye. Analytical techniques for the determination and quantification of surface functional groups like amines, azides, and peptides were also developed and established. Thus, in the first part of the work, a TGF-1 binding peptide and allatostatin 1 (ALST1), both supposed to act as active tumour targeting vectors, were synthesized by solid-phase peptide synthesis (SPPS) and characterized by high pressure liquid chromatography (HPLC) and mass spectrometry. Then, azide-functionalized silica nanoparticles were synthesized by the St{\"o}ber process and characterized by transmission electron microscopy (TEM) and infrared spectroscopy (IR). The surface loading of amine and azide groups was determined by a new protocol. The azide groups were reduced with sodium boronhydride to amine and then functionalized with Fmoc-Rink Amide linker according to a standard SPPS protocol. Upon cleavage of Fmoc by piperidine, the resulting dibenzofulvene and its piperidine adduct were quantified by UV/Vis spectroscopy and used to determine the amount of amine groups on the nanoparticle surface. Then, ALST1 and related tyrosine- and phenylalanine substituted model peptides were conjugated to the azide-functionalized silica nanoparticles by copper(I)-catalyzed azide-alkyne dipolar cycloaddition (CuAAC). The successful peptide conjugation was demonstrated by the Pauly reaction, which however is only sensitive to histidine- and tyrosine-containing peptides. As a more general alternative, the acid hydrolysis of the peptides to their individual amino acid building blocks followed by derivatization with phenyl isothiocyanate (PITC) allowed the separation, determination, and quantification of the constituent amino acids by HPLC. In the second part of the work, amine- and azide-functionalized silica-coated superparamagnetic iron oxide nanoparticles (SPIONs) were synthesized by co-precipitation and subsequent silica-coated based on the St{\"o}ber process and characterized by TEM and IR. The amine surface loading was determined by the method already established for the pure silica systems. The azide surface loading could also be quantified by reduction with sodium boronhydride to amine groups and then conjugation to Fmoc-Rink amide linker. Upon cleavage of Fmoc with piperidine, the total amine surface loading was obtained. The amount of azide surface groups was then determined from the difference of the total amine surface loading and the amine surface loading. Thus, it was possible to quantify both amine and azide surface groups on a single nanoparticle system. Superparamagnetic iron oxide nanoparticles (SPIONs) are potent T2 contrast agents for magnetic resonance imaging (MRI). Due to their natural metabolism after injection into the blood stream, SPIONs mostly end up inside macrophages, liver, spleen or kidneys. To generate a potential target-specific SPION-based T2 contrast agent for MRI, the neurohormone peptide ALST1 was conjugated by CuAAC to the azide- and amine functionalized superparamagnetic iron oxide nanoparticles, since ALST1 is supposed to target difficult-to-treat neuroendocrinic tumours due to its analogy to galanin and somastatin receptor ligands. The organic fluorescent dye cyanine 5 (Cy5) was also conjugated to the silica-coated superparamagnetic iron oxide nanoparticles (SPIONs) via a NHS-ester to the amines to enable cell uptake studies by fluorescence microscopy. These constructs were characterized by TEM, dynamic light scattering (DLS), and IR. The amino acids of the conjugated ALST1 were determined by the HPLC method as described before for peptide-modified silica nanoparticle surfaces. Then, the relaxivity r2 was measured at 7 T. However, a r2 value of 27 L/mmolFe·s for the dual ALST1-/Cy5-functionalized silica-coated SPIONs was not comparable to T2 contrast agents in clinical use, since their relaxivity is commonly determined at 1.5 T, and no such instrument was available. However, it can be assumed that the synthesized dual ALST1-/Cy5-functionalized silica-coated SPION would show a lower r2 at 1.5 T than at 7T. Commercial T2 MRI contrast agents like VSOP-C184 from Ferropharm show at r2 values of about 30 L/mmolFe·s at 1.5 T. Still, the relaxivity of the new material has some potential for application as a T2 contrast agent. Then, the material was used in cell uptake studies by fluorescence microscopy with the conjugated Cy5 dye as a probe. The dual ALST1-/Cy5-functionalized silica-coated SPION showed a high degree of agglomeration with no cellular uptake unlike described for ALST1-functionalized nanoparticles in literature. It is assumed that upon agglomeration of the particles, constructs form which are unable to be internalized by the cellular endocytotic pathways anymore. As a future perspective, the tendency of the particle to agglomerate should be reduced by changing the coating material to polyethylene glycol (PEG) or chitosan, which are known to be bio-compatible, bio-degradable and prevent agglomeration. In the third part of the work, the rhenium compound [ReBr(CO)3(L)] with L = 2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline and its manganese analogue were synthesized by heating the ligand and rhenium pentacarbonyl bromide or and manganese pentacarbonyl bromide respectively, in toluene. However, [MnBr(CO)3(L)] was unstable upon illumination by UV light at 365 nm. Thus, it was dismissed for further application. The photophysical properties of [ReBr(CO)3(L)] were explored, by determination of the excited-state life time by the time-correlated single-photon counting (TCSPC) method and the quantum yield by a fluorescence spectrometer equipped with an integration sphere. A value of  = 455 ns, a Stokes shift of 197 nm and a rather low quantum yield =were found. Metal complexes are supposed to have superior properties compared to organic dyes due to their large Stokes shifts, long excited-state life times, and high quantum yields. Thus, amine- and azide-functionalized detonation nanodiamond (DND) as an alternative biological inert carrier system was functionalized with ALST1 to enhance its cell uptake properties. A luminescent probe for cell uptake studies using fluorescence microscopy was also attached, either based on the new rhenium complex or the commercially available organic dye Cy5, respectively. The aldehyde-functionalized rhenium complex was conjugated to the DND via oxime ligation, which is known to be a mild and catalyst-free conjugation method. The amount of peptide ALST1 on the DND was analyzed and quantified after acid hydrolysis and PITC derivatization by HPLC as described before. Then, the ALST1-/luminescent probe-functionalized DND was investigated for its photophysical properties by fluorescence spectroscopy. The Cy5-functionalized material showed a slightly lower fluorescence performance in aqueous solution than reported in literature and commercial suppliers with a life time  < 0.4 ns and quantum yields not determinable by integration sphere due to the week signal intensity. The rhenium complex-functionalized material had a very low signal intensity in only aqueous medium, and thus determination of life times and quantum yield by fluorescence spectroscopy was not possible. After incubation with MDA-MB 231 cells, the Cy5-functionalized DND could easily be detected due to its red fluorescence. However, it was not possible to visualize the rhenium complex-functionalized DND with fluorescence microscopy due to the low fluorescence intensity of the complex in aqueous medium and the lack of proper filters for the fluorescence microscope. Cy5-functionalized DND did not show any cellular uptake in fluorescence microscopy after conjugation with ALST1. Since the nanodiamond surface is known to strongly adsorb peptides and proteins, it is assumed that the peptide chain is oriented perpendicular to the nanoparticle surface and thus not able to interact with cell membrane receptors to promote cell uptake of the particles. As a future perspective, the ALST1-promoted cellular uptake of the DND should be improved by using different linker systems for peptide conjugation to prevent adsorption of the peptide chain on the particle surface. The new analytical methods for amino-, azide-, and peptide-functionalized nanoparticles have great potential to assist in the quantification of nanoparticle surface modifications by UV/Vis spectroscopy and HPLC. The determination of surface amine and azide groups based on the cleavage of conjugated Fmoc-Rink amide linker and detected by UV/Vis spectroscopy is applicable to all amine-/azide-functionalized nanomaterials. However, particles which form very stable suspension with the cleavage mixture can cause quantification problems due to scattering, making an accurate quantification of dibenzofulvene and its piperidine adduct impossible. The detection of tyrosine- and histidine-containing peptides based on the Pauly reaction is well-suited as a fast and easy-to-perform qualitative demonstration of successful peptide surface conjugation. However, its major drawback as a colourimetric approach is that coloured particles cannot be evaluated by this method. The amino acid analysis based on HPLC after acid hydrolysis of peptides conjugated to nanoparticle surfaces to its individual building blocks and subsequent derivatization with PITC, can be used on all nanomaterials with peptide or protein surface modification. It allows detection of amino acids down to picomolar concentrations and even enables analysis of very small peptide surface loadings. However, the resulting HPLC traces are difficult to analyze. Three new analytical methods based on UV/Vis and HPLC techniques have been developed and established. They assisted in the characterization of the synthesized DND and SPIONs with dual functionalization by ALST1 and Cy5 or [ReBr(CO)3(L)], respectively. However, the nanomaterials showed no cellular uptake due to a high tendency to agglomerate. The cellular uptake should be improved and the tendency to agglomerate of the SPIONs should be reduced by changing the surface coating from silica to either PEG or chitosan. Furthermore, different linker systems for connecting peptides to DND surfaces should be synthesized and evaluated to reduce potential peptide chain adsorption.}, subject = {Nanopartikel}, language = {en} } @phdthesis{Nagel2015, author = {Nagel, Christoph}, title = {Novel manganese- and molybdenum-based photoactivatable CO-releasing molecules: synthesis and biological activity}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-120376}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2015}, abstract = {Since its discovery as a small signaling molecule in the human body, researchers have tried to utilize the beneficial cytoprotective properties of carbon monoxide in therapeutic applications. Initial work focused on the controlled direct application of CO gas. However, to circumvent the disadvantages of this method such as requirement for special equipment, hospitalization of the patient and the risk of overdosing, metal-carbonyl complexes were developed as CO-releasing molecules (CORMs) which are able to deliver CO in a tissue-specific manner. However, upon the release of CO from the metal coordination sphere, complex fragments termed inactivated CORMs (iCORMs) with free coordination sites remain which can undergo nonspecific follow-up reactions under physiological conditions. Thus, the first aim of the present thesis was the coordination of tetradentate ligands such as tris(2-pyridylmethyl)amine (tpa), bis(2-pyridylmethyl)(2-quinolylmethyl)amine (bpqa), bis(2-quinolylmethyl)(2-pyridylmethyl)amine (bqpa) and tris(2-quinolylmethyl) amine (tmqa) in a tridentate facial manner to a fac-Mn(CO)3 moiety previously established as a photoactivatable CO-releasing molecule (PhotoCORM). The desired coordination of the pedant donor group upon photolytic CO release at 365 nm was demonstrated by UV/Vis-, IR- und 1H NMR experiments and verified by DFT calculations. All complexes of the series showed long-term dark stability in phosphate-buffered saline (PBS), but released between two and three equivalents of carbon monoxide with half-lives of around 5-10 minutes upon illumination at 365 nm. Although the photolytic properties of the complexes were quite similar besides the differences in type of hetereoaromatic ligands, the determination of the logP values showed an increase of lipophilicity with the number of quinoline groups, which might enable tissue-specific uptake. A significant cellular manganese uptake as well as the binding of CO released upon photolysis to the cytochrome c oxidases in E. coli cells was demonstrated for [Mn(CO)3(tpa)]+. Furthermore, this complex exhibited photoinduced bactericidal activity when the cells were grown in succinate-containing medium and thus unable to change their metabolism to mixed acid fermentation. In the second part of the project, the hexadentate ligand 1,4,7-tris(2-pyridylmethyl)-1,4,7-triazacyclononane (py3tacn) was coordinated to a facial Mn(CO)3 moiety. The resulting [Mn(CO)3(py3tacn-3N)]+ complex has one pedant donor group per labile carbonyl ligand and thus is a significant improvement over the 1st generation tpa-complexes. The metal-coligand inactivated CORM (iCORM) fragment expected to be generated upon complete photolytic CO release, [Mn(py3tacn-6N)]2+, was synthesized independently and will serve as a well-defined negative control in upcoming biological tests. The corresponding CORM has long-term dark stability in pure dimethylsulfoxide or phosphate-buffered myoglobin solution, with three equivalents of CO released with a half-life of 22 minutes upon illumination at 412 nm. The photolysis was also followed by IR spectroscopy and the intermediates, in line with a stepwise release of carbon monoxide, and occupation of vacated sites by the pedant pyridine group were verified by DFT calculations. Due to possible tissue damage by energy-rich light and the inverse correlation of tissue penetration depth and illumination wavelength, the absorption maxima of PhotoCORMs should ideally be in the phototherapeutic window between 600 and 1200 nm. Thus, in the third part of this work, a series of heterobinuclear Mn(CO)3/Ru(bpy)2 PhotoCORMs was prepared to shift the absorption of these compounds into the red region of the UV/Vis spectrum. For the synthesis of such Mn(I)/Ru(II) complexes, the bridging ligands 2,3-di(2-pyridyl)quinoxaline (dpx) and 3-(pyridin-2-yl)-1,2,4-triazine[5,6-f]-1,10-phenanthroline (pytp) were prepared and the two binding pockets subsequently filled with a Ru(bpy)2 and a fac-Mn(CO)3 moiety. The resulting two heterobinuclear metal complexes [Ru(bpy)2(dpx)MnBr(CO)3]2+ and [Ru(bpy)2(pytp)MnBr(CO)3]2+ as well as [Ru(etx)(tbx)MnBr(CO)3]2+ with etx = ethyl(2,2':6',2''-terpyridine)-4'-carboxylate and tbx = N-((2,2':6',2''-terpyridin)-4'-yl)2,2'-bipyridine-5-carboxamide which was prepared by a metal precursor provided by the group of Prof. Dr. Katja Heinze showed a significant shift of the main absorption bands to higher wavelengths as well as two times higher extinction coefficients than the analogous mononuclear Mn(I) compounds. However, both the Mn(I)/Ru(II) and Mn(I) complexes had a reduced stability in phosphate-buffered myoglobin solution even in the absence of light. The efficiency of the CO-release from [Ru(etx)(tbx)MnBr(CO)3]2+ and [Ru(bpy)2(dpx)MnBr(CO)3]2+ could be controlled by proper choice of the excitation wavelength. A change from 468 to 525 nm or even 660 nm led to a decrease of the number of CO equivalents released from two to one and an elongation of the half-lives. Finally, since nitric oxide also serves as a small messenger molecule in the human body with its signaling pathways interacting with those of CO, a mixed-ligand CO/NO metal complex was sought. [Mo(CO)2(NO)(iPr3tacn)]+ with iPr3tacn = 1,4,7-triisopropyl-1,4,7-triazacyclonane was selected from the literature and its molecular structure determined by single crystal diffraction, demonstrating the presence of an NO+ ligand in the coordination sphere as indicated by a MO-N-O angle close to 180°. Photolysis of [Mo(CO)2(NO)(iPr3tacn)]+ required high-energy UV light, which prevented a quantification of the CO release due to photolytic decomposition of the myoglobin. However, solution IR experiments showed that the complex lost the two carbon monoxide ligands upon illumination at 254 nm while the NO remained tightly bound to the metal. The structures observed of the intermediates were also verified by DFT calculations. In conclusion, in this project, four different classes of novel transition metal-based photoactivatable CO-releasing molecules (PhotoCORMs) were prepared and studied. The first group incorporated one additional free donor group per LMn(CO)3 moiety but varied in the number of coordinated pyridyl and quinolinyl groups which allows the control of the lipophilicity of these compounds. As an extension of this concept, the second series incorporated one free donor group per labile carbonyl ligand which gives rise to well-defined photolysis products that can be independently prepared and assayed. The third class was based on a Ru(II) photosensitizer unit connected to a MnBr(CO)3 PhotoCORM moiety. This shifts the absorption maximum from 500 nm to about 585 nm in [Ru(bpy)2(dpx)MnBr(CO)3]2+. Finally, a first mixed-ligand CO/NO carrier molecule was evaluated for its photolytic behavior. However, while the carbonyl ligands were photolabile at low excitation wavelengths, release of the NO ligand was not observed under the conditions studied. In a next step, detailed studies on the bioactivity of the different classes of PhotoCORMs need to be carried out with partner groups from biochemistry to fully explore their biomedical potential.}, subject = {Kohlenmonoxid}, language = {en} } @phdthesis{Roth2021, author = {Roth, Patrick}, title = {Metalltricarbonyl-basierte CO-releasing molecules (CORMs): Variation der Freisetzungskinetik und Biokonjugation}, doi = {10.25972/OPUS-24017}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-240171}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2021}, abstract = {Kohlenstoffmonoxid ist ein wichtiges kleines Signalmolek{\"u}l das im menschlichen K{\"o}rper durch die enzymatische Wirkung von H{\"a}m-Oxygenase (HO) auf H{\"a}m produziert wird. F{\"u}r eine thera-peutische Anwendung werden Metallcarbonyl-Komplexe als CO-releasing molecules (CORMs) untersucht, die eine kontrollierte Freisetzung in biologischen Zielstrukturen erlauben. Daf{\"u}r wird entweder die Ligandenperipherie ("drug sphere") modifiziert oder die CORMs an bio-molekulare Tr{\"a}gersysteme konjugiert. Im Rahmen dieser Arbeit stand dabei die lichtinduzierte Freisetzung von Kohlenstoffmonoxid aus Mangan(I)tricarbonyl-Komplexen im Vordergrund. Die oktaedrische Koordinationssph{\"a}re des Metallzentrums wurde dabei durch verschiedene faciale tridentate Liganden komplettiert, welche außerdem eine einfache und modulare Verkn{\"u}pfung mit biologischen Tr{\"a}ger-molek{\"u}len erm{\"o}glichen sollten. Als Chelatoren wurden Derivate von N,N-Bis(pyridin-2-ylmethyl)amin (bpa) ausgew{\"a}hlt, in denen das zentrale Stickstoffatom mit Alkylaminen unterschiedlicher Kettenl{\"a}nge funktionalisiert ist, welche {\"u}ber Amid-Bindungen mit Carboxylat-modifizierten Tr{\"a}germolek{\"u}len verkn{\"u}pft werden k{\"o}nnen. Diesen bpa-Liganden sollte ein neuartiges Ligandensystem auf der Basis von N-(Phenanthridin-6-ylmethyl)-N-(chinolin-2-ylmethyl)ethan-1,2-diamin (pqen) gegen{\"u}bergestellt werden, in denen die Phenanthridin-Gruppe interessante photophysikalische und photochemische Eigenschaften erwarten l{\"a}sst. Die CO-releasing molecules sollten zudem mit den isostrukturellen Rhenium(I)tricarbonyl-Komplexen verglichen werden, die als Marker f{\"u}r die Fluoreszenz-mikroskopie dienen.}, subject = {Metallcarbonyle}, language = {de} } @phdthesis{Roehrig2015, author = {R{\"o}hrig, Florian}, title = {Verbesserung der Medikamenteneinbringung in solide Tumoren durch Modifikation der extrazellul{\"a}ren Matrix}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-117381}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2015}, abstract = {Bei der Behandlung solider Tumoren spielen systemisch verabreichte Chemotherapeutika eine wich- tige Rolle. Allerdings akkumulieren diese Therapeutika besser in normalem Gewebe als in Tumoren. Als Ursache f{\"u}r diesen unzureichenden Transport von Medikamenten in den Tumor wurde bisher vor allem die dysfunktionale Tumorvaskulatur diskutiert. Diese befindet sich in einem chaotischen und unreifen Zustand ohne ausreichende Bedeckung der Gef{\"a}ße mit stabilisierenden Perizyten. Aus dem Zustand der Vaskulatur resultierend erreichen Medikamente den Tumor nur in geringem Ausmaß und werden dort heterogen verteilt. Als Grund f{\"u}r den Zustand der Vaskulatur wur- de ein großer {\"U}berschuss an pro-angiogenetischen Faktoren im Tumor ausgemacht. Durch eine anti-angiogenetische Behandlung konnte in pr{\"a}klinischen Modellen f{\"u}r einen gewissen Zeitraum die Tumorvaskulatur „normalisiert" werden. Dies zeichnete sich vor allem durch Ver{\"a}nderung von zwei wichtigen Parametern f{\"u}r die Medikamenteneinbringung aus: zum Einen kommt es zu einer Reduktion der Gef{\"a}ßdichte. Zum Anderen zu einer Reifung der Blutgef{\"a}ße. In einem Teil von Pati- enten scheint dabei der Effekt der Gef{\"a}ßverbesserung zu {\"u}berwiegen und es kann eine verbesserte Perfusion detektiert werden. Mutmaßlich f{\"u}hrt dies auch zu einer verbesserten Einbringung von Therapeutika in den Tumor und so zu einer erh{\"o}hten Effizienz der Therapie. In einem weiteren Teil der Patienten scheint jedoch der Effekt der Gef{\"a}ßreduktion zu {\"u}berwiegen und die detektierte Perfusion im Tumor wird durch die Behandlung verringert. Das in dieser Arbeit verwendete MT6-Fibrosarkom-Modell reagierte auf eine anti-angiogenetische Therapie nicht mit einer sonst in murinen Modellen beobachteten Wachstumsreduktion. Die- se erm{\"o}glichte eine so bisher nicht m{\"o}gliche Untersuchung der sekund{\"a}ren Effekte einer anti- angiogenetischen Therapie wie die Medikamenteneinbringung in den Tumor. Die Vaskulatur in MT6-Tumoren zeigte dabei nach einer anti-angiogenetischen Vorbehandlung, die erwarteten Merk-male einer „normalisierten" Vaskulatur wie eine Reduktion der Gef{\"a}ßdichte bei gleichzeitiger Rei- fung der verbleibenden Gef{\"a}ße. Dies f{\"u}hrte jedoch nicht zu einer verbesserten Effizienz einer subsequenten Chemotherapie. Durch Vergleich mit einem weiteren Tumor-Modell, dem 4T1-Modell f{\"u}r ein metastasierendes Mammakarzinom, konnten signifikante Unterschiede im Gef{\"a}ßbild beider Modelle ausgeschlossen werden. Durch mikroskopische Methoden konnte dabei beobachtet werden, dass die Diffusion von Medikamenten aus den Blutgef{\"a}ßen des MT6-Modells im Vergleich zum 4T1-Modell verringert war. Weitere Untersuchungen deuten auf eine Differenz in der Qualit{\"a}t der extrazellul{\"a}ren Matrix der verwendeten Tumor-Modelle. Durch mRNA-Expressionsanalysen konnte die Enzymfamilie der Lysyloxidasen als m{\"o}gliche Ursache f{\"u}r diesen Diffusionsunterschied identi- fiziert werden. Lysyloxidasen katalysieren vor allem die Quervernetzung von Proteinen der Extra- zellul{\"a}rmatrix. Im Weiteren konnte gezeigt werden, dass die Quervernetzung von Matrixproteinen durch Lysyloxidasen urs{\"a}chlich f{\"u}r die Diffusions-Inhibierung kleiner Molek{\"u}le wie das Chemo- therapeutikum Doxorubicin sein kann. Durch spezifische Inhibition der Lysyloxidasen mittels des Inhibitors βAPN konnte diese Diffusions-Inhibition sowohl in vitro als auch im MT6-Tumor-Modell nahezu vollst{\"a}ndig verhindert werden. Die hohe Aktivit{\"a}t von Lysyloxidasen im MT6-Modell stell- te allerdings kein Alleinstellungsmerkmal dieses Modells dar. In weiteren Untersuchungen konnte gezeigt werden, dass Lysyloxidasen in einer Vielzahl von murinen und humanen Tumorzelllinien {\"u}berexprimiert wird. Die Inhibition von Lysyloxidasen durch βAPN konnte dabei in allen unter- suchten Modellen die Einbringung von Medikamenten in den Tumor erh{\"o}hen und k{\"o}nnte so eine sinnvolle adjuvante Maßnahme zur Verbesserung bestehender Chemotherapien darstellen.}, subject = {Lysin-Oxidase}, language = {de} } @phdthesis{MawambaKemo2020, author = {Mawamba Kemo, Viviane}, title = {Beladung von Microbubbles mit Platin(II)- oder Palladium(II)-Komplexen und Freisetzung unter Einwirkung von Ultraschall zur Behandlung von Glioblastomen}, doi = {10.25972/OPUS-21616}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-216165}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2020}, abstract = {Metall-basierte Antitumorwirkstoffe wie Cisplatin, Carboplatin und Oxaliplatin sind weltweit f{\"u}r die Behandlung verschiedener Krebsarten zugelassen. Resistenzbildung, starke Nebenwirkungen und ein eingeschr{\"a}nktes Spektrum responsiver Tumoren schr{\"a}nken jedoch ihren Anwendungsbereich ein. Daher ist die Suche nach neuen Platinverbindungen mit verbesserten Eigenschaften sowie Antitumor-aktiven Metallkomplexen anderer Metalle ein aktuelles Forschungsthema. Durch die Einbettung der Wirkstoffe in entsprechende Tr{\"a}germaterialien und eine Freisetzung mit pr{\"a}ziser zeitlicher und r{\"a}umlicher Kontrolle sollten sich zudem die Nebenwirkungen deutlich reduzieren lassen. Im Rahmen dieser Doktorarbeit wurde daher eine Serie von quadratisch-planaren Platin(II)- und Palladium(II)-Komplexen mit N^N^S-Chelatliganden auf der Basis von N-Phenyl-2-(pyridin-2-ylmethylen)hydrazin-1-carbothioamid und N-Phenyl-2-(chinolin-2-ylmethylen)hydrazin-1-carbothioamid synthetisiert, die mit l{\"a}ngeren Alkylketten funktionalisiert wurden, um eine hohe Affinit{\"a}t f{\"u}r Lipid-basierte Microbubbles als Tr{\"a}ger zu erreichen, aus denen die Metallkomplexe dann unter Einwirkung von Ultraschall freigesetzt werden sollten. Es wurden drei verschiedene Ligandenfamilien ausgehend von der Grundstruktur L = R1-CR2=N-NH-C(S)-NH-R3 synthetisiert, wobei R1 = 2-Pyridyl oder 2-Chinolinyl, R2 = H, CH3, C8H17 oder C10H21 und R3 = CH3 oder C6H5 gew{\"a}hlt wurden. Die Umsetzung der Liganden mit Kaliumtetrachloridoplatinat(II), Natriumtetrachloridopalladat(II) oder [PdCl2(cod)] mit cod = 1,5-Cyclooctadien f{\"u}hrte zu neutralen N^N^S-Komplexen [MCl(L)] mit M = Pd, Pt in allgemein guter Ausbeute. Die Kristallstruktur von [PtCl(L)] mit R1 = 2-Pyridyl, R2 = C10H21 und R3 = CH3 best{\"a}tigte zudem die quadratisch-planar Koordination des Metalls durch den N^N^S-koordinierten Liganden und ein Chlorid-Anion. Die Verbindungen mit R3 = CH3 zeigen im 195Pt NMR zwei Peaks, was auf das Vorliegen eines Isomerengemischs hindeutet, wobei die Daten vermuten lassen, dass neben der N^N^S-gebundenen Hauptspezies noch eine weitere mit N^N^N-koordiniertem Liganden und freier SH-Gruppe vorliegt. Solche Isomerengemische sind f{\"u}r biologischen Anwendungen ungeeignet, da die Isomere eine unterschiedliche Aktivit{\"a}t aufweisen k{\"o}nnen. Die anderen Platin(II)-Komplexe zeigen dagegen im 195Pt NMR nur einen Peak und sind somit f{\"u}r Cytotoxizit{\"a}tsstudien geeignet. Mit Hilfe des MTT-Assays wurden EC50-Werte an verschiedenen Gliablastom-Zellinien f{\"u}r zw{\"o}lf einheitliche Komplexe bestimmt. F{\"u}r die potentesten Verbindungen wurden EC50-Werte im unteren mikromolaren Bereich ermittelt (2-9 µM), so dass die Aktivit{\"a}t teilweise sogar die von Cisplatin als Referenzverbindung {\"u}bertraf. Insbesondere die Variation der aromatischen Oberfl{\"a}che in den Pyridyl- vs. Chinolinylverbindungen hatte jedoch keinen wesentlichen Einfluss auf die EC50-Werte. Zudem f{\"u}hrte eine Verl{\"a}ngerung der R2-Seitenkette bei den Palladium(II)-Verbindungen zu einer niedrigeren Aktivit{\"a}t. Die Verteilungskoeffizienten logP ergaben f{\"u}r alle Verbindungen recht {\"a}hnliche positive Werte, was die Lipophilie der Neutralkomplexe belegte. F{\"u}r eine weitere Strukturvariation wurden außerdem zwei Azido-Komplexe [M(N3)(L)] mit M = Pd, Pt in moderater Ausbeute synthetisiert. Diese wurden dann in einer „iClick"-Reaktion unter sehr milden Bedingungen mit Dimethylacetylendicarboxylat (DMAD) und 4,4,4-Trifluorobut-2-ins{\"a}ureethylester zu den Triazolat-Komplexen [M(triazolateR,R')(L)] umgesetzt. Durch 1H NMR- und 19F NMR-spektroskopische Untersuchungen wurden gezeigt, dass diese teilweise als Isomerengemische vorliegen, da das Triazolat entweder {\"u}ber das N1-, N2- oder N3-Stickstoffatom an das Metall gebunden sein kann. F{\"u}r eine ausgew{\"a}hlte Verbindung mit M = Pt und DMAD als Alkin wurde die Kinetik der „iClick"-Reaktion mit Hilfe der 1H NMR Spektroskopie untersucht. Die ermittelte Geschwindigkeitskonstante 2. Ordnung k2 = (1.82 ± 0.05).10-1 L mol-1 s 1 ist vergleichbar zum Beispiel der der etablierten strain-promoted azide-alkyne cycloaddition (SPAAC). F{\"u}r die Einbettung der lipophilen Metallkomplexe in ein Ultraschall-aktivierbares Tr{\"a}gersystem wurden aus Dipalmytoylphosphatidylcholin, 1,2-Dipalmitoyl-sn-glycero-3-phosphat, 1,2-Distearoyl-sn-glycero-3-phosphoethanolamin-N-methoxy(polyethylenglycol)-2000 Ammonium Salz und Komplexl{\"o}sung unter Zusatz von Octafluopropan gasgef{\"u}llte Microbubbles hergestellt. Stabilit{\"a}tsversuche zeigten, dass die Bl{\"a}schenzahl selbst unter Normalbedingungen innerhalb von 2 h um 50\% abnimmt. Daher sollten die Microbubbles vor jeder Verwendung t{\"a}glich frisch hergestellt und m{\"o}glichst unmittelbar danach verwendet werden. Ein unmittelbares Zerplatzen der Bl{\"a}schen wurde durch Behandlung mit Ultraschall bei 500 Hz erreicht. Die Viabilit{\"a}t der verwendeten GaMG-Zellen wudre unter diesen Bedingungen jedoch nicht beeintr{\"a}chtigt. Dennoch war die auf die Microbubbles geladene Platin-Konzentration zu niedrig, um mit dem MTT-Assay einen signifikanten Unterschied zwischen beladenen und unbeladenen Bl{\"a}schen zu erreichen, so dass hier in Zukunft noch weitere Optimierungen erforderlich sein werden.}, subject = {Glioblastom}, language = {de} } @phdthesis{Pai2014, author = {Pai, Sandesh}, title = {Synthesis of manganese tricarbonyl PhotoCORM conjugates - from small molecules to peptides and dendrimers}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-104824}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2014}, abstract = {Novel manganese(I) tricarbonyl complexes based on the tridentate bis(pyrazolyl)ethylamine (bpea) ligand with pendant functionalized phenyl groups were synthesized and conjugated to biological carrier systems like peptides and dendrimers. Their dark stability establishes them as CORM prodrugs. The monomers show a faster CO-release compared to the peptide and dendrimer conjugates. However, both monomers and peptide conjugates release two equivalents of CO upon photoactivation at 365 nm. The dendrimer conjugates can deliver up to seven equivalents of CO due to the higher number of Mn(CO)3 moieties per molecular unit. In the future, the biological activity of the conjugates needs to be further explored to establish the targeted delivery of CO to cells and tissues.}, subject = {Mangankomplexe}, language = {en} } @phdthesis{Rudolf2014, author = {Rudolf, Philipp Benjamin}, title = {Uncovering photoinduced chemical reaction pathways in the liquid phase with ultrafast vibrational spectroscopy}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-96200}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2014}, abstract = {The experimental technique predominantly employed within the scope of this Thesis constitutes one subarea of femtochemistry: the time-resolved spectroscopy of photoin- duced chemical reactions in the liquid phase by means of molecular signatures in the mid-infrared (MIR) spectral range. Probing transient vibrational states, i.e., dynamic changes in the vibrational motion of speci� c molecular subunits or functional Groups allows for a distinct separation and assignment of measured signals to emerging molecular species. For this purpose, one key building block is indispensable, which most of the investigations carried out within the � eld of femtochemistry have in common: a coherent light source delivering ultrashort laser pulses with a temporal duration that matches the femtosecond time scale on which molecular motions typically occur. This instrumentation enables the observation of photoinduced chemical reactions from the starting point|the excitation event to the appearance of intermediates to the nal formation of stable photoproducts after several pico- or nanoseconds. This work comprises the acquisition and presentation of time-resolved spectroscopic data related to promising molecular systems upon photoexcitation as well as the im- plementation and testing of experimental optical techniques both for the presented experiments but as well for experiments conceivable in the future. In addition, linear spectroscopy measurements and quantum-chemical simulations on the emerging chemical species have been carried out. In so doing, the primary processes and subse- quently emerging reaction products of two compounds on a timescale of several nanoseconds after photoexcitation have been elucidated in great detail. Both compounds, the [Mn(CO)3(tpm)]+ (tpm = tris(2-pyrazolyl)methane) CO-releasing molecule (CORM) and the 5-diazo Meldrum's acid (DMA), are of academic interest but in addition belong to molecular classes that might be utilized in the near future as dark-stable prodrugs under physiological conditions or that are already utilized in industrial chemistry procedures, respectively. The � ndings of both studies gave rise to implement and examine two techniques for prospective transient absorption experiments, namely the shaping and characterization of ultraviolet (UV) laser pulses and the recording of two-photon excitation spectra. Beyond that, since each of the depicted experiments is based on the detection of weak transient absorption signals in the MIR spectral region, two dif- ferent detection schemes, via chirped-pulse upconversion (CPU) on the one hand and via direct multichannel MCT detection on the other hand, have been juxtaposed at the conclusion of this work. Since both techniques are suitable in femtosecond pump-probe measurements but thereby exhibit individual strengths and weaknesses, a comparative study provides clari� cation of the respective pros and cons. The � first study introduced within this work investigates the complex photochemistry of DMA, a photoactive compound used in lithography and industrial chemistry. By femtosecond MIR transient absorption spectroscopy covering several nanoseconds, the light-induced dynamics and ultrafast formation of several photoproducts from the manifold of reaction pathways have been disclosed to form a coherent picture of the overall reaction scheme. After UV excitation of DMA dissolved in methanol to the second excited state S2, 70\% of excited molecules relax back to the S0 ground state. In compet- ing processes, they can either undergo an intramolecular Wolff rearrangement to form ketene, which reacts with a solvent molecule to an enol intermediate and further to carboxylate ester, or they � rst relax to the DMA S1 state, from where they can isomerize to a diazirine. The third competing reaction channel, having the lowest quantum efficiency with respect to the � rst two channels, is the formation of a singlet carbene out of the S1 state. From there an ylide can arise or, via an intersystem crossing, the triplet form of the carbene follows. Whereas the primary reaction steps occur on a picosecond timescale, the subsequently arising intermediates and stable photoproducts are formed within a few hundreds to thousands of picoseconds. For a reliable identi� cation of the involved compounds, density functional theory calculations on the normal modes and Fourier-transform infrared spectroscopy of the reactant and the photoproducts in the chemical equilibrium accompany the analysis of the transient spectra. Additional experiments in ethanol and isopropanol led to slight spectral shifts as well as elongated time constants due to steric hindrance in transient spectra connected with the ester Formation channel, further substantiating the assignment of the occurring reaction pathways and photoproducts. The study demonstrated that the combination of linear and time-resolved spectroscopic measurements in conjunction with quantum-chemical calculations constitutes a powerful tool to unravel even highly complex photoreactions exhibiting multiple consecutive intermediate states within parallel reaction pathways. Although some of the individual reaction steps, for example the ketene formation via Wolff rearrangement, have been observed on ultrashort time scales before, this work encompassed the Observation of the whole set of appearing photoproducts of DMA in different alcohol solutions within several nanoseconds. In this sense, the ultrafast photochemistry of DMA represents a prototype example for a multisequential reaction scheme, elucidated by the capabilities of femtosecond MIR spectroscopy. With a modi� fied instrumentation concerning amongst others the system delivering the fundamental laser pulses or the generation of the UV pump pulses, the next ob- jective within this work was to elucidate the primary processes upon UV Irradiation of a manganese tricarbonyl CORM in aqueous environment. The time-resolved experiment was performed with two different pump wavelengths and furthermore supported by linear spectroscopy methods and time-dependent density functional theory (TDDFT) calculations on the excited states as well as DFT calculations on the ground states. The measurements revealed that irradiating the compound with UV excitation pulses primarily leads to ultrafast photolysis of one CO ligand. Geminate recombination may occur within one picosecond but it remains a minor process as the photolyzed CO group is liberated and the unoccupied coordination site is predominantly fi� lled by an incoming solvent molecule. There was no evidence for hot CO bands, i.e., the remaining CO ligands|in the dicarbonyl photoproduct as well as in the intact CORM are not vibrationally excited through the UV excitation of the CORM. According to this, the excess energy merges into low-frequency vibrational modes associated with the molecule as a whole. Since studies on a macroscopic scale at irradiation times of several minutes prove that UV irradiation eventually leads to the release of two or even all three CO ligands, further loss of CO most likely necessitates manganese oxidation or another interaction with light. To clarify the latter, a consecutive UV pulse was employed in order to excite the photoproducts subsequent to the initial pump interaction. However, the data obtained was not instructive enough to de� nitely exclude the manganese oxidation being responsible for the loss of further CO groups. Besides the exchange of a CO Group by a solvent molecule or the geminate recombination, the employment of two different excitation wavelengths in combination with � ndings derived from the TDDFT calculations suggested another reaction process, namely the possibility that the excitation does not lead to any bond cleavage at all. As the CORM under investigation is tissue-selective and cytotoxic against cancer cells, knowledge of these � rst photoinduced reaction steps is essential for a full understanding of its biological activity. Inspired by these two studies, experimental techniques for prospective transient absorption measurements have been implemented and tested within preparative measure- ments. First, in the course of a UV-pump-MIR-probe experiment with speci� cally tailored excitation pulses, one could pursue the aim of coherently controlling the outcome of a photoreaction in the liquid phase. Out of the rich photochemistry of DMA the vibrational signature of a particular molecular species might thereby serve as a feedback signal, which is a central part of a learning loop that adaptively determines the pulse shape that steers the quantum mechanical system upon photoexcitation into a desired direction. This motivated the installation and testing of devices by means of which the shaping and characterization of ultrashort laser pulses in the UV could be performed. Second, motivated by the biological applications of CORMs, one can imagine a scenario where a certain amount of CORMs is deposited inside cancerous tissue. Since the activation of CO loss by means of UV pulses is not possible due to the absorption characteristics of biological tissue, the simultaneous excitation via two photons from the visible spectral regime seems appealing. However, success or failure of such an application depends on whether the deposited compound efficiently absorbs two photons simultaneously, i.e., whether the two-photon absorption cross section is large enough. Therefore, a setup to record two-photon excitation spectra under full consideration of the crucial laser pulse parameters like the pulse duration, energy and central wavelength was arranged and tested. The � rst results were obtained with a commercially available reference system (Mn2CO10) but the setup as well as the described measurement and data analysis procedure can easily be applied to record the two-photon absorption cross section of more promising molecular systems. Third, as the detection of probe pulses in the MIR spectral region is part of each time-resolved measurement throughout this thesis, a comparison between the newly established technique of CPU and direct multi- channel MCT detection is presented by means of pump{probe experiments on Mn2CO10 and Co4CO12 with a 1 kHz shot-to-shot data acquisition. It was shown that the CPU detection technique scores with its high spectral resolution and coverage of the easy-to-handle and more cost-effective CCD detectors. On the other hand, in the course of the additional nonlinear upconversion process intensity fluctuations of the chirped fundamental pulses are transferred to the probe spectrum in the visible regime. This entails a lower signal-to-noise ratio than the direct MCT detection, which can be compensated by an additional normalization procedure applied to the CPU probe pulses. As a consequence, the CPU detection scheme offers more flexibility for future investigations employing MIR probe pulses. This is of great importance for many applications within the presented � eld of femtochemistry as a huge variety of time-resolved investigations on a multitude of systems in the liquid phase is based on the detection of weak transient absorption signals in the MIR spectral region.}, subject = {Ultrakurzzeitspektroskopie}, language = {en} } @phdthesis{Graf2022, author = {Graf, Dominic}, title = {Surface and active site modification of proteins with organometallic markers and inhibitors}, doi = {10.25972/OPUS-28742}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-287424}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2022}, abstract = {After implementing a reliable mass spectrometry based kinetic study the indole conjugation with different organometallic indoles led to questions about the electronical and sterical influences on reactivity. The substitution pattern of the ferrocene functionalized indoles at the six-membered ring determines the electron density on the C3 atom, which reacts with the formed Schiff base. Since the experimental results showed the exact opposite trend, covalent docking studies were performed elucidating the importance of surface interactions. These studies were in harmony with the experimental results and determined lysine 33 as most preferable conjugation site as well as substitution in 6-position as most favourable pattern. The amine motif in compounds 6, 7 and 8 proofed to be easily fragmented by the ESI method used. The amide linker in 10 remains intact but shows a lower conversion. Those two inherent characteristics are however preferable for well-defined and site-specific bioconjugation. The synthesis and evaluation of piano stool complex derivatives with manganese and rhenium metal centre 15, 16, 18 and 22 gave additional guidance by the interpretation of applicable structural motifs. The electron-withdrawing carbonyl groups lead to the hindrance of fulvene formation and thus to no fragmentation as seen with the ferrocene group. The total conversion is low compared to 8, only 22 shows a good enough conversion to mainly monoconjugate of 45\% and a possible radio-labelling application as 99mTc analogue. As consequence manganese complexes with a stable facial tricarbonyl unit and a tridentate chelator with 4-, 5- and 6-substituted aminomethylindole conjugated through an amide bond were synthesized and consecutively evaluated. The resulting organometallic indole derivatives 29, 30 and 31 all showed a total conversion around 40\% similar to 16, but at the same time a rate constant in the range of 10-4 s-1 like the organic indole. Besides the similar conversion, the rate constants followed the trend of the 6-substituted derivative as fastest and then 5- and 4- substituted derivative with decreasing reactivity. For underlining the usage as technetium label for the best out of the series 31, a rhenium analogue was prepared. The resulting compound 32 was especially interesting, because the conversion was even higher than the 70\% of 8 with a total of 88\%. Additionally, the rate constant was a tenfold higher as well. This rendered compound 32 as best possible 99mTc analogue for further application as radio-label. After the success of 32 and realizing the sterical benefits resulting from the flexible tridentate ligand-system, substitution at the five-membered ring was explored. The complexes 33, 34 and 35 are based on indole-2-carboxylic acid and with the difference of the length of the alkyl spacer between amide and complex to probe for the influence and sterical hindrance, but all three derivatives showed no conjugation which excludes functionalization in 2-position. As the C3 is used for the actual bioconjugation, the last possible derivatization was realized on the indole-N1 by using 1-(3-bromopropyl)indole as building block during the synthesis of the ligand-system. The corresponding manganese 36 and rhenium 37 complexes both showed similar properties of a moderate conversion like 22 and a rate constant in the range of 10-5 s-1. In conclusion the rhenium complex 32 with the 6-substitution pattern at the tridentate indole-bearing ligand remains the most promising structure. The here developed liquid chromatography coupled mass spectrometry-based assay for the determination of inhibitory activity of drug candidates against the 3CLpro of the sever acute respiratory syndrome coronavirus type 2 was successfully implemented and especially designed to give, due to the available absorption spectra and corresponding mass traces, further insight in the otherwise through fluorescence resonance energy transfer-based assays neglected influences on the inhibition results. Starting with a literature-known quinolone containing covalent inhibitor 42 an N1-methylated derivative 43 and their analogues 44 and 45 in which the benzoic acid was exchanged for ferrocene carboxylic acid were synthesized. The inhibition of 3CLpro was evaluated by the concentration of initial 15mer peptide left after incubation and for that purpose the for 280 nm defined molar attenuation coefficient of (26.41±0.59) L*mol-1*cm-1 determined and used. The results showed a reaction of DL dithiothreitol with the less stable benzoic acid esters leading to a moderate inhibitory effect. The methylation in N1-position showed an increase in stability. The methylated and with ferrocene carboxylic acid functionalized derivative showed a complete inhibition during the timeframe of the assay. In search of a fluorescent and therefore traceable inhibitor, 4 hydroxycoumarin was used to synthesize the analogue with benzoic acid 49 and ferrocene carboxylic acid 50. Both derivatives were less stable than their analogues but exhibited the same trend of a more stable ferrocene-derived compound, which exerted a higher inhibition as well. After preparing and testing the model thioester 53 and showing an inactivation of the established inhibitor ebselen, it was concluded that the reaction with DL dithiothreitol reduces the concentration of active intact inhibitor and therefore decreases the inhibition rate during the assay. The next step was proofing the reducing agent as non-essential for the fast assay conducted in a timeframe of 5 min to circumvent the negative influence of DL dithiothreitol. By excluding every inhibition-altering part, the resulting method is the perfect tool for precise statements in relation of inhibitory activity. Then the inhibition assay was repeated for ebselen and the best out of the here introduced organometallic inhibitors 45. Both give equivalent results of a complete inhibition during the measurement. The implemented liquid chromatography coupled mass spectrometry-based assay has many advantages over the fluorescence resonance energy transfer-based assays in which all the information and insight accumulated by the evaluation of uv/vis traces and mass spectra are not available leading to wrong or deviating results regarding the inhibitory capacity of inhibitor candidates.}, subject = {Inhibition}, language = {en} } @phdthesis{Feizy2019, author = {Feizy, Nilab}, title = {iClick-Reaktionen von Palladium(II)azid- und Platin(II)azid-Komplexen mit tridentaten N,N,N-Chelatliganden und elektronenarmen Alkinen}, doi = {10.25972/OPUS-17893}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-178938}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2019}, abstract = {Katalysatorfreie [3+2]-Cycloadditionen von Aziden mit Alkinen werden in der bioorthogonalen Chemie h{\"a}ufig verwendet und haben großes Potential zur milden Synthese von Biokonjugaten. W{\"a}hrend solche Reaktionen in der Ligandenperipherie von Metallkomplexen h{\"a}ufiger angewendet werden ist, sind solche Reaktionen direkt in der inneren Koordinationssph{\"a}re von Metallzentren bisher nur wenig erforscht. Die neue Beispiele daf{\"u}r sind die Synthese und Untersuchungen der Kinetik und Reaktivit{\"a}t einer Reihe von Rhodium(III)azid-Halbsandwichkomplexen der allgemeinen Formel [Rh(Cp*)(N3)(bpyR,R)]+ oder von isoelektronische und isostrukturelle Molybd{\"a}n(II)azid- und Wolfram(II)azid-Komplexe mit verschiedenen elektronenarme Alkine. Das Ziel der vorliegenden Arbeit waren daher iClick-Reaktionen (engl. inorganic click, „iClick") von Palladium(II)azid- und Platin(II)azid-Komplexen der allgemeinen Formel [M(N3)(L)]+ und [M(N3)(L)] mit elektronenarmen Alkinen Dimethylacetylendicarboxylat (DMAD) und 4,4,4-Trifluorobut-2-ins{\"a}ureethylester. Als Liganden kamen die N,N,N-Chelatoren 1,3-Bis(arylimino)isoindolin (HL1-4) die sich nur im Bezug auf die Position der Methylgruppen in den Pyridinringen unterscheiden, 6',6"-Dimethyl-2',2:6,2"-terpyridin (L5) und 2,6-Bis(3-pyridazinyl)pyridin (L6) zum Einsatz. Die Reaktionen von L1-L4 mit [MCl2(cod)] (M = Pd, Pt) liefert neutrale Komplexe [MCl(L1-3)] und f{\"u}r L5 einfach geladene [MCl(L5)]+. Das koordinierte Chlorid wurde dann mit Natriumazid substituiert. Im abschließenden Teil der Arbeit wurde die zwei Alkinen in iClick-Reaktion verwendet um Palladium(II)- und Platin-Triazolat-Komplexe zu synthetisieren. F{\"u}r die resultierenden Triazolat-Komplexe wurde eine N2-koordinierten des Triazolat-Liganden durch R{\"o}ntgenstrukturanalyse f{\"u}r baii-Triazolat-Komplexe best{\"a}tigt. Besonderes Merkmal dieser Verbindungen ist, dass der Triazolat-Ligand aus Platzmangel senkrecht zum 1,3-Bis(arylimino)isoindolin-Ligand steht. In verwandten Terpyridin-Komplexen sind der mono- und tridentate Ligand dagegen coplanar. Mit 1,3-Bis(6-methyl-2-pyridylimino)isoindolin als Ligand konnten man keine Metall-Komplexe hergestellt werden, da die zus{\"a}tzlichen Methylgruppem in 6',6"-Positionen aus sterische Gr{\"u}nden eine Reaktion mit [MCl2(cod)] verhindern. Auch der in drei Stufen synthetisierte Ligand 6',6"-Dimethyl-2',2:6,2"-terpyridin der im Vergleich zu Terpyridin zwei zus{\"a}tzliche Methylgruppen in 6',6"-Position besitzen reagiert nur mit [PdCl2(cod)] nicht aber mit [PtCl2(cod)], da der Ionenradius von Pt(II) gr{\"o}ßer als der von Pd(II) ist. Die hergestellte Chlorid-, Azid- und Triazolat-Komplexe mit L5 als N,N,N-Chelator waren nur in DMSO L{\"o}slich. Darin zersetzt es sich jedoch teilweise wieder in den freien Liganden. Die zus{\"a}tzlichen Methylgruppem in 6',6"-Positionen verhindern aus sterische Gr{\"u}nden die Chlorid-, Azid- und Triazolat-Komplexe stabil zu bleiben. Ligand L6 konnte nur in sehr niedrige Ausbeute isoliert werden, da in der letzten Stufe bzw. bei Stille-Kupplung zwischen 2,6-Bis(trimethylstannyl)pyridin und 3-Iodopyridazin die Homokupplungsprodukte von 3-Iodopyridazin entsteht, sodass die nicht getrennt werden konnten. Aufgrund der niedrigen Ausbeute wurden dann mit L6 keine Metall-Komplexe hergestellt. Die Kinetik der iClick-Reaktion ist ein entscheidender Faktor, wenn diese f{\"u}r die Markierung von Bio(makro)molek{\"u}len eingesetzt werden soll, da die Markierungsreaktion schneller als der interessierende biologische Prozess ablaufen muss. Daher wurden mit IR- und UV/Vis-Spektroskopie die Geschwindigkeitskonstanten pseudoerster Ordnung f{\"u}r die iClick-Reaktion der verschiedenen baii-Palladium(II)azid- und baii-Platin(II)azid-Komplexe mit Dimethylacetylendicarboxylat (DMAD) und 4,4,4-Trifluorobut-2-ins{\"a}ureethylester bestimmt. Hier sollte insbesondere der Einfluss der zus{\"a}tzlichen Methylgruppen in 4',4"- bzw. 5',5"-Positionen am 1,3-Bis(arylimino)isoindolin-Liganden sowie die Variation des Metallzentrums und Alkins auf die Geschwindigkeit der iClick-Reaktionen untersucht werden. Mit IR-Spektroskopie wurden Geschwindigkeitskonstanten um (2.8-4.9)⋅10-4 s-1 an Alkinen erhalten. Die Einf{\"u}hrung elektronenschiebender Methylgruppen in 4',4"- bzw. 5',5"-Positionen am 1,3-Bis(arylimino)isoindolin-Liganden f{\"u}hrt zu einer Erh{\"o}hung der Geschwindigkeitskonstant einem Faktor von 1.3 bzw. 1.2 gegen{\"u}ber 1,3-Bis(2-pyridylimino)isoindolin. Die iClick-Reaktion mit Platin als Metall ist 1.3-mal schneller als mit Palladium. Elektronenarme Alkine wie 4,4,4-Trifluorobut-2-ins{\"a}ureethylester f{\"u}hren im Vergleich zu Dimethylacetylendicarboxylat (DMAD) zu einer 1.8-fachen Erh{\"o}hung der Reaktionsgeschwindigkeit. Mit UV/Vis-Spektroskopie wurden niedrigere Geschwindigkeitskonstanten um 8.9·10-6 - 3.3·10-5 s-1 nur f{\"u}r die iClick-Reaktion der 1,3-Bis(arylimino)isoindolinplatin(II)azid-Komplexe mit Dimethylacetylendicarboxylat (DMAD) und 4,4,4-Trifluorobut-2-ins{\"a}ureethylester bestimmt, weil die Spektralen Unterschiede zwischen Azid-Vorstufe und Triazolat-Produkt mit Palladium als Metallzentren zu gering sind. Auch hier konnte die Erh{\"o}hung der Geschwindigkeitskonstanten durch Verwendung elektronen{\"a}rmerer Alkine best{\"a}tigt werden. Hier sollte die iClick-Reaktion in Zukunft f{\"u}r gr{\"o}ßere Auswahlm{\"o}glichkeiten an Chelatoren optimiert und außerdem die Geschwindigkeitskonstanten der Bildung von iClick-Produkten mit anderen Methoden untersucht werden, bevor biologische Tests durchgef{\"u}hrt werden.}, subject = {Alkine}, language = {de} } @phdthesis{Peng2020, author = {Peng, Kun}, title = {iClick reactions as a modular access to palladium(II) and platinum(II) triazolato complexes: Trends in kinetics and biological activity}, doi = {10.25972/OPUS-21161}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-211613}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2020}, abstract = {In the context of this work, important trends in the influence of the metal center, coligand, and alkyne reaction partner on the iClick reaction of square-planar palladium(II) and platinum(II) complexes with a N^N^N, C^N^N, or S^N^N coordination sphere and a number of internal as well as terminal alkynes were elaborated. Preliminary bioactivity studies on a human cancer cell line gave low micromolar EC50 values, for the most promising compound comparable to cisplatin serving as a reference drug. The further application of the iClick reaction to bioconjugation will be explored in future work.}, subject = {Click-Chemie}, language = {en} } @phdthesis{Berger2022, author = {Berger, Sarina Maria}, title = {Influence of Charge and Its Distribution on Biological Applications of Bis-Triarylboranes and Preliminary Investigations on H\(_2\)O\(_2\)-Cleavable Aryl Boronate Esters}, doi = {10.25972/OPUS-24314}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-243147}, school = {Universit{\"a}t W{\"u}rzburg}, year = {2022}, abstract = {This dissertation describes the synthesis of an unsymmetrically-substituted triarylborane. This term describes a three-coordinate boron atom that is bound to three different aromatic systems, namely 2,6-dimethylphenyl, mesityl, and 4-(N,N-dimethylamino)-2,6-dimethylphenyl. It is also demonstrated that the amine functionality can be converted with methyl triflate into an ammonium moiety. The investigation of photophysical and electrochemical properties of this compound in comparison with the non-aminated and di-aminated analogues of the triarylborane is described besides other investigations of e. g. singlet oxygen sensitization, rotational barriers, and fundamental DFT calculations. Based on these investigations, selectively mono-, bis- and tris-dimethylamino- and trimethylammonium-substituted bis-triarylborane bithiophene chromophores were synthesized and their photophysical, and electrochemical properties were investigated together with the water solubility and singlet oxygen sensitizing efficiency of the cationic compounds Cat1+, Cat2+, Cat(i)2+, and Cat3+. Comparing these properties with the results obtained for the mono-triarylboranes reveals a large influence of the bridging unit on the investigated properties of the bis-triarylboranes. In addition, the interaction of the cationic bis-triarylboranes with different polynucleotides were investigated in buffered solutions as well as the ability of these selectively charged compounds to enter and localize within organelles of human lung carcinoma and normal lung cells. All these investigations demonstrate that the number of charges and their distribution influences the interactions and staining properties as well as most of the other properties investigated. In addition, preliminary investigations on H2O2-cleavable boronate esters in the presence of stochiometric amounts of H2O2 are described for three different aryl boronate esters.}, subject = {Triarylborane}, language = {en} }