Nltrogen-15 isotope effects have been measured In the simple bond formation reactions of triethylamine with methyl iodide, triethylamine with ethyl iodide and triethylamine with ethyl bromide. Tests have been performed, to establish that the reactions were unidirectional under the experimental conditions and that the chemical procedures used to convert the quaternary ammonium salts to molecular nitrogen proceeded quantitatively. The mass spectrometer data were examined by rigorous statistical tests. At 6°C. the values of the ratio of rate constants, k14/k15, were: triethylamine-methyl iodide, 1.0009 +/- 0.0005; triethylamine-ethyl iodide, 1.0007 ±0.0006; trimethylamine-ethyl bromide, 0.9994 ±0.0006. For the reaction of triethylamine with ethyl iodide the value of k14/k15 increased slightly with increasing temperature. The results have been discussed in terms of the Bigeleisen theory of isotope effects with particular reference to the correct form for the effective mass term. / Thesis / Doctor of Philosophy (PhD)

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Metal-catalyzed, hydrogenative methods for carbon-carbon bond formation are attractive alternatives to traditional carbonyl addition reactions. Through in situ generation of aldehyde and organometallic species, these redox-triggered reactions circumvent the need for preactivation of reactive partners, thereby providing a more atom economic, efficient approach to carbonyl addition products. Efforts have been focused on the development of ruthenium-catalyzed coupling reactions of primary and secondary alcohols to basic feedstock chemicals and easily accessible and stable unsaturated compounds. To perform highly stereoselective reactions, investigation into the factors that control stereoselectivity in ruthenium catalyzed transfer hydrogenative couplings was undertaken. As a critical tool for the construction of organic molecules, modernizing methods for carbonyl addition can contribute to the evolution of synthetic organic methodology. / text

An increasing global energy demand requires alternative fuel sources. A promising method is artificial photosynthesis. Although, the artificial processes are different from the natural photosynthetic process, the basic principles are the same, i.e. to split water and to convert solar energy into chemical energy. The energy is stored in bonds, which can at a later stage be released upon combustion. The bottleneck in the artificial systems is the water oxidation. The aim of this research has been to develop catalysts for water oxidation that are stable, yet efficient. The molecular catalysts are comprised of organic ligands that ultimately are responsible for the catalyst structure and activity. These ligands are often based on polypyridines or other nitrogen-containing aromatic compounds. This thesis describes the development of molecular ruthenium catalysts and the evaluation of their ability to mediate chemical and photochemical oxidation of water. Previous work from our group has shown that the introduction of negatively charged groups into the ligand frameworks lowers the redox potentials of the metal complexes. This is beneficial as it makes it possible to drive water oxidation with [Ru(bpy)3]3+-type oxidants (bpy = 2,2’-bipyridine), which can be photochemically generated from the corresponding [Ru(bpy)3]2+ complex. Hence, all the designed ligands herein contain negatively charged groups in the coordination site for ruthenium. The first part of this thesis describes the development of two mononuclear ruthenium complexes and the evaluation of these for water oxidation. Both complexes displayed low redox potentials, allowing for water oxidation to be driven either chemically or photochemically using the mild one-electron oxidant [Ru(bpy)3]3+. The second part is a structure–activity relationship study on several analogues of mononuclear ruthenium complexes. The complexes were active for water oxidation and the redox potentials of the analogues displayed a linear relationship with the Hammet σmeta parameter. It was also found that the complexes form high-valent Ru(VI) species, which are responsible for mediating O–O bond formation. The last part of the thesis describes the development of a dinuclear ruthenium complex and the catalytic performance for chemical and photochemical water oxidation. It was found that the complex undergoes O–O bond formation via a bridging peroxide intermediate, i.e. an I2M–type mechanism.

We report the application of our rhodium-catalyzed nucleophilic ring-opening methodology to the enantioselective synthesis of nitrogen-substituted polycyclic heterocycles. By using a cationic Rh(I) triflate catalyst in the presence of the chiral Josiphos ligand PPF-PtBu2, the ring opening reactions on dihydrooxaquinoline and dihydrooxaisoquinoline using different nucleophiles afford access to multiple dihydroquinolines and dihydroisoquinolones in high yield and high enantioselectivity (up to 99% total yield and >99%ee). A variety of nucleophiles were shown to be compatible with the catalytic system. The electronic effects in the new ring opening reactions were investigated using a variety of nucleophiles. It was found that reactivity and enantioselectivity of the ring opening products depends on the electronic effects as well as the position of the substituents on the substrates. Good yields and high ee of regioisomeric products are obtained using electron donating substituents, whereas electron withdrawing substituents decelerate the reactions.

We report the application of our rhodium-catalyzed nucleophilic ring-opening methodology to the enantioselective synthesis of nitrogen-substituted polycyclic heterocycles. By using a cationic Rh(I) triflate catalyst in the presence of the chiral Josiphos ligand PPF-PtBu2, the ring opening reactions on dihydrooxaquinoline and dihydrooxaisoquinoline using different nucleophiles afford access to multiple dihydroquinolines and dihydroisoquinolones in high yield and high enantioselectivity (up to 99% total yield and >99%ee). A variety of nucleophiles were shown to be compatible with the catalytic system. The electronic effects in the new ring opening reactions were investigated using a variety of nucleophiles. It was found that reactivity and enantioselectivity of the ring opening products depends on the electronic effects as well as the position of the substituents on the substrates. Good yields and high ee of regioisomeric products are obtained using electron donating substituents, whereas electron withdrawing substituents decelerate the reactions.

The formation of aberrant disulfide bonds is a structural consideration for the manufacturing of the extracellular domain of human CD83 (hCD83ext), a potential therapeutic protein. In certain instances, hCD83ext protein products, even when stored frozen, tend to dimerize or even multimerize through the formation of aberrant intermolecular disulfide bonds. Herein, we discovered an analytical inconsistency and applied a modified sample preparation protocol for proper structural analysis of hCD83ext products which are heterologously expressed in Escherichia coli and subsequently purified. In addition, a mutant derivative with the Cys100Ser mutation was identified as an improved version which did not form dimers or multimers. The identification of this mutant variant as a more potent therapeutic protein than other hCD83ext species demonstrated that the structural variation associated with disulfide bond formation can be a critical issue for rigorous control of the quality and bioactivity of therapeutic proteins. The application of this mutant variant for protein therapeutic is currently under exploration. As a comparative study, the hCD83ext was expressed as a glutathione-S-transferase (GST) fusion in two E. coli B strains, i.e. BL21 and Origami B having a reductive and oxidative cytoplasm. The final therapeutic products of hCD83ext produced by the two expression hosts exhibited significant differences in protein conformation and molecular properties, which presumably resulted from different disulfide patterns. The study highlights the importance of developing proper host/vector systems and biomanufacturing conditions for the production of recombinant therapeutic proteins with a consistent product quality. Cys27 in the hCD83ext was identified as a target for molecular manipulation. Two E. coli strains of BL21(DE3) and Origami B(DE3) were used as the expression host to produce the Cys27 mutants. It was observed that Cys27 was involved in the in vivo formation of intramolecular disulfide bonds when hCD83ext was expressed in Origami B(DE3). The Origami-derived protein products had a higher tendency than the BL21-derived counterparts for multimerization via the in vitro formation of intermolecular disulfide bonds. Various analyses were conducted to identify the structural differences among these mutant variants. Most importantly, molecular stability was enhanced by the Cys27 mutations since the Cys27 mutants derived from either BL21 or Origami were much less susceptible to degradation compared to wild-type hCD83ext. This study highlights the implications of aberrant disulfide bond formation on the production of therapeutic proteins. To address an inconsistent bioactivity issue that is primarily due to the aberrant formation of disulfide bonds associated with the presence of five cysteine residues, i.e. AA 27, 35, 100, 107, and 129, the molecular role that each cysteine plays upon the formation of intramolecular or intermolecular disulfide bonds was characterized, using various hCD83ext mutant variants derived by two E. coli expression hosts, i.e. BL21(DE3) and Origami B(DE3). Among the five cysteines, Cys100 and Cys129 can act as a bridging cysteine for in vitro multimerization via the formation of intermolecular disulfide bonds. The multimerization can be alleviated to some extent with less free Cys129 residues, associated with the possible formation of Cys27-Cys129 intramolecular disulfide bond. As a result, introducing the Cys27 mutation can increase the multimerization presumably via freeing more Cys129 residues. In addition, protein stability can be improved in the presence of the Cys27 mutation. The formation of the Cys27-Cys129 intramolecular disulfide bond appears to be more effective in the presence of the Cys100 mutation, resulting in the suppression of multimerization. The two conserved cysteine residues, i.e. Cys35 and Cys107, can be potentially linked to form an intramolecular disulfide bond, particularly when the protein is produced in Origami B(DE3).

Dichloromethane is one solvent that is used widely in laboratory. It is so stable that it seldom reacts with other materials. There are only a few reports that involves dichloromehane as a reactant. Previous in our laboratory, we discovered a dichloromethane activation product. To study the details of the methylene insertion prouct, [(SCH2S)PS]2Zn form (Et4N)2 (PS3Zn)2, we used monodentate thiols in different condition as models to understand what condition thiols may react with dichloro methane. We found out thiols can react with dichloromethane under strong base. When there is aromatic ring in a thiol, that thiol will react with dichloromethane easily. Adding zinc salts showed that zinc ion is an inhibitor in thiolate/dichloromethane reactions. We synthesized Tris(3-trimethylsilyl-2-thiophenyl)phosphine [H3SiPS3] and SiPS3Zn complex to study the parallel effect of adding bulky silyl substitaents on PS3 ligand towards reaction with dichloromethane. Most conclusion are similar to that of monodentate thiols. There is one thing that differs from the results of monedentate thiols: the SiPS3Zn complex does react with dichloromethane.