Montmorillonite-catalyzed allylation of aldehydes with potassium allyltrifluoroborate is a convenient method for the synthesis of homoallylic alcohols. This chemistry was applied to various unsaturated aldehydes, and the homoallylic alcohols produced were used as common intermediates for two separate but related synthetic routes to α,β-unsaturated ketones. In the first route, oxidation of the alcohol to the ketone was followed by base-catalyzed isomerization of the olefin to the α,β-unsaturated ketone. This was subjected to ring closing metathesis conditions to afford the cyclic enone of ring size n. In the other route, ring-closing metathesis was performed first, followed by oxidation of the alcohol and isomerization of the olefin to the cyclic enone of ring size n+1.

Montmorillonite-catalyzed allylation of aldehydes with potassium allyltrifluoroborate is a convenient method for the synthesis of homoallylic alcohols. This chemistry was applied to various unsaturated aldehydes, and the homoallylic alcohols produced were used as common intermediates for two separate but related synthetic routes to α,β-unsaturated ketones. In the first route, oxidation of the alcohol to the ketone was followed by base-catalyzed isomerization of the olefin to the α,β-unsaturated ketone. This was subjected to ring closing metathesis conditions to afford the cyclic enone of ring size n. In the other route, ring-closing metathesis was performed first, followed by oxidation of the alcohol and isomerization of the olefin to the cyclic enone of ring size n+1.

Two alternative compounds have been prepared and studied, as alternatives to the benzyl amine in the preparation of hemoglobin bis-tetramers. The reactivity of these compounds towards the activated ester groups on trimesoyl tris(3,5-dibromosalicylate), and 3,5-dibromosalicylate benzyl ester has been studied, as well as their functionality in a range of acidic solutions. Initial rate kinetics was used to determine rate constants for hydrazinolysis, and hydrolysis of the two esters. The objective was to find a suitable functional group that would be less basic than the benzyl amine yet still exhibit good nucleophilicity in the preparation of hemoglobin bis-tetramers. Hydrolysis being the main side reaction limiting the yield, could be slowed down in an acidic solution, but the nucleophile used could not be protonated in the process. It has been found that the benzyl hydrazine was the most effective and able to react with cross-linked hemoglobin to give the most product.

Two alternative compounds have been prepared and studied, as alternatives to the benzyl amine in the preparation of hemoglobin bis-tetramers. The reactivity of these compounds towards the activated ester groups on trimesoyl tris(3,5-dibromosalicylate), and 3,5-dibromosalicylate benzyl ester has been studied, as well as their functionality in a range of acidic solutions. Initial rate kinetics was used to determine rate constants for hydrazinolysis, and hydrolysis of the two esters. The objective was to find a suitable functional group that would be less basic than the benzyl amine yet still exhibit good nucleophilicity in the preparation of hemoglobin bis-tetramers. Hydrolysis being the main side reaction limiting the yield, could be slowed down in an acidic solution, but the nucleophile used could not be protonated in the process. It has been found that the benzyl hydrazine was the most effective and able to react with cross-linked hemoglobin to give the most product.

The addition of an aldehyde C−H bond across a ketone functionality, formally a hydroacylation, has emerged as an atom-economical approach to the synthesis of esters. While this is an efficient strategy for producing biologically-relevant materials, the field of transition metal-catalyzed ketone hydroacylation is currently limited to intramolecular systems. The development of a new rhodium catalyst will be presented, and its application to intermolecular ketone hydroacylation will be discussed. Ester products were synthesized from unfunctionalized, aliphatic aldehydes and chelating ketones in excellent yields under relatively mild reaction conditions. Efforts towards an asymmetric intermolecular ketone hydroacylation will be described, including the application of known chiral catalysts and the development of novel chiral phosphine ligands for asymmetric catalysis. Ester products were obtained in as high as 78% enantiomeric excess.

The synthesis of glycoconjugates has facilitated a wide variety of techniques for the detailed study of carbohydrates and their interactions in biological systems. However, when only small amounts of the isolated oligosaccharide are available, multistep synthetic approaches are not possible. This thesis explores new synthetic methods for the preparation of glycoconjugates without protecting group manipulations. A new glycosidation method was developed which introduces N-glycopyranosylsulfonohydrazides as glycosyl donors for the protecting group-free synthesis of O-glycosides, glycosyl azides and oxazolines. The glycosyl donors were synthesized in a single chemical step by condensing p-toluenesulfonylhydrazide with the corresponding mono- and disaccharides. The N-glycopyranosylsulfonohydrazides were activated with NBS and subsequently glycosylated with the desired alcohol or transformed to the oxazoline or glycosyl azide. Recent advances in chemoselective ligation methods for the functionalization of unprotected carbohydrates have provided new routes towards complex glycoconjugates. Despite the wide use of those chemoselective methods, the properties of these linkages have not been thoroughly investigated. Characterization of a series of glycoconjugates formed by chemoselective ligation of xylose, glucose and N-acetylglucosamine with either an acyl hydrazide, a p-toluenesulfonylhydrazide or an N-methylhydroxylamine were carried out to gain further insight into the optimal conditions for the formation and the stability of these useful conjugates. Their apparent association constants (9-74 M-1) at pD 4.5, as well as rate constants for hydrolysis were determined at pH 4.0, 5.0 and 6.0. The half-lives of the conjugates varied between 1 h and 300 days. All the compounds were increasingly stable as the pH approached neutrality. Finally, selective chemical modification of a glycosaminoglycan chondroitin sulfate was attempted at the non-reducing end by utilizing the Δ4-uronic acid functional group formed upon cleavage of the glycosaminoglycan with a bacterial lyase enzyme. The captodative double bond of the unique Δ4-uronic acid functionality was unreactive towards Michael addition, even if the carboxylate was methylated. Trials towards radical addition using thiyl radicals were unsuccessful, although a synthesized model phenyl Δ4-uronic acid monosaccharide was successfully functionalized under the same conditions.

The increasing environmental impact of society has created the need for the modification of current and the implementation of new industrial processes which are less environmentally harmful. However, these new or modified processes must be less material-, time-, and cost-intensive such that they are more economically beneficial than the processes they are to supplant. The described research was inspired by these two ideas and is comprised of two projects, both focused on the creation of recyclable, CO2-switchable methods of separating organic compounds. The development and optimization of switchable water, a CO2-switchable ionic strength aqueous solvent is described. The solvent system, an amine/aqueous mixture, had the ability to switch from low to high ionic strength via the application and removal of CO2. This solvent system was able to achieve salting-out of water-miscible organics in comparable amounts to several inorganic salts typically used for salting-out. The switchable water system was explored for use in several industrial applications. A homogeneous catalysis recycling system was developed for the hydroformylation of styrene. A catalyst was able to be recovered and recycled five times with minimal loss of activity. The use of switchable water to expedite the settling of clay suspensions was also explored. Switchable water, when used as process water did not settle bulk clay solids as quickly as a CO2-only treatment, but did however increase the settling rate of small clay fines resulting in lower turbidities of the supernatant. The solvent could be recovered from settled clay suspensions and recycled up to three times. Finally, efforts towards the realization of CO2-switchable chiral resolving agents are presented. It is hypothesized that chiral nitrogenous bases could be used as switchable resolving agents by forming diastereomeric salt pairs with racemic alcohols via the application of CO2. After separation of the diastereomers, removal of CO2 would afford the resolved alcohol enantiomers and the chiral base. Efforts towards the synthesis of a library of chiral nitrogenous bases and the screening of their reactivity with CO2-treated alcohols are described. Several bases were generated, but the necessary reactivity between the bases and the racemic alcohols in the presence of CO2 was not observed. / Thesis (Ph.D, Chemistry) -- Queen's University, 2012-12-20 16:26:50.635

Our group has developed a strategy for the synthesis of benzofused carbocycles and heterocycles through tandem palladium-catalyzed reactions of gem-dibromoolefins. In these syntheses, one bromide undergoes a Pd-catalyzed cyclization reaction, and the other participates in an orthogonal inter- or intramolecular Pd-catalyzed reaction to functionalize or annulate that ring, respectively. Chapter 1 describes the pairing of an intramolecular C–N bond forming reaction (the Buchwald-Hartwig coupling) with an intramolecular direct arylation for the synthesis of fused indole derivatives. A range of previously unknown heterocycles were synthesized through this method. Chapter 2 describes the synthesis of benzothiophenes through the combination of a Pd-catalyzed C–S coupling with an orthogonal Suzuki, Heck, or Sonogashira reaction. This represents the first example of the incorporation of Pd-catalyzed C–S coupling into a tandem reaction. In Chapter 3, a tandem intramolecular Heck / intermolecular Suzuki reaction is described for the synthesis of methyleneindenes. Studies on this reaction have provided advanced understanding of the mechanism, including how variation of the ligand controls the regioselectivity of the reaction.

The structural complexity of glycosaminoglycans (GAGs) such as heparin and heparan sulfate (HS) and their numerous biological roles, brings forth the need to develop new methods, capable of studying GAGs and their interactions with peptides and proteins under native settings. This thesis explores the development of chemical tools to study heparin/HS binding interactions under physiologically relevant conditions using fluorescence. In chapter 2, we designed peptide-based quinolinium probes to study the structural requirements of cationic peptides required for high affinity peptide-heparin interactions. These fluorescent probes enabled the study of peptide-heparin interactions at nM concentrations allowing the calculation of peptide-heparin binding constants. It was observed that peptides with positive charge displayed on one face of an α-helix in a continuous arrangement bound to heparin with the highest affinity and that heparin likely prefers to bind to these peptides while remaining in an extended conformation. In chapter 3, we set out to study an important biological role of HS which involves the binding and sequestering of proteins at the cell surface, facilitating endocytosis. HS has been implicated in the mechanism of cell penetrating peptide (CPP) cell uptake, with different CPPs showing different degrees of HS dependence on uptake as well as different mechanisms of entry. The role of HS in the mechanism of CPP uptake was investigated in chapter 3 using fluorescent peptide-based probes incorporating fluorophore/quencher pairs. These were used to identify and characterize the ability of heparin/HS to bind and cluster with CPPs to form colloidally stable aggregates. It was shown that the CPP Antp formed much more stable clusters with heparin than the TAT peptide despite both peptides having similar binding affinity for a single heparin chain. These findings were used to explain the cell surface HS dependence of Antp on cell uptake via endocytosis in contrast to the low dependance of TAT on HS and its uptake via translocation. A general model relating the ability of a CPP to cluster surface HS to its preferred mechanism of cell entry was proposed. In chapter 4, a strategy to selectively, and site specifically acylate carbohydrate binding proteins was developed using thioester-based affinity conjugates. It was possible to label maltose binding protein, a periplasmic protein, with high yield and selectivity at a single lysine residue proximal to the maltose binding site. Selective protein labeling could be carried out in bacterial cell extracts and in live bacterial cells. This strategy can potentially be applied to develop protein-based carbohydrate biosensors as well as profile carbohydrate binding proteins in biological samples.

Photoisomerization of azobenzene can be used to reversibly photo-control peptide and protein structure, thereby offering the potential to probe peptide and protein function directly in living systems. Most azobenzene photo-switches to date however require the use of UV light, which suffers from poor tissue penetration and can lead to cell damage. In addition, the unknown stability of azobenzene photo-switches within the reducing intracellular environment has limited these switches to extracellular applications. The ability to red-shift the photo-switching wavelength and tune the thermal cis-to-trans rate independently from one another has been a central challenge. Incorporating a piperazine-like moiety at the 4,4′-para positions red-shifts the π-π* band of an azobenzene-based cross-linker such that trans-to-cis photoisomerization can be triggered with blue light. Accompanying this red-shift was a faster cis-to-trans thermal rate, which allowed for fast secondary structural changes of the attached peptides (τ½ ~2s). Piperazine-like moieties at the 2,2′-ortho positions of an azobenzene-based cross-linker resulted in a similar red-shift in the π-π* band but a longer-lived cis isomer relative to its para-counterpart (τ½ ~minutes). These results suggest that the thermal rate could be tuned independently from the photo-switching wavelength by appropriate para or ortho substitution. The effect of 2,2′,6,6′-ortho-tetramethoxysubstitution provided an alternative approach to red-shifting the photo-switching wavelength. These groups caused an unconventional red-shift in the n-π* band of the trans isomer allowing for trans-to-cis and cis-to-trans photoisomerization to occur with green and blue light, respectively. In this case, the half-life of the cis isomer was not shortened, but rather extended relative to its parent compound (τ½ ~days versus minutes). These results provide progress in tuning the photo-switching wavelength independently from the thermal rate. A fluorescent reporter was developed to determine the stability of a commonly employed 4,4′-diamido derivative in vivo. Photoisomerization was found to cause time-dependent changes in fluorescein emission intensity. The reporter was microinjected in zebrafish embryos and photo-switching could be imaged for at least two days. This work provides the first direct evidence of azobenzene photo-switching in vivo, and indicates that it will be possible, in general, to photo-control peptide and protein function in living systems.