This thesis is concerned with the synthesis of functionalised piperidines. The piperidine unit is found in numerous alkaloids and other natural products, and has also been incorporated into synthetic compounds of significant therapeutic or strategic synthetic potential. Although numerous syntheses of individual piperidine compounds have been published, there are few general routes to this class of compounds, and there is therefore a need for such routes to be developed. Bicyclic lactams derived from (S)-pyroglutamic acid have been shown to be useful synthons in the synthesis of pyrrolidines. An analogous bicyclic lactam was therefore synthesised from (S)-lysine. Alkylations at the carbon atom a- to the lactam carbonyl group were studied in detail. A number of different groups could be introduced in good to excellent yield, and with moderate to good stereoselectivity. The selectivity of these reactions was believed to depend on the shape of the bicyclic system, and a stereoelectronic effect of the nitrogen lone pair. The bicyclic lactam was further elaborated to an activated enone. This was shown to be relatively unreactive to cycloaddition reactions. It is believed that this is because the activating ester group is not conjugated with the carbon-carbon double bond. However, conjugate addition could be achieved in high yield and excellent stereoselectivity using a zinc enolate Reformatsky reagent. An epoxide could also be formed in excellent yield and with excellent stereoselectivity. The Reformatsky adduct was shown to be a suitable substrate for a Pb(IV)-mediated arylation reaction. Examples of the functionalised lactams were deprotected to give hydroxymethyl piperidinones and cyclic amino acids. This route therefore allows access to the desired novel compounds.

Submitted in complete fulfillment for the Degree of Master of Applied Sciences in Biotechnology, Durban University of Technology, Durban, South Africa, 2016. / Piperidine derivatives are essential heterocyclic compounds that have beneficial roles in the medical and commercial sector. They can be isolated from plant material and can be chemically synthesised using simple cost efficient methods. Piperidines and their derivatives are clinically used to prevent postoperative vomiting, facilitate radiological evaluation, correct gastrointestinal function as well as speed up gastric emptying before anaesthesia. Piperidine derivatives also demonstrate a wide spectrum of biological activities which include; antimicrobial, anticancer, anti- TB, anti-HIV, anti-inflammatory, analgesic, anti-influenza, anti-inflammatory and antitumor activity. The properties of piperidine derivatives depend on the nature of the side chains and their orientation. Based on the promising data that demonstrated the synergistic effects of biological agents with piperidine derivatives, the aim of our research is to determine the pharmacological activities, i.e. (i) antimicrobial activity, (ii) anti-inflammatory, (iii) anti-oxidant activity, (iv) cytotoxicity, and (v) biosafety of six piperidine derivatives, PM1 to PM6. All six piperidine derivatives (PM1-PM6) screened for antimicrobial activity exhibit characteristics of varying degrees of microbial inhibition against some Gram-positive and Gram-negative bacteria (B. cereus, B. subtilis, E. coli, S. aureus, Kl. Pneumonia, M. liuteus and P. aurenginosa) with the exception of B. polymixa, S. marcescens and S. faecalis. Certain piperidine derivatives did not demonstrate high inhibition activity towards the fungal strains, with inhibition only shown against four fungal species; A. niger, A. flavus, C. albicans and S. cerevisiae. Thus it is proposed that minor changes could be made to the structure of the compounds so that they can alter the effect that the compounds have on the specific fungi strains. With regard to antioxidant activity it is noted that the concentrations of the test compounds are directly proportional to the percentage of scavenging capacity. In comparison of the piperidine derivatives (PM1-PM6) to Rutin (reference standard), it was illustrated that Rutin displayed the best antioxidant activity. All six piperidine derivatives (PM1-PM6) showed greater than 50% anti-inflammatory activity, whilst the anti-inflammatory reference standard NCGA displayed the greatest activity in comparison to the piperidine derivatives tested. The safety of the piperidine derivatives was tested by assaying cytotoxicity, against melanoma, MCF7 cancer cells and normal fibroblasts as well as Brine shrimp lethality assay. All piperidine derivatives demonstrated high cytotoxicity activity against both cancer cell lines (melanoma and MCF7) and around 50 – 52% cytotoxicity against healthy cells. Chloro substitution of the phenyl ring increases cytotoxicity of compounds (Aerluri et al., 2012). This compound can be used in the treatment of cancer cells while inhibiting 50% of normal cells. All six piperidine derivatives (PM1-PM6) were also tested for toxicity against Artemia salina in a brine shrimp lethality assay. Piperidine derivatives exhibited varying degree of toxic activity towards the shrimp, with all derivatives displaying ± 50% toxic activity at 1000 µg/mL. These results reveal a directly proportional relationship between concentration of drug and toxicity. It remains a future research objective to modify these piperidine compounds (PM1-PM6) chemically to produce more derivatives for further biological evaluation. All the studied piperidine compounds have possible leads for optimization to carry out pre-clinical trials. We can conclude that the substitution of different side chains on the piperidine nucleus results in varying degree of pharmacological activity. Also, compounds containing the substitution of a chloro group at position 4 and a fluoro group at position 2 on the phenyl ring attached to carbon 2 and 6 on the piperidine nucleus resulted in high pharmacological activity. This good pharmacological activity was also exhibited by compounds containing substitutions of a methoxy group at position 3 on the phenyl ring attached to carbon 1 and 6 on the piperidine nucleus. Compounds containing a methoxy group positioned at carbon 4 on the phenyl ring which is attached to carbon 1 and 4 on the piperidine nuleus presented low pharmacological activity. Low activity was also exhibited by compounds containing substitution of a cyano group at position 4 on the phenyl ring which is attached to carbon 2 and 6 on the piperidine ring and a methyl group at position 4 on the phenyl group attached to a nitrogen at position 1 on the piperidine nucleus. / M

Dans ce mémoire, il sera question de la formation de dérivés pipéridines en utilisant la fragmentation de Grob. Tout d’abord, une introduction sur les alcaloïdes ainsi que sur l’expertise du groupe Charette associée à leur formation démontrera l’importance de ces composés dans le domaine de la chimie organique. Cela sera suivi par un résumé de la fragmentation de Grob incluant les conditions de réactions utilisées, l’importance de la structure de la molécule initiale, les prérequis stéréoélectroniques ainsi que les modifications qui y ont été apportées. Le chapitre 2 sera dédié au développement de la méthodologie c’est-à-dire, à l’optimisation de tous les paramètres jouant un rôle dans la fragmentation de Grob. Par la suite, l’étendue de la réaction ainsi que des explications sur la régiosélectivité et la diastéréosélectivité de la réaction seront fournies. La méthodologie peut être exploitée dans un contexte de synthèse qui sera démontré dans le chapitre 3. De plus, elle servira pour une étude mécanistique qui est encore d’actualité à partir du concept d’effet frangomérique. Finalement, quelques projets futurs, notamment des améliorations possibles de la méthodologie, seront présentés dans le dernier chapitre. Le tout sera suivi d’une conclusion résumant l’ensemble des travaux effectués. / This thesis discusses the formation of piperidine derivatives using the Grob fragmentation. Firstly, an introduction of the important alkaloid family as well as previous work completed by the Charette group towards the synthesis of these compounds will be demonstrated. This will be followed by a summary of the Grob fragmentation including a discussion of the reaction conditions, molecular structures, stereoelectronic requirements and modifications of the Grob fragmentation. Chapter 2 will be dedicated to the development of the methodology and more precisely, to the optimization of all parameters necessary to the reaction. Furthermore, the scope of the reaction and some explanation of the regioselectivity and the diastereoselectivity of the reaction will be discussed. The developed methodology can be used in a total synthesis and will be demonstrated in Chapter 3. Moreover, using the frangomeric effect concept, a mechanistic study on the Grob fragmentation will be discussed. Finally, some future projects, especially possible improvement of the methodology, will be presented in the last chapter. This is followed by a conclusion and a summary of the work completed on this project.

Dans ce mémoire, il sera question de la formation de dérivés pipéridines en utilisant la fragmentation de Grob. Tout d’abord, une introduction sur les alcaloïdes ainsi que sur l’expertise du groupe Charette associée à leur formation démontrera l’importance de ces composés dans le domaine de la chimie organique. Cela sera suivi par un résumé de la fragmentation de Grob incluant les conditions de réactions utilisées, l’importance de la structure de la molécule initiale, les prérequis stéréoélectroniques ainsi que les modifications qui y ont été apportées. Le chapitre 2 sera dédié au développement de la méthodologie c’est-à-dire, à l’optimisation de tous les paramètres jouant un rôle dans la fragmentation de Grob. Par la suite, l’étendue de la réaction ainsi que des explications sur la régiosélectivité et la diastéréosélectivité de la réaction seront fournies. La méthodologie peut être exploitée dans un contexte de synthèse qui sera démontré dans le chapitre 3. De plus, elle servira pour une étude mécanistique qui est encore d’actualité à partir du concept d’effet frangomérique. Finalement, quelques projets futurs, notamment des améliorations possibles de la méthodologie, seront présentés dans le dernier chapitre. Le tout sera suivi d’une conclusion résumant l’ensemble des travaux effectués. / This thesis discusses the formation of piperidine derivatives using the Grob fragmentation. Firstly, an introduction of the important alkaloid family as well as previous work completed by the Charette group towards the synthesis of these compounds will be demonstrated. This will be followed by a summary of the Grob fragmentation including a discussion of the reaction conditions, molecular structures, stereoelectronic requirements and modifications of the Grob fragmentation. Chapter 2 will be dedicated to the development of the methodology and more precisely, to the optimization of all parameters necessary to the reaction. Furthermore, the scope of the reaction and some explanation of the regioselectivity and the diastereoselectivity of the reaction will be discussed. The developed methodology can be used in a total synthesis and will be demonstrated in Chapter 3. Moreover, using the frangomeric effect concept, a mechanistic study on the Grob fragmentation will be discussed. Finally, some future projects, especially possible improvement of the methodology, will be presented in the last chapter. This is followed by a conclusion and a summary of the work completed on this project.

Doctor of Philosophy / Department of Chemistry / Duy H. Hua / Three research projects are described in this dissertation, and they are: (i) discovery of piperidine derivatives as T-type calcium channel inhibitors for the treatment of epilepsy and neuropathic pain and as protein disulfide isomerase inhibitors for the treatment of influenza viral infection; (ii) discovery of peptide-based calcitonin gene-related peptide receptor antagonists for the treatment of inflammatory pain; and (iii) synthesis of substituted 6-(dimethylamino)-2-phenylisoindolin-1-ones for the inhibition of luciferase. T-type calcium channels are important regulators of nervous system, and upregulated T-type calcium channel activities have been found to link to various types of neurological disorders, such as epilepsy and neuropathic pain. To discover novel T-type calcium channel blockers, a series of 1,4-disubstituted piperidine derivatives were designed and synthesized. Among them, compound 1-4 was found to be a good T-type calcium channel inhibitor with an IC₅₀ of 1 nM for Ca[subscript v]3.2 inhibition. It also showed 86% suppression of seizure induced death in mice and good in vivo analgesic effects on both thermal and mechanical pain thresholds in Spared Nerve Injury rat models. Therefore 1-4 can potentially be used as a T-type calcium channel blocker in the treatment of epilepsy and neuropathic pain. Influenza is a respiratory viral infection. Since viruses rely on host cell proteins for their entry, survival and replication, development of drugs targeting host cell proteins has identified as an effective strategy in controlling viral infections. We synthesized a series of 1,4-disubstituted piperidine derivatives for the inhibition of protein disulfide isomerase enzyme and influenza. Among them, 1-29 was found to possess strong anti-influenza activity (EC₅₀ = 2.5 µM). This suggests the potential use of piperidine scaffold in designing anti-influenza drugs in future. Calcitonin gene-related peptide (CGRP) receptor antagonism has been identified as a successful approach for the treatment of inflammatory pain. Therefore, a novel class of peptide antagonists of CGRP receptor was synthesized and screened for their binding affinities to the CGRP receptor and their analgesic effects on inflammatory-induced pain in rats. Among them, peptide 2-3 showed a higher binding affinity towards the CGRP receptor than previously reported peptide antagonists and exhibited analgesic effects up to 2 h in both Aδ and c-fiber pain tests. Therefore 2-3 indicates its potential use as a CGRP receptor antagonist in the treatment of inflammatory pain. Firefly luciferase is commonly used as a reporter in cells expressing a luciferase gene or its enzymatic activity under the control of a promoter of interest to assess its transcriptional activity. It has been found that some molecules such as molecules with carboxylic acid moiety can directly inhibit luciferase activity in cells. However, it is suggested that carboxylic acid moiety of the compounds may also be associated with side reactions in cells. Therefore, to study whether carboxylic acid moiety causes side effects, we designed two probe molecules, 3-1 and 3-2. Synthesis of probe molecule 3-2 is discussed. Synthesis of probe molecule 3-1 and further investigation of its luciferase inhibition will therefore be useful to understand the toxicity of carboxylic acid containing drugs in future.

No / Forumla chem. . A variety of 5-chloro-2(H)-1,4-oxazin-2-ones bearing a range of substituents at their 3- and 6-positions undergo Diels-Alder cycloaddition as a 2-azadiene component with electron-rich, electron-deficient, and electron-neutral dienophiles. These reactions proceed with moderate regio- and stereoselectivity to afford relatively stable and readily isolable bridged bicyclic lactone cycloadducts. Chemical manipulation of these cycloadducts affords highly substituted and functionally rich piperidines. The regio- and stereochemical preferences of the cycloadditions of 5-chloro-2(H)-1,4-oxazin-2-ones are investigated computationally using density functional theory (B3LYP/6-31G*).

Chapter 1 gives an overview of the fluorine chemistry field, from its early developments to recent applications in medicinal chemistry. The development of asymmetric electrophilic or nucleophilic installation of fluorine in organic molecules is highlighten. Chapter 2 of this thesis discusses the enantioselective synthesis of α-fluoroamides. The study is applied to the synthesis of fluoroenantiomers of the bioactive molecule capsaicin and short-chain analogues. The biological activity of these compounds is assayed with the TRPV1 receptor. Results show that enantioselective α-fluoroamides (R)-97, (R)-99 and (S)-99 can generate differentiated biological responses, from TRPV1 agonists to TRPV1 antagonists. Chapter 3 focuses on the optimisation and development of 2-(aminomethyl)piperidine (R)-251 dihydrochloride. The development of 2-(aminomethyl)piperidine (R)-251 as its ditetrafluoroborate salt proved to offer excellent reactivity and solubility for the preparation of derivatives. This tetrafluoroborate salt was used to improve the syntheses of organocatalysts 2,2,2-trifluoro-N-(piperidin-2-ylmethyl)acetamide 363 and 4-methyl-N-(piperidin-2-ylmethyl)benzenesulfonamide 364.The catalytic properties of these latter two molecules for asymmetric Mannich reaction is demonstrated. Both (R)-363 and (R)-364 show up to 86% ee, in a typical 20 mol% loading, but loading of (R)-363 as low as 5 mol% still induces the catalysis.

No / A novel series of 1,4-disubstituted aminoanthraquinones were prepared by ipso-displacement of 1,4-difluoro-5,8-dihydroxyanthraquinones by hydroxylated piperidinyl- or pyrrolidinylalkyl-amino side chains. One aminoanthraquinone (13) was further derivatized to a chloropropyl-amino analogue by treatment with triphenylphosphine-carbon tetrachloride. The compounds were evaluated in the A2780 ovarian cancer cell line and its cisplatin-resistant variants (A2780/ cp70 and A2780/MCP1). The novel anthraquinones were shown to possess up to 5-fold increased potency against the cisplatin-resistant cells compared to the wild-type cells. Growth curve analysis of the hydroxyethylaminoanthraquinone 8 in the osteosarcoma cell line U-2 OS showed that the cell cycle is not frozen, rather there is a late cell cycle arrest consistent with the action of a DNA-damaging topoisomerase II inhibitor. Accumulative apoptotic events, using time lapse photography, indicate that 8 is capable of fully engaging cell cycle arrest pathways in G2 in the absence of early apoptotic commitment. 8 and its chloropropyl analogue 13 retained significant activity against human A2780/cp70 xenografted tumors in mice.