The use of peptide drugs has gained popularity recently. Peptides are attractive drug targets due to their high specificity and potency towards their biological targets. A drawback for peptide drugs is a lack of stability for oral delivery. Two classes of disulfide-rich peptides, conotoxins and cyclotides, have been shown to have higher stability than linear peptides thanks to their disulfide connectivity. Conotoxins are present in the venom of cone snails, a carnivorous marine mollusk that preys upon fish, worms, or other mollusks. Conotoxins are promising drugs leads with great prospects in the treatment of diseases and disorders such as chronic pain, multiple sclerosis and Parkinson’s and Alzheimer’s diseases. Cyclotides, which are cyclic cysteine knot containing peptides, isolated from the Violaceae (violet), Rubiaceae (coffee), and Cucurbitaceae (cucurbit) families and they have a wide range of biological activities, such as anti-HIV, uterotonic, and antimicrobial. P-superfamily framework IX conotoxins (C-C- C-CXC- C) contain the same cysteine framework, homologous sequences, and similar 3D structures to cyclotides. The knot containing conotoxins have been identified in several Conus species, but this work focuses on those from Conus brunneus, Conus purpurascens, and Conus gloriamaris. The cysteine knot motif of cyclotides and P-superfamily conotoxins is characterized by a cyclic backbone and six-conserved cysteine residues that form the three-disulfide bridges of the “knot”. This motif provides cyclotides and conotoxins with superior stability against thermal, chemical, and enzymatic degradation; marking them as potential frameworks for peptide drug delivery. Presented are details on the isolation of conotoxins and cyclotides, from Viola tricolor, and the characterization of their activity in the well-characterized Drosophila melanogaster giant fiber system (GFS) neuronal circuit, which contains GAP, acetylcholine, and glutamate synapses. The transcriptomes of two Conus brunneus specimens were assembled and mined for P-superfamily framework IX conotoxins. Eleven mature P-superfamily framework IX conotoxins were identified in the crude venom. / Includes bibliography. / Dissertation (Ph.D.)--Florida Atlantic University, 2018. / FAU Electronic Theses and Dissertations Collection

An investigation has been conducted into factors affecting the interparticulate cohesion profile of three micronised drugs, as a function of humidity. An atomic force microscope (AFM) colloid probe technique was correlated with physico-chemical properties and in-vitro performance. Briefly, micronised drug particles of salbutamol sulphate, triamcinolone acetonide (TAA) and disodium cromoglycate (DSCG) were mounted onto Vshaped tipless cantilevers using a developed micromanipulation technique. Interactions between the AFM ‘drug probes’ and a series of model drug surfaces were conducted at a 15,30, 45, 60 and 75% relative humidity using a custom built perfusion apparatus connected to the AFM. As expected, separation energy distributions for drug probe interactions were dependent on the surface rugosity of the drug model surfaces. Separation energy measurements conducted between drug probes and individual micronised drug particles (mounted in polymer resin) suggested large variations in separation energy. Further analysis of such data suggested a lognormal separation energy distribution, however, limitations in individual particle measurements (finite particle measurements per experiment) allowed restricted statistical analysis.

Over the past decade, the illicit drug market has experienced an explosion of designer drugs being produced by clandestine laboratories that include modifications of illicit drugs that dominated drug markets for long periods of time (e.g. cathinones and MDMA). These designer drugs, which are commonly known as “legal highs”, are popular due to that fact that they are legally obtainable and not currently controlled. Examples of these include the phenylalkylamine analogues 5-iodo-2-aminoindane (5-IAI) and 5, 6-methylenedixoy-2-amonindane (MDAI), which have similar biological effects to MDMA. In spite of the unknown risk factors associated with these substances, it is believed that “legal highs” continue to have high levels of interest among recreational users. As such, the potential for abuse is high, and 5-IAI and MDAI are under consideration in numerous jurisdictions for regulation. Many of these novel compounds have never been analyzed previously within a forensic setting. The chemical and physical properties of 5-IAI and MDAI are not fully understood. As a result, the analytical analysis of “legal highs” can be challenging. Color test kits provide a quick screening method for law enforcement officials looking to presumptively identify a substance in the field. The difficulty with this form of analysis is that most of the active ingredients present in “legal highs” are not detected by standard presumptive tests, or the results when the tests are used are ambiguous. Gas chromatography-mass spectrometry (GC-MS) is one of the most utilized analytical instruments in forensic laboratories for the identification of drugs of abuse. However, due to the rapid development and commercialization of “legal highs,” the limited availability of certified reference standards and mass spectral data make the confirmatory analysis of “legal highs” challenging. The primary aims of this research were two-fold. The first was to evaluate selected commercially available Narcotics Analysis Reagent Kits (NARK® II) and color reagent formulations recommended by the National Institute of Justice (Color Tests Reagents/ Kits for Preliminary Identification of Drugs of Abuse) to determine if the phenylalkylamine analogues 5-IAI and MDAI generate a color development. If a color was generated using a particular reagent, further testing was conducted to establish if the observed color would be detectable in the presence of various adulterants. The second aim of this research was to develop a rapid GC-MS method for the detection of 5-IAI and MDAI in contrived multi-component mixtures of selected adulterants. Standard color tests provided consistent results for 5-IAI and MDAI pure samples as well as mixtures with adulterants. 5-IAI produced a light brown color with both the Marquis and the methylenedioxypyrovalerone (MDPV) color reagent tests. The Mandelin reagent from the NARK® II test kit produced a greenish brown color and a light green color with the In-House preparations of the same reagent when tested with MDAI. Confirmatory analysis was performed using GC-MS with a temperature gradient. The analysis was performed on a non- polar (5% phenyl) methylpolysiloxane column with a total run time of 10 minutes. 5-IAI and MDAI were chromatographically separated and distinguishable from various adulterants based on retention time and mass to charge ratio.

1 - Total Syntheses of Helicterin B, Helisorin, and Helisterculin A - The total synthesis of three members of the helicterin family of natural products, helicterin B, helisorin and helisterculin A, was completed from a common, non-biomimetic intermediate. This work featured the development of a complex retro Diels-Alder/Diels-Alder sequence, a Friedel-Crafts ring closure resulting in a highly strained ring-system, an acid-catalyzed hydroxyketone rearrangement, a Lewis acid-promoted dimerization of a bicyclooctene monomer, and the use of a novel phenol protecting group. 2 - Synthetic Studies of an Alternative Diels-Alder Approach Towards the Helicterin and the Yunnaneic Acid Families of Natural Products - Efforts towards the development of an enantioselective synthesis of the helicterin natural products are presented. Given the failure of known methods to achieve this goal, a novel Diels-Alder approach based on a revised biosynthetic proposal was explored. This method achieved a bicyclooctene framework with good stereoselectivity, but its conversion to the natural product core proved challenging. Additionally, a Diels-Alder approach to the core of the yunnaneic acids was developed, one in which the effective reversal of the regiochemical preference of the reaction was achieved. 3 - Small Molecule Inhibitors of Cell Death in a Huntington's Disease Model - A compound based on the core of the helicterin natural products with the ability to prevent cell death in a Huntington's Disease model system was identified. Although its mechanism of action is unknown, its cellular target appears to be distinct from that of prior inhibitors. A number of analogs based on this initial hit were synthesized, and structure-activity relationships were explored. 4 - Synthetic Studies Towards a Biomimetic Approach to the Myrmicarin Alkaloids - The shortest and highest yielding synthesis of the monomeric myrmicarin alkaloids to date has been completed, and the stereochemical complexity of the upper half of the dimeric myrmicarin 430A has been achieved for the first time. Although the dienamine building blocks readily afforded the monomeric natural products under presumed biological conditions, the results of synthetic and theoretical experiments suggest that their dimers are either non-biomimetic or that enzymes are required in the biosynthesis of the higher-order myrmicarins. 5 - Studies on The Mechanism of the Knorr Pyrrole Cyclization - A mechanistic study of the Knorr pyrrole cyclization of an indolizidine-based dienamine to myrmicarin 215B was completed. This cyclization was found to occur exclusively in polar protic solvents, and two solvent molecules were required in the rate-determining step. A revised mechanism for this cyclization is proposed, in which a slow ketone protonation step precedes the cyclization and dehydration steps.

Biological processes often depend on protein-ligand binding events, so that accurate prediction of protein-ligand binding affinities is of central importance in structural based drug design. Although many techniques exist for calculating protein-ligand binding affinities, ranging from techniques that should be accurate in principle, such as free energy perturbation (FEP) theory, to relatively simple approximations based on empirically derived scoring functions, the counterbalancing demands of speed and accuracy have left us with no completely satisfactory solution thus far. This thesis will be focused on the methodology development towards more robust and reliable Protein-Ligand binding affinity calculation. In Part I, we will present the WaterMap method, which will bridge the gap between the efficiency of empirical scoring functions and the accuracy of rigorous FEP methods. Unlike most other methods with the main focus on the direct interaction between the protein and the ligand, the WaterMap method we developed considers the explicit driving force from the solvent, in which several individual water molecules in the binding pocket play an active role in the binding process. We demonstrate that protein may adopt active site geometries that will destabilize the water molecules in the binding pocket through hydrophobic enclosure and/or correlated hydrogen bonds, and displacement of these water molecules by ligand groups complementary to protein surface will provide the driving force for ligand binding. In some extreme cases, the interactions are so unfavorable for water molecules that a void is formed in the binding pocket of protein. Our method also considers the contribution from occupation of ligand atoms in the dry regions of binding pocket, which in some cases provides the driving force for ligand binding. FEP provides an in-principle rigorous method to calculate protein-ligand binding affinities within the limitations of the potential energy model and it may have a potentially large impact on structure based drug design projects especially during late stage lead optimization when productive decisions about compound modification are made . However, converging explicit solvent simulations to the desired precision is far from trivial, especially when there are large structural reorganizations in the protein or in the ligand upon the formation of the binding complex or upon the alchemical transformation from one ligand to another. In these cases, there can be large energy barriers separating the different conformations and the ligand or the protein may remain kinetically trapped in the starting configuration for a very long time during brute-force FEP/MD simulations. The incomplete sampling of the configuration space results in the computed binding free energies being dependent on the starting protein or ligand configurations, thus giving rise to the well known quasi-nonergodicity problem in FEP. In Part II, we will present a new protocol called FEP/REST, which combines the recently developed enhanced sampling technique REST (Replica Exchange with Solute Tempering) into normal FEP to solve the sampling problem in brute force FEP calculation. The computational cost of this method is comparable with normal FEP, and it can be very easily generalized to more complicated systems of pharmaceutical interest. We apply this method to two modifications of protein-ligand complexes which lead to significant conformational changes, the first in the protein and the second in the ligand. The new approach is shown to facilitate sampling in these challenging cases where high free energy barriers separate the initial and final conformations, and leads to superior convergence of the free energy as demonstrated both by consistency of the results (independence from the starting conformation) and agreement with experimental binding affinity data. Part III focus on two topics towards the foundational understanding of hydrophobic interactions and electrostatic interactions. To be specific, the nonadditivity effect of hydrophobic interactions in model enclosures is studied in Chapter 9, and the competition between hydrophobic interaction and electrostatic interaction between a hydrophobe and model enclosure is studied in Chapter 10. The approximations in popular implicit solvent models, like the surface area model in hydrophobic interaction, and the quadratic dependence of electrostatic interaction on the magnitude of charge are investigated. Six of the Chapters (Chapter 2-4, Chapter 6, and Chapter 9-10) have been published and the other one (Chapter 7) has been accepted for publication and currently is in press. Each Part begins with its own introduction. Each chapter also contains its own abstract and introduction, and focus on one specific topic. They all share the common theme, that is to develop more robust and reliable methods to calculate protein-ligand binding affinities. The conclusions and discussions about future research directions are presented in Part IV.

No description available.

This research report will engage in the debate surrounding decriminalization of drug use and whether it is a possible solution to the problem of drug use in South Africa. This is a question becoming more prevalent in global discussions regarding drug policy and its efficacy. It is held in the report that when evaluating policy two aspects must be addressed; namely the philosophical justification for the policy and the efficacy of the policy. Regarding criminalization it is found that policy may be justified by the public harm principle but that it does not effectively achieve the purpose of preventing and decreasing drug use and associated burdens. Thus, it is argued that prohibition is a constitutional limitation, but does not necessarily achieve its purpose in the least repressive or most effective way. Finally it is suggested that the solution to the drug problem will be one which addresses the background, particularly socio-economic, to drug use. Decriminalization has the theoretical potential to address this context but further empirical research is required in order to establish evidential grounds for continued discussion.

Thesis (MSc(Med)(Pharmacy))--University of Limpopo, 2012. / ABSTRACT South Africa, 80% of the population is dependent on the vernment to provide for their health care needs, mainly ugh primary health care facilities. In the health objectives of the National Drug Policy, the government of South Africa outlines its commitment to ensuring availability and accessibility of medicines which are effective, affordable, safe and of good quality in all sectors of the health care system ( N a t ion a IDe par t men t of He a It h, 1 996) . In o rd e r to assess the availability of d ru g s and identify ch a II en g e s w hi c h . ex is tin the Emf u Ie n i sub - d is t r i c t wi t hi nth e Sedibeng district, a questionnaire was administered to 21 primary health care facility managers/store managers, fo u r Community Health Centre managers and five transport officers in the district. In addition, a document review process was conducted to verify aspects of th e facility managers' and store managers' responses. Bin cards and primary health care order files were also examined in conjunction with a checklist to establish whether stock control systems were in place. There was a 100% response with all primary health care centres and community health care centres completing th e questionnaires. It was established that drugs at primary and community health care clinics were procured from the Sedibeng district pharmacy. In each of these clin ics there were specific individuals responsible for medicine supply management. Only four primary health care clinics had full-time pharmacist assistants employed, and 14 clinics were visited by the assistants on a weekly/bi-weekly basis. There were no employees that have received training in drug supply management in the last 12 months in 88% of the clinics interviewed. Nineteen clinics claimed that the storage area was not large e n 0 ugh to s tor e a II the s toe k f or a m 0 nth's sup ply and 0 n I yon e clinic had a secure delivery area for their medication. It was established that 24 facilities received stock by two specific procedures namely; that the number of boxes were checked and the driver's note was then signed, and stock received was checked against the invoice. Of the interviewed cl i nics, 20% admitted that the re-order level had not been calculated for all tracer items in the store. Standard Operating Procedures, Standard Treatment Guidelines and the Essential Drugs List were also not available at all facilities. The results indicate inadequacies and weaknesses in procurement, quantification, stock control, storage and record keeping. It clearly demonstrates that inadequately-trained staff was a ma j 0 reo n t rib uti n g fa c tor to d rug s h 0 r tag e s. The r e was a I a c k 0 f monitoring and evaluation by th e district pharmacy as pharmacists did not manage to visit all the clinics each month. Most of the inadequacies and weaknesses can be addressed at facility level with pro per supervision, in-service training, mentoring and support of staff and the reinforcement of drug supply management training. Regular supervisory visits together with updating the monitoring too I in terms of th e problems identified will improve th e management of drugs and ultimately decrease the number of out of stocks where problems have been identified at primary health care level.

No description available.

Amendements: leaves 252-254. Copies of author's previously published articles inserted. Bibliography: leaves 226-251. The CYP2C19 and CYP2D6 genetic polymorphisms control the oxidative metabolism of many different drug classes. Populations are separated into groups of extensive metabolisers (EM), poor metabolisers (PM), and in the case of CYP2D6, ultra-rapid metabolisers (UM). In vitro studies using human liver microsomes were conducted to examine the kinetics of the oxidative metabolism of flunitrazepam, and which CYP450 enzymes mediate the oxidative metabolism of flunitrazepam, (S)-mephenytoin and proguanil.