Dissertation submitted in partial compliance with the requirements for the Master's Degree in Technology: Homoeopathy, Technikon Natal, 1999. / The purpose of this study was to investigate the Nuclear Magnetic Resonance spectra of samples of LM6; LM14 and LM22 homoeopathic quinquagenimillesimal (LM) potencies of tin (Stannum metallicum), lead (Plumbum metallicum) and two lactose based controls: one control which is prepared by the means of potentisation and one control prepared without the means of potentisation. It was hypothesised that in terms of the effect of different substances (Stannum metallicum and Plumbum metallicum, and the lactose controls) and of different dilutions (LM6, LM14, LM22) of these substances that significant differences exist between the chemical shift and relative integration values of the CH3, CH2, OH and H20 signals of these homoeopathic substances. / M

There has been a lot of interest on solid lipid nanoparticles (SLNs) as these colloidal submicron drug dosage forms present a promising frontier in drug delivery. It is possible to incorporate susceptible drugs such as protein intended for oral delivery. Here, we aim to develop an oral delivery system based on SLNs to deliver the peptide hormone, insulin using the double emulsion (W/O/W) solvent evaporation technique for formulating the SLNs. The choice of lipids was carefully selected to incorporate acceptability to biological milieu. The main purpose of the work was to formulate SLNs to achieve different localisation of insulin within the SLNs, based on the three hypothetical models proposed by Muller et al. (2000). Following that, the effect of this localisation on the propensity of the SLNs to be taken up by absorptive cells was investigated. SLNs was successfully fabricated to achieve two insulin localisation models, namely the solid solution model and the core-shell model with drug-enriched shell. The zeta potential measurements was used to indirectly indicate the appropriate insulin localisation model. The zeta potential of the unloaded SLNs, insulin-loaded SLNs and surface-adsorbed insulin SLNs were recorded as -51.7 ± 1 mV, -45.8 ± 1 mV and -40.8 ± 1 mV respectively. In vitro cell studies showed a notable difference in the Caco-2 cell lines when the cells were exposed to SLNs of the two different insulin localisation models. Thus, different effects seen on the Caco-2 cells suggests that the localisation of insulin within SLNs can potentially influence its uptake, stressing the importance of characterising drug localisation in nanoparticles, as this eventually affects drug bioavailability.

Successful delivery of pharmaceuticals orally requires a firm understanding of how dosage forms behave during their passage through the gastrointestinal (GI) tract. In this study, the GI transit time and absorption of amphotericin B (AmB) solid lipid nanoparticles (SLN) were investigated in rats, using paracetamol (PAR) and sulphapyridine (SP) as indirect markers. A high encapsulation efficiency of 91.2% was obtained for the AmB SLNs. The SLNs were exhaustively characterised with regards to size, zeta potential (ZP), viscosity, density, migration propensity within agarose gel, in vitro drug release and morphology, to ensure similar disposition in the GI tract after simultaneous oral administration. Freeze-drying did not significantly alter the size or ZP of the AmB SLNs, and in vitro drug release from fresh and freeze-dried SLNs were identical. AmB, PAR and sulphasalazine (SSZ) (the latter being the prodrug of SP) were individually formulated into SLNs using beeswax and theobroma oil as the lipid matrix. The z-averages, polydispersity indices and ZPs of the SLNs ranged from 206.5-224.8 nm, 0.161-0.218 and |61.90|-|71.90| mV, respectively. Gel electrophoresis studies indicated a similar movement propensity among the three SLNs as their migration distances were identical (22.2-22.4 mm) within agarose gel. Scanning electron and atomic force microscopy studies revealed that all three SLNs were spherical in morphology and with similar surface characteristics. The SLNs were assessed for changes in size and surface charge on exposure to simulated GI fluids using dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA). On contact with the fluids, the particles had a slight increase in size due to ingress of the dissolution media. NTA results were found to be more beneficial than DLS as the latter was biased towards larger particles that were present possibly due to aggregation. After incubation in simulated gastric fluid followed by simulated intestinal fluid (mimicking gastric emptying), all the SLNs were found to be less than 350 nm in size and neutral in charge, which are optimal attributes for intestinal absorption. Time-of-flight secondary ion mass spectroscopic (ToF-SIMS) analyses revealed minimal drug amounts on the surfaces of the particles indicating that drug location was in the core of the SLNs. A developed and validated high-performance liquid chromatography (HPLC) method for simultaneous assay of the drugs in rat plasma using piroxicam as internal standard was found to be sensitive, accurate and precise, with drug recovery from plasma exceeding 92% in each case. A pilot GI transit study conducted in rats showed that the HPLC method was appropriate for the study. In the main study, the effects of food on the transit and absorption of the AmB SLNs were investigated. The presence of food slowed the transit of the SLNs in the GI tract. The gastric transit time of the AmB SLNs was estimated indirectly using PAR and was obtained as 1.71-2.25 hr. Caecal arrival time (CAT) of the AmB SLNs was estimated using SP detection in plasma as SSZ metabolism to produce SP occurs predominantly by the activity of colonic flora. In both fasted and fed states, CAT was 1.80-1.90 hr whereas transit time through the small intestine was 1.65-1.79 hr. A delayed rate of AmB absorption was observed in the fed state however, the extent of absorption was not affected by food. The percentage AmB absorption during the fasted state in the stomach, small intestine and colon were not significantly different from absorption within the respective regions in the fed state. In both states however, absorption was highest in the colon and appeared to be a summation of small intestinal absorption plus absorption proper within the colon. The study indicated that, AmB SLNs irrespective of food status were slowly but predominantly taken up via the lymphatic route and the small intestine was the most favourable site for their absorption. The data obtained indicate that it is possible to enhance the bioavailability of AmB through its incorporation into SLNs. Further enhancement of AmB bioavailability can be achieved through appropriate formulation interventions aimed at slowing transit of the SLNs in the small intestine. Finally, being a lipid-based system, the SLNs may have a potential to reduce the nephrotoxic effects of AmB.

Solid dispersions are commonly used to overcome bioavailability issues of poorly water soluble drugs. Various preparation methods along with carrier systems have been used to develop solid dispersions. However, this study investigates the application of microwave heating methods in formulation development alongside associated analytical investigations. Formulations of poorly soluble drugs, namely, fenofibrate, gemfibrozil, ibuprofen, ibuprofen (+) S and phenylbutazone were prepared using a microwave technique and compared with standard formulation techniques. Mesoporous silicas and polyethylene glycol were used as excipients. Then in vitro dissolution analysis was carried out for the performance evaluation of the resultant formulations. It was found that effective products were produced as a result of microwave processing compared with the traditional techniques. Analytical techniques such as differential scanning calorimetry (DSC), X-ray diffraction (XRD), scanning electron microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR) were employed to determine the solid state properties, i.e. thermal stability, crystalline state, physical appearance and chemical stability of developed formulations. The overall findings indicate that successful formulation can be achieved using microwave heating. Isothermal titration calorimetry (ITC) was used to probe the interactions of model drugs, namely, caffeine, diprophylline, etofylline, paracetamol and theophylline with excipients such as sodium dodecyl sulphate (SDS), sodium deoxycholate (NaDC) and PEG. Thermodynamic data suggests the successful use of ITC to investigate drug-excipient interactions. In summary, the potential of microwave heating in formulation development and ITC to characterise drug-excipient interactions was thoroughly investigated and both found as potential alternatives to more traditional techniques.

No description available.

The antimycobacterial activity of twenty South African medicinal plants were investigated using two methods commonly used; the conventional agar plate method and the BACTEC radiometric method. Fourteen of the twenty acetone extracts of medicinal plants used to treat pulmonary diseases showed inhibitory activity at a concentration of 0.5 mg/ml against a sensitive strain of Mycobacterium tuberculosis using the conventional agar plate method. These fourteen extracts were also tested against M tuberculosis by the BACTEC radiometric method against a sensitive as well as a strain resistant to the drugs isoniazid and rifampin. Eight plants showed activity against both the strains at a concentration of 1.0 mg/ml. Susceptibility testing of M tuberculosis by the agar plate method is reliable, economical, and reproducible whereas the BACTEC radiometric method is much faster and probably more accurate than the agar plate method. A cytotoxicity assay of the fourteen plants on primary vervet monkey kidney cells showed that the crude acetone extracts of E. natalensis was the least cytotoxic extract with significant antimycobacterial properties. It was therefore, chosen for the isolation of active compound(s). An antibacterial assay of the water and acetone extracts of the roots of E. natalensis showed that they inhibited the growth of Gram-positive bacteria at concentrations ranging between 0.1 and 6.0 mg/ml. The water extract did not exert any inhibitory action on Gram-negative bacteria while the acetone extract showed inhibitory activity at a concentration of 5.0 mg/ml. The MIC of diospyrin, isolated from E. natalensis, was found to be 100 µg/ml for a drug-sensitive and a number of drug-resistant strains of M. tuberculosis and Gram-positive bacterial species. An antiviral investigation of the crude extracts of E. natalensis showed that the water extract of the roots of the plant inhibited the replication of herpes simplex virus type 1 moderately at a concentration of 0.2 mg/ml whereas, acetone extract at concentrations ranging from 0.1 to 0.02 mg/ml. Diospyrin exhibited no inhibitory effect against the virus. The MIC of 7-methyljuglone, isolated from E. natalensis, was found to be 50 µg/ml for both drug-sensitive and drug-resistant strains of M. tuberculosis. The compound inhibited the growth of Gram-positive bacterial species at concentrations ranging from 50 to 100 µg/ml. No inhibitory effect of the compound was observed on any Gram-negative bacteria at the highest concentration tested. A significant synergistic effect of the two naphthoquinones was observed against M tuberculosis and some of the bacterial species. MICs obtained were 10 µg/ml and 50 µg/ml for M tuberculosis and the bacterial species respectively. No synergistic effect was observed on any Gram-negative bacterial species investigated. In view of the encouraging results obtained from this study on the biological activity of the two naphthoquinones; diospyrin and 7 -methyljuglone, it appears that the compounds deserve further investigation in order to explore its potential as antimycobacterial agents. / Thesis (DPhil (Plant Physiology))--University of Pretoria, 2007. / Plant Science / unrestricted

Recent progress in synthetic chemistry has enabled the preparation of new highly-defined polymers that exhibit changes in their structure in response to environmental changes. These responsive nanomaterials may be desirable as carriers of drugs to deliver at the cellular and sub-cellular level. However, the endocytic pathways used by these nanoparticles to access cells must be defined. Carboxylated polystyrene beads (C-PB) of 50 and 100 nm size were chosen as ‘model’ nanomedicines and their route of uptake into cells characterised and compared to thermoresponsive PLGA-b-(PEGMA-co-PPGMA) and PLA-b-(DEGMA-co-OEGMA) block copolymers of 50-150 nm (‘candidate’ drug delivery systems) uptake. A number of protocols were optimised for endocytosis inhibition studies. Results reported that the inhibition of clathrin mediated endocytosis (CME) with chlorpromazine (CPZ) was cell- and time-dependent. After the maximal effect of the inhibitor, the endocytosis of human transferrin (Htf), a marker of CME, recovered up to uninhibited levels in 3T3 and HCT116 cells. Furthermore, high passage number and ageing of cells showed a resistance towards the inhibition of the uptake of Htf with CPZ. Both PLGA-b-(PEGMA-co-PPGMA) and PLA-co-(DEGMA-co-OEGMA) thermoresponsive block copolymers presented colloidal instability and aggregation that impeded further endocytic pathway internalization experiments. However, the results reported in this thesis question some of the interpretation in the literature of the susceptibility of cells to CPZ in the internalization of nanomaterials. New experimental settings for CPZ inhibition studies should be considered and protocols optimised in order to avoid incorrect and potentially misleading outcomes.

A high number of new chemical entities emerging from the drug development process show pharmacological activity, but at the same time are characterised by poor dissolution and solubility profiles. As a result, there is a strong push to develop innovative formulations for the delivery of such compounds so that the desired oral bioavailability and pharmacological effects are achieved. An increasingly popular class of formulations to improve the dissolution properties of poorly soluble drugs is represented by amorphous solid dispersions, whereby the drug is molecularly dispersed in a carrier matrix. One of the key challenges for developing amorphous solid dispersions in real-world formulations is the understanding of the dissolution performance. Although this is very important, due to the fact that the dissolution performance limits the in vivo efficacy, the dissolution mechanisms by which the amorphous solid dispersions dissolve is still not fully understood. This thesis investigates the dissolution performance of three solid dispersions model systems including felodipine, bicalutamide and indomethacin, all poorly soluble drugs, with copovidone, a water-soluble polymer. The complexity of the model systems increases through the chapters, starting by testing the dissolution of a well-documented poorly soluble drug model, i.e. felodipine, as a function of the drug loading (5% and 50% w/w) (Chapter 3). In Chapters 4 and 5 the dissolution of bicalutamide, which is known to exist in at least two different polymorphic forms (form I and form II), is investigated as a function of three drug loadings (5%, 30% and 50% w/w). Finally, the dissolution of indomethacin, which presents a pH-variable solubility and dissolution rate due to its weakly acidic nature (pKa of 4.5), is probed as a function of both dissolution medium pH (pH 2 and 6.8) and drug loading (5%, 15%, 30%, 50%, 70% and 90% w/w) (Chapter 6). The dissolution of amorphous solid dispersions and other oral dosage forms is commonly tested using the USP dissolution apparatuses (types I, II and IV). These methods present a significant limitation, i.e. they can not provide any directly spatially-resolved chemical information on potential changes occurring to the solid form (e.g. amorphous to crystalline, polymorphs or solvation-related transformations). In this thesis Raman spectroscopy is employed as primary analytical technique in an attempt to fill the gaps related to the understanding of the dissolution mechanisms of amorphous solid dispersions and the limitations of the conventional USP apparatuses. The novelty of this approach derives from collecting Raman data directly from the dosage form in real time and in situ during the course of the dissolution test using a flow-through cell placed below the Raman microscope. Temporally- and spatially-resolved chemical Raman maps are generated using a novel mathematical approach which derives from the use of concatenated maps to explicitly probe the chemical and physical changes as a function of time as well as space. In-line ultraviolet spectroscopy is also integrated to the Raman system to directly relate changes in dissolution behaviour to physicochemical changes that occur to the solid form during the dissolution test. A wide range of other state-of-the-art analytical techniques is also used to complement the Raman data to obtain a clear picture of drug release from amorphous solid dispersions. This includes a combined magnetic resonance imaging/ultraviolet flow cell system to allow, similarly to the Raman/ultraviolet method, changes in dissolution profile to be related to physical changes occurring in the solid material, and for the first time quantitative suppressed-water proton nuclear magnetic resonance spectroscopy was applied to amorphous solid dispersions. Proton nuclear magnetic resonance, due to the high chemical selectivity, provides quantitative data on both the drug and the polymer. Finally, the rotating disk dissolution rate test, i.e. a modified version of the conventional intrinsic dissolution rate test, is developed and employed for the first time to gain information, similarly to proton nuclear magnetic resonance spectroscopy, on the dissolution rates of both the drug and the polymer. The dissolution performance of all amorphous solid dispersion model systems is shown to be strongly affected by the drug loading. At low drug loading the drug and the polymer dissolve with the same rate from the molecular dispersion, pointing to a drug release dependent on the high water solubility of copovidone. At high drug loading, the dissolution rates of both the drug and the polymer are significantly slower and this is shown to be ascribed to the formation of an amorphous drug-rich shell around the compact, followed by the drug re-crystallisation. For the high drug loaded amorphous solid dispersions, the dissolution performance is strongly dependent on the physicochemical properties of the drug, i.e. low aqueous solubility and high hydrophobicity. The dissolution behaviour of the amorphous indomethacin solid dispersions is also found to be affected by the dissolution medium pH. Indomethacin from the amorphous solid dispersions with 15% or higher drug loading is released only at pH 6.8 due to the significant increase in its aqueous solubility at this pH.

There are numerous chemotherapeutic agents available today that treat a wide array of tumours. The majority of these compounds are administered via intravenous (i.v.) infusion in large doses (1000 mg/m2), necessary to sustain a desired therapeutic effect. The systemic nature of the drug delivery, the dosage size and non-specific nature of many chemotherapeutic agents however means that they attack any rapidly dividing tissue system, leading to the commonly observed side effects e.g. alopecia, nausea, neutropenia and thrombocytopenia. Low molecular weight gelators (LMWGs) are increasing in popularity as an alternative platform for drug delivery. They are typically small amphiphilic organic molecules which self-assemble in water forming a 3D gel network; they offer advantages over other drug delivery platforms as they are typically derived from biological polymers and are therefore inherently biocompatible. Using the nucleoside gemcitabine; a first-line treatment for the treatment of gastric and pancreatic cancer as a model drug, two localised delivery systems were developed. The first, an inert LMWG matrix for the encapsulation and passive release of gemcitabine and the second, a therapeutic molecular gel derived from the chemotherapeutic itself. Cytidine, an inert analogue of gemcitabine was used to develop a passive delivery system. Regioselective synthesis of N-acylated derivatives of varying chain lengths was achieved via an activated triazine ester. Using a minimal amount of ethanol and an ‘anti-solvent’ switch gelation method a gelating system derived from the N-myristoyl derivative, containing a solvent volume fraction (ΦSOL) of 0.40 was found to have the most advantageous mechanical and structural properties; a crosslinked nanofibrillar network, established by rheological measurements and microscopy (TEM). The gel was validated as a drug delivery platform via encapsulation and release low molecular weight fluorescein and high molecular weight FITC Dextran, with the gelator matrix releasing the smaller fluorescein and retarding the release of the higher molecular weight dextran. Further modification and optimisation of the passive system afforded an N-octanoyl 2’-deoxycytidine conjugate that underwent molecular reorganisation into a cross-linked nanofibrillar structure in a 100 % aqueous environment. This gel was the first of its kind to assemble in this manner and the rheological measurements demonstrate its self-healing properties, whilst encapsulation of fluorescents once again demonstrated controlled release of low molecular weight fluorescein over a 24 h period. In vitro growth inhibition assays validated the platform as biologically compatible against gastric (MKN-7) and pancreatic (MIA PaCa-2) cell lines. Additional modification of the chemotherapeutic itself laid the foundation for an intra-tumoural targeted therapeutic delivery system. Enzyme cleavable pro-drugs of gemcitabine were created with amphiphilic properties, linkages designed to undergo varying rates of hydrolysis from both the N-amino and 5ʹ-hydroxyl positions. Whilst no successful gelating entities were achieved, the N-amide and 5’-ester prodrugs were found to have comparable potencies to the parent compound in vitro when tested against gastric and pancreatic cell lines. From the results obtained during this work, it can be concluded that with further chemical modification based upon parameters discussed here on in that a LMWG system could be a viable platform for drug delivery in the future.

With increased worries of our Nations’ reliance on fossil fuels and their deleterious effects on the environment, researchers are concentrating on developing sustainable alternative sources for energy and chemicals. One potential starting resource that is worldwide distributed and renewable is biomass. Cellulose, the most plentiful source of biomass on earth, can be hydrolyzed into biofuel precursors such as 5-hydroxymethylfurfural (HMF). However, due to the poor solubility of cellulose and its robust crystalline structure, current methods available to degrade cellulose into these biofuel precursors are costly, result in low yields along with a large amount of waste. Generally, fructose is the preferred feedstock for the synthesis of HMF with high efficiency and selectivity. However, the large-scale production of HMF from fructose is limited due to the scarcity and the high cost of fructose. Therefore, it is desirable to use a cheaper renewable starting material for the synthesis of HMF such as sucrose. This study is conducted to develop an efficient one-pot process to synthesize HMF from biomass, particularly sucrose, using various sulfonated heterogeneous catalysts such as ordered mesoporous silica, bridged periodic mesoporous organosilicas (PMO) and carbon materials. The HMF yields in the presence of such acidic nanoporous materials were comparable to those using much less environmentally-friendly metal-based catalysts.