The marine surface-associated bacterium Pseudoaltermonas tunicata, produces a range of compounds that inhibit fouling organisms, including invertebrate larvae, bacteria, algal spores and fungi. In addition to these antifouling compounds P. tunicata cells produce both a yellow and a purple pigment. The aim of this study was to further characterise the antifouling activities, their regulation and relationship with pigmentation, and the ecological significance of P. tunicata and related organisms. It was discovered that the anti-algal compound was extracellular, heat sensitive, polar and between 3 and 10 kDa in size. The anti-fungal compound was found to be the yellow pigment and active against a wide range of fungal and yeast isolates. Chemical analysis suggests that this compound consists of a carbon ring bound to a fatty-acid side chain. Genetic analysis supports the chemical data for the active compound as a mutant in a gene encoding for a long-chain fatty-acid CoA ligase was deficient for anti-fungal activity. To address the regulation of antifouling compounds and their relationship to pigmentation transposon mutagenesis of P. tunicata was performed. Mutants lacking the yellow pigment displayed a reduced ability to inhibit fouling organisms. Further analysis of these mutants identified genes involved with the synthesis and regulation of synthesis of pigment and antifouling compounds. One of these mutants was disrupted in a gene (wmpR) with similarity to the transcriptional regulators ToxR from Vibrio cholerae and CadC from Escherichia coli. Analysis of global protein expression using two-dimensional gel electrophoresis showed that WmpR is essential for the expression of at least fifteen proteins important for the synthesis of fouling inhibitors. The ecological significance of antifouling bacteria was addressed by assessing the antifouling capabilities of a collection of bacteria isolated from different marine surfaces. Overall, isolates from living surfaces displayed more antifouling traits then strains isolated from non-living surfaces. Five dark-pigmented strains originating from the alga Ulva lactuca were further studied. Phylogenetic and phenotypic analysis revealed that they were all members of the genus Pseudoalteromonas and were closely related to P. tunicata. Two strains represented a novel species within the genus and were taxonomically defined as P. ulvae sp. nov.

Secondary metabolites from natural sources are fast growing as popular drug leads. The structural novelty and favourable biological activity that these compounds display contribute to their popularity as drugs of the future. Examples of such compounds include the potent anticancer drug paclitaxel isolated from the bark of a yew tree as well as the more commonly known analgesic aspirin which stems from the bark of the willow tree. The biological activities exhibited by these secondary metabolites are vast and range from antimicrobial to anticancer activity to mention but a few. As a result, the isolation of novel compounds from natural sources is on the rise. The South African seaboard is home to a wealth of various marine algal species which produce fascinating secondary metabolites. For example, Portierria hornemanii was shown to produce halomon, a halogenated monoterpene which has displayed promising cytotoxic activity. This study thus focused primarily on pursuing novel compounds from three endemic South African marine algal species which have never been analysed previously from a chemical perspective. These are Plocamium rigidum (Bory de Saint-Vincent), Laurencia natalensis (Kylin) and Delisea flaccida (Suhr) Papenfuss. Four known compounds and one new halogenated monoterpene, (2E,5E,7Z)-8-chloro- 7-(dichloromethyl)-4-hydroxy-3-methylocta-2,5,7-trienal, were isolated from Plocamium rigidum. The breast cancer (MCF-7 cell line) inhibitory activity for these compounds was assessed and it was observed that an increase in the lipophilic nature of the compounds produced more favourable IC50 values. A pre-cursor to bromofucin type compounds, cis-laurencenyne, was isolated from Laurencia natalensis, as well as a new acetoxy chamigrane type compound, 4-bromo- 3,10-dichloro-7-hydroxy-3,7,11,11-tetramethylspiro [6.6] undec-1-yl acetate. Delisea flaccida was seen to contain two known bromofuranone type compounds isolated as an isomeric mixture, 1-[(5Z)-4-bromo-5-(bromomethylidene)-2-oxo-2,5- dihydrofuran-3-yl] butyl acetate and 1-[(5E)-4-bromo-5-(bromomethylidene)-2- oxo-2,5-dihydrofuran-3-yl]butyl acetate. These compounds are famous for their ability to inhibit bacterial biofilm production and they have been isolated before from an Australian Delisea spp / Adobe Acrobat 9.53 Paper Capture Plug-in

Background: Pancreatic disease is a global problem. Severe acute pancreatitis (AP) carries a 30-50% mortality. Current scoring systems fall short in predictive accuracy, sensitivity, specificity and availability. Pancreatic cancer (PC) is a leading cause of cancer-related mortality, most patients die within one year of diagnosis. Late presentation and lack of effective oncological treatment determine a desperate need to focus on early detection of the pancreatic cancer. Current biomarkers fall short in accessibility, sensitivity and specificity and ability to distinguish malignant from benign conditions. Metabolomics aims to decipher molecular signatures that will distinguish disease from controls, ultimately leading to novel targets for diagnosis and treatment. Initial studies are discovery-based, hypothesis-generating and typically aim to establish a snapshot of the metabolism of an individual by metabolite profile. Aims: Establish a prospective phenotypic and demographic database of patients with acute pancreatitis. Determine urinary and serum metabolomic profiles of AP and PC in comparison to controls and establish if metabolomic profiling can distinguish severity of each disease in order to identify potential novel bio-markers. Methods: Urine and serum samples from 73 AP, 32 PC, 62 Healthy Controls, 8 Chronic pancreatitis and 8 Benign jaundice participants were analysed using GC-MS and UPLCMS. Metabolite identification was subject to univariate and multivariate analysis (p<0.05). Results: The differentiation of metabolite profiles was most distinct with AP. There was no differentiation by AP aetiology. AP severity was distinquished by metabolite profile. Profiles of resectable patients were distinct form non-resectable PC. Fatty acids(FA), glycerophoshocholines, eicosanoids, TCA cycle intermediates and melatonin levels were altered in AP. PC was defined by altered concentrations of FAs, eicosanoids, glycerophoshocholines, sphingomyelins, folates and amino acids and peptides (e.g. glutamine). Altered levels of UFAs, neuromedins, Vitamin D3 determined stage of PC. Conclusion: Urinary and serum metabolomic signatures may provide future biomarker panels for grading AP and PC.

No description available.

No description available.

published_or_final_version / Anatomy / Doctoral / Doctor of Philosophy

Fungi have been an important source for producing a wide range of secondary metabolites of widely differing chemical structures, as well as biological activities. Many of their metabolites now play a major role in pharmaceutical and agricultural industries. A number of fungi were isolated from soil and leaf litter collected from Arthur’s Pass, West Coast and Kaituna Valley using a particle filtration technique. Fungi were selected based on their unusual morphology or observed cytotoxicity and antimicrobial activity for large scale culture and extraction. A pale yellow compound was isolated from cytotoxic extracts from the culture of Aspergillus versicolor. This compound was identified as sterigmatocystin and the identity confirmed by UV profile and mass spectrometry. Five compounds were isolated from extracts prepared from two different species of Penicillium of which three were active against P388 cells (mycophenolic acid, cycloaspeptide A and mevastatin), one was active against dermatophytes (griseofulvin) and one was not active (3,4,6,8-tetrahydroxy-3-methyl-3,4-dihydroisocoumarin). Two compounds were isolated from extracts prepared from two different species of Phoma. A dark red compound was found to be novel and showed activity against P388 cells and Bacillus subtilis. A second compound also showing cytotoxicity was identified as the known compound phomenone. A further new compound was isolated from extracts of an identified dematiaceous fungus. This alkyl glucoside, however, was not bioactive.

3,4-(±)-Methylenedioxymethamphetamine (MDMA) is a synthetic amphetamine derivative commonly used as a recreational drug. Although the selectivity of MDMA for the serotonergic system in rat and humans is well established, the specific mechanism associated with MDMA-induced neurotoxicity is not fully understood. The long-term neurotoxicity of MDMA appears to be dependent upon systemic metabolism since direct administration of MDMA into the brain fails to reproduce the neurotoxic effects seen following peripheral administration, indicating that the parent compound alone is unlikely to be responsible for the neurotoxicity. MDMA is O-demethylenated to the catechol metabolite N-methyl-α-methyldopamine (N-Me-α-MeDA) and N-demethylated to MDA by cytochrome (s) P450 (CYP450). Thioether (glutathione and N-acetylcysteine) metabolites of N-Me-α-MeDA and α-MeDA are neurotoxic and can be found in rat brain following s.c. injection of MDMA. Because multidose administration of MDMA is typical of drug intake during rave parties, we investigated the effects of multiple doses of MDMA on the concentration of neurotoxic thioether metabolites in rat brain. Administration of MDMA at 12-h intervals for a total of four injections led to a significant accumulation of the N-Me-α-MeDA thioether metabolites in striatal dialysate. In contrast, acute release of 5-HT concentrations was decreased. Since isoenzymes of the CYP2D subfamily (30% metabolism), and the CYP2B or CYP3A1 isoforms, catalyze the low and high KM O-demethylenation reactions, respectively, we subsequently examined the potential role of CYP2D1 in both a genetic and pharmacological model. The data is consistent with the hypothesis that systemic metabolism of MDMA contributes to MDMA-induced serotonergic neurotoxicity via the 20) generation of reactive metabolites. In both the genetic and pharmacological models of CYP2D1 deficiency, attenuation of MDMA-mediated decreases in brain 5-HT concentrations were in the same range (30-40%). Finally, we examined the contribution of various transporters using genetic and pharmacological models to investigate the mechanisms regulating the concentration of thioether metabolites in MDMA neurotoxicity. The data suggest that by regulating various transporters and brain concentrations of the neurotoxic thioether metabolites of MDMA, may subsequently modulate the degree of neurotoxicity. However, further studies are necessary to understand the precise mechanism by which Mrp’s and Oat1 transporters modulate MDMA-neurotoxicity. Taken together, these studies are consistent with the view that neurotoxicity of MDMA requires systemic metabolism to form α-MeDA and N-Me-α- MeDA by CYP2D6. Therefore, It is likely that neurotoxicity is mediated by the formation of systemic neurotoxic metabolites.

No description available.

No description available.