Australian macrofungi as a source of antimicrobial compounds

Neeraj Bala (2011). Australian macrofungi as a source of antimicrobial compounds PhD Thesis, School of Pharmacy, The University of Queensland.

       
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Author Neeraj Bala
Thesis Title Australian macrofungi as a source of antimicrobial compounds
School, Centre or Institute School of Pharmacy
Institution The University of Queensland
Publication date 2011-10
Thesis type PhD Thesis
Supervisor Kathryn Steadman
Elizabeth Aitken
Benjamin Ross
Total pages 180
Total colour pages 4
Total black and white pages 176
Language eng
Subjects 11 Medical and Health Sciences
Abstract/Summary Plant-derived natural products have an ancient history of use in traditional as well as modern medicine. However, the therapeutic potential of natural products derived from fungi became evident only after the discovery of the first antibiotic, penicillin. Most of the fungi-derived pharmaceuticals have been sourced from microfungi such as Penicillium and Aspergillus. Recent interest in the discovery and development of new antimicrobials from natural and diverse sources has attracted wide-scale scientific attention towards macrofungi, owing to their well-acknowledged pharmacological potential in Traditional Chinese Medicine. Therefore, this study aimed to find new antimicrobial compounds from macrofungi. Limited research efforts have involved the evaluation of antimicrobial activities of Australian macrofungi. Therefore, the present study investigated 47 Australian macrofungi for their antibiotic potential. The fruiting bodies were collected from across Queensland, Australia. Collected specimens were freeze-dried and sequentially extracted using three solvents viz. water, ethanol and hexane. These extracts were initially tested against sensitive strains of one Gram+ve (Staphylococcus aureus) and one Gram-ve bacterium (Escherichia coli) in a high throughput 96-well microplate bioassay. Overall water and ethanol extracts were more effective against S. aureus than E. coli whereas a small number of hexane extracts showed better results for their antimicrobial potential against E. coli at higher concentrations only. From these preliminary data, the water and ethanol extracts of 12 macrofungi were identified, showing promising results against the two test pathogens. Therefore, these macrofungi were selected for further evaluation of their antimicrobial potential against a range of clinically important micro-organisms including two Gram+ve (Bacillus cereus, Listeria monocytogenes) and two Gram-ve (Pseudomonas aeruginosa, Acinetobacter baumannii) bacteria and two fungi (Geotrichum candidum, Saccharomyces cerevisiae) along with their cytotoxic potential against human cancer cell lines. A degree of specificity in antimicrobial activity was exhibited by the water extract of Ramaria sp. and the ethanol extracts of Psathyrella sp. and Hohenbuehelia sp. which inhibited the growth of two fungal pathogens used in the assay. Interestingly, the water extract of Hohenbuehelia sp. and the ethanol extract of Ramaria sp. showed wider activity against one Gram+ve and one Gram-ve bacterium and both fungi. Therefore, these two macrofungi were carried forward towards the identification of the active component(s) responsible for activity via bioassay-guided fractionation. The water and ethanol extracts of the two selected macrofungi were fractionated and purified by reverse phase high performance liquid chromatography (RP-HPLC) yielding 25 fractions in total, from all the four extracts. The next step to be accomplished was to again test these fractions for their bio-activity. Therefore, different fractions from water and ethanol extracts were tested against the pathogens of interest i.e. against which the crude extracts had exhibited activity. The water extract of Ramaria sp. did not reveal any activity against any of the tested pathogens whereas fractions from water and ethanol extracts of Hohenbuehelia sp. revealed antifungal activity against either one or both the fungi. On the other hand, encouraging results were observed with the ethanol extract of Ramaria sp. where four fractions exhibited antibacterial activity against Gram+ve bacterium, B. cereus in addition to antifungal activity against both fungi. The purity of these fractions was confirmed using analytical HPLC. Remarkably, the fraction (F5) from Hohenbuehelia water extract and two fractions (F3 and F4) from Ramaria ethanol extract appeared to be almost pure, comprising one major component in addition to a few minor components and hence, were carried forward for the identification of the active components using analytical techniques (LC-MS and NMR). The active components in F5 turned out to be a mixture of phenylalanine and dipeptides while saturated and unsaturated fatty acids were identified in the bioactive fractions from Ramaria ethanol extract. However, due to limiting amounts of the samples available, it was not feasible to isolate individual components. Hence, to confirm their bioactivity further extraction and purification would be needed on a greater scale to facilitate isolation and identification of all the components and then individually testing those components.
Keyword Macrofungi, crude extracts, 96-well microplate bioassay, Hohenbuehelia, Ramaria, bioassay-guided fractionation, antimicrobial activity, mushrooms, S. aureus, E. coli
Additional Notes Coloured pages: Title page, 34, 120, 140 Landscape pages: 33, 73, 92, 94

 
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Created: Wed, 01 Feb 2012, 09:51:48 EST by Neeraj Bala on behalf of Library - Information Access Service