Dissertation
Isolation and structure elucidation of secondary metabolites from selected fungi
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Autumn 2022
DOI: 10.25820/etd.006785
Abstract
Secondary metabolites are organic compounds that are produced by a wide variety of organisms from the building blocks of primary metabolism. They are not essential for growth, but may foster organism survival in various ways via involvement in antagonism, competition, communication, regulation, etc. Fungi are prominent producers of secondary metabolites that have value as herbicides, insecticides, antifungal agents, antibiotics, and other types of pharmaceuticals. The majority of past research on fungal metabolites has employed fungi isolated at random from soil samples. The studies described here are part of a long-term program that takes ecological aspects into account when choosing fungi to study because environmental factors suggest that some fungi may benefit in some respects from the production of bioactive secondary metabolites. The work described in this thesis included studies of members of various fungal ecological groups, including coprophilous, fungicolous, and plant-colonizing fungi. These types are targeted based on the idea that such fungi are especially likely to show antagonism toward other microorganisms.
Chemical investigation of four coprophilous isolates collectively yielded two new compounds and 12 previously known compounds. Studies of a coprophilous isolate of Penicillium sacculum led to the identification of a new decarestrictine-derived metabolite, along with the known compounds decarestrictine B, decarestrictine I, and four dioxopiperazines. The new compound consisted of a decarestrictine subunit linked to an unusual tricyclic tetrahydrofuroindole moiety. The structure of this new metabolite was assigned mainly by analysis of nuclear magnetic resonance (NMR) and high resolution electrospray ionization mass spectrometry (HRESIMS) data. Although the original extract showed antifungal and antibacterial activity against the human pathogenic fungi Candida albicans and Cryptococcus neoformans, as well as the bacterium Staphylococcus aureus, the new compound did not show significant activity in these assays. Another coprophilous isolate afforded a new analogue of the known compounds PF1052 and vermisporin.
Studies of fermentation extracts of nine fungicolous fungal isolates yielded a total of five new compounds and 15 known compounds. Studies of fermentation cultures of an isolate of Sphaerostilbella broomeana led to the identification of a new cyclic octapeptide along with two closely related analogues and four dioxopiperazines. The structure of the lead compound, broomeanamide A, was assigned mainly by analysis of NMR and HRESIMS data. The structure consisted of one unit each of N-MeVal, Ala, N-MePhe, Pro, Val, and Ile and two N-MeLeu units. The amino acid sequence was determined on the basis of NMR and HRESIMSMS data. NMR and HRMS data revealed that the other two new peptides have the same amino acid composition except that the Ile unit was replaced with Val in one instance and the N-MeVal unit was replaced with Val in the other. The absolute configuration of broomeanamide A was assigned by analysis of the acid hydrolyzate by application of Marfey’s method using both C18 and C3 bonded-phase columns. Broomeanamide A showed antifungal activity against C. neoformans and C. albicans, with minimum inhibitory concentration (MIC) values of 8.0 and 64 μg/mL, respectively.
The majority of the studies described in this thesis arose from chemical investigation of fungal colonists of various plant materials. These efforts resulted in the identification of two new compounds and 30 known compounds. Chemical investigation of a leaf litter isolates of Eupenicillium sp. afforded a new symmetrical diamide along with the known compound trichodermamide C and three known tryptoquivaline analogues. The identification of the new diamide was complicated by poor NMR properties, which were ascribed to conformational issues in solution associated with the amide groups. The structure was proposed by analysis of 1H NMR, 13C NMR, and HRESIMS data, but ultimately required confirmation by X-ray diffraction analysis. An isolate of Pleonectria cucurbitula obtained from a pinecone, produced a new cyclodepsipeptide. The structure was independently assigned by analysis of NMR and HRMS data and found to be similar to known compounds trichodepsipeptides A and B.
In addition to these studies, the structure revision of a previously reported antifungal agent, campafungin, as well as the assignment of its relative stereochemistry and the identification of three new analogues was also described. These compounds were obtained from a strain of Plenodomus enteroleucus originally isolated from leaf litter.
In summary, several new fungal secondary metabolites, as well as numerous known natural products, some of which exhibited antifungal activity, were isolated and identified in this work. The research presented here shows that the selected types of fungi have the potential to produce a wide range of bioactive natural products with diverse structural features, such as polyketides, peptides, and terpenoids. 1D and 2D NMR data, mass spectrometry, and/or X-ray crystallography were employed to characterize the structures. Chapters 2-6 of this thesis provide details of the isolation and structure elucidation of the compounds described here.
Details
- Title: Subtitle
- Isolation and structure elucidation of secondary metabolites from selected fungi
- Creators
- Dulamini Indika Ekanayake
- Contributors
- James B Gloer (Advisor)Renee S Cole (Committee Member)F Christopher Pigge (Committee Member)Elizabeth A Stone (Committee Member)David F Wiemer (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Chemistry
- Date degree season
- Autumn 2022
- Publisher
- University of Iowa
- DOI
- 10.25820/etd.006785
- Number of pages
- xxi, 187 pages
- Copyright
- Copyright 2022 Dulamini Indika Ekanayake
- Language
- English
- Description illustrations
- Illustrations, charts, graphs, tables
- Description bibliographic
- Includes bibliographical references (pages 178-187).
- Public Abstract (ETD)
Secondary metabolites are organic compounds that are produced by a wide variety of organisms from the building blocks of primary metabolism. They are not essential for growth, but may foster organism survival in various ways via involvement in antagonism, competition, communication, regulation, etc. Such compounds can have useful properties. Fungi are prominent producers of secondary metabolites that have value as herbicides, insecticides, antifungal agents, antibiotics, and other types of pharmaceuticals. The research presented here focuses on the investigation of coprophilous fungi (colonists of herbivore dung), mycoparasitic/fungicolous fungi (colonists of other fungi), and fungal colonists of plant material. Given the need for new antimicrobial agents with novel modes of action, together with the track record of fungi as metabolite producers, we have targeted these groups of organisms as potential sources of new antibacterial and antifungal agents. The majority of these fungi compete with other organisms for nutrients and other resources. As a result, the likelihood of encountering antimicrobial agents from such fungi is hypothesized to be greater than that of random soil isolates. Moreover, these types of fungi are more likely to be chemically unexplored.
Chemical investigations of selected members of these fungal groups resulted in the discovery of a number of new compounds, some of which showed antifungal properties. The structures of these compounds were determined primarily by analysis of various kinds of spectroscopic data. In addition to these new metabolites, many previously reported compounds were encountered and identified, the majority of which have been reported to have biological activities such as antifungal, antibacterial, and cytotoxic effects.
- Academic Unit
- Chemistry
- Record Identifier
- 9984362857602771
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