Date of Award
Winter 12-4-2024
Degree Type
Dissertation/Thesis
Degree Name
Masters of Science in Integrative Biology
Department
Biology
Committee Chair/First Advisor
Christopher Cornelison
Second Advisor
Mario Bretfeld
Third Advisor
Martin Hudson
Abstract
Melanins are enigmatic pigments produced by a wide variety of macro and microorganisms, including numerous species of bacteria and fungi. Sepia officinalis (cuttlefish) is the most well-known and sought-after source of non-synthetic eumelanin, but its harvest is limited by the availability of the cuttlefish fishery and its extraction from an animal source gives rise to ethical concerns. In addition, cuttlefish may harbor gram-negative bacteria leading to possible negative interactions without proper purification if the melanin is used in biomedical applications. Food-grade fungal-derived melanin does not exhibit the same complications. The high value of these molecules is derived from their current lack of stable supply methods and greatly increased demand in a variety of industries such as cosmetics, biomedical, semiconductors, novel UV protectants, with a myriad of additional potential applications. Thus, the ability to produce these molecules, on demand, without animal inputs using fungi grown on globally abundant, low-cost feedstock while applying a circular economic approach could have ramifications across these economic sectors.
Research surrounding melanin-producing fungi dates back more than 50 years. Melanin can be bound within the chitin of the cell wall, but some fungi can produce melanin extracellularly as metabolic exudates. Conventionally, L-tyrosine and L-DOPA are used as precursor molecules to bolster eumelanin production from fungi, however these chemical constituents can be expensive, rendering the process cost prohibitive. However, a quantification of the ability for most edible fungi to produce these molecules when grown upon agricultural byproducts is yet unavailable. This project will also address this lack of knowledge and evaluate the feasibility of producing melanin using food-grade fungi grown on agricultural and industrial byproducts. Trials in this research included an L-DOPA only standard, 4% malt extract + 1mM L-DOPA, Precursor (tyrosine) and Inducer (ethanol), filtered spent media, heat treatment, and finally alternative feedstocks including lignin sulfonate with urea and velvet bean extract.
The results of this research when compared to the standard non-nutritive liquid fermentation culture of 1 mM L-DOPA are as follows. The trials containing 4% malt extract and 1 mM of L-DOPA produced less melanin than that of the L-DOPA only standard when comparing the mushroom mycelium trials. Melanin concentrations were greater in both the tyrosine trials compared to the L-DOPA only trials. Time was the greatest driving factor with melanin concentrations increasing over time with the different tyrosine trials. Melanin concentrations were greater in both ethanol trials compared to the L-DOPA only trials. The filtered spent media trials had no effect on melanin production when compared to the L-DOPA trials. Melanin concentrations were greater in the solutions that underwent heating via the hot plate when compared to that of the autoclave. No discernable melanin was observed from all the lignin sulfonate and urea trials. Melanin concentrations were much higher in the velvet bean extract with mushroom mycelium trials compared to the standard L-DOPA treatments and the velvet bean trials without mushroom mycelium. Overall, the results suggest that production of melanin using the proposed methodologies using a food grade fungi are viable alternatives to current methods using pathogenic fungi. Futher testing needs to be explored to look at possible synergistic effects of treatments that showed increases in melanin production including ethanol inducers, using a hot plate for heating the liquid fermentation, and using velvet bean extract as the feedstock.