Global food production management has become a challenge with an anticipated population of 10 billion people by 2050 and the ongoing COVID-19 epidemic. Seafood is a vital food source due to its widespread consumption, excellent nutrient profile, and low feed conversion ratio, rendering its sustainable production quintessential. Cellular agriculture or cultured meat can increase seafood production; however, the conventional use of Fetal Bovine Serum (FBS) in culture media restricts its utilization at an industrial level. FBS is effective but has many limitations: unethical animal extraction, high demand and low supply, poorly defined ingredients, variable performance, and high cost that impedes the feasibility and commercial viability of cellular agriculture. Thus, employing serum-free media becomes a quintessential need for cellular agriculture. This project aims to replace or reduce the typical 10% serum usage in Zebrafish embryonic stem cell (ESC) production media with protein hydrolysates derived from low-cost natural sources with high protein content. Enzymatic hydrolysis was performed on nine sources: insects (black army fly and cricket), plants (pea), fungi (mushroom and yeast), algae, and marine invertebrates (oyster, mussel, and lugworm). The resulting hydrolysates were evaluated for serum replacement in zebrafish ESCs. All hydrolysates were used at five different concentrations (10, 1 0.1, 0.01 and 0.001 mg/mL) in serum concentrations of 10%, 5%, and 0% with four biological replicates. The best hydrolysate sources and concentrations were selected for further testing at 2.5% and 1% serum concentrations. All hydrolysates, except for cricket, could restore or significantly increase cell growth with 50% less serum at a concentration of 0.1-0.001mg/mL. Protein hydrolysate concentration of 10 and 1mg/mL was toxic for cells. Additionally, the eight hydrolysates could reduce serum concentrations up to 75–90%. However, no protein hydrolysate could completely replace serum, as cells using only protein hydrolysates exhibited morphological aberrations and decreased growth. Replacing serum with protein hydrolysates lowers cellular agriculture's overall cost, thus enabling the commercialization of cultured meat and the development of a sustainable food system. In the future, blending various protein hydrolysate sources with or without the addition of conventional growth factors could be done to create the ideal serum-free media. / Doctor of Philosophy / With a predicted population of 10 billion by 2050 and the ongoing COVID-19 outbreak, the management of global food production has become a dilemma. However, due to its widespread consumption and good nutrient profile, seafood is an essential food supply, making its sustainable production indispensable. Both capture fisheries and aquaculture are conventional ways to produce seafood. However, they are under tremendous pressure and require alternatives that can alleviate this demand and contribute to the sustainable growth of seafood. In-vitro cultured meat, also known as lab-grown meat, is a novel technique with the potential to supplement the traditional fish sector. It appears a great option, as it completely imitates meat and offers numerous environmental, financial, and health advantages. A culture medium supports the existence, survival, growth, and multiplication of meat-producing cells and tissues in cell-based meat. However, the culture medium uses a Fetal Bovine Serum (FBS) supplement, which dramatically increases the cost and raises many ethical concerns as it is derived from a cow's fetus. In this thesis, we substitute FBS with protein hydrolysates derived from nine distinct sources. Hydrolysing proteins with enzymes produce protein hydrolysates, rich in nutrients and peptides that promote cell development. Enzymes were used to hydrolyse nine unique and protein-rich sources, including insects (black army fly and cricket), plants (pea), fungi (mushroom and yeast), algae, and marine creatures (oyster, mussel, and lugworm). The resultant hydrolysates were investigated for replacement of serum in cell culture. Eight protein hydrolysates successfully replaced 90% of serum without impairing cell growth and structure but could not replace serum entirely. In the future, serum-free media could be created by combining these various protein hydrolysates with or without adding other growth-promoting components.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/111775 |
| Date | 08 September 2022 |
| Creators | Batish, Inayat |
| Contributors | Food Science and Technology, Ovissipour, Reza, Kuhn, David D., O'Keefe, Sean F., Bomkamp, Claire |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf |
| Rights | In Copyright, http://rightsstatements.org/vocab/InC/1.0/ |
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