- Raw Materials, Ingredients, & Inputs
- Target molecule selection
- Host strain development
- Ingredient optimization
Animal-free meat often struggles to mimic the flavor, aroma, and mouthfeel of animal-based meat due to the unique chemical structure and unique molecular arrangement of animal fats compared to plant-based fats. Strategies to address each of these challenges include the synthesis of animal-like fats by fermentation and the use of encapsulation techniques to improve properties like melt temperature and stability. Accumulation of high concentrations of lipids during fermentation as well as scalable methods for extraction, storage, and delivery of those lipids to food matrices remain considerable technical hurdles.
The microencapsulation of lipids within yeast cells is a well-described technique. In an aqueous solution, the yeast cell wall and membrane allow lipid molecules to pass through and accumulate inside the cell. The lipids then are protected against damage from heat (e.g., in cooking) or degradation by light and oxygen during storage. The stored lipids then may be extracted before subsequent use or, in food applications, may be delivered within the yeast cell. The microencapsulation technique is effective for both live and inactivated yeast cells from a variety of species, including yeasts commonly used in food products, such as S. cerevisiae.
While microencapsulation of lipids into yeast cells has been shown to be effective, generating and encapsulating lipids in the same step could increase efficiency. Oleaginous yeast species such as C. curvatus and Y. lipolytica accumulate a high percentage of lipids (20-75% dry weight) in the form of triacylglycerides (TAGs) when subjected to nutrient depletion. Those lipid-containing cells can then be harvested after fermentation, de-watered, and stored directly, without a separate microencapsulation step. These yeast may be used directly in a dried, whole-cell form as a food-grade, lipid-containing food ingredient. Future research in this area could screen wild-type oleaginous yeast strains for superior accumulation of desired types of lipids (e.g., animal-like lipids) and for the ability to retain and protect lipids during downstream drying, storage, and incorporation into alternative protein end products, including extruded plant-based meats. While culture conditions and media formulations conducive to the production of certain types of lipids have been identified, future research should optimize culture conditions specifically for the ideal fat profiles for alt protein applications. Research in this area should also identify parameters of the encapsulation process that work best for each of the major oleaginous yeast strains of interest, in order to start developing standard operating procedures that can be applied to commercial production.
In addition to leveraging strains that naturally produce high levels of desirable lipids, genetically tractable oleaginous yeast species such as Y. lipolytica may be engineered to overproduce lipids or to generate target lipid molecules (e.g., animal-like lipids). Extensive research over the past few decades on Y. lipolytica has developed a molecular toolbox—including holistic omics tools—that can be used to improve lipid biosynthesis and accumulation in this model species. Depending on the target food product or market, a synthetic biology approach may require downstream extraction and purification of the lipids produced, in order to separate them from the genetically modified yeast biomass. The purified lipids then could be re-encapsulated in a GRAS, food-grade yeast such as S. cerevisiae, as described above. This synthetic biology approach should be compared in its entirety with the approach described above to select for high-lipid-producing wild-type strains with respect to the degree of processing required as well as regulatory and labeling considerations. Comparing the entire process, including any downstream isolation or encapsulation steps, will provide direction on which approach offers the most promise for commercial production.
For cases where a yeast cell is used as the carrier for delivery of encapsulated lipids, the lipid release mechanism in alternative protein products should be characterized. Precise descriptions of the kinetics of lipid release by yeast cells under expected temperatures and water activities will allow for the design of foods in which the encapsulated lipids are released appropriately during cooking and consumption, approximating lipid release in animal-derived meats. Understanding the conditions for lipid release from yeast cells will also allow standards for storage and handling to be developed for commercial production, in order to prevent early lipid release and product spoilage.For cases where lipids will be removed from the yeast cell—either from a genetically modified yeast for re-encapsulation or for direct application of the lipid to food—extraction and recovery of lipids from yeast biomass will be required. Extensive research has addressed lipid extraction from oleaginous yeast using various techniques such as ultrasound, microwaves, organic solvents, supercritical CO2, enzymes, and combination methods. These techniques each present challenges for scale-up to commercial production, mostly due to the recalcitrance of the rigid cell wall of oleaginous yeast, which contains high amounts of lignin and mannan. Because each oleaginous yeast has a slightly different cell wall composition, extraction methods specific to each yeast will be required. Also, many of the existing methods for lipid extraction were designed to extract lipids for biofuel usage; thus, the solvents and processing steps may not be appropriate for preparing food-grade lipids. Future research should focus on either finding food-suitable methods for lipid extraction from oleaginous yeast or genetically engineering strains with less rigid cell wall structures. The challenge of developing these solutions could inform the comparison between the use of wild-type or engineered oleaginous yeast with respect to cost and scalability.
This research is expected to address fundamental questions on the feasibility of using oleaginous yeast for the production and encapsulation of lipids for alternative protein products. Methods to either encapsulate lipids inside the oleaginous yeast producing them or to re-encapsulate them after extraction from the host will allow lipids to be protected from light and oxygen that might otherwise degrade them. This research will also assess whether a fermentation strategy using wild-type oleaginous yeast or genetically engineered yeast would be preferred from a cost, feasibility, regulatory, and scale-up perspective. The research also has the potential to lay the foundation for developing scalable, commercially viable methods for encapsulation methods specific to oleaginous yeast.
- Eden Research
- Pharmaceutical Encapsulation
- Oleaginous yeast for biofuels and biochemicals
- Fat and moisture encapsulation techniques for alternative protein applications
- Fat uptake & biosynthesis in cultivated meat cells
- Plants as a recombinant protein expression platform for functional food ingredients
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