Project Description

Environmental Challenge

Food waste, often overlooked as a tiny fragment of human life, has evolved into one of the most pressing global environmental challenges. It comprises 8-10% of yearly greenhouse gas emissions globally. Food waste contributes to nearly five times the total emissions compared to the aviation sector. This contributes to a significant biodiversity loss that uses up almost one-third of the world's agricultural land (UNFCCC, 2021). On top of this, from production and transportation to decomposition, emissions accumulate across the entire supply chain.

Economic Challenge

Economically, food waste is a major burden. The U.S. Environmental Protection Agency reports that the cost of food waste to American consumers averages about $728 per year, and for a household of four, the annual cost is $2913, which highly contributes to the inefficiency of the economy (2025). In 2022, according to Katarzyna Slopiecka, a researcher at the University of Perugia, Department of Engineering, there is an inverse relationship between the national GDP and the composition of organic food waste (2022). In high-income countries, advanced packaging, cold-chain logistics, and efficient distribution effectively reduce the amount of organic food waste. In contrast, lower-income neighborhoods often experience higher proportions of organic waste because of limited access to preservation technologies, inefficient collection systems, and reliance on fresh produce markets. These provide insights into targeting our solution towards varied socioeconomic contexts.

Current Solution and Problem

The composition of food waste is primarily made up of carbohydrates (41-62%), proteins (15-25%), and lipids (13-30%). Carbohydrates in food waste mainly consist of cellulose, hemicellulose, and starch, while lignin represents a separate class of aromatic polymers often associated with them (Katarzyna Slopiecka, 2022). These organic components make up a significant portion of the total compounds in food waste and contribute to the main goal of our project.

Connection to the BSFL and gap within the Current Solution

Black soldier fly larvae (BSFL), which is seen as the most efficient solution for breaking down, it is polyphagous, consuming all types of organic materials of animals and plants (Bava et al., 2019). It can be used to convert municipal waste with a reduction rate ranging from 65.6% to 78.9% (Diener et al., 2011). However, the natural gut microbiota of BSFL do not produce enough cellulase to fully break down plant cell walls, creating a barrier to digesting cellulose and limiting nutrient extraction. Through detailed analysis of the problem, we came up with a collective approach of using synthetic biology as the foundation to renovate Escherichia coli (E. coli) to enhance the pre-digestion of plant-based food waste, ultimately optimizing BSFL-based upcycling.

Our Experimental Solution

To complement public acceptance strategies, our team advances a bioengineering approach centered on Endo-1,4-β-glucanase (abbreviated as endoglucanase). This enzyme, naturally produced by Bacillus subtilis JA18, plays a crucial role in breaking down cellulose into smaller glucose polymers, which boost the degradation of plant-based food waste. We designed to engineer this enzyme gene into the pET22b (+) plasmid, which provides both a T7 promoter for strong expression and the potential for downstream protein purification. For system stability, the recombinant plasmid is then maintained in E. coli DH5α for plasmid propagation and cloning. Subsequently, the construct is transferred into E. coli BL21, enabling high-level protein expression following IPTG induction.

This strategy ensures reliability, as DH5α safeguards plasmid fidelity while BL21 drives efficient protein production. Once expressed, endoglucanase can be purified and tested for its ability to degrade food waste substrates, mainly composed of cellulose-rich residues. In addition, our design includes secretion tags of YebF, which are also integrated in the designed endoglucanase strain to enhance extracellular enzyme release. Normally, recombinant proteins expressed in E. coli BL21 remain inside the cytoplasm or periplasm, limiting their direct contact with substrates such as cellulose. By fusing endoglucanase with YebF, we harness its secretion pathway to transport the enzyme outside the cell, allowing it to act directly on cellulose. As a result, food waste decomposition could become more scalable by coupling BSF larvae's natural waste-recycling capacity with our targeted enzymatic pre-treatment.

This pre-treatment strategy not only accelerates the bioconversion rate but also enhances the nutritional yield of the resulting biomass. Once introduced into the food waste, the biologically engineered material will break down the large and cellulose into smaller and bioavailable fragments. This process is beneficial for the BSFL as it makes it easier for the organism to digest and speeds up the overall waste degradation speed, which optimizes conversion into high-protein by-product for the biomass to use.

Scientific role of fusing Endo-1,4-β-glucanase with E. coli

Endo-1,4-β-glucanase plays the central role in the breakdown of cellulose. Cellulose consists of long chains of glucose molecules linked by β-1,4-glycosidic bonds, which are highly crystalline and resistant to degradation. Endoglucanase functions by cleaving these β-1,4 linkages at random internal sites along the cellulose chain, producing shorter polysaccharides such as cellooligosaccharides and ultimately glucose units. This random internal hydrolysis differentiates endoglucanase from exoglucanases, which act only at chain ends, and makes it particularly effective for initiating cellulose depolymerization.

When fused with E. coli, endoglucanase can be expressed in large quantities in a controlled and renewable manner. Using vector pET22b (+), expression can be regulated with IPTG induction to optimize enzyme yield. The recombinant E. coli acts as a cell factory, producing endoglucanase that can then attack the rigid cellulose in food waste. In short, endoglucanase reduces the structural complexity of cellulose waste and improves accessibility for further enzymatic digestion or nutrient assimilation by fragmenting the polysaccharide network into smaller units.

Implementation

Conventional enzymatic treatments can be expensive due to extraction, purification, and stability concerns. However, recombinant expression in E. coli offers a low-cost, modular, and sustainable platform. By adding an enhancement to the efficiency of the degradation process of cellulose for the BSFL, we have not only reduced the cost but also made it more scalable, which makes it easy to implement in different parts of the world. In comparison to the traditional method, which makes it ineffective for BSFL digestion due to the composition and variability of food waste, our solution guarantees a more stable, accelerated, and more efficient degradation process by directly addressing both substrate inconsistency and enzymatic accessibility. This makes the large-scale production of endoglucanase practical for community or industrial waste treatment, ensuring accessibility and broad adoption. In conclusion, by implementing the LarVase plan, food waste management could contribute to the world vision on minimizing food waste and improving our environmental conditions.

Conclusion

Our project creates a brand new vision in the world of waste management. Traditional solutions of feeding waste to pigs or relying on BSFL digestion face challenges. Pigs, for instance, cannot be widely adopted in Taiwan due to disease risks like African swine fever. BSFL, while effective in reducing food waste, struggles with digesting cellulose, which limits its efficiency.

To overcome this issue, our project takes a synthetic biology approach by engineering E. coli to express Endo-1,4-β-glucanase, an enzyme that hydrolyzes cellulose into smaller, more easily digestible sugars. This highlights our advantage of accelerating decomposition and increasing the value of BSFL growth. By doing so, our project can evolve into a green and circular economic system for the greater good.

"Food Waste Research | US EPA." US EPA, 16 Nov. 2022, www.epa.gov/land-research/food-waste-research.
Slopiecka, Katarzyna, et al. "Chemical and Physical Characterization of Food Waste to Improve Its Use in Anaerobic Digestion Plants." Energy Nexus, vol. 5, Mar. 2022, p. 100049, https://doi.org/10.1016/j.nexus.2022.100049.
United Nations Framework Convention on Climate Change. "Food Loss and Waste Account for 8-10% of Annual Global Greenhouse Gas Emissions; Cost USD 1 Trillion Annually | UNFCCC." Unfccc.int, 30 Sept. 2024, unfccc.int/news/food-loss-and-waste-account-for-8-10-of-annual-global-greenhouse-gas-emissions-cost-usd-1-trillion.