This article addresses the rising environmental concerns related to organic waste management and explores innovative ways to transform waste into valuable resources. Conventional methods like composting and incineration contribute significantly to greenhouse gas emissions. However, biological approaches such as converting organic waste into biofuel and biodegradable plastics are gaining traction. For instance, bioethanol and biogas production effectively manage waste but have limitations in carbon sequestration. A novel method involves using microorganisms to produce biodegradable plastics like PHAs, although cost remains a barrier. Additionally, recombinant spider silk emerges as a promising carbon-sequestering biomaterial. The development of spider silk biomaterials using waste could reduce production costs and environmental impact, fostering a low-carbon circular economy.

Against the backdrop of escalating public health concerns and a deepening environmental consciousness, the effective management and utilization of waste streams and renewable resources emerge as a shared focal point of interest and study within the scientific and industrial spheres. The mismanagement of high organic waste could potentially lead to the release of potent greenhouse gases such as carbon dioxide, methane, and nitrous oxide—a significant driver of global warming (Nordahl et al., 2023). Conventional methods for treating high organic waste encompass composting, incineration, landfilling, and anaerobic digestion (Hogg et al., 2008; Lou & Nair, 2009; Shen, Huang, et al., 2022). Notably, the greenhouse gas emissions resulting from waste treatment activities contribute approximately 5% to the global greenhouse gas inventory, predominantly stemming from methane emissions arising from the anaerobic degradation of solid waste and carbon dioxide emissions resultant from the breakdown of wastewater constituents (Eggleston et al., 2006). Hence, the imperative to develop more efficient mechanisms for the treatment of high organic waste emerge as a pressing societal and environmental challenge (Nordahl et al., 2023).

Embracing biological methodologies to metabolize organic constituents within high organic waste streams and converting them into bioenergy or usable commodities represents the prevailing trend in addressing this challenge (Evans, 2014). An example of this is the process of producing bioethanol, which entails converting organic waste with a high organic content into fermentable sugars such as glucose, fructose, and sucrose. This is achieved by pretreatment procedures, followed by fermentation using specialized strains of yeast to produce biofuel (Bhuyar et al., 2021; Jayakumar et al., 2023). In the same way, the production of biogas involves the anaerobic fermentation of high-quality organic waste. This process involves the sequential hydrolysis, acidification, acetification, and methanation of complex hydrocarbons to produce hydrogen and methane, which can be utilized as biofuels (Shen, Huang, et al., 2022; Shen, Torre, et al., 2022; Souvannasouk et al., 2021). The conversion of waste streams into biofuels is an effective option for waste management. However, it is important to point out that the long-term carbon sequestration advantages are limited due to the short shelf life and operational cycles of biofuel consumption. On the other hand, using different groups of microorganisms to produce polyhydroxyalkanoates (PHAs), which are biodegradable plastics, from large amounts of organic waste is a novel approach to dealing with the disposal of organic waste and increasing the lifespan of the resulting products (M.-Y. Shen et al., 2023). However, the high production costs of PHAs compared to the low market pricing create extended payback times, which require a reevaluation of economic value models for waste-to-product conversions (Lai et al., 2022; M. Shen et al., 2023). The production of recombinant spider silk involves the use of culture media rich in organic carbon and nitrogen sources as fermentation substrates, subsequently undergoing biosynthesis (Hsu et al., 2024). If high organic waste could replace the culture media used in producing recombinant spider silk, it could enable the conversion of waste into high-value, long-term carbon-sequestering biomaterials.

To enhance the commercial competitiveness of spider silk biomaterials and simultaneously develop applications using spider silk materials as carbon-negative feedstock and product-oriented innovations, our team aims to integrate innovative biotechnologies and next-generation bioprocess development. This endeavor seeks to achieve a carbon conversion process that transforms biomass waste into high-value spider silk materials, significantly reducing production costs while substantially enhancing the carbon sequestration benefits of spider silk. Additionally, the durability and biodegradability of recombinant spider silk help extend product lifespan and reduce waste, thus further lowering carbon emissions and creating a new bioprocess for materials within a low (negative) carbon circular economy model. Specifically, this project will develop recombinant spider silk materials and renewable indigestible biopolymers as our carbon sequestration materials, demonstrating the potential and advantages in lowering carbon emissions, extending the carbon sequestration cycle, and promoting resource recycling.

In our iGEM project, the Dry Lab team was tasked with analyzing and expanding the experimental data from the Wet Lab, focusing specifically on carbon emission impacts. Our main goal was to upscale the results from a microbioreactor to a larger 330L reactor to conduct a comprehensive life cycle assessment (LCA) and quantify the carbon emissions.

Utilizing data from the Wet Lab, we calculated the carbon footprint involved in producing recombinant spider silk protein. When we compared conventional commercial carbon sources with high-organic-content wastewater as raw materials, our analysis clearly indicated that using wastewater significantly reduced carbon emissions. This reduction occurs because wastewater is rich in organic compounds, effectively decreasing the need for external carbon sources.

Importantly, the recombinant spider silk produced from wastewater not only exhibits excellent material properties but also serves as a form of long-term carbon storage. This dual benefit not only reduces carbon emissions but also sequesters carbon within the fiber material, contributing to environmental protection and sustainability. Our Dry Lab analysis highlights the importance and potential of waste reuse in carbon reduction strategies, offering valuable insights for future environmentally friendly production methods.

A carbon audit is a systematic approach to measuring the greenhouse gas emissions produced by an organization or activity. The goal is to understand and manage the carbon footprint, enabling more effective emission reduction strategies. Here's a typical process for conducting a carbon audit:

1. Define Scope: First, determine the scope and boundaries of the audit, including direct emissions (Scope 1), indirect energy emissions (Scope 2), and other indirect emissions (Scope 3). This helps identify specific sources of carbon emissions.
2. Data Collection: This step involves gathering data related to energy use, transportation, manufacturing processes, waste management, etc. Sources of data might include bills detailing electricity, natural gas, and fuel usage, transportation records, and logistics data.
3. Emission Calculation: Use standardized emission factors and calculation tools, such as the GHG Protocol or ISO 14064, to convert the collected data into specific greenhouse gas emissions. This quantifies the contribution of each activity to the overall carbon emissions.
4. Analysis and reporting: Analyze the data to identify major sources of emissions and potential reduction opportunities. Compile a report to provide stakeholders with a clear understanding of the organization's carbon emission status and areas for improvement.
5. Develop a Plan: Based on the analysis, establish specific emission reduction targets and strategies. These might include enhancing energy efficiency, adopting renewable energy, or utilizing carbon offsets.

By conducting such a carbon audit, an organization can more accurately assess its environmental impact and effectively advance sustainable development goals.

We will calculate the carbon emissions for both the Commercial Medium and the High Organic Wastewater Medium by expressing the calcula- tions in matrix form. Each element in the matrices represents specific data, which we will explain in detail. We will use matrix operations to compute the emissions where possible.

Let:

1. Q: Combined concentration matrix (g/L) of chemicals used.
2. C: Emission coefficient vector (kg CO$_2$e/kg) for each chemical.
3. V: Volume vector (L) for Initial Medium and Feeding Medium.
4. M: Mass matrix (kg) of chemicals used: \[ \mathbf{M} = \frac{\mathbf{Q} \mathbf{V}}{1000} \]


5. E: Emission matrix (kg CO$_2$e) for each chemical: \[ \mathbf{E} = \mathbf{M} \circ \mathbf{C} \] Note: where $\circ$ denotes element-wise multiplication.


6. $E_{\text{total}}$: Total carbon emissions (kg CO$_2$e): \[ E_{\text{total}} = \sum_{i} E_i \]

Combined Concentration Matrix

Volume Vector V (L):

\[ \mathbf{V} = \begin{bmatrix} 2.5 \\ 1.0 \end{bmatrix} \]

Emission Coefficient Vector $\mathbf{C}$ (kg CO$_2$e/kg):

\[ \mathbf{C} = \begin{bmatrix} 1.84 \\ 2.16 \\ 3.36 \\ 18.7 \\ 3.03 \\ 4.54 \\ 1.31 \\ 0.47 \\ 0.18 \end{bmatrix} \]

We sum the masses from the Initial Medium and Feeding Medium:

\[ \mathbf{M}_{\text{total}} = \mathbf{M}_{\text{initial}} + \mathbf{M}_{\text{feeding}} \] Compute total mass for each component:

We compute the emissions for each component: \[ \mathbf{E} = \mathbf{M}_{\text{total}} \circ \mathbf{C} \] Computing the emissions:

\[ E_{\text{medium}} = \sum_{i=1}^{9} E_i = 2.221536 \, \text{kg CO}_2\text{e} \]

We calculate the emissions from the process chemicals used in the production process.

\[ E_{\text{process}} = 0.006875 + 0.019341 + 0.0514 + 0.001542 + 0.24684 + 0.0315 \]

\[ E_{\text{process}} = 0.357498 \, \text{kg CO}_2\text{e} \]

\[ V_{\text{water}} = 426.925 \, \text{L} = 0.426925 \, \text{m}^3 \]

\[ EF_{\text{water}} = 0.0948 \, \text{kg CO}_2\text{e/m}^3 \]

\[ E_{\text{water}} = V_{\text{water}} \times EF_{\text{water}} = 0.426925 \times 0.0948 = 0.040472 \, \text{kg CO}_2\text{e} \]

Process Chemicals Emissions: $E_{\text{process}} = 0.356498 \, \text{kg CO}_2\text{e}$

Water Usage Emissions: $E_{\text{water}} = 0.040472 \, \text{kg CO}_2\text{e}$

\[ E_{\text{total, commercial}} = E_{\text{medium}} + E_{\text{process}} + E_{\text{water}} \]

\[ E_{\text{total, commercial}} = 2.221536 + 0.356498 + 0.040472 = 2.618506 \, \text{kg CO}_2\text{e} \]

Combined Concentration Matrix

Volume Vector $\mathbf{V}$ (L):

\[ \mathbf{V} = \begin{bmatrix} 2.5 \\ 1.0 \end{bmatrix} \]

Emission Coefficient Vector $\mathbf{C}$:

\[ \mathbf{C} = \begin{bmatrix} 0.43 \\ 3.48 \\ 3.36 \\ 18.7 \\ 3.03 \\ 4.54 \\ 1.31 \\ 0.47 \\ 0.18 \end{bmatrix} \]

We sum the masses from the Initial Medium and Feeding Medium:

\[ \mathbf{M}_{\text{total}} = \mathbf{M}_{\text{initial}} + \mathbf{M}_{\text{feeding}} \]

We compute the emissions for each component: \[ \mathbf{E} = \mathbf{M}_{\text{total}} \circ \mathbf{C} \]

Note:Since the wastewater carbon source (Crude Glycerol) has no preprocessing, we can subtract the purification emission of PMP (3.46 kg CO$_2$e) from its emission coefficient.

\[ E_{\text{medium}} = \sum_{i=1}^{9} E_i = 1.2263955 \, \text{kg CO}_2\text{e} \]

We calculate the emissions from the process chemicals used in the production process.

Total Process Chemicals Emissions:

\[ E_{\text{process}} = 0.006875 + 0.019341 + 0.0514 + 0.001542 + 0.24684 + 0.0315 \]

\[ E_{\text{process}} = 0.357498 \, \text{kg CO}_2\text{e} \]

\[ V_{\text{water}} = 426.925 \, \text{L} = 0.426925 \, \text{m}^3 \]

\[ EF_{\text{water}} = 0.0948 \, \text{kg CO}_2\text{e/m}^3 \]

\[ E_{\text{water}} = V_{\text{water}} \times EF_{\text{water}} = 0.426925 \times 0.0948 = 0.040472 \, \text{kg CO}_2\text{e} \]

\[ E_{\text{total, wastewater}} = E_{\text{medium}} + E_{\text{process}} + E_{\text{water}} \]

\[ E_{\text{total, wastewater}} = 1.2263955 + 0.357498 + 0.040472 = 1.6243655 \, \text{kg CO}_2\text{e} \]

We calculate the spider silk protein carbon footprint:

\[ \text{Carbon Footprint} = \frac{E_{\text{total}}}{\text{Protein Yield (kg)}} \]

Given:

This study highlights the significant potential of transforming high organic waste into valuable products, such as recombinant spider silk production, while addressing the challenges of carbon emissions and environmental impact. The carbon footprint analysis of two media types, commercial and high organic wastewater, reveals the possibilities of reducing carbon emissions through alternative methods.

Recombinant spider silk, in particular, stands out as a promising carbon-sequestering biomaterial that could play a key role in fostering a low-carbon circular economy. By utilizing high organic waste as a culture medium for producing spider silk, the environmental and economic benefits can be substantial, including cost reduction, enhanced carbon sequestration, and longer product lifespans. This approach not only offers a sustainable solution for waste management but also creates new opportunities in carbon-sequestration bioprocesses.

Future work should focus on improving the efficiency of waste-to-product conversion processes and addressing the economic barriers associated with the production of biodegradable materials like PHAs and recombinant spider silk. This will be crucial in enabling the large-scale adoption of these biotechnological solutions and promoting a sustainable, low-carbon economy.