Product Design

Fig 1. Flow chart of E. coli carbon utilization system

1. Overview

In this project, we, the NCKU Tainan Team, have proposed an alternative way to reduce the emission of Carbon dioxide (CO₂). Referring to the opinions and feedbacks from many industry experts and professors, we design a new factory flow to capture CO₂ by E. coli Not only our device meets the specs to commercialize, but it also demonstrates high cost performance.

The emission of CO₂ has been a serious problem for a century that causes global warming and severe climate change. Even though many ways have been tried to reduce it, the generation of CO₂ primarily from industry is still overwhelming. Therefore, scientists and governments have been working hard to find solutions to tackle the problem.

2. Control System


Fig 2. Overview of the control system

There are many aspects we need to consider. First, we calculate the emission velocity of CO₂ from the factory, as well as the medium exchange rate and the growth rate of E. coli.

Fig 1. is a process of whole E. coli carbon utilization that we design for industrial application. We simplify it into three parts which shows in Fig 2. to explain more clearly. Three switches control three parts, named A, B and C. Basically, the factory replaces the medium twice a day. At one hour before replacing the medium, the user needs to turn on switch C to discharge ninety percent of the medium. When it is time to replace the medium, switch C will be turned off and switch B will be turned on to refill medium. When sufficient medium is added, switch B will be turned off and switch A will be turned on to let CO₂ in. Just like the animation showed on Fig 1..

Considering the cost, the growth time of our E. coli and the floor area, we optimized replace time of the medium, replace it every twelve hours and with 72 parallel bioreactors. Next, we are going to have more detail description on three parts, which are Gas preparation system and flow system, Medium preparation, and Downstream products purification and biosafety.

A. Gas preparation system and flow system



Fig 3. Diagram of gas preparation system and flow system

According to IGCC (Integrate Gasification Combined Cycle) flow diagram, the fuel is first converted to syngas which is a mixture of H₂ and CO. The syngas is then burned in a combined cycle consisting of a gas turbine and a steam turbine with a heat recovery steam generator (HRSG). After CO₂ / H₂ separation, IGCC can reach the demand of CO₂ purity including low SOx and NOx emission fraction of allowable limits of bacteria. Finally, the produced flue gas could enter the pipeline leading to the bioreactor.

In E. coli utilization system, the air is pumped in to neutralize the concentration of CO₂. A controlled valve is used to control flow rate and split distribution. When the switch a is turned on, the switch b will be turned off, and vice versa. As for the CO₂ inlet and outlet, it will maintain an open system of bioreactor. In other words, CO₂ will enter continuously and cause some non-reacted CO₂ emitted.



Fig 4. IGCC process flow diagram. Source: Vattenfall. (2010)

Syngas has been treated by sulfur and nitrogen removal, as well as heavy metal removal and cooling tank. Through IGCC process, purified CO₂ in flue gas is allowable for E. coli CO₂ utilizing.

B. Medium preparation



Fig 5. Diagram of medium preparation

At this stage, we have two sections to consider, medium storage and medium preparation before replacing time.

The medium is composed of M9 salt and xylose. For storage, we will convert it into powder with the required proportion. At one hour before replacing time, pour the powder into the medium tank and turn on the water injection switch. Turn on the stirrer of medium tank to have medium powder and water perfect mixing. The outlet of bioreactor (switch c) will be turned on at the same time, letting ninety percent of the medium in the bioreactor flow out . When the medium have prepared well, turn on the switch a and switch b for replacing medium in bioreactor, while the switch c will be turned off.

We also consider the process of raw materials, especially xylose, which is the key source of our pathway. Since xylose is one of the products of agricultural waste degradation, we visited the 2018 Tainan Biotechnology and Green Energy Expo to consulted with researchers from National Energy Program-Phase II, whose projects was biofuel and biodegradable plastic production via agricultural waste. They had developed technique that degrade cellulose and semi-cellulose by ion solution.

Besides, we have opportunity to collaborate with UESTC-Chian team . They work for degrading straw with synthetic biology and convert the product into bio-fuel. One of the product from straw degradation is xylose. These techniques are eco-friendly and low-energy-require. Therefore, the process development of xylose production will be a low-carbon-emission process.

C. Downstream products purification and biosafety



Fig 6. Diagram of downstream process

We will discharge 90% of the used medium in the bioreactor one hour before new medium flows in. Which means that we let 10% of the culture remain in the bioreactor as seed culture. The effluent medium will be sterilized and filtered in the downstream clean-up tank. At this step, we harvest the bacteria and extracting the by-product such as amino acids, proteins, medicine or bio-fuel. Different extracting process designed depends on different by-product.

Besides, we try to reuse the waste heat of factories for sterilizing. The waste water can be recycled as well through removing toxins and adjusting pH value the effluent could return to the medium tank. As for energy require for this system, renewable energy helps us to reach near -zero carbon emission process.

Furthermore, we would like to set up membrane bioreactor (MBR) system, which use a hollow filter membrane that is able to filter most of bacteria in the sewage sludge. We use the system to concentrate the used medium before extracting by-product. And the water went through the system is able to recycle back to the medium tank.



Fig 7. Picture of waste water recycle system



Fig 8. Picture of MBR from KME technology Inc.

Application : China Steel

Fig 9. Picture of CSC interview

Meeting with experts and stakeholders is important in shaping our project to fulfill the needs of our target user. China Steel Corporation is the largest integrated steel Manufacturer in Taiwan. Also, they had been adopting the algal bio-sequestration by cooperating with the research group at our university.

Process

We were given the opportunity to meet with the senior executive of China Steel Corporation to gain invaluable insight for our research. The meeting commenced with our presentation. During the presentation, we introduced our project, including the bioreactor design and the industrial model. By listing out all the aspects we had considered, we would like to obtain advice on the practical and social considerations involved in the application of our project in industry.

Suggestion and Question

Will the high concentration of CO₂ retard growth of engineered bacteria?

Microalgae is reported resistant to SOx and NOx. Does E. coli survive under such conditions?

The two questions above were the main concern of CSC. Basically,the best condition for engineered E. coli to capture CO₂ is a lower CO₂ concentration without too much SOx and NOx particles. However, we won’t be able to provide an ideal culture condition in Industrial application. After testing the tolerance of E. coli, we conclude that E. coli is possible to survive under that kind of condition in factory and the only effects its expression. It may not capture as much CO₂ as culture in the lab.

It is important to define a specific commercial product that can be truly produced since your user may consider its economic viability. They stated that a product that can be widely used is better. At the same time, we should consider current GMO legislation if we want to commercialize those products. The actual condition is not as ideal as in the laboratory, we should optimize the condition to maximize the carbon fixation ability of the microbes.

Interview record

The record can be separated into two parts. One is about their feedback after interview, another one is our customer investigate questions. We use CSC represent China Steel.

Click to see complete interview

Business Model

The business model describes how an organization creates, delivers, and captures value in an economic, social, cultural, or other environment. Therefore, we introduce this business model as the basis for assessing the integrity and effectiveness of our ideas to work with our industry and even national research. First, we ask questions about this, and beyond the solution, we also explain why we chose this question. Second, we analyzed future developments, including the advantages of using this approach. Next, we introduce our plan to many relevant departments and discuss with the national research. I hope that this plan can be used to promote this plan in the future.

Target issue

More and more people are now paying attention to the impact of CO₂. The trend of environmental degradation is gradually increasing. Scientist and national worldwide contribute to capture those excessive CO₂. However, how to reduce carbon and use it has become a major problem today. Challenges against carbon process are complicate. Except the technique and implement problem, social acceptability and policy are other key factors about carbon process technology.

In general, planting is a method of carbon process, and the current use of green algae as a method of carbon utilization. This year, we hope to combine synthetic biology with the most advanced technologies. We want to draw people's attention to the environment and reuse these environmentally stimulating projects.

Business model analysis

Cost Evaluation

The cost evaluation is always crucial for product being on the market. To compare our engineered E. coli to microalgae, we calculate how much the cost it would be when capturing 1 ton of CO₂.

Volume

Table 1 Volume required in capturing 1 ton of CO₂

Organisms	CO2-fixation rate (mg/L*hr)	Biomass concentration (gDCW/L)	Specific CO2-fixation rate	Volume requiredd (L)
Engineered E. coli	19.6	0.87	22.5	51000
Chlorella vulgaris	53	5.7	9.3	19000

Fig 6. Different volume required between micralgae and engineered E. coli

For capturing 1kg of CO₂ in one hour, 51000 L is required with engineered E. coli carbon utilization. It seems that the difference volume required for utilizing same amount of CO₂ is disadvantage of E. coli carbon utilization system. At this situation, we have to look into the design of the different bioreactor. For microalgae culture, it requires a large surface area to increase light intensity. As usual, the height of the microalgae culture pond cannot exceed 0.5 m. In other words, we have to build a 7 m diameter culture pond with the volume of 19000L. In constrast, engineered E. coli is not limited by light. The bioreactor of E. coli can be built with any height in the indoor or outdoor. To scale up the bioreactor, a 5.8 m diameted with 1.9 m height equals to 51000 L which has lower floor area required.

As a result,the bioreactor of engineered E. coli can save more than 30% floor area compared with micoralgae culture pond. Take the floor area of Taiwan as an example, we can build 94 billions of microalgae culture pond to uilize 10% of annual emission with 12 operation hours. However, 1 over 3 of floor area will be save if we replace them with E. coli bioreactor. E. coli bioreactor is more flexible on spacing using, and is less sensitive to weather effect.

Cost

The most expensive source in the medium of our engineered E. coli is xylose. 1 mole of xylose will capture 0.17 mole of CO₂. Therefore, we need 20.0535 kg of xylose while 1 kg of xylose costs 2 USD. The total cost for our engineered E. coli requires 40.107 USD for capture 1 ton of CO₂. In contrast, microalgae needs 1000 liter to capture 250 g of CO₂, so it needs 4000 liter (about 4 tons) water while 1 ton costs 9.78 USD. The total cost for microalgae is 39.13 USD.

Table 2 Cost required in capturing 1 ton of CO₂

Future Work

For industrial application design, we focus on manufacturing valuable products using pyruvate and the linkage between our engineered E. coli between factory. We have designed a device containing our recombinant E. coli, constructed a system which links with factory. However, we still look forward to more modifications of our biological pathway and system.

The most important intermediate product, pyruvate, is also possible to be converted to other compounds by E. coli native enzymes or constructed enzymes which is clone into E. coli from other organism. For future work of pyruvate, we expect that it is predicable to produce amino acid, fatty acid, biofuel and even biodegradable plastic. Pyruvate is crucial for central metabolism pathway, the TCA cycle, of most organism and has the potential to become vary biochemistry compounds.

We set our first future goal at producing glutamine, an essential amino acid for human and some animals. We can simply purify it as a nutrient supply. Not only for medical and daily usage for people, but also for animal husbandry. Furthermore, glutamine can easily convert to other amino acid, and potentially produce other proteins.

Fig 10. Diagram of pyruvate in central carbon metabolism

Furthermore, researchers have successfully constructed pathways produced cellulose and Poly 3-Hydroxybutyrate-co-3-Hydroxyvalerate through the TCA cycle. We are confident of manufacturing more valuable and diverse products from pyruvate.

WAs for the device we designed, we expect that it is possible to modify our device for power generator and other industry. Our device can utilize CO₂ and convert it into various valuable products. With our system, companies can not only reduce CO₂ emission but also make profits.