Medium-chain fatty acids, unlike long-chain fatty acids, have no negative effect on cholesterol. Medium-chain fatty acids help reduce the risk of atherosclerosis and heart disease, both diseases, and help prevent heart disease. No GMO risk, no allergen risk. The lampblack is significantly lower than the common cooking oil on the market, which can reduce the impact of lampblack on the family environment.
So MCFA is considered to be a good helper in controlling inner body fat accumulation and can be added to food as a supplement to protect gut health. The most common thing we know containing MCFA is coconut oil, but since natural MCFA is relatively rare, we hope to improve existing technologies through our project to achieve an increase in the production of MCFA. Technically, we aimed to intensify the MCFAs synthesis pathway by over expressing key enzymes in the reversal of the β-oxidation cycle in E. coli and optimize the T7 promoter to increase MCFAs production from 1.3g/L.
We also did a market-oriented background analysis of the biotechnology industry, based on Porter’s Five Forces Model. To summarise, our challenges come mainly from two aspects, one is our identity as a new entrant will face many restrictions, and the other is the present competitors. For the threat of substitutes, the medium chain fatty acids are special commodities and the substitutes are mainly products of new technologies or new mechanisms, the process of developing and putting substitutes into use will be extremely long, so substitutes pose little threat to competition in the biotechnology industry. More specific information will be explained in the Challenge part and our business plan.
One of our proposed products is healthy cooking oil by adding MCFAs we produced into cooking oil. Our products have less soot, are not easy to deposit into fat, are easy to digest, high conversion rate of suitable for middle-aged and elderly people's cooking oil, but also suitable for people in the period of fat reduction.
Invetigation on cooking oil and coffee products
In addition, MCFAs we produced can not only be added to the cooking oil but also other products like bulletproof coffee. One of the latest healthy trends in fashion circles in the US and Europe is to 'drink oil', but instead of going home and drinking it from a bottle as you might expect, Bulletproof Coffee is the Bulletproof Coffee that contains a variety of functional oils. Bulletproof Coffee is a fatty, over 460 calorie, carb-free coffee. Its main formula is low mold coffee beans, salt - free grass feed butter, medium chain triglyceride oil.
Therefore, based on the different functions of MCFAs, we have summarized two categories of target customers.
Function 1: For those who need "Weight management" and "sports nutrition": Metabolic pathways distinct from long-chain fat are important for patients with poor gastrointestinal function and are therefore widely used in enteral nutrition preparations.
Function 2: It can also be used for special patients: it can be used as food for patients with fat malabsorption, AIDS patients, cancer patients, and diabetics, it can be used for the treatment of prostatic hyperplasia, cholesterol reduction, prevention of hyperlipidemia, etc. It can also be used as special food for patients after surgery, infection, and skin burns.
Concept drawing of our product
Availability and environmental protection: Prepared from crude glycerol, less in tropical China, so far, the only oils with medium chain fatty acids are coconut oil and palm kernel oil. Our technology can break down the influence of geography.
Controllability and replicability: The technical difficulty is relatively low, and the replicability is strong. The existing production chain of relevant enterprises can achieve large-scale production without purchasing large heavy assets.
We presented a new approach to producing MCFAs using an engineered reversal of the β-oxidation (r-BOX) cycle as shown below, and we use the Escherichia coli strain to set up the platform.
The fatty acid β-oxidation cycle
Traditionally, the bacteriophage T7 promoter has been widely used for the expression of heterologous protein, but its use in metabolic engineering applications is often limited because the overburden of cells, which reduces the translation of essential intrinsic mRNAs, thereby making the cell unhealthier. The t7 promoter is prone to leaky expression even under T7-lac control. By site-directed mutagenesis of the reported promoter region, t7 promoter mutants with reduced strength can be obtained, reducing the cellular burden and thus increasing MCFAs production.
HPLC profiles of BL21(DE3) strain expressing r-BOX. As shown above, after the HPLC analysis of C4-C12 fatty acid standards and BL21_bktB-ydiI-fadB-ter, at the same location of the peak as the standard MCFAs, B represented the successful production of 1.08 g/L MCFAs by strain BL21_ bktB- ydiI- fadB-ter. So that our platform for producing MCFAs was successfully established.
Safety is always the most important during the experiment. Since doing the experiments, we have strictly followed the security policies required by iGEM. So in the whole experiment, safety has been completely guaranteed. Our experiments are conducted in the ATLATL laboratory, and the entire laboratory is strictly equipped with safety equipment to protect our safety. Besides, during our experiment, all materials we used are uncontaminated and pose little risk to our health. Thus the safety of our process of producing medium chain fatty acid can be ensured which will not cause any pollution. Regarding the safety of our product. The final product we eventually obtained will be purified and assessed until it reaches the food standard jus like what we can get in natural palm oil.
Challenges come from all aspects. As required by the biotechnology industry technical threshold is relatively high, new entrants will have to face the test of economies of scale, low input cannot guarantee the quality of products and high investment means the markets need a longer time to buffer, short-term returns cannot achieve the desired effect. At the same time, new entrants need to win the trust of customers and establish their brand image and their enterprises. Another challenge is to have a complete set of sales routes (from the production department to the sales department of the biotechnology company to the factories). At the national level, the biotechnology industry is also greatly affected by national policies, which will make the price of drugs and the technology for building factories more difficult for new entrants.
About our product development, we will further explore the optimizing condition of our engineered strain for higher production. Besides, we need to check with related food standards to ensure our product could be applied to cooking oil products and bulletproof coffee, more tests will be conducted in the future.
Per our investigation, MCFAs can be widely used in food additives, nutritional AIDS, cosmetics, and chemical materials. It can be seen that the prospect of our project is very good. In the future, we also plan to do a more detailed investigation to solidify our business plan, and cooperate with companies that are willing to help us to see how possible it is to put our technology into real production work.
