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Disruptive platform for sustainable and low-cost amino acid production

Periodic Reporting for period 1 - BioBAA (Disruptive platform for sustainable and low-cost amino acid production)

Reporting period: 2020-12-01 to 2021-11-30

The BioBAA project is about achieving a more economical, more sustainable and environmentally friendly method for producing two amino acids that are used in the feed food and pharma industries. The proprietary technology developed by Cysbio consists of genetically modified bacterial strains that can make the desired biochemical products by simple fermentation from renewable carbon sources. The current commercial production processes are either less effective biobased methods, or they rely on fossil resource based raw materials and involves harsh chemical treatments with undesirable environmental impact. The Cysbio bacterial strains have high production yield and titers and will therefore lead to production methods that greatly reduces both production costs and the carbon footprint.

The objective of the BioBAA is to further improve the amino acid production strains to reach a production level in terms of yield and productivity and thereby lead to costs that allows entering new large markets within especially the feed and nutritional health sectors, thereby having a large economical, health and environmental impact on society. In this project, Cysbio is pursuing successful demonstration and documentation of the value and benefits provided using these amino acids primarily in the feed sector. The use of amino acids as nutritional supplements will also be explored. The data generated will be used for certification and regulatory approvals and will feed into the life cycle assessment analysis, documenting the socioeconomic and environmental impacts of the products and methods.
Push the limits of AA platform technology performance

So far, the main focus of the project has been to further improve the amino acid production platform with respect to sustainability and cost effectiveness. By using state of art metabolic engineering tools, Cysbio have further optimized the expression of the relevant biosynthetic pathways. We have also tuned other metabolic pathways in the cell which helped in enhancing the yields and titers of the amino acid production. These efforts have led to 10 % increase in yield and titer. We have also optimized the production process to further reduce production cost of another amino acid and make it more sustainable. We are filing more patent applications to further strengthen our proprietary technologies. We have also successfully developed antibiotic marker free strains to avoid the use of antibiotic resistant genes in the production strain. This will enable the use of the biomass for feed and agriculture applications thus closing the carbon loop (Figure 1). We will further publish the work in coming year to facilitate other research groups and companies for such approach. We have also examined and successfully demonstrated production of amino acids on crude industrial substrate such as glycerol which is a byproduct from biodiesel production. However, due to current increase in demand of glycerol for use in hand sanitizers and the subsequent increase in price of this raw material, we have tested and successfully demonstrated use of other cheap renewable carbon sources such as maltose (C-40) sugars and glucose syrup from corn starch production.

Together with our production partner we have developed downstream processes for purification of the targeted amino acids. These processes are cost effective and helps to reduce byproduct formation. This development has gone through several iterations and have taken more time than originally planned, but it has been pivotal to reach a sustainable and cost-effective production method. As a proof of concept, we have purified several kilos of all the targeted products with high purity, and the material is currently used for the efficacy and performance testing at different testing institutes in both UK and Denmark. Due to the longer than expected development times of these downstream processes, some of the work tasks in the project have been extended with up to 6 months.

Perform the evidence-based results of the AA-platform and AA’s feature

To validate amino acids for replacing less environmentally friendly feed ingredients in animal feed, a large-scale feed trial has been established through a contracting company using the purified amino acids from our production. The results will provide information on growth/feed value and health parameters. The trial will be performed to the EU required standards that ensure a quality allowing the data to be submissible as part of a feed registration dossier. In order to promote the nutraceutical applications of one of the products, we have collaborated with contract research company to establish practical formulations that can be used as nutritional supplements, the efficacy of which will be tested in clinical trials. A collaboration with a food testing institute has also been established to further assess the applicability of the products for various food additive uses, assessing functionality and likelihood of consumer acceptance. The initial results look promising, more detailed investigations will be available during 2022.

Ensure regulatory compliance of Cysbio process and products

A regulatory expert company has been engaged to provide the service ensuring a full REACH registration which will be needed to get the Cysbio amino acids on the market. The company has so far provided a roadmap for how to ensure REACH and for what is necessary to get a full feed and food registration for the amino acid products in Europe. The REACH registration work will be initiated when the pilot scale material is ready to be used for the registration. The quality test of the downstream purified products will also be initiated.
Due to the challenges faced in downstream purification of the amino acids the project is delayed by six months as many of the demonstration activities, regulatory approval and LCA analysis is dependent on purified pilot scale material that adequately represent the final products. With the granted extension, Cysbio is confident to achieve all the mentioned milestones by end of this project. We have already improved the performance of production strains through optimizations of pathways and fluxes towards the precursor. For further improvements we are establishing state of art omics and synthetic biology tools to enable identification of the unknown bottlenecks in the production strains and to rapidly engineer them to achieve a further boost in productivity and yield. As a result of this work, further 10 to 20 % improvements in yields are expected and this will translate into similar cost improvements. Cysbio therefore feels confident to be able to introduce potent and attractive feed additives that will benefit European farmers with a cost effective and more environmentally friendly alternative that can replace some of the currently used less attractive feed components. It is also very likely that the work will lead to new nutritional products that can be used to improve the health of many people, by addressing nutritional imbalances that affect up to 20 % of the global population. Overall, the broad use of the Cysbio products will contribute to reducing CO2 emissions and it will further reduce our society’s dependence on fossil resources.
Figure 1. Schematic representation of production of amino acids by Cysbio in a circular economy