2,4-dihydroxybutyric acid as a biomonomer precursor for new biocompatible and biodegradable polymers – POLYDHB

To meet the POLYDHB project's objective, complementary chemo-catalytic and biotechnological approaches were carried out in parallel, each constituting a separate work package and being overseen by a package comprising methods for the synthesis and production of the monomers of interest, i.e. the racemic mixture DHB, the L and D enantiomers DHB and the lactone derivative of DHB, namely the 2-hydroxy-?-butyrolactone (2-HBL). Overall, the project did not call for the development of new technologies, but made good use of the existing, well-mastered methods of the partner teams.

The major results obtained in this project were
• Chemocatalysis has demonstrated that the lactone form of DHB (2-HBL) can be incorporated at different rates with other monomers such as ?-caprolactone and lactide by ROP to produce co-polymers with novel physico chemical properties.
• The biotechnological approach has provided the first indications that DHB can be used in the enzymatic synthesis of polyhydroxyalkanoate-type polyesters
• While to date the fermentative process only produces the L- form of DHB, an enzymatic system capable of specifically producing the D-DHB form has been identified and characterized.

The partners are keen to pursue this project by broadening its scientific, technical and industrial ambitions. To this end, we have submitted a project as part of the ANR/DFG 2024 call concerning the production of DHB derivatives by biotechnological means and their use in (co)polymer synthesis by chemocatalysis. In parallel, we will be submitting a more ambitious project before the end of 2024 as part of the European «EIC Pathfinder 2024« call in 'Nature inspired alternatives for food packaging and films for agriculture' section, the aim of which will be to produce a wide range of biopolymers based on DHB and its derivatives and to test them for both their physical (solubility, degradability) and technological properties.

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At date, the project led to 2 publications on the use of DHB's lactone derivative (2-HBL) in the production of co-polymers with ?-caprolactone. A patent was filed for a technological improvement in the microbial DHB production method. The project also enabled two students to complete their doctoral theses, one in polymer chemistry and the other in microbial and enzymatic engineering. Finally, the project was disseminated through 6 posters in international meetings, 3 oral communications and 4 invited conferences.

One of the cornerstones of the 'Bioeconomy' will rest on our ability to exploit renewable carbon resources to produce environmental eco-friendly fuels and chemicals that will profitably replace those derived at present from fossil resources. The POLYDHB project is in frame with this endeavour. This project originality stands on previous works carried out by two partners of this proposal which have exploited synthetic biology toolbox combined to metabolic and enzymes engineering to construct a synthetic pathway that leads to the microbial production of a non-naturally metabolite 2,4-dihydroxybutyric acid (DHB) from renewable carbon sources (i.e. sugars). Initially conceived as a precursor for the synthesis of the methionine to target the field of animal nutrition, this molecule actually turns out to be a unique ‘green’ platform chemical for the production of other bio-based products with application in chemical and pharmaceutical industries.
The purpose of the POLYDHB project is here to demonstrate that DHB can be used as an original non-natural monomer for the production of new bio-sourced and biodegradable polymers. The scientific and technical challenges of this project will be realized through three workpackages: (I) production of pure enantiomers and lactide /lactone derived from DHB, (II) development of a chemocatalytic process of (co)polymerisation of DHB and/or its lactone and lactide derivatives alone or with other monomers, and (III) conception of a microbial process for the synthesis of DHB-based polymers. In each of these workpackages, scientific risks have been identified and contingency solutions clearly proposed. To succeed in this objective, a multidisciplinary and complementary core of expert in the field of Systems and Synthetic Biology (LISBP, Toulouse), Polymer Chemistry (LCPO, Bordeaux) with the participation of a Japanese team expert in molecular biology of bio-sourced polymers has been set-up. Moreover, the strong commitment of the industrial partner Adisseo in this project is not solely justified by its indispensable position in mastering the chemical and microbial process for DHB production, but it is also an asset for the industrial exploitation of this molecule on markets other than nutrition animal that can be opened from the results obtained in this project. The ambition of POLYDHB project is to reach the technology readiness level of 4 within the 3 years period.