GLYCOSIDE PHOSPHORYLASES AS INDUSTRIAL GLYCOSYLATION BIOCATALYSTS

Biocatalysts are engineered for optimized conversion of the glucose part from sucrose into innovative functional glucosides.

The optimisation concept of CARBAFIN is to engineer flexible cell factories (preferred: Escherichia coli) for high-level expression of coupled phosphorylases to obtain whole-cell glycosylation biocatalysts sufficiently active and robust for use in the production processes of the target glucosides. Focus is put on enzyme engineering, strain optimisation and synthetic biology for balanced co-expression.

Partners involved:

ACIB, TUG, UGent

Lead contact:

Tom Desmet (UGent)

DIRECT OR INDIRECT GLYCOSYLATION TECHNOLOGY APPROACH

Single enzyme or enzyme cascade reactions are optimized in terms of product titer, space-time yield and catalyst turnover number.

Cell factories are produced by batch cultivations and are applied for biosynthesis in a non-living (non-growing) state to avoid use of reaction substrates for biomass formation, a main problem of fermentative processes. Using reaction optimisation guided by kinetic modelling, economically viable process metrics (yield, productivity etc.), which are implemented with the help of process-oriented LCA, are obtained. Furthermore, possibility of biocatalyst recycling supports transition from batch to continuous production processes.

Partners involved:

ACIB, TUG, P&L

Lead contact:

Chao Zhong (TUG)

TAILORED DOWNSTREAM PROCESSING OPERATIONS

Membrane and filtration technologies are evaluated and optimized.

Despite the variable chemical nature of glucoside products made by CARBAFIN technology, the DSP converges on the common task of removing fructose because yield and selectivity are both very high in the reactions. This is achieved by combination of different membrane technologies, resulting in a concentrated retentate from which the glucoside is ideally obtained by crystallisation, or alternatively by precipitation. Solid product might also be obtained simply by solvent (water) removal. The filtration permeate contains fructose as a highly valuable co-product. Although specific characteristics of each glucoside must be taken into account, the overall DSP is adaptable and therefore constitutes a second important platform development of CARBAFIN.

Partners involved:

ACIB, P&L, AVA, bitop

Lead contact:

Andreas Bilstein (bitop)

INTEGRATION OF UP- AND DOWNSTREAM PROCESSING PARTS

Biocatalysis is integrated with downstream processing to ensure cost efficiency.

The process integration work package connects up- and downstream technology for biocatalytic glucoside production and feeds fructose streams into AVA’s HMF production technology. CARBAFIN aims at development of a fully continuous production process but also repetitive batch production will be evaluated. Lab scale results will be transferred to demonstration activities in WP5 (Demonstration in industrial environment). Mass and energy-balance datasets are fed into environmental and economic hot spot analysis in WP6 (Process and product evaluation). The tight integration of the up- and downstream processing parts of the glucosylation process, guided by results from process oriented environmental and economic sustainability evaluations, is a hallmark of CARBAFIN’s holistic conceptual approach.

Partners involved:

ACIB, TUG, AVA

Lead contact:

Bernd Nidetzky (ACIB) 

BIO-BASED PRODUCTION AT DEMONSTRATION SCALE

Demonstration activities are carried out to prove techno-economic viability of the developed processes.

CARBAFIN aims at demonstrating the industrial and commercial feasibility of the cell factory-based glucosylation technology by bringing the biocatalytic production of two platform glucosides (e.g., glucosylglycerol and cellodextrin) and a fructose-derived base chemical (HMF) to a proven techno-economic viability. CARBAFIN will thus bridge the gap between lab and industrial scale for biocatalytic glucosylations. CARBAFIN will have the whole process development guided by environmental and economic analyses. The process technology CARBAFIN aims for is simple by design in order to facilitate process scale up and implementation in a fast and reliable fashion. Multipurpose stirred tank reactors (STRs) broadly available in the industry will be used in demonstrations without requirement of costly modifications. Integration of STRs with membrane filtration is also well established at demonstration scale or larger. Knowhow and technology for packed bed reactors is also available at industrial partner’s sites.

Partners involved:

P&L, AVA, bitop, GALAB

Lead contact:

Thomas Häßler (P&L) 

PROCESS DEVELOPMENT GUIDED BY ENVIRONMENTAL AND ECONOMIC ANALYSES

Evaluation of environmental impact and economic feasibility of developed processes and products will accompany, not just post-evaluate, the design of the process.

The economic feasibility of CARBAFIN processes and the competitiveness of CARBAFIN products in their respective markets will be evaluated by a techno-economic analysis. Moreover, each of the two main value chains (including fructose exploitation) will undergo process oriented environmental sustainability evaluations. The environmental hot spots will be identified and a reduction potential of the environmental impact will be illustrated. Evaluation results will guide improvement during process development and provide feedback to WP1-4. A combined final evaluation of optimized processes as best practice examples for bio-based chemical production will be done. Improved data sets will substantially assist future evaluation of bio-based products. CARBAFIN products will be evaluated regarding their purity, efficacy and fulfilment of regulatory benchmarks.

Partners involved:

ACIB, TUG, P&L, bitop, GALAB

Lead contact:

Barbara Petschacher (TUG)

Alexander Passer (TUG)

VALUE CHAINS OF TARGET GLUCOSIDES BENEFIT FROM FRUCTOSE

Next to our target glucosides (glucosylglycerol, cellodextrin) also the fructose part of sucrose will be exploited as substrate for hydroxymethylfurfural (HMF), a valuable platform chemical for the bio-based economy.

An important feature of CARBAFIN is that the cell factories developed in the project can be introduced and implemented directly in industrial operations, thus making them of immediate relevance for application by the company partners. An open-innovation strategy will be pursued in the development of platform cell factories and DSP technologies (to the extent applicable). Technology assessment for commercialisation and business plan development will be done case-specific. Marketing activities will be based on business plans. Outreach activities include trade fairs, promotional tours in and outside Europe, flyers, publication in scientific journals and at conferences, press releases, websites, technology offers but also more specific measures like stakeholder workshops with industry for replication (e.g. detergents sector), open days at the industrial partners to educate consumers on bio-based economy and educational events with schools.

Partners involved:

ACIB, TUG, P&L, AVA, UGent, bitop, GALAB, PNO

Lead contact:

Katrin Weinhandl (ACIB)

Martin Walpot (ACIB)

Christiane Luley (ACIB)

This work package sets out the 'ethics requirements' that the project must comply with.

Partners involved:

ACIB, AVA, bitop, GALAB, UGent, P&L, PNO, TUG

Lead contact:

Christiane Luley (ACIB)

This work package supports the integral management of CARBAFIN and enables and stimulates collaboration and efficient exchange of information between all partners and the Commission to ensure successful implementation of the proposed work plan. ACIB has the overall project management responsibility of monitoring the project progress and guiding the CARBAFIN consortium according to the EC requests. The management tasks and the contribution of all the partners are described as follows:

• Establishment of efficient overall project management and communication structure

• Organisation of project meetings

• Monitoring and ensuring of thorough documentation and progress control

• Financial guidance and controlling

• Continuous risk management

• Coordination of stakeholder advisory board

Partners involved:

ACIB, TUG, P&L, UGent, bitop (representing the coordinator and all work package leaders)

Lead contact:

Christiane Luley (ACIB)