Final Report Summary - SYNPOL (Biopolymers from syngas fermentation)
Executive Summary:
Every day we waste money in transporting municipal solid waste (MSW) miles out of cities or states to be landfilled. Long term environmental and economic problems created by landfilling prove that it is not the solution to the waste problem, and now waste conversion technologies are becoming a massive growth industry worldwide. In this context, the significance of the SYNPOL project was remarkable considering the high amount of available relevant waste produced solely in Europe every year.
Complex organic wastes such as municipal and agricultural residues, which were firstly pyrolyzed to produce synthesis gas (“syngas”) and then fermented by microorganisms, were the starting feedstock of the SYNPOL project which aimed at the transformation of wastes into 100% biodegradable bioplastic materials and further chemical building block compounds (e.g. butanediol, succinic acid, hydroxybutyric acid and crotonic acid).
Syngas fermentation is already an attractive technology for the production of biofuels and in fact several industrial processes for ethanol production from syngas are now under development, but these promising industrial processes have not been implemented in Europe yet and this is the challenge that SYNPOL was addressing. SYNPOL´s syngas fermentation technology helped to open a new window for the rational design of an innovative process to convert complex wastes into new biopolymers and other high added value compounds.
SYNPOL has done this waste conversion in three major steps:
• Pyrolysis of different organic waste streams to produce syngas (CO, H2 and CO2)
• Fermentation of syngas by using different bacteria to produce chemical building blocks and polyhydroxyalkanoates (PHA)
• Synthesis of bio-based plastic prototypes with well-defined structures and improved properties for wide commercial use via chemical and enzymatic catalysis by utilizing the monomers and polymers produced during syngas fermentation
Novel processing technologies, like the combination of novel microwave induced pyrolysis (MIP) of wastes with improved syngas fermentation bioreactors, are additional relevant outcomes that have been derived from SYNPOL. To fulfill its aims, SYNPOL established an integrated platform for biopolymers production reducing energy input and optimizing purification of waste streams, thus contributing to the economic viability of the whole process.
Specific main objectives were defined and carried out to achieve the above-mentioned general objectives: i) Waste samples preparation as feedstock for pyrolysis and application of innovative gasification methods to produce syngas containing high percentages of CO and H2, ii) Improvement of bacterial strains for syngas fermentation by genetic and metabolic engineering, iii) Development of novel fermentation design for effective and safe syngas fermentations at lab-scale, iv) Development of biopolymer design by characterizing novel biopolymers/copolyesters from chemical catalysis and synthesis of novel biopolymer blends with PHAs, plasticizers and nanoclays using building blocks produced from syngas fermentations, v) Biopolymer degradation and life cycle assessment (LCA), and vi) Demonstration of the success of the SYNPOL platform technologies by presenting three novel prototype biopolymers for different industrial applications and a newly developed biopolymer production plant design taking into account the best scale-up of fermentation and downstream processes.
Summarizing, SYNPOL has linked the notion of waste valorization with representative members of the European bioindustries which are distinguished for their interest in the development of biotechnological processes to produce biopolymers by using biowastes as starting materials. The ultimate scope of SYNPOL was to establish a win-win situation between these bioindustries and the polymer industry, as a branch of the chemical industry.
Project Context and Objectives:
Too much waste around us
There are many waste resources hidden in our communities. For example, municipal solid waste (MSW), agricultural residues and sewage sludge from water treatment plants contain lots of reusable carbon fractions. An eco-efficient waste conversion process means recovering most of the waste as valuable products in an environmentally friendly manner. In this context, the significance of the SYNPOL project is remarkable considering the high amount of available relevant waste produced solely in Europe every year (approx. 261million tons (MT) of MSW including approx. 25 MT of plastics, approx. 120 MT of agricultural residues (e.g. straw) and >10 MT of sewage sludge). [1-4]
The basic idea and work flow scheme of the project is presented in Figure 1.
Polyhydroxyalkanoates (PHA): an alternative for petrol-based plastics
Polyhydroxyalkanoates (PHA) are a well-known family of polyesters accumulated as cell granules by naturally occurring microorganisms serving as carbon and energy reserve. Figure 2 shows bacterial cells containing PHA granules and some short chain length PHA isolated from bacteria. Many different PHAs of diverse desired properties including biodegradability can be produced from renewable resources by the biosynthetic action of selected prokaryotes. Currently, PHAs are one of the alternatives to petrol-based plastics, elastomers or latexes.
Non-food competing feedstock materials
The need for alternative materials, because of the finite sources of fossil reserves is generally accepted. In order to become a competitive alternative on the market, the price of bioplastics for a certain application must be in the same range as the competing petroleum-based plastic. Hence, the costs of PHAs have to be reduced considerably despite the current unstable price of crude mineral oil. As an alternative solution, diverse waste streams exist which currently constitute severe disposal problems and, at the same time, do not interfere with the nutrition chain. This is especially true for the daily produced household waste that ends up in landfills or the sewage sludge loadings from water treatment plants. The utilization of these waste streams is a viable strategy to overcome a potential ethical conflict. Indeed, it can be considered as the most promising approach in making PHAs economically more competitive.
Philosophy, roadmap and main objectives
The SYNPOL project aimed at converting these waste streams via refined gasification processes (pyrolysis supported by innovative microwave technology) into synthesis gas (syngas) that contains high amounts of carbon monoxide (CO) and hydrogen (H2). These gaseous carbon fractions were biotechnologically converted towards PHA biopolymers and further chemical building block compounds by applying bacterial fermentation processes using different bacterial strains. The selected bacterial strains were naturally occurring microorganisms that were optimized for higher biopolymer production efficiency by genetic and metabolic engineering techniques based on the newest findings from systems biology.
The project brought together public research centers with waste producers from different processing industries (landfilling, agricultural and water treatment industries), gasification industry and polymer processing biotechnological companies. This synergism was benefiting all players and resulted in new value creations. The project´s time roadmap is illustrated in Figure 3 and summarizes the main working lines of the consortium.
These working lines were created to cover the following main objectives of the project:
1. Optimize syngas production from different feedstocks by using new gasification technologies.
2. Characterize syngas generated from feedstocks via a combination of state-of-the art techniques.
3. Analyse energy costs required to generate syngas from different feedstocks.
4. Enhance naturally occurring pathways of acetogenic bacteria by metabolic engineering supported by systems biology tools with the aim of generating by syngas fermentation large amounts of intermediates that can be used as building blocks for the synthesis of new biopolymers.
5. Design new metabolic pathways by using systems biology tools to produce polyhydroxyalkanoates in acetogenic bacteria.
6. Enhance naturally occurring pathways of purple bacteria by metabolic engineering, again supported by systems biology to generate by syngas fermentation large amounts of polyhydroxyalkanoates.
7. Design transferable genetic systems to confer the ability to metabolize CO to other bacteria under aerobic conditions.
8. Design new recombinant bacteria endowed with programmed autolytic systems to facilitate the downstream processing of biopolymers.
9. Establish a cost-effective consolidated syngas fermentation technology using the natural and recombinant microorganisms by improving the operational conditions.
10. Integrate interdisciplinary knowledge to consolidate an efficient downstream processing, to decrease energy inputs, reduce environmental impacts, increase purification efficiency and diminish the final cost of the products.
11. Develop cost effective technologies for the chemical synthesis of new biopolymers based on the “green” building blocks and the polyhydroxyalkanoates produced by syngas fermentation, utilizing state-of-the art of chemical processes.
12. Develop blend, plasticizer, and composite formulations based on a PHA matrix using fermented and waste stream products.
13. Reduce the environmental impact of the integrated SYNPOL technology by recycling the residues.
14. Demonstrate the chemical and organic recyclability of the developed biopolymers.
15. Develop life cycle and environmental impact analyses as a valuable tool to provide technical and economic advances in the development of environmental friendly biopolymers.
16. Plant production design, scale-up and optimization of consolidated SYNPOL technology targeting on the increase of the products yield, improving the overall production economics as well as the upscaling capabilities.
17. Establish portfolios for the exploitation of the process and the products derived from SYNPOL project.
18. Disseminate the scientific knowledge acquired during SYNPOL project to the society by developing multiple training and diffusion activities.
A project of great expectations
From the perspective of the SYNPOL project, important progress was achieved in terms of combining the environmental benefit of future-oriented biopolymers with the economic viability of their production. This should finally facilitate the decision of responsible policy-makers from the waste-generating industrial sectors and from the polymer industry to break new ground in sustainable production. In the future, PHA production from different waste streams applying gasification technology should be integrated into existing process lines of biotechnological bioplastic companies, where the feedstock material directly accrues. By taking profit of synergistic effects, this can be considered a viable strategy to minimize production costs. The 48 month SYNPOL project has secured almost 7.5 million € in funding under the Food, Agriculture and Fisheries, and Biotechnology theme of the European Union Seventh Framework Program (FP7/2007-2013; grant agreement nº 311815). The project was launched in October 2012 and ended in September 2016 and was coordinated by the Biological Research Center (CIB) in Madrid (Spain) which is part of the Spanish National Research Council (CSIC).
The project team players:
From Industry
▪▪ Biopolis S. L. (Spain) – Industrial fermentation
▪▪ Bioplastech Ltd. (Ireland) – Biopolymer synthesis
▪▪ Organic Waste Systems NV (Belgium) – Biodegradation & LCA analysis
▪▪ Bionet Servicios Técnicos S. L. (Spain) – Simulations & Pilot plant design
▪▪ Infors AG (Switzerland) – Fermentation development & Bioreactor design
▪▪ Befesa Gestión de Residuos Industriales S. L., later substituted by Abengoa Research (Spain) – Waste management
From Academia
▪▪ Consejo Superior de Investigaciones Científicas (Spain) – Project coordination, Syngas production, Bacterial Research & Downstream processing
▪▪ University of Manchester (United Kingdom) – Bacterial Systems Biology
▪▪ University of Amsterdam (The Netherlands) – Bacterial Systems Biology
▪▪ Universität Ulm (Germany) – Bacterial fermentation & Recombinant strains
▪▪ University College Dublin (Ireland) – Proteomics & Molecular biology
▪▪ Haute Ecole Spécialisée de Suisse Occidentale (Switzerland) – Fermentation reactor design & Downstream processing
▪▪ Kungliga Tekniska Högskolan (Sweden) – Biopolymer synthesis
▪▪ Westfälische Wilhelms-Universität Münster (Germany) - Bacterial fermentation & Recombinant strains
▪▪ Université de Strasbourg (France) – Biopolymer design
References:
[1] http://bit.ly/12ZOjgs
[2] http://bit.ly/12kWe1y
[3] http://bit.ly/1351OwK
[4] http://bit.ly/12ZOQPy
Project Results:
The most outstanding non-confidential S&T results obtained in the course of the SYNPOL project are described, with a mention of those already disseminated by scientific publications (and other means).
The following sections include a description of the results and achievements in the 5 RTD work packages and in the Demonstration work package conforming the SYNPOL project scheme (Fig. 1) and work plan.
Results of the works on syngas production from organic wastes:
First of all, it was necessary to identify appropriate feedstock for the pyrolysis process and to determine their characteristics and compositions, since the efficiency of the global process will be affected by these parameters. The different waste samples (municipal solid waste, agricultural residues and sewage sludge) have been characterized for their use as feedstock for subsequent pyrolysis by application of standardized methods.
The following general results have been obtained:
- A major conclusion of these experiments was that the best results (highest syngas production and with a high CO + H2 content) have been obtained with agricultural residues (straw) and with the organic fractions of the municipal solid waste as feedstock for pyrolysis. From an environmental point of view, municipal solid wastes face an important challenge, such as the need for new technologies to manage them through valorisation strategies. Hence, this substrate has been selected to study the process of microwave-induced pyrolysis to produce syngas.
- Concerning the syngas production and composition, it can be concluded that the results obtained in the microwave-induced pyrolysis are clearly superior to those obtained in the conventional pyrolysis.
- Since the organic residues are poor microwave absorbers, it was necessary to use a microwave receptor, which could be directly heated by microwaves and so heating the biomass by conduction, convection and radiation processes. The best material to play this role is the carbonaceous residue obtained in a previous pyrolysis. A minimum microwave receptor-to-waste ratio of 0.2:1 is needed to induce the pyrolysis.
- Higher microwave power favours the production of syngas but this increases the energy consumption.
- Depending on the initial moisture content and pyrolysis time, it is possible to obtain different syngas compositions. The moisture content was found to improve the volumetric gas production by almost 50% at the expense of the energy consumption needed for the water evaporation. Nevertheless, 10-20 wt% of moisture in waste enhance the microwave absorption by biomass and allows for a maximum production of syngas.
- The use of different carrier gases (N2, He) affects the yields of the fractions obtained during the pyrolysis of organic residues only slightly. In the case of microwave pyrolysis, and especially at high temperature, no significant difference in the gas composition was observed when using different carriers. Furthermore, it was observed that the implementation of microwave pyrolysis can be overcome without the need for a carrier gas.
- Different experiments on CO2-reactive atmosphere were conducted. The syngas concentration can be slightly improved due to the partial gasification of the carbonaceous residue.
- Microwave pyrolysis leads to a reduction in the liquid fraction production. It has been found that hazardous PAHs production from heterogeneous wastes (organic fraction from municipal solid wastes and sewage sludge) can be reduced by microwave pyrolysis.
-The scale up parameters of the microwave induced pyrolysis (MIP) process were established as follows:
✓ The organic fraction of Municipal Solid Waste (MSW) is a good substrate (best among all the residues studied) to produce high yields of syngas (up to 90 vol% of the gas fraction).
✓ Temperature about 800ºC is the best option to balance maximum gas production/minimum energy consumption.
✓ A ratio of microwave susceptor/substrate of about 0.2:1 is the best option. Below this ratio is almost impossible to reach the 800ºC (lots of energy consumed), but higher ratios impair penetration of microwaves resulting in a worse homogeneity.
✓ Certain amount of moisture is not bad since it helps to the heating in the first stages and contributes to gasification. Substrates with about 12 wt% of moisture are a good choice.
✓ It is possible to carry out MIP without using any carrier gas. In any case the influence of the carrier gas is minimal.
✓ The energy consumed by the MIP of MSW is estimated to be about 1.5 kW/kg.
Given that the solid fraction (chars) obtained in MIP is of low quality for using it as a fuel (high inorganic matter content containing heavy metals) it was studied the possibility of use this fraction as soil amendment. The maximum amount of char that can be used in soils was determined by means of standard ecotoxicity tests.
Given that the tars produced in the MIP process need to be upgraded (by removing its moisture, heavy metals and other undesirable compounds and reducing its oxygen content) before to be used as fuel; a new concept of pyrolysis that does not produce oils was developed, tested at laboratory scale and patented.
These results have been published and are accessible at:
http://dx.doi.org/10.1016/j.jaap.2013.11.009
http://dx.doi.org/10.1016/j.fuel.2014.04.064
http://dx.doi.org/10.1016/j.jaap.2014.12.005
http://dx.doi.org/10.1016/j.jaap.2014.12.011
http://dx.doi.org/10.1016/j.cep.2015.05.001
http://dx.doi.org/doi:10.1016/j.jaap.2015.06.006
http://dx.doi.org/10.1016/j.fuel.2016.01.016
http://dx.doi.org/10.1016/j.jaap.2016.08.013
http://dx.doi.org/10.1111/1751-7915.12411
http://dx.doi.org/10.1016/j.biortech.2016.07.019
http://link.springer.com/chapter/10.1007/978-94-017-9612-5_6
Results of the works on improvement of microorganisms for syngas fermentation:
This working field aimed to i) analyse the capacity of various bacteria to ferment syngas, ii) create genetically modified strains, iii) transfer the capacity of using CO as a carbon source to aerobic bacteria, and iv) find generic ways to improve this type of engineering.
The following general results have been obtained:
Systems Biology studies:
In Systems Biology studies, a general Rhodospirillum metabolic model was developed by applying an existing metabolic map for the three bacterial species Rhodospirillum rubrum, Rhodobacter sphaeroides and Rhodopseudomonas palustris, and by adapting it to the special case of R. rubrum using syngas as carbon and energy source. The results obtained will help in the near future to adjust bioreactor fermentation processes (e.g. metabolic fluxes of key substrates such as CO, CO2 and H2) more precisely towards the desired end product.
In addition to the reconstructions of metabolic maps for C. ljungdahlii and R. rubrum further genome-wide reconstructions for Acetobacterium woodii, Clostridium acetobutylicum, Clostridium cellulovorans and Oligotropha carboxidovorans were produced in the project.
Studies with bacteria of the genus Clostridium:
The wild type bacterial strain Clostridium ljungdahlii showed the highest 2,3-butanediol production on syngas, therefore the genes for 2,3-butanediol synthesis starting from pyruvate alsS (acetolactate synthase), budA (acetolactate decarboxylase), and 2,3bdh (2,3-butanediol dehydrogenase) of this organism were chosen to create two 2,3-butanediol overexpression plasmids. The overexpression of the three genes alsS, budA, and 2,3bdh improved the 2,3-butanediol production in some other model clostridia strains.
Additionally it was shown that succinate formation can be naturally achieved with C. aceticum and C. ljungdahlii.
Studies with bacteria of the genus Rhodospirillum:
While the wild type strain R. rubrum S1 accumulated a maximum of 26% of cell dry weight (cdw) polyhydroxybutyrate (PHB), the complemented (recombinant) strain R. rubrum ∆phaC2 pBBR1MCS-2::phaC2R. rubrum showed the highest PHB synthesis of all tested strains with 32% PHB (cdw). Thus, recombinant R. rubrum strains accumulated up to 25 % more PHB than the wild type strain.
A R. rubrum strain able to synthesize solely medium chain length PHA (PHAMCL) from an unrelated carbon source has been generated. It is potentially the first microorganism able to accumulate these polyesters from syngas. Recombinant R. rubrum ΔphaC1ΔphaC2 pBBR1MCS-2::phaGPp::phaCPp accumulated a total amount of 2.5 % (w/w of cell dry weight) of PHAMCL. No other PHAs, as for example PHB, were synthesized by this recombinant strain. Currently, further work focuses on increasing the amount of synthesized PHAMCL.
R. rubrum S1 was successfully grown with syngas (Fig. 4) and a second carbon source under anaerobic conditions in the dark. In order to increase the carbon flux towards PHB production, the effect of syngas fermentation and PHB accumulation on central metabolism of R. rubrum has been analyzed by using transcriptomic approaches such as reverse-transcription quantitative PCR. Results show an activity increase of more than four times in CO-metabolic genes of R. rubrum independently of the carbon source used (malate or acetate), thus demonstrating the stimulating effect of the CO from syngas.
R. rubrum was successfully grown with syngas and poly(3HB-co-3HV) was formed from syngas by recombinant strains.
Different media and carbon sources were tested and optimal conditions for aerobic growth of R. rubrum S1 established. The addition of acetate to the cells grown on malate and fructose improved PHB yields in R. rubrum 3-fold and 8-fold, respectively.
Proteomic analysis of aerobically grown Rhodospirillum rubrum S1 cells generated 2077 high-confidence (≥0.99 Protein Prophet score) protein identifications. The number of exclusive proteins detected in fructose/acetate and in malate/acetate grown cells was only 33 and 21, respectively. These proteins are currently investigated in more detail and will be compared to anaerobically grown R. rubrum cells.
Based on the comparison of the proteome of R. rubrum grown on different substrates under PHB accumulating conditions, a potential target protein and pathway possibly affecting PHB accumulation was identified. Additionally, to completely understand the physiology of PHB accumulation and the natural role of this biopolymer in R. rubrum, a deletion mutant with impaired PHB accumulation is actually investigated.
Studies with bacteria of the genus Oligotropha and Ralstonia:
The genome of Oligotropha carboxidovorans OM5 encodes two potential PHA synthases with the locus tags OCAR_4862 and OCAR_6012. RT-PCR studies revealed the active expression of the PHA synthase gen of the locus tag OCAR_6012. This result is important for the general target to genetically engineer O. carboxidovorans to accumulate a higher amount of PHB than the wild type strain. Further cloning and expression studies are currently on the way to achieve this target.
Overexpression of the phaC gene from O. carboxidovorans apparently leads to a significant increase in accumulated PHB.
Three genes encoding mcl-PHA synthesis related enzymes in pseudomonades were heterologously expressed in O. carboxidovorans. Since the efforts to create a PHB negative mutant of O. carboxidovorans were unsuccessful, it was not possible to generate a strain which would accumulate solely mcl-PHA. O. carboxidovorans pBBR1MCS-2::phaGPp::phaC1Pp::pp0763Pp expressing Pseudomonas putida KT2440 genes phaG, phaC1 and PP_0763 accumulated traces of polyhydroxydecanoate (PHD) when cultivated in NB medium with added sodium acetate.
Transfer of the capacity of using CO as a carbon source to aerobic bacteria:
Design and construction of a cox cassette (a genetic cassette including the genes necessary to use CO as carbon source) in a broad-host range plasmid (pBBR1-cox) that can be transferred to different bacterial hosts. In that way, a newly created recombinant Ralstonia eutropha strain H16-cox (containing the transferrable cox cassette) was able to grow with CO as sole carbon source. Without this cassette the strain is not able to use CO as substrate. This demonstrated that the cox cassette is functional. The cox cassette is the first genetic device that allows the expansion of the catabolic potential of some bacteria to use CO as sole carbon and energy source.
Additionally, identification and study of the transcriptional promoters and regulators of the cox cluster from O. carboxidovorans OM5 are driven further to design and construct a new CO biosensor.
Part of these results have already been published and are accessible at:
http://dx.doi.org/10.3389/fmicb.2014.00379
http://dx.doi.org/10.1093/femsle/fnv038
http://dx.doi.org/10.1016/j.copbio.2015.03.008
http://dx.doi.org/10.1002/jctb.4721
http://dx.doi.org/10.15255/CABEQ.2014.2249
http://dx.doi.org/10.1111/1462-2920.13087
http://dx.doi.org/10.1038/srep24381
http://dx.doi.org/10.1111/1751-7915.12409
http://dx.doi.org/10.1111/1751-7915.12393
http://dx.doi.org/10.1111/1751-7915.12411
http://dx.doi.org/10.1016/j.mimet.2016.10.003
http://dx.doi.org/10.1007/s00253-016-7711-5
http://dx.doi.org/10.1128/AEM.01744-16
http://dx.doi.org/10.1007/8623_2015_168
Results of the works on fermentation design:
These works were addressed to increase the efficiency of the bioconversion of syngas produced into two types of bioproducts: building blocks such as succinate, butanediol and hydroxybuturate and biopolymers such as polyhydroxyalkanoates (PHA). These works were therefore focused in the development of different bioreactor configurations and operational conditions for the anaerobic fermentation of syngas using two different types of microorganisms Clostridium and Rhodospirillum. To orient and optimize the operational conditions we used also the recommendations provided by the results from Systems Biology analyses.
The following general results have been obtained:
- A safe lab zone environment (including fume hoods and gas detectors) for fermentations with syngas was designed and implemented (Fig. 5). Two Labfors 5 bioreactors (3.6 and 13 L) were built and equipped with gas analyzer and operating systems for fermentations with R. rubrum. Standardized working procedures for fermenting with syngas were established. Later progress was achieved with respect to fermentation infrastructure (e.g. characterization of gas transfer, cleaning in place), the fermentation itself (growth on synthetic acetate containing medium), and fermentation analytics (e.g. online PHB quantification, determination of dissolved CO).
- A dual-stain method for measuring PHA in the bacterial cultures by use of the flow-cytometer was established.
- SYNPOL partners demonstrated the need to add acetate to the cultivation medium to achieve PHB accumulation during syngas fermentation with R. rubrum anaerobically in the dark. With acetate, the PHA content reached up to 27% CDW (cellular dry weight) at early stationary phase and was composed of mainly PHB with traces of PHV. Yeast extract addition is not required for growing of R. rubrum in syngas under the same conditions.
- SYNPOL provided a basic engineering design for a syngas simulator, including a pre-treatment unit allowing syngas conditioning regarding the requirements of subsequent syngas fermentation processes.
- The fermentation with a recombinant Clostridium strain resulted in a first production of 0.1 g x L-1 2,3-butanediol in addition to 0.75 g x L-1 ethanol.
- Very important results were found for the biosynthesis of poly(3-hydroxyalkanoate) by R. rubrum from syngas. A detailed study carried out revealed that a direct production of PHB from syngas. Nevertheless, for a successful PHB production acetate needs to be added. Moreover, syngas assimilation into biomass and PHB is demonstrated by metabolomic analyses.
Part of these results have already been published and are accessible at:
http://ac.els-cdn.com/S187167841631576X/dx.doi.org/10.1016/j.nbt.2016.06.793
http://dx.doi.org/10.1016/j.mimet.2016.10.003
Results of the works on biopolymer design:
These works had the key objective to synthesize biopolymers with well-defined structures and improved properties by utilizing the monomers and polymers created during syngas fermentation by using chemical and enzymatic catalysis. For the biopolymer synthesis two different approaches were utilized; firstly polymerization of the monomers generated in the SYNPOL fermentation processes and secondly polymerization of functional monomers and oligomers created through chemical recycling of PHAs. These oligomers were subsequently homo- or copolymerized into new materials, minimizing the need of virgin chemicals. In addition, the inherent PHA properties were improved through plasticization, blending, and by creating composites. Polymers with new properties were obtained while maintaining the green character to target new applications and markets.
The following general results have been obtained:
- Successful syntheses of the following three different homopolyesters and one copolyester was conducted by SYNPOL partners: poly(butylene succinate), PBS; poly(propylene succinate), PPS; poly(butylene adipate), PBA; and poly(butylene succinate-co-butylene adipate), PBSA. They were performed by a two-stage (esterification and trans-esterification) melt polycondensation catalyzed by titanium (IV) isopropoxide. The characteristics of these polyesters were shown. For example, PBSA copolyesters have very different properties (melting temperature, glass transition temperature [Tg], crystallinity, mechanical properties, biodegradability, etc.) depending on the succinic acid:adipic acid (SA:AA) molar ratio in the reactor. Solubility tests showed that the polyesters synthesized (PBS, PBA, PPS and PBSA) are only soluble at a concentration of 20 mg/mL in few halogenated solvents such as chloroform, dichloromethane and hexafluoropropan-2-ol (HFIP). PPS, PBA and PBSA [50:50] are also soluble in 1,2-dichlorobenzene.
- SYNPOL partner have successfully synthesized poly-and oligoesters with different combinations of building block monomers such as 1,4-butanediol, adipic acid, and succinic acid by utilizing both a green organic catalyst (1,5,7-Triazabicyclo[4.4.0]dec-5-ene) and a conventional catalyst (titanium isopropoxide). The kinetics of the homo- and copolymerizations showed a typical two-step behavior and the desired molecular weight range could be achieved using both catalysts. Based on the kinetics PBA, PBS, and PBSA with two different theoretical molecular weights i.e. 1000 and 2000 g/mol were synthesized.
- In addition, the plasticization efficiency was evaluated in PVC blends. Good plasticization effect for PBA and PBSA as indicated both by the decrease in Tg and increased elongation-at-break was determined. The order of increased plasticization effect was PBS << PBSA < PBA.
- In other studies, the molecular weight, dispersity and thermal properties of acrylic acid (AA) and crotonic acid (CA) grafted PLA and PVC materials has been determined. It is worth to mention here, that for the PVC-based materials the strain-at-break increased more than 25 times the value for the grafted polymers as compared to the neat PVC although the Tg was virtually unchanged. This is highly interesting and unusual since the maximum usage temperature for an amorphous material is generally dictated by the materials Tg.
- Additionally, we determined the mechanical properties and glass transition temperature (Tg) for the PVC films with anchored plasticizers using either grafted AA or CA as anchoring sites and PBSA as plasticizer. All materials displayed smooth surfaces indicating homogenous and amorphous materials that can be used for different applications.
- The same partner has completed at the end of the project the full characterization of previously synthesized poly(butylene succinate-co-adipate) (PBSA) grafted PLA (PLA-g-PBSA) and the physical blends of PLA/PBSA. The grafted materials illustrated promising mechanical properties, crystallization behavior and improved migration resistance.
- Finally, this partner developed a gradient NMR method to give final proof of the successful grafting of PHB oligomers into PLA chains.
The study of PHA plasticized with different plasticizers with the aim of understanding the influence of the type of plasticizer and its concentration in the formulation over the final properties of the material was also carried out. Poly-3-hydroxybutyrate-co-4-hydroxybutyrate (P(3HB-co-4HB) was chosen as model PHA for its interesting mechanical properties and wide range of applications including biomedical and drug delivery systems. Three commercial plasticizers were studied for the P(3HB-co-4HB) matrix: Citroflex A4, Polysorb ID37, and Grindsted Soft-n-Safe at various compositions. The chemical nature of the plasticizers was found to affect the polymer-plasticizer interactions and hence, the final properties of the material. Interestingly, the long chain fatty acid moieties of both bio-based plasticizers Polysorb ID37 and Soft-n-Safe Danisco were found to provide better interactions with the polymer and therefore result in better plasticization as observed from the tensile strength studies and negligible weight loss at constant temperature. The Grindsted Soft-n-Safe was slightly better than Polysorb ID37 given its higher temperatures of degradation that allow minimal loss in degradation of P(3HB-co-4HB) during processing. Citroflex A4 was found to be least effective of the three plasticizers as it made the PHA susceptible to degradation at lower temperatures and also resulted in phase separation and exudation upon ageing (with heating). The preferred compositions for the plasticizers were found to be 13% for Polysorb ID37 and Citroflex A4 while for Soft-n-Safe Danisco the 13% and 17% compositions were both very good.
In order to evaluate various bio-based polymers on the mechanical and barrier properties and biodegradation of commercial PHB polymers, one industrial SYNPOL partner prepared blends with polycaprolactone (PCL), polylactic acid (PLA) and mcl-PHA produced by Bioplastech Ltd. company (Dublin, Ireland). Blends prepared with 50:50 PLA and Biomer PHB showed maximum improvement of the mechanical properties. In particular the % strain for 50:50 PLA/PHB blend increased to 232% compared to pure PHB (24%) and pure PLA (12%).
The same SYNPOL partner also evaluated the effect of various nucleating agents on the mechanical and barrier properties of PHB. Selected nucleating agents were melt-processed with the commercial PHB ENMAT Y100. The initial results indicated that addition of bio-based polymers such as mcl-PHA and PLA would improve the mechanical properties of PHB in particular % strain which is one of critical parameter to produce packaging films.
One SYNPOL partner developed PHB-PLA blends by melt mixing that showed drastically improved mechanical properties when plasticized by acetyl tributyl citrate. Plasticized PHB-PLA blends properties could be improved even more by addition of nano-crystalline cellulose or organo-modified nanoclay.
Later in the project, one SYNPOL partner successfully synthesized copolyesters and terpolyesters from SYNPOL building blocks using both organometallic and enzymatic catalysts.
Another partner has used SYNPOL building block crotonic acid obtained from PHB recycling to graft biopolyesters segments on PLA. In addition, the oligo-PHB building blocks from PHB recycling/degradation were successfully used as grafted plasticizer for PLA.
We also investigated the efficiency of a bio-based plasticizer for PHB compared to non-renewable analogues. In order to improve PHB properties, a SYNPOL partner prepared nanocomposites with various types of nanofillers.
One SYNPOL partner synthesized copolyesters from SYNPOL building blocks (adipic acid, 1,4-butanediol and 2,3-butanediol) using both organometallic and enzymatic catalysis. Despite lower reactivity of 2,3 BDO, optimized polymerization conditions allow to control and tune copolyesters composition and properties.
PHB diol oligomers were successfully used as building blocks to synthesize copolyesters and poly(ester urethanes) using both organometallic and enzymatic catalysis and it was demonstrated the possibility to control copolyesters structure (random or blocky tendency) and properties according to the PHB diol length and polymerization conditions.
Finally, PHA-based nanocomposites based on montmorillonite and sepiolite clays were elaborated and organo-modified sepiolite gave the best mechanical properties as a result of improved dispersion and strong interactions with the matrix.
Part of these results have already been published and are accessible at:
http://dx.doi.org/10.1021/sc500397h
http://dx.doi.org/10.1021/acs.macromol.5b00235
http://dx.doi.org/10.1002/mame.201500026
http://dx.doi.org/10.3390/ma9050313
http://dx.doi.org/10.1016/j.polymer.2016.07.022
http://dx.doi.org/10.1021/acs.biomac.6b01494
Results of the works on biopolymer degradation and life cycle:
These studies analyzed the degradation of the polymers synthesized in the SYNPOL project using different approaches, taking into account environmental conditions as well as the use of enzymes or organisms that could be useful for the biodegradation and/or recycling. Further works studied the development of processes for chemical and biological recycling of biopolymers for a waste-free society.
The following general results have been obtained:
- In a task called “Chemical recycling of PHAs to functional building blocks”, an effective and rapid process was developed for the chemical recycling of PHB through microwave assisted reactions in alkaline methanol, which resulted mainly in the monomeric degradation products, 3-hydroxubutyric acid and 3-methoxybutyric acid. The most effective and efficient microwave-assisted degradation conditions were obtained in 0.5% (w/w) alkaline methanol solution heated at 110°C. Additionally, PHB has been thermally degraded to crotonic acid and oligomers with crotonic acid end groups.
- In the same task, a potential route for feedstock recycling of PHB and a highly promising approach for fully biobased and biodegradable toughened PLA through covalently anchored oligomeric PHB plasticizers has been developed and demonstrated. This process consisted of a commercially viable and green two step extrusion process. PLA with 20% (w/w) grafted dPHB demonstrated an impressive elongation at break of 538%, 66 times higher than that of pure PLA.
- Biodegradation tests within the task “Biodegradability, organic recycling and ecotoxicity evaluation of developed biopolymers” were started on materials delivered by SYNPOL partners. The materials consisted of PHA mixed with nanoclays. Therefore, the degradation of neat PHA, PHA + 3% Sepiolite clay, PHA + 3% Cloisite Na clay and PHA + 3% C30B clay in freshwater and soil was evaluated. The tests are still running, but first observations show that biodegradation rate in soil of the PHA mixed with nanoclays is quite similar to neat PHA and after 14 days a biodegradation around 50% was observed. These soil biodegradation tests have a standard duration of 120 days. In freshwater conditions also high biodegradation was observed for the different materials as neat PHA reached a biodegradation of 79.7% after 28 days, while a value of 79.1% and 75.8% was measured for PHA + Cloisite Na clay and PHA + C30B clay. For PHA + Sepiolite clay a lower degradation rate was measured, reaching a biodegradation of 68.1% after 28 days.
- In two further (out of many) biodegradability examples, PLA/o-PHB and PLA-g-o-PHB (25 g each sample) were used for composting experiments. The oligomeric PHB was prepared by thermal recycling of PHB, a model SYNPOL polymer and the material was prepared under an earlier task. Under controlled composting conditions (58°C) complete biodegradation could be established on compounds PLA20G (PLA-graft-PHB) and PLA20B (PLA/PHB blend).
- In the task about ecotoxicity, the solid fraction (char) after the pyrolysis of wet and dried municipal solid waste and straw was examined for plant toxicity on barley and cress plants. The porous structure of these chars has been found out to be one of the possible factors related to the germination rates. This alternative has resulted to be highly promising since the toxic effect on plants germination was very limited or non-existing regardless of the municipal waste char composition.
- 11 data sheets from SYNPOL biopolymer products have been made by three SYNPOL partners. These data sheets are for the following novel bioplastic products: PLA-graft-PHB, PLA/dPHB-blend, PB67B’33A, Poly(3-hydroxybutyrate-co-4-hydroxybutyrate)-sepiolite nano-biocomposites, BIOPLASTECH 1, BIOPLASTECH 2, BIOPLASTECH 3, BIOPLASTECH 4, BIOPLASTECH 5, PHO, and PSA70). The data sheets contain information on production, thermal and mechanical properties, biodegradation, advantages and possible applications and will be publicly available soon.
- The environmental impact of the novel biotechnological production pathways developed within the framework of the SYNPOL project was also examined in LCA studies (Fig. 6). This impact was assessed in 16 effect categories in line with ILCD midpoint method (EC-JRC 2011) and ISO 14040/44 and ILCD Handbook guidelines. Both the total life cycle impact (cradle-to-grave, excluding use phase) and the life cycle hot spots (cradle-to-factory gate) are identified. The total impact results of the SYNPOL processes are compared to their conventional equivalents.
- There are two products developed in the project: a building block 2,3-butanediol (2,3-BD) and a biopolymer polyhydroxybutyrate (PHB). The common step of both production processes is microwave induced pyrolysis (MIP), which transfers pre-dried municipal solid waste (MSWd) into pyrolysis gas (mixture of mainly H2, CO, CO2). This gas is fed to the fermenter where bacteria (Clostridium ljungdahlii for 2,3-BD and Rhodospirillum rubrum for PHB) use it as a carbon source in order to metabolically produce desired substances. Subsequently, the fermentation broth follows different downstream pathways.
- The functional unit (FU) chosen for the whole production chain is 1 kg produced chemical. Despite very complex downstream pathway, production of pure 2,3-BD was unsuccessful. The final stream (70% moisture) contains still a vast amount of impurities (81% dry mass basis). Hence, 1 kg 2,3-BD diluted in the final stream was chosen as FU. The downstream pathway for PHB production was developed and pure PHB was successfully obtained at the end. It was observed that in both cases the biggest hotspot is associated with low mass transfer between CO and water and low efficiency of fermentation process. Consequently, a lot of gas needs to be produced via MIP (to provide enough CO to microorganisms), which is directly linked to the relatively high electricity consumption of this novel technique. At the same time a vast amount of unused gases needs to be flared, as a result of a small CO mass transfer into water, which leads to significant CO2 release to the atmosphere. MIP is very important contributor to the life cycle impact of the SYNPOL production process, but also in terms of possible benefits, associated with avoided municipal waste incineration. This is especially visible in human toxicity and freshwater ecotoxicity impact.
- The sensitivity analysis was performed in order to look for possible improvements. It showed that the model is sensitive to changes in MIP electricity consumption and waste gas treatment methods.
- The environmental impacts associated with the studied building block and biopolymer production can be effectively improved by optimization of CO mass transfer into water, which will increase the amount of CO available for microorganism enhancing their efficiency and thus yields of interesting chemicals. At the end of fermentation process, the waste gas should be flared, best with electricity recovery so conventional electricity production could be avoided. Additionally, further research is recommended in order to allow recycling of pyrolysis gas at the fermentation stage. This goes hand in hand with better CO mass transfer into water and would lower the impact of MIP and reduce gas emissions. Furthermore, it should be investigated if reduction in electricity consumption by MIP is possible in order to make it more environmentally interesting than conventional techniques.
- From the life cycle assessment it can be concluded that several factors should be improved (also when compared with alternative production processes). Within the life cycle of 2,3-BD production, flash evaporation is mainly responsible for the impact on photochemical ozone formation due to significant energy consumption.
- Within the life cycle of PHB production, used solvent (dichloromethane) carries significant impact on ozone depletion. It is advised to replace it with more environmentally friendly solvent in further research. The remaining impact of the solvent and chemicals use was avoided due to introduction of partial recovery and recycling step at the end of the downstream process. Final filtration and chemicals recovery process are mainly responsible for the impact on photochemical ozone formation due to significant energy consumption.
- Both processes have the same main bottleneck. Mass transfer between CO and water is very low, hence very little amount of CO from pyrolysis gas is available for microorganisms during fermentation. Consequently the final yields are very low and the fermentation process is not efficient. This is especially visible when compared to reference scenarios. This means that at this point the SYNPOL processes cannot compete with conventional production pathways. Both processes are sensitive to changes in electricity consumption by MIP and to treatment of waste gas.
- Pyrolysis is very important contributor to the life cycle impact of the SYNPOL production process, but also in terms of possible benefits, associated with avoided municipal waste incineration. This is especially visible in human toxicity and freshwater ecotoxicity mitigated impact. Electricity consumption is the most important factor determining the impact of MIP. Four different feedstocks were analysed and it turned out that MSWd is the most interesting one for pyrolysis, due to small amount of produced waste (solid and liquid) and high CO flowrate. Additionally, it was observed that MIP at 800°C is more environmentally friendly than at 400°C.
- The impact of conventional pyrolysis (CP) on climate change is smaller than of MIP, mainly due to higher electricity consumption by the latter. However, taking into account all 16 effect categories, MIP scores better in most of them (11 out of 16), mainly due to much lower pollution associated with by-products (chars and tars).
Part of the ecotoxicity results have already been published and are accessible at:
http://dx.doi.org/10.1016/j.jaap.2016.08.013
Results of the works on demonstration of SYNPOL biopolymers:
The objectives of the demonstration part of the project were:
➢ to determine the commercial application of the novel PHA nanocomposites and PHA blends
➢ the best performing materials will be processed in sufficient quantities to allow full materials characterization study
➢ preparation of a biopolymer production plant design taking into account:
• optimal process performance with fermentation and downstream operational data at large scale
• minimum energy requirements
• reduced environmental impacts
• end-to-end economical evaluation
The following results were obtained during the final phase of the project:
- The up-scaling of fermentation and downstream processes for the production of 2,3-butanediol from syngas was explained. In a first stage, laboratory scale-up was developed by one SYNPOL partner. Later on, projection on industrial scale was carried out by another SYNPOL partner. A basic process diagram, equipment, utilities and ancillary equipment are described. Further on, analysis of bottlenecks of the processes is provided in the public available deliverable D8.5 (“Fermentation and downstream operational data at large scale”).
- A demonstration plant capacity for the production of 2,3-butanediol has been designed according to the usual ratios for products with a low economic value. For details see the public available deliverable D8.4 (“Pilot plant - Biopolymer production plant design and economic viability”).
- Three SYNPOL prototypes have been successfully produced by SYNPOL partners and are described in the public available deliverables D8.1 (“Film for packaging applications made of Bioplastech mclPHA blended with SYNPOL PHB”). D8.2 (“An electrospun composite of SYNPOL PBS and Bioplastech mclPHA to form a scaffold for biomedical applications”) and D8.3 (“Biodegradable adhesive based on a blend of Bioplastech mclPHA and SYNPOL polyester urethanes”). As an example, Fig.7 shows a compostable paper bag made of crafting paper and cellulose film window utilizing the 100% biobased adhesive (from SYNPOL biopolymers) for its assembling.
- The novel SYNPOL biopolymers prototypes will be patented in the future and therefore, no official publications have been produced within this demonstration task of the SYNPOL project.
Potential Impact:
Description of the potential impact
Waste is actually a pressing environmental, social and economic issue. Increasing consumption and a developing economy continue to generate large amounts of waste - with more effort required to reduce and prevent it. While waste was viewed as disposable in the past, today it is increasingly recognized as a resource; this is reflected in the waste management shift away from disposal towards recycling and recovery. Therefore, SYNPOL’s valorisation of anthropogenic derived wastes, as raw materials for the industrial production of polymers and chemicals, provided an effective response to the challenge of sustainable production of these materials, while in parallel handled the vast environmental and economic problem of wastes disposal.
Within the context of an ‘integrated operation’ there are multiple objectives satisfied in parallel at the benefit of environment, sustainability and bioeconomy. Therefore, the socioeconomic impact of SYNPOL project can be evaluated from different perspectives:
ϖ - Utilization/valorisation of the complex biowastes for the production of high-added value products like biopolymers
ϖ - Replacement of common chemically catalysed production processes for the transformation of syngas with eco-efficient microbial ones
ϖ - Reduction of the volume and polluting load of the complex biowastes
ϖ - Decrease of the waste treatment costs
ϖ - Establishment of the technological developments for the formation of renewable raw materials for the polymer industry contributing to its modernization, sustainability, consolidation and independence from the availability of fossil feedstocks
ϖ - Replacement of conventional polymer materials with biodegradable ones extending the range of polymer applications to food, biomedicine and pharmaceutics
ϖ - Production of novel biopolymer building blocks from renewable sources opening an increasing window-of-opportunity for bio-based products for the chemical industry and creating new markets to the manufacturers of pharmaceutical and nutritional care products
ϖ - Bridging the gap between the novel biomaterial production technologies and their industrial/commercial application and contribution to the faster satisfaction of the humanity need for renewable biopolymers that can be composted after use in contrast to conventional plastics
Enhanced environmental care and ecosystem protection both facilitating the safe and within the regulations discharge of wastes from colossal industries and contributing to the preservation of carbon balance in the atmosphere. This is due to the fact that the carbon sources used for the biotechnological production of biopolymers derived mainly from carbon dioxide that was previously fixed by plants. Hence, the release of CO2 by degradation of biopolymers is just the final step of natural mineralization.
Perhaps the greatest benefit generated by this project was traced with respect to the environment and the global climate through the reduced emission of fossil-carbon derived CO2. By the industrial implementation of the biochemical routes for the waste bioconversion to biopolymers and biochemicals, substantial reductions in greenhouse gas emissions could be realized. The more the produced technology is established and the proposed bio-based products replace the conventional existing ones, we could assume that the emission reduction can be of the order of 15-50%, which is a step closer to the satisfaction of global Kyoto target. Obviously, all these promises of the project with great impact on different sectors can only be realised provided the recognition and the adaptation of the produced technology by the industry, hence, the developments over the scaling-up capability of the produced technologies and the market value of the new products is the biggest challenge. The know-how gained from SYNPOL project can support any plans for large scale implementation of biopolymers and “green” chemicals.
To further appraise the project impact through the proposed cost- and eco-efficient production of bio-based polymeric materials we should contemplate the problems of the greenhouse effect and the global warming that the conventional polymers are highly linked with due to the utilization of constrained fossil resources for their production.
The main problem arising from incineration of plastics is the same as for energy recovery from fossil feedstocks: carbon that was fixed during millions of years and within this time was not part of the natural carbon cycle is converted to CO2, which can accumulate in the atmosphere, contributing to the mentioned climatic effects. In addition, incineration of plastics often generates toxic compounds. Besides, more and more waste of highly resistant plastics that are not incinerated is piled up every year. Recycling systems demand a certain degree of purity and a high sorting accuracy. In addition, the collection costs are fairly high, and recycling has a negative impact on the quality of the materials, such as an increase in brittleness of the recycled material.
In addition to these ecological considerations, the price of crude oil is unpredictably fluctuating, not least due to miscellaneous developments in the global political situation. This constitutes a factor of immense uncertainty especially for the highly petrol-dependent polymer industry. The technological developments produced by this project can contribute to overcome these problems and confront in parallel the increasingly posed question of how we will derive the materials we will need in the twenty-first century when the contingency of the depletion of fossil feedstock become one day a reality. In front of this contingency, the involved industrial branches (i.e. polymer and chemical industry), which are already aware of the necessity for promoting novel production techniques based on renewable resources, will turn to solutions also provided by the SYNPOL project.
The impact of the project can be also appraised from the perspective of the produced bio-based materials. It should be noted that the EU polymers and chemicals production sector is an industry in transition. This is a consequence of changing domestic demand and the pressures of globalization, both of which are driving the industry towards the manufacture of higher added value products and sustainable production. “Green” and thus biodegradable polymers and biochemicals from renewable resources is the technological response to the above challenge.
In addition to the project’s impact on the polymer industry, the contribution to a sustainable production of building blocks will strengthen the link of the biochemical industry with the pharmaceutical, nutritional and agricultural products industries creating an opportunity for the fine chemicals companies for growth and invigoration. These high added value biochemical products have the potential to become a significant part of EU’s economy in the future.
There is little doubt that the waste-based bioprocessing technology has great market potential both in replacement for current processes related to fossil resources, as well as in the creation of new markets through its sustainable, efficient and economically viable production of biopolymers and high value-added chemicals. Moreover, a successful research outcome in the production of these bio-based polymers and chemicals will enormously enhance the competitiveness and sustainability of the European biopolymer and biochemical industry and will generate new business opportunities. As “green” technology is making the industry more sustainable, it is expected that benefits will also be seen across a range of critical society-based areas representing other profound fields of economy.
The SYNPOL project responded to all these objectives with the technological advances described above. Precisely, in addition to cost reduction achieved by the efficient transformation of the low-cost complex biowastes in syngas as fermentation substrate, there are several lines of evidence supporting the feasibility of this objective, such as the genomic modification of the selected bacteria and the fermentation strategies properly designed for the selected bacteria. The synergistic action of the project approaches along all the routes together, (i.e. raw materials, microbial producers and operation strategies) generate the best outcome with respect to the low-cost production of biopolymers and bioconversion of wastes into high value-added products (Fig. 8).
To ensure the successful accomplishment of the project objectives, a multidisciplinary partnership has been established, consisting of academic and research scientists covering various disciplines and complementary scientific and technical areas (i.e. biotechnology, molecular biology, biochemistry, biosystems and bioprocess engineering, nanotechnology, chemical and process systems engineering, environmental engineering, etc.). These FP7 partnerships are also fruitful for future cooperation in further European projects, and indeed, some partners are actually working together again in other H2020 projects concerning the use of gases as feedstock or in related bioeconomy projects.
European dimension
The core idea of the SYNPOL project for a profitable utilization of selected complex biowastes for the production of bio-based products represents a strategic initiative for Europe in the area of biopolymers and high-added value biochemicals. The tactical decision on the complex biowastes, as raw materials, and the way to convert them into targeted products by fermentation of syngas reflects the needs, the capabilities and deficiencies as well of Europe, as a continent with low fossil-oil resources, high fossil-oil requirements though, abundant amount of the selected biowastes, legislations with intense environmental concern, political desire for imported-oil independence and capability to lead the world biopolymer and biochemical industry.
The various works for development of a novel and efficient industrial biowaste processing and valorisation technology, performed within the framework of SYNPOL project, successfully addressed community societal issues pertaining to the quality of life of European citizens by protecting the environment, climate, and increasing the energy resources via the use of innovative waste processing systems.
In order to be able to sustain the EU position, huge advances in the environmental control must be achieved. Such advances (i.e. CO2 mitigation and GHG reduction) would make it possible for the European members to successfully face the challenge of effective protection of the environment at affordable cost. In this sense, the SYNPOL project had a truly European dimension, as it served to protect and possibly enhance the European members’ position as the most ecologically sensitized.
Biobased products can replace 1:1 their chemically derived counterpart if the cost is competitive. Moreover, biobased products can bring new properties and functions to materials and chemical intermediates. This characteristic can enable a chain growth and development to new sectors as well as to already existing highly profitable ones such as the pharmaceutical and nutritional products industry, with the European representatives being of the pioneer ones world widely.
Community societal objectives
The current challenges in the area of renewable biodegradable polymers and biochemicals are to design, produce at a competitive cost and use industrial waste sources that satisfy the energy needs of the society and ensure an improved quality of life, whilst maintaining the capacity of the natural environment to renew itself. As a result of project exploitation, the industrial partners involved in the SYNPOL project benefit from the novel technological solutions and related markets that will improve their short and long-term profit. The research and academic centres scientifically benefited from the collaboration with each partner of the consortium exchanging know-how and creating background knowledge in a field that is anticipated to be at the edge of the technology. Finally, all partners had synergistically worked with common target the extroversion of the consortium as a whole, in order to make the scientific and technological developments known to the related industries and attract their investing interest for the commercial application of the produced technology. Innovation and intellectual property rights provide economic opportunities such as establishment of spin-off companies. The society at large will benefit from the innovations in different areas (e.g. energy, environment, and climate).
Additionally, the present project provided the core knowledge for potential exploitation of different industrial biowastes further promoting to the “integration” and link of heterogeneous industrial sectors and expanding their business opportunities. Moreover, it will have drastic impacts on all societal, economic, ethical, legislative and educational aspects of everyday life, since it supports innovative, fundamental research in the science and technology of the production of biobased materials leading to potential breakthroughs primarily to the directly involved industrial sector (i.e. biopolymer and biochemical industry) as well as to the indirectly involved one as recipient of these technological developments (e.g. pharmaceutical and food industry).
Main dissemination activities and the exploitation of results
Translation of basic foreground into popularised communications or executive summaries was handled by the dissemination work package of the project and freely distributed to all concerned stakeholders within and outside the EU (see summary of dissemination activities in the automatic generated table from the Participant Portal).
The scientific and technological expertise gained was disseminated through scientific publications, patents, presentations at conferences and press releases. Knowledge acquired was also used for advanced training purposes and as an input to further research and industrial applications. In order to promote academic and industrial cross-fertilization within the partners’ organizations and towards the ERA we have established a number of dissemination activities. In this sense it is worth mentioning that special attention will be devoted to the dissemination of the social benefits derived from the SYNPOL technology concerning to the transformation of different biowastes into recyclable bioplastics. We are aware of the enormous social, economical and technical problem involving the accumulation of wastes and of the great benefices that will provide the technology developed in this project.
To fulfil these goals the following dissemination activities have been carried out:
1. We created different Communication Materials (website, flyer & brochure) that was useful for improving not only partners’ communications but also the communication with other R&D initiatives and with potential partners interested in participating in or supporting the project activities:
- A specific internet website for the project (www.synpol.org). This tool was one of the elements to integrate the complete dissemination plan of the project. This internet web site was used to communicate the advantages of SYNPOL technology to scientific or other stakeholders at both European and international levels. The web page displayed links with other related and collaborating EU projects. Apart from being an effective way of advertising our research and main achievements, the website was used to promote events and publications or even to distribute key messages on the home page. The partners contributed scientific contents in order to ensure its continuous update and extension. The web page, which was updated more or less weekly, allowed and still allows the interested persons accessing the website to be informed on the project developments. Thus, the website supported a close co-operation between the consortium partners and helped raise public awareness.
- Information brochures and flyer on SYNPOL technology. A brochure and a flyer were produced presenting the SYNPOL project. It was intended to support the presentations at events and the individual meetings carried out and it served as a communication tool for potential mailings to target audiences. Flyer and brochure were project oriented, as both materials described the project, its objectives, the participating organisations, the expected results and the activities planned.
- SYNPOL updates were also published in the institutional or company web sites of nearly all beneficiaries. The CSIC website for example and the other available web sites have a great exposure to public and they constitute an excellent complementary dissemination mechanism apart from our own SYNPOL web site.
2. The SYNPOL web site also offered links to open the project to Social Networks. Therefore, we created SYNPOL accounts in the most important social networks such as Facebook and Twitter. The social networks provided us the opportunity to connect not only with the citizens but also with the companies and scientists interested in this technology.
3. Plenary lectures and workshops. We organized as satellite events of the annual General Assembly meetings (12, 24, 36 and 48 months) four specific workshops to invite technology users of SYNPOL achievements, including keynote lectures from partners as well as for specially invited speakers. Additionally, 3 further workshops (one in Spain [2014] and two in Switzerland [2014 & 2016]) on biotechnology issues were organized and carried out by SYNPOL beneficiaries.
4. The activities developed in the project were disseminated also by stimulating the presentation of results with poster and oral presentations at scientific international conferences, congresses, workshops, and meetings. The absolute scientific dissemination highlight for the SYNPOL project can be seen in the 15th edition of the International Symposium on Biopolymers 2016 (www.isbp2016.com) that was co-organized (and financially supported) by the SYNPOL project and that was held during the final week of the project from 26th to 29th of September 2016 in Madrid, Spain. The ISBP2016 was the final and main dissemination platform for the SYNPOL project and more than half of all SYNPOL consortium partners have presented their results and perspectives at this congress.
5. The publication of results in high impact scientific journals and books was also encouraged and successfully done by many SYNPOL partners (see automatically generated list of publications in the Participant Portal).
6. Theses; at the end of the project time (30th of September 2016) one PhD thesis can be announced as terminated. Title: “Strategies for utilizing biobased and recycled resources for polylactide plasticization”, Author: Xi Yang, Date of approval: 16/09/2016 at KTH - Royal Institute of Technology - School of Chemical Science & Engineering10044 Stockholm (Sweden). Further PhD theses will be defended late 2016 or in 2017 by doctoral candidates from the SYNPOL partner universities of Ulm (Germany), Münster (Germany), Manchester (UK), Amsterdam (The Netherlands), Strasbourg (France), HES-SO (Sion, Switzerland), and Dublin (Ireland).
7. In coordination with the press offices of the partners´ institutions we tried to transmit the results and advantages of SYNPOL technology to the journalists through radio interviews (2), videos (2) and specialized articles (5). A few press releases in different languages have been made and monitored as an indicator of dissemination of project results.
8. To improve the training of the personnel involved in the project, to stimulate and promote the transfer of knowledge and to increase the cooperation within the partners, we established 15 specific exchange/mobility activities for interchanging personnel between the academic institutions and the companies. This interaction favoured the flow of ideas across the partner laboratories.
9. Announcements of vacant research positions within the project at partner’s laboratories have been published on the website a few times.
10. General public open-doors activities. Some partners promoted participation of SYNPOL in activities to promote as “open days”, “science fairs” and “meet the scientist” presentations and and participated in workshops to train high-school teachers. These activities targeted to the public led to a better understanding of the efforts carried out by the project and the EU Commission to develop clean and sustainable technologies such as SYNPOL to efficiently recycle biowastes.
Dissemination & Exploitation outcomes of the SYNPOL project at a glance:
Patents: 2 (from academic partners, but there are more in the pipeline from the industrial partners)
Spin-off companies: 1 is planned by one industrial partner by the name of CEWAS, Consulting and Engineering technologies for Wastes (Seville, Spain)
Publications: 39 peer-reviewed scientific papers and a few more general project presentation papers (from all academic partners and one industrial partner; there are at least 3 more publications in the pipeline)
Workshops: 3 (2 in Sion, Switzerland and 1 in Madrid, Spain)
Annual SYNPOL courses: 4 (in Valencia, Oviedo, Murcia and Madrid, all Spain) with 50 – 75 participants from many European countries
Congresses: 1 (ISBP2016 from 26th to 29th of September 2016, thus in the final project week)
Open door activities: 4 (3 in Madrid & 1 in Sion, Switzerland)
Poster presentations on congresses: 16
Oral presentations on congresses: 29
PhD theses: 1 defended and thus terminated (7-8 ending soon)
List of Websites:
Website at www.synpol.org
Primary Coordinator Contact: Prof. Dr. José Luis García López (jlgarcia@cib.csic.es)
Coordinator Contact/Project manager: Dr. Oliver Drzyzga (drzyzga@cib.csic.es)
Both at: Centro de Investigaciones Biológicas (CIB) of the Agencia Estatal Consejo Superior de Investigaciones Científicas (CSIC), C/Ramiro de Maeztu 9, 28040 Madrid, Spain; Tel. +34 91 8373 112
Every day we waste money in transporting municipal solid waste (MSW) miles out of cities or states to be landfilled. Long term environmental and economic problems created by landfilling prove that it is not the solution to the waste problem, and now waste conversion technologies are becoming a massive growth industry worldwide. In this context, the significance of the SYNPOL project was remarkable considering the high amount of available relevant waste produced solely in Europe every year.
Complex organic wastes such as municipal and agricultural residues, which were firstly pyrolyzed to produce synthesis gas (“syngas”) and then fermented by microorganisms, were the starting feedstock of the SYNPOL project which aimed at the transformation of wastes into 100% biodegradable bioplastic materials and further chemical building block compounds (e.g. butanediol, succinic acid, hydroxybutyric acid and crotonic acid).
Syngas fermentation is already an attractive technology for the production of biofuels and in fact several industrial processes for ethanol production from syngas are now under development, but these promising industrial processes have not been implemented in Europe yet and this is the challenge that SYNPOL was addressing. SYNPOL´s syngas fermentation technology helped to open a new window for the rational design of an innovative process to convert complex wastes into new biopolymers and other high added value compounds.
SYNPOL has done this waste conversion in three major steps:
• Pyrolysis of different organic waste streams to produce syngas (CO, H2 and CO2)
• Fermentation of syngas by using different bacteria to produce chemical building blocks and polyhydroxyalkanoates (PHA)
• Synthesis of bio-based plastic prototypes with well-defined structures and improved properties for wide commercial use via chemical and enzymatic catalysis by utilizing the monomers and polymers produced during syngas fermentation
Novel processing technologies, like the combination of novel microwave induced pyrolysis (MIP) of wastes with improved syngas fermentation bioreactors, are additional relevant outcomes that have been derived from SYNPOL. To fulfill its aims, SYNPOL established an integrated platform for biopolymers production reducing energy input and optimizing purification of waste streams, thus contributing to the economic viability of the whole process.
Specific main objectives were defined and carried out to achieve the above-mentioned general objectives: i) Waste samples preparation as feedstock for pyrolysis and application of innovative gasification methods to produce syngas containing high percentages of CO and H2, ii) Improvement of bacterial strains for syngas fermentation by genetic and metabolic engineering, iii) Development of novel fermentation design for effective and safe syngas fermentations at lab-scale, iv) Development of biopolymer design by characterizing novel biopolymers/copolyesters from chemical catalysis and synthesis of novel biopolymer blends with PHAs, plasticizers and nanoclays using building blocks produced from syngas fermentations, v) Biopolymer degradation and life cycle assessment (LCA), and vi) Demonstration of the success of the SYNPOL platform technologies by presenting three novel prototype biopolymers for different industrial applications and a newly developed biopolymer production plant design taking into account the best scale-up of fermentation and downstream processes.
Summarizing, SYNPOL has linked the notion of waste valorization with representative members of the European bioindustries which are distinguished for their interest in the development of biotechnological processes to produce biopolymers by using biowastes as starting materials. The ultimate scope of SYNPOL was to establish a win-win situation between these bioindustries and the polymer industry, as a branch of the chemical industry.
Project Context and Objectives:
Too much waste around us
There are many waste resources hidden in our communities. For example, municipal solid waste (MSW), agricultural residues and sewage sludge from water treatment plants contain lots of reusable carbon fractions. An eco-efficient waste conversion process means recovering most of the waste as valuable products in an environmentally friendly manner. In this context, the significance of the SYNPOL project is remarkable considering the high amount of available relevant waste produced solely in Europe every year (approx. 261million tons (MT) of MSW including approx. 25 MT of plastics, approx. 120 MT of agricultural residues (e.g. straw) and >10 MT of sewage sludge). [1-4]
The basic idea and work flow scheme of the project is presented in Figure 1.
Polyhydroxyalkanoates (PHA): an alternative for petrol-based plastics
Polyhydroxyalkanoates (PHA) are a well-known family of polyesters accumulated as cell granules by naturally occurring microorganisms serving as carbon and energy reserve. Figure 2 shows bacterial cells containing PHA granules and some short chain length PHA isolated from bacteria. Many different PHAs of diverse desired properties including biodegradability can be produced from renewable resources by the biosynthetic action of selected prokaryotes. Currently, PHAs are one of the alternatives to petrol-based plastics, elastomers or latexes.
Non-food competing feedstock materials
The need for alternative materials, because of the finite sources of fossil reserves is generally accepted. In order to become a competitive alternative on the market, the price of bioplastics for a certain application must be in the same range as the competing petroleum-based plastic. Hence, the costs of PHAs have to be reduced considerably despite the current unstable price of crude mineral oil. As an alternative solution, diverse waste streams exist which currently constitute severe disposal problems and, at the same time, do not interfere with the nutrition chain. This is especially true for the daily produced household waste that ends up in landfills or the sewage sludge loadings from water treatment plants. The utilization of these waste streams is a viable strategy to overcome a potential ethical conflict. Indeed, it can be considered as the most promising approach in making PHAs economically more competitive.
Philosophy, roadmap and main objectives
The SYNPOL project aimed at converting these waste streams via refined gasification processes (pyrolysis supported by innovative microwave technology) into synthesis gas (syngas) that contains high amounts of carbon monoxide (CO) and hydrogen (H2). These gaseous carbon fractions were biotechnologically converted towards PHA biopolymers and further chemical building block compounds by applying bacterial fermentation processes using different bacterial strains. The selected bacterial strains were naturally occurring microorganisms that were optimized for higher biopolymer production efficiency by genetic and metabolic engineering techniques based on the newest findings from systems biology.
The project brought together public research centers with waste producers from different processing industries (landfilling, agricultural and water treatment industries), gasification industry and polymer processing biotechnological companies. This synergism was benefiting all players and resulted in new value creations. The project´s time roadmap is illustrated in Figure 3 and summarizes the main working lines of the consortium.
These working lines were created to cover the following main objectives of the project:
1. Optimize syngas production from different feedstocks by using new gasification technologies.
2. Characterize syngas generated from feedstocks via a combination of state-of-the art techniques.
3. Analyse energy costs required to generate syngas from different feedstocks.
4. Enhance naturally occurring pathways of acetogenic bacteria by metabolic engineering supported by systems biology tools with the aim of generating by syngas fermentation large amounts of intermediates that can be used as building blocks for the synthesis of new biopolymers.
5. Design new metabolic pathways by using systems biology tools to produce polyhydroxyalkanoates in acetogenic bacteria.
6. Enhance naturally occurring pathways of purple bacteria by metabolic engineering, again supported by systems biology to generate by syngas fermentation large amounts of polyhydroxyalkanoates.
7. Design transferable genetic systems to confer the ability to metabolize CO to other bacteria under aerobic conditions.
8. Design new recombinant bacteria endowed with programmed autolytic systems to facilitate the downstream processing of biopolymers.
9. Establish a cost-effective consolidated syngas fermentation technology using the natural and recombinant microorganisms by improving the operational conditions.
10. Integrate interdisciplinary knowledge to consolidate an efficient downstream processing, to decrease energy inputs, reduce environmental impacts, increase purification efficiency and diminish the final cost of the products.
11. Develop cost effective technologies for the chemical synthesis of new biopolymers based on the “green” building blocks and the polyhydroxyalkanoates produced by syngas fermentation, utilizing state-of-the art of chemical processes.
12. Develop blend, plasticizer, and composite formulations based on a PHA matrix using fermented and waste stream products.
13. Reduce the environmental impact of the integrated SYNPOL technology by recycling the residues.
14. Demonstrate the chemical and organic recyclability of the developed biopolymers.
15. Develop life cycle and environmental impact analyses as a valuable tool to provide technical and economic advances in the development of environmental friendly biopolymers.
16. Plant production design, scale-up and optimization of consolidated SYNPOL technology targeting on the increase of the products yield, improving the overall production economics as well as the upscaling capabilities.
17. Establish portfolios for the exploitation of the process and the products derived from SYNPOL project.
18. Disseminate the scientific knowledge acquired during SYNPOL project to the society by developing multiple training and diffusion activities.
A project of great expectations
From the perspective of the SYNPOL project, important progress was achieved in terms of combining the environmental benefit of future-oriented biopolymers with the economic viability of their production. This should finally facilitate the decision of responsible policy-makers from the waste-generating industrial sectors and from the polymer industry to break new ground in sustainable production. In the future, PHA production from different waste streams applying gasification technology should be integrated into existing process lines of biotechnological bioplastic companies, where the feedstock material directly accrues. By taking profit of synergistic effects, this can be considered a viable strategy to minimize production costs. The 48 month SYNPOL project has secured almost 7.5 million € in funding under the Food, Agriculture and Fisheries, and Biotechnology theme of the European Union Seventh Framework Program (FP7/2007-2013; grant agreement nº 311815). The project was launched in October 2012 and ended in September 2016 and was coordinated by the Biological Research Center (CIB) in Madrid (Spain) which is part of the Spanish National Research Council (CSIC).
The project team players:
From Industry
▪▪ Biopolis S. L. (Spain) – Industrial fermentation
▪▪ Bioplastech Ltd. (Ireland) – Biopolymer synthesis
▪▪ Organic Waste Systems NV (Belgium) – Biodegradation & LCA analysis
▪▪ Bionet Servicios Técnicos S. L. (Spain) – Simulations & Pilot plant design
▪▪ Infors AG (Switzerland) – Fermentation development & Bioreactor design
▪▪ Befesa Gestión de Residuos Industriales S. L., later substituted by Abengoa Research (Spain) – Waste management
From Academia
▪▪ Consejo Superior de Investigaciones Científicas (Spain) – Project coordination, Syngas production, Bacterial Research & Downstream processing
▪▪ University of Manchester (United Kingdom) – Bacterial Systems Biology
▪▪ University of Amsterdam (The Netherlands) – Bacterial Systems Biology
▪▪ Universität Ulm (Germany) – Bacterial fermentation & Recombinant strains
▪▪ University College Dublin (Ireland) – Proteomics & Molecular biology
▪▪ Haute Ecole Spécialisée de Suisse Occidentale (Switzerland) – Fermentation reactor design & Downstream processing
▪▪ Kungliga Tekniska Högskolan (Sweden) – Biopolymer synthesis
▪▪ Westfälische Wilhelms-Universität Münster (Germany) - Bacterial fermentation & Recombinant strains
▪▪ Université de Strasbourg (France) – Biopolymer design
References:
[1] http://bit.ly/12ZOjgs
[2] http://bit.ly/12kWe1y
[3] http://bit.ly/1351OwK
[4] http://bit.ly/12ZOQPy
Project Results:
The most outstanding non-confidential S&T results obtained in the course of the SYNPOL project are described, with a mention of those already disseminated by scientific publications (and other means).
The following sections include a description of the results and achievements in the 5 RTD work packages and in the Demonstration work package conforming the SYNPOL project scheme (Fig. 1) and work plan.
Results of the works on syngas production from organic wastes:
First of all, it was necessary to identify appropriate feedstock for the pyrolysis process and to determine their characteristics and compositions, since the efficiency of the global process will be affected by these parameters. The different waste samples (municipal solid waste, agricultural residues and sewage sludge) have been characterized for their use as feedstock for subsequent pyrolysis by application of standardized methods.
The following general results have been obtained:
- A major conclusion of these experiments was that the best results (highest syngas production and with a high CO + H2 content) have been obtained with agricultural residues (straw) and with the organic fractions of the municipal solid waste as feedstock for pyrolysis. From an environmental point of view, municipal solid wastes face an important challenge, such as the need for new technologies to manage them through valorisation strategies. Hence, this substrate has been selected to study the process of microwave-induced pyrolysis to produce syngas.
- Concerning the syngas production and composition, it can be concluded that the results obtained in the microwave-induced pyrolysis are clearly superior to those obtained in the conventional pyrolysis.
- Since the organic residues are poor microwave absorbers, it was necessary to use a microwave receptor, which could be directly heated by microwaves and so heating the biomass by conduction, convection and radiation processes. The best material to play this role is the carbonaceous residue obtained in a previous pyrolysis. A minimum microwave receptor-to-waste ratio of 0.2:1 is needed to induce the pyrolysis.
- Higher microwave power favours the production of syngas but this increases the energy consumption.
- Depending on the initial moisture content and pyrolysis time, it is possible to obtain different syngas compositions. The moisture content was found to improve the volumetric gas production by almost 50% at the expense of the energy consumption needed for the water evaporation. Nevertheless, 10-20 wt% of moisture in waste enhance the microwave absorption by biomass and allows for a maximum production of syngas.
- The use of different carrier gases (N2, He) affects the yields of the fractions obtained during the pyrolysis of organic residues only slightly. In the case of microwave pyrolysis, and especially at high temperature, no significant difference in the gas composition was observed when using different carriers. Furthermore, it was observed that the implementation of microwave pyrolysis can be overcome without the need for a carrier gas.
- Different experiments on CO2-reactive atmosphere were conducted. The syngas concentration can be slightly improved due to the partial gasification of the carbonaceous residue.
- Microwave pyrolysis leads to a reduction in the liquid fraction production. It has been found that hazardous PAHs production from heterogeneous wastes (organic fraction from municipal solid wastes and sewage sludge) can be reduced by microwave pyrolysis.
-The scale up parameters of the microwave induced pyrolysis (MIP) process were established as follows:
✓ The organic fraction of Municipal Solid Waste (MSW) is a good substrate (best among all the residues studied) to produce high yields of syngas (up to 90 vol% of the gas fraction).
✓ Temperature about 800ºC is the best option to balance maximum gas production/minimum energy consumption.
✓ A ratio of microwave susceptor/substrate of about 0.2:1 is the best option. Below this ratio is almost impossible to reach the 800ºC (lots of energy consumed), but higher ratios impair penetration of microwaves resulting in a worse homogeneity.
✓ Certain amount of moisture is not bad since it helps to the heating in the first stages and contributes to gasification. Substrates with about 12 wt% of moisture are a good choice.
✓ It is possible to carry out MIP without using any carrier gas. In any case the influence of the carrier gas is minimal.
✓ The energy consumed by the MIP of MSW is estimated to be about 1.5 kW/kg.
Given that the solid fraction (chars) obtained in MIP is of low quality for using it as a fuel (high inorganic matter content containing heavy metals) it was studied the possibility of use this fraction as soil amendment. The maximum amount of char that can be used in soils was determined by means of standard ecotoxicity tests.
Given that the tars produced in the MIP process need to be upgraded (by removing its moisture, heavy metals and other undesirable compounds and reducing its oxygen content) before to be used as fuel; a new concept of pyrolysis that does not produce oils was developed, tested at laboratory scale and patented.
These results have been published and are accessible at:
http://dx.doi.org/10.1016/j.jaap.2013.11.009
http://dx.doi.org/10.1016/j.fuel.2014.04.064
http://dx.doi.org/10.1016/j.jaap.2014.12.005
http://dx.doi.org/10.1016/j.jaap.2014.12.011
http://dx.doi.org/10.1016/j.cep.2015.05.001
http://dx.doi.org/doi:10.1016/j.jaap.2015.06.006
http://dx.doi.org/10.1016/j.fuel.2016.01.016
http://dx.doi.org/10.1016/j.jaap.2016.08.013
http://dx.doi.org/10.1111/1751-7915.12411
http://dx.doi.org/10.1016/j.biortech.2016.07.019
http://link.springer.com/chapter/10.1007/978-94-017-9612-5_6
Results of the works on improvement of microorganisms for syngas fermentation:
This working field aimed to i) analyse the capacity of various bacteria to ferment syngas, ii) create genetically modified strains, iii) transfer the capacity of using CO as a carbon source to aerobic bacteria, and iv) find generic ways to improve this type of engineering.
The following general results have been obtained:
Systems Biology studies:
In Systems Biology studies, a general Rhodospirillum metabolic model was developed by applying an existing metabolic map for the three bacterial species Rhodospirillum rubrum, Rhodobacter sphaeroides and Rhodopseudomonas palustris, and by adapting it to the special case of R. rubrum using syngas as carbon and energy source. The results obtained will help in the near future to adjust bioreactor fermentation processes (e.g. metabolic fluxes of key substrates such as CO, CO2 and H2) more precisely towards the desired end product.
In addition to the reconstructions of metabolic maps for C. ljungdahlii and R. rubrum further genome-wide reconstructions for Acetobacterium woodii, Clostridium acetobutylicum, Clostridium cellulovorans and Oligotropha carboxidovorans were produced in the project.
Studies with bacteria of the genus Clostridium:
The wild type bacterial strain Clostridium ljungdahlii showed the highest 2,3-butanediol production on syngas, therefore the genes for 2,3-butanediol synthesis starting from pyruvate alsS (acetolactate synthase), budA (acetolactate decarboxylase), and 2,3bdh (2,3-butanediol dehydrogenase) of this organism were chosen to create two 2,3-butanediol overexpression plasmids. The overexpression of the three genes alsS, budA, and 2,3bdh improved the 2,3-butanediol production in some other model clostridia strains.
Additionally it was shown that succinate formation can be naturally achieved with C. aceticum and C. ljungdahlii.
Studies with bacteria of the genus Rhodospirillum:
While the wild type strain R. rubrum S1 accumulated a maximum of 26% of cell dry weight (cdw) polyhydroxybutyrate (PHB), the complemented (recombinant) strain R. rubrum ∆phaC2 pBBR1MCS-2::phaC2R. rubrum showed the highest PHB synthesis of all tested strains with 32% PHB (cdw). Thus, recombinant R. rubrum strains accumulated up to 25 % more PHB than the wild type strain.
A R. rubrum strain able to synthesize solely medium chain length PHA (PHAMCL) from an unrelated carbon source has been generated. It is potentially the first microorganism able to accumulate these polyesters from syngas. Recombinant R. rubrum ΔphaC1ΔphaC2 pBBR1MCS-2::phaGPp::phaCPp accumulated a total amount of 2.5 % (w/w of cell dry weight) of PHAMCL. No other PHAs, as for example PHB, were synthesized by this recombinant strain. Currently, further work focuses on increasing the amount of synthesized PHAMCL.
R. rubrum S1 was successfully grown with syngas (Fig. 4) and a second carbon source under anaerobic conditions in the dark. In order to increase the carbon flux towards PHB production, the effect of syngas fermentation and PHB accumulation on central metabolism of R. rubrum has been analyzed by using transcriptomic approaches such as reverse-transcription quantitative PCR. Results show an activity increase of more than four times in CO-metabolic genes of R. rubrum independently of the carbon source used (malate or acetate), thus demonstrating the stimulating effect of the CO from syngas.
R. rubrum was successfully grown with syngas and poly(3HB-co-3HV) was formed from syngas by recombinant strains.
Different media and carbon sources were tested and optimal conditions for aerobic growth of R. rubrum S1 established. The addition of acetate to the cells grown on malate and fructose improved PHB yields in R. rubrum 3-fold and 8-fold, respectively.
Proteomic analysis of aerobically grown Rhodospirillum rubrum S1 cells generated 2077 high-confidence (≥0.99 Protein Prophet score) protein identifications. The number of exclusive proteins detected in fructose/acetate and in malate/acetate grown cells was only 33 and 21, respectively. These proteins are currently investigated in more detail and will be compared to anaerobically grown R. rubrum cells.
Based on the comparison of the proteome of R. rubrum grown on different substrates under PHB accumulating conditions, a potential target protein and pathway possibly affecting PHB accumulation was identified. Additionally, to completely understand the physiology of PHB accumulation and the natural role of this biopolymer in R. rubrum, a deletion mutant with impaired PHB accumulation is actually investigated.
Studies with bacteria of the genus Oligotropha and Ralstonia:
The genome of Oligotropha carboxidovorans OM5 encodes two potential PHA synthases with the locus tags OCAR_4862 and OCAR_6012. RT-PCR studies revealed the active expression of the PHA synthase gen of the locus tag OCAR_6012. This result is important for the general target to genetically engineer O. carboxidovorans to accumulate a higher amount of PHB than the wild type strain. Further cloning and expression studies are currently on the way to achieve this target.
Overexpression of the phaC gene from O. carboxidovorans apparently leads to a significant increase in accumulated PHB.
Three genes encoding mcl-PHA synthesis related enzymes in pseudomonades were heterologously expressed in O. carboxidovorans. Since the efforts to create a PHB negative mutant of O. carboxidovorans were unsuccessful, it was not possible to generate a strain which would accumulate solely mcl-PHA. O. carboxidovorans pBBR1MCS-2::phaGPp::phaC1Pp::pp0763Pp expressing Pseudomonas putida KT2440 genes phaG, phaC1 and PP_0763 accumulated traces of polyhydroxydecanoate (PHD) when cultivated in NB medium with added sodium acetate.
Transfer of the capacity of using CO as a carbon source to aerobic bacteria:
Design and construction of a cox cassette (a genetic cassette including the genes necessary to use CO as carbon source) in a broad-host range plasmid (pBBR1-cox) that can be transferred to different bacterial hosts. In that way, a newly created recombinant Ralstonia eutropha strain H16-cox (containing the transferrable cox cassette) was able to grow with CO as sole carbon source. Without this cassette the strain is not able to use CO as substrate. This demonstrated that the cox cassette is functional. The cox cassette is the first genetic device that allows the expansion of the catabolic potential of some bacteria to use CO as sole carbon and energy source.
Additionally, identification and study of the transcriptional promoters and regulators of the cox cluster from O. carboxidovorans OM5 are driven further to design and construct a new CO biosensor.
Part of these results have already been published and are accessible at:
http://dx.doi.org/10.3389/fmicb.2014.00379
http://dx.doi.org/10.1093/femsle/fnv038
http://dx.doi.org/10.1016/j.copbio.2015.03.008
http://dx.doi.org/10.1002/jctb.4721
http://dx.doi.org/10.15255/CABEQ.2014.2249
http://dx.doi.org/10.1111/1462-2920.13087
http://dx.doi.org/10.1038/srep24381
http://dx.doi.org/10.1111/1751-7915.12409
http://dx.doi.org/10.1111/1751-7915.12393
http://dx.doi.org/10.1111/1751-7915.12411
http://dx.doi.org/10.1016/j.mimet.2016.10.003
http://dx.doi.org/10.1007/s00253-016-7711-5
http://dx.doi.org/10.1128/AEM.01744-16
http://dx.doi.org/10.1007/8623_2015_168
Results of the works on fermentation design:
These works were addressed to increase the efficiency of the bioconversion of syngas produced into two types of bioproducts: building blocks such as succinate, butanediol and hydroxybuturate and biopolymers such as polyhydroxyalkanoates (PHA). These works were therefore focused in the development of different bioreactor configurations and operational conditions for the anaerobic fermentation of syngas using two different types of microorganisms Clostridium and Rhodospirillum. To orient and optimize the operational conditions we used also the recommendations provided by the results from Systems Biology analyses.
The following general results have been obtained:
- A safe lab zone environment (including fume hoods and gas detectors) for fermentations with syngas was designed and implemented (Fig. 5). Two Labfors 5 bioreactors (3.6 and 13 L) were built and equipped with gas analyzer and operating systems for fermentations with R. rubrum. Standardized working procedures for fermenting with syngas were established. Later progress was achieved with respect to fermentation infrastructure (e.g. characterization of gas transfer, cleaning in place), the fermentation itself (growth on synthetic acetate containing medium), and fermentation analytics (e.g. online PHB quantification, determination of dissolved CO).
- A dual-stain method for measuring PHA in the bacterial cultures by use of the flow-cytometer was established.
- SYNPOL partners demonstrated the need to add acetate to the cultivation medium to achieve PHB accumulation during syngas fermentation with R. rubrum anaerobically in the dark. With acetate, the PHA content reached up to 27% CDW (cellular dry weight) at early stationary phase and was composed of mainly PHB with traces of PHV. Yeast extract addition is not required for growing of R. rubrum in syngas under the same conditions.
- SYNPOL provided a basic engineering design for a syngas simulator, including a pre-treatment unit allowing syngas conditioning regarding the requirements of subsequent syngas fermentation processes.
- The fermentation with a recombinant Clostridium strain resulted in a first production of 0.1 g x L-1 2,3-butanediol in addition to 0.75 g x L-1 ethanol.
- Very important results were found for the biosynthesis of poly(3-hydroxyalkanoate) by R. rubrum from syngas. A detailed study carried out revealed that a direct production of PHB from syngas. Nevertheless, for a successful PHB production acetate needs to be added. Moreover, syngas assimilation into biomass and PHB is demonstrated by metabolomic analyses.
Part of these results have already been published and are accessible at:
http://ac.els-cdn.com/S187167841631576X/dx.doi.org/10.1016/j.nbt.2016.06.793
http://dx.doi.org/10.1016/j.mimet.2016.10.003
Results of the works on biopolymer design:
These works had the key objective to synthesize biopolymers with well-defined structures and improved properties by utilizing the monomers and polymers created during syngas fermentation by using chemical and enzymatic catalysis. For the biopolymer synthesis two different approaches were utilized; firstly polymerization of the monomers generated in the SYNPOL fermentation processes and secondly polymerization of functional monomers and oligomers created through chemical recycling of PHAs. These oligomers were subsequently homo- or copolymerized into new materials, minimizing the need of virgin chemicals. In addition, the inherent PHA properties were improved through plasticization, blending, and by creating composites. Polymers with new properties were obtained while maintaining the green character to target new applications and markets.
The following general results have been obtained:
- Successful syntheses of the following three different homopolyesters and one copolyester was conducted by SYNPOL partners: poly(butylene succinate), PBS; poly(propylene succinate), PPS; poly(butylene adipate), PBA; and poly(butylene succinate-co-butylene adipate), PBSA. They were performed by a two-stage (esterification and trans-esterification) melt polycondensation catalyzed by titanium (IV) isopropoxide. The characteristics of these polyesters were shown. For example, PBSA copolyesters have very different properties (melting temperature, glass transition temperature [Tg], crystallinity, mechanical properties, biodegradability, etc.) depending on the succinic acid:adipic acid (SA:AA) molar ratio in the reactor. Solubility tests showed that the polyesters synthesized (PBS, PBA, PPS and PBSA) are only soluble at a concentration of 20 mg/mL in few halogenated solvents such as chloroform, dichloromethane and hexafluoropropan-2-ol (HFIP). PPS, PBA and PBSA [50:50] are also soluble in 1,2-dichlorobenzene.
- SYNPOL partner have successfully synthesized poly-and oligoesters with different combinations of building block monomers such as 1,4-butanediol, adipic acid, and succinic acid by utilizing both a green organic catalyst (1,5,7-Triazabicyclo[4.4.0]dec-5-ene) and a conventional catalyst (titanium isopropoxide). The kinetics of the homo- and copolymerizations showed a typical two-step behavior and the desired molecular weight range could be achieved using both catalysts. Based on the kinetics PBA, PBS, and PBSA with two different theoretical molecular weights i.e. 1000 and 2000 g/mol were synthesized.
- In addition, the plasticization efficiency was evaluated in PVC blends. Good plasticization effect for PBA and PBSA as indicated both by the decrease in Tg and increased elongation-at-break was determined. The order of increased plasticization effect was PBS << PBSA < PBA.
- In other studies, the molecular weight, dispersity and thermal properties of acrylic acid (AA) and crotonic acid (CA) grafted PLA and PVC materials has been determined. It is worth to mention here, that for the PVC-based materials the strain-at-break increased more than 25 times the value for the grafted polymers as compared to the neat PVC although the Tg was virtually unchanged. This is highly interesting and unusual since the maximum usage temperature for an amorphous material is generally dictated by the materials Tg.
- Additionally, we determined the mechanical properties and glass transition temperature (Tg) for the PVC films with anchored plasticizers using either grafted AA or CA as anchoring sites and PBSA as plasticizer. All materials displayed smooth surfaces indicating homogenous and amorphous materials that can be used for different applications.
- The same partner has completed at the end of the project the full characterization of previously synthesized poly(butylene succinate-co-adipate) (PBSA) grafted PLA (PLA-g-PBSA) and the physical blends of PLA/PBSA. The grafted materials illustrated promising mechanical properties, crystallization behavior and improved migration resistance.
- Finally, this partner developed a gradient NMR method to give final proof of the successful grafting of PHB oligomers into PLA chains.
The study of PHA plasticized with different plasticizers with the aim of understanding the influence of the type of plasticizer and its concentration in the formulation over the final properties of the material was also carried out. Poly-3-hydroxybutyrate-co-4-hydroxybutyrate (P(3HB-co-4HB) was chosen as model PHA for its interesting mechanical properties and wide range of applications including biomedical and drug delivery systems. Three commercial plasticizers were studied for the P(3HB-co-4HB) matrix: Citroflex A4, Polysorb ID37, and Grindsted Soft-n-Safe at various compositions. The chemical nature of the plasticizers was found to affect the polymer-plasticizer interactions and hence, the final properties of the material. Interestingly, the long chain fatty acid moieties of both bio-based plasticizers Polysorb ID37 and Soft-n-Safe Danisco were found to provide better interactions with the polymer and therefore result in better plasticization as observed from the tensile strength studies and negligible weight loss at constant temperature. The Grindsted Soft-n-Safe was slightly better than Polysorb ID37 given its higher temperatures of degradation that allow minimal loss in degradation of P(3HB-co-4HB) during processing. Citroflex A4 was found to be least effective of the three plasticizers as it made the PHA susceptible to degradation at lower temperatures and also resulted in phase separation and exudation upon ageing (with heating). The preferred compositions for the plasticizers were found to be 13% for Polysorb ID37 and Citroflex A4 while for Soft-n-Safe Danisco the 13% and 17% compositions were both very good.
In order to evaluate various bio-based polymers on the mechanical and barrier properties and biodegradation of commercial PHB polymers, one industrial SYNPOL partner prepared blends with polycaprolactone (PCL), polylactic acid (PLA) and mcl-PHA produced by Bioplastech Ltd. company (Dublin, Ireland). Blends prepared with 50:50 PLA and Biomer PHB showed maximum improvement of the mechanical properties. In particular the % strain for 50:50 PLA/PHB blend increased to 232% compared to pure PHB (24%) and pure PLA (12%).
The same SYNPOL partner also evaluated the effect of various nucleating agents on the mechanical and barrier properties of PHB. Selected nucleating agents were melt-processed with the commercial PHB ENMAT Y100. The initial results indicated that addition of bio-based polymers such as mcl-PHA and PLA would improve the mechanical properties of PHB in particular % strain which is one of critical parameter to produce packaging films.
One SYNPOL partner developed PHB-PLA blends by melt mixing that showed drastically improved mechanical properties when plasticized by acetyl tributyl citrate. Plasticized PHB-PLA blends properties could be improved even more by addition of nano-crystalline cellulose or organo-modified nanoclay.
Later in the project, one SYNPOL partner successfully synthesized copolyesters and terpolyesters from SYNPOL building blocks using both organometallic and enzymatic catalysts.
Another partner has used SYNPOL building block crotonic acid obtained from PHB recycling to graft biopolyesters segments on PLA. In addition, the oligo-PHB building blocks from PHB recycling/degradation were successfully used as grafted plasticizer for PLA.
We also investigated the efficiency of a bio-based plasticizer for PHB compared to non-renewable analogues. In order to improve PHB properties, a SYNPOL partner prepared nanocomposites with various types of nanofillers.
One SYNPOL partner synthesized copolyesters from SYNPOL building blocks (adipic acid, 1,4-butanediol and 2,3-butanediol) using both organometallic and enzymatic catalysis. Despite lower reactivity of 2,3 BDO, optimized polymerization conditions allow to control and tune copolyesters composition and properties.
PHB diol oligomers were successfully used as building blocks to synthesize copolyesters and poly(ester urethanes) using both organometallic and enzymatic catalysis and it was demonstrated the possibility to control copolyesters structure (random or blocky tendency) and properties according to the PHB diol length and polymerization conditions.
Finally, PHA-based nanocomposites based on montmorillonite and sepiolite clays were elaborated and organo-modified sepiolite gave the best mechanical properties as a result of improved dispersion and strong interactions with the matrix.
Part of these results have already been published and are accessible at:
http://dx.doi.org/10.1021/sc500397h
http://dx.doi.org/10.1021/acs.macromol.5b00235
http://dx.doi.org/10.1002/mame.201500026
http://dx.doi.org/10.3390/ma9050313
http://dx.doi.org/10.1016/j.polymer.2016.07.022
http://dx.doi.org/10.1021/acs.biomac.6b01494
Results of the works on biopolymer degradation and life cycle:
These studies analyzed the degradation of the polymers synthesized in the SYNPOL project using different approaches, taking into account environmental conditions as well as the use of enzymes or organisms that could be useful for the biodegradation and/or recycling. Further works studied the development of processes for chemical and biological recycling of biopolymers for a waste-free society.
The following general results have been obtained:
- In a task called “Chemical recycling of PHAs to functional building blocks”, an effective and rapid process was developed for the chemical recycling of PHB through microwave assisted reactions in alkaline methanol, which resulted mainly in the monomeric degradation products, 3-hydroxubutyric acid and 3-methoxybutyric acid. The most effective and efficient microwave-assisted degradation conditions were obtained in 0.5% (w/w) alkaline methanol solution heated at 110°C. Additionally, PHB has been thermally degraded to crotonic acid and oligomers with crotonic acid end groups.
- In the same task, a potential route for feedstock recycling of PHB and a highly promising approach for fully biobased and biodegradable toughened PLA through covalently anchored oligomeric PHB plasticizers has been developed and demonstrated. This process consisted of a commercially viable and green two step extrusion process. PLA with 20% (w/w) grafted dPHB demonstrated an impressive elongation at break of 538%, 66 times higher than that of pure PLA.
- Biodegradation tests within the task “Biodegradability, organic recycling and ecotoxicity evaluation of developed biopolymers” were started on materials delivered by SYNPOL partners. The materials consisted of PHA mixed with nanoclays. Therefore, the degradation of neat PHA, PHA + 3% Sepiolite clay, PHA + 3% Cloisite Na clay and PHA + 3% C30B clay in freshwater and soil was evaluated. The tests are still running, but first observations show that biodegradation rate in soil of the PHA mixed with nanoclays is quite similar to neat PHA and after 14 days a biodegradation around 50% was observed. These soil biodegradation tests have a standard duration of 120 days. In freshwater conditions also high biodegradation was observed for the different materials as neat PHA reached a biodegradation of 79.7% after 28 days, while a value of 79.1% and 75.8% was measured for PHA + Cloisite Na clay and PHA + C30B clay. For PHA + Sepiolite clay a lower degradation rate was measured, reaching a biodegradation of 68.1% after 28 days.
- In two further (out of many) biodegradability examples, PLA/o-PHB and PLA-g-o-PHB (25 g each sample) were used for composting experiments. The oligomeric PHB was prepared by thermal recycling of PHB, a model SYNPOL polymer and the material was prepared under an earlier task. Under controlled composting conditions (58°C) complete biodegradation could be established on compounds PLA20G (PLA-graft-PHB) and PLA20B (PLA/PHB blend).
- In the task about ecotoxicity, the solid fraction (char) after the pyrolysis of wet and dried municipal solid waste and straw was examined for plant toxicity on barley and cress plants. The porous structure of these chars has been found out to be one of the possible factors related to the germination rates. This alternative has resulted to be highly promising since the toxic effect on plants germination was very limited or non-existing regardless of the municipal waste char composition.
- 11 data sheets from SYNPOL biopolymer products have been made by three SYNPOL partners. These data sheets are for the following novel bioplastic products: PLA-graft-PHB, PLA/dPHB-blend, PB67B’33A, Poly(3-hydroxybutyrate-co-4-hydroxybutyrate)-sepiolite nano-biocomposites, BIOPLASTECH 1, BIOPLASTECH 2, BIOPLASTECH 3, BIOPLASTECH 4, BIOPLASTECH 5, PHO, and PSA70). The data sheets contain information on production, thermal and mechanical properties, biodegradation, advantages and possible applications and will be publicly available soon.
- The environmental impact of the novel biotechnological production pathways developed within the framework of the SYNPOL project was also examined in LCA studies (Fig. 6). This impact was assessed in 16 effect categories in line with ILCD midpoint method (EC-JRC 2011) and ISO 14040/44 and ILCD Handbook guidelines. Both the total life cycle impact (cradle-to-grave, excluding use phase) and the life cycle hot spots (cradle-to-factory gate) are identified. The total impact results of the SYNPOL processes are compared to their conventional equivalents.
- There are two products developed in the project: a building block 2,3-butanediol (2,3-BD) and a biopolymer polyhydroxybutyrate (PHB). The common step of both production processes is microwave induced pyrolysis (MIP), which transfers pre-dried municipal solid waste (MSWd) into pyrolysis gas (mixture of mainly H2, CO, CO2). This gas is fed to the fermenter where bacteria (Clostridium ljungdahlii for 2,3-BD and Rhodospirillum rubrum for PHB) use it as a carbon source in order to metabolically produce desired substances. Subsequently, the fermentation broth follows different downstream pathways.
- The functional unit (FU) chosen for the whole production chain is 1 kg produced chemical. Despite very complex downstream pathway, production of pure 2,3-BD was unsuccessful. The final stream (70% moisture) contains still a vast amount of impurities (81% dry mass basis). Hence, 1 kg 2,3-BD diluted in the final stream was chosen as FU. The downstream pathway for PHB production was developed and pure PHB was successfully obtained at the end. It was observed that in both cases the biggest hotspot is associated with low mass transfer between CO and water and low efficiency of fermentation process. Consequently, a lot of gas needs to be produced via MIP (to provide enough CO to microorganisms), which is directly linked to the relatively high electricity consumption of this novel technique. At the same time a vast amount of unused gases needs to be flared, as a result of a small CO mass transfer into water, which leads to significant CO2 release to the atmosphere. MIP is very important contributor to the life cycle impact of the SYNPOL production process, but also in terms of possible benefits, associated with avoided municipal waste incineration. This is especially visible in human toxicity and freshwater ecotoxicity impact.
- The sensitivity analysis was performed in order to look for possible improvements. It showed that the model is sensitive to changes in MIP electricity consumption and waste gas treatment methods.
- The environmental impacts associated with the studied building block and biopolymer production can be effectively improved by optimization of CO mass transfer into water, which will increase the amount of CO available for microorganism enhancing their efficiency and thus yields of interesting chemicals. At the end of fermentation process, the waste gas should be flared, best with electricity recovery so conventional electricity production could be avoided. Additionally, further research is recommended in order to allow recycling of pyrolysis gas at the fermentation stage. This goes hand in hand with better CO mass transfer into water and would lower the impact of MIP and reduce gas emissions. Furthermore, it should be investigated if reduction in electricity consumption by MIP is possible in order to make it more environmentally interesting than conventional techniques.
- From the life cycle assessment it can be concluded that several factors should be improved (also when compared with alternative production processes). Within the life cycle of 2,3-BD production, flash evaporation is mainly responsible for the impact on photochemical ozone formation due to significant energy consumption.
- Within the life cycle of PHB production, used solvent (dichloromethane) carries significant impact on ozone depletion. It is advised to replace it with more environmentally friendly solvent in further research. The remaining impact of the solvent and chemicals use was avoided due to introduction of partial recovery and recycling step at the end of the downstream process. Final filtration and chemicals recovery process are mainly responsible for the impact on photochemical ozone formation due to significant energy consumption.
- Both processes have the same main bottleneck. Mass transfer between CO and water is very low, hence very little amount of CO from pyrolysis gas is available for microorganisms during fermentation. Consequently the final yields are very low and the fermentation process is not efficient. This is especially visible when compared to reference scenarios. This means that at this point the SYNPOL processes cannot compete with conventional production pathways. Both processes are sensitive to changes in electricity consumption by MIP and to treatment of waste gas.
- Pyrolysis is very important contributor to the life cycle impact of the SYNPOL production process, but also in terms of possible benefits, associated with avoided municipal waste incineration. This is especially visible in human toxicity and freshwater ecotoxicity mitigated impact. Electricity consumption is the most important factor determining the impact of MIP. Four different feedstocks were analysed and it turned out that MSWd is the most interesting one for pyrolysis, due to small amount of produced waste (solid and liquid) and high CO flowrate. Additionally, it was observed that MIP at 800°C is more environmentally friendly than at 400°C.
- The impact of conventional pyrolysis (CP) on climate change is smaller than of MIP, mainly due to higher electricity consumption by the latter. However, taking into account all 16 effect categories, MIP scores better in most of them (11 out of 16), mainly due to much lower pollution associated with by-products (chars and tars).
Part of the ecotoxicity results have already been published and are accessible at:
http://dx.doi.org/10.1016/j.jaap.2016.08.013
Results of the works on demonstration of SYNPOL biopolymers:
The objectives of the demonstration part of the project were:
➢ to determine the commercial application of the novel PHA nanocomposites and PHA blends
➢ the best performing materials will be processed in sufficient quantities to allow full materials characterization study
➢ preparation of a biopolymer production plant design taking into account:
• optimal process performance with fermentation and downstream operational data at large scale
• minimum energy requirements
• reduced environmental impacts
• end-to-end economical evaluation
The following results were obtained during the final phase of the project:
- The up-scaling of fermentation and downstream processes for the production of 2,3-butanediol from syngas was explained. In a first stage, laboratory scale-up was developed by one SYNPOL partner. Later on, projection on industrial scale was carried out by another SYNPOL partner. A basic process diagram, equipment, utilities and ancillary equipment are described. Further on, analysis of bottlenecks of the processes is provided in the public available deliverable D8.5 (“Fermentation and downstream operational data at large scale”).
- A demonstration plant capacity for the production of 2,3-butanediol has been designed according to the usual ratios for products with a low economic value. For details see the public available deliverable D8.4 (“Pilot plant - Biopolymer production plant design and economic viability”).
- Three SYNPOL prototypes have been successfully produced by SYNPOL partners and are described in the public available deliverables D8.1 (“Film for packaging applications made of Bioplastech mclPHA blended with SYNPOL PHB”). D8.2 (“An electrospun composite of SYNPOL PBS and Bioplastech mclPHA to form a scaffold for biomedical applications”) and D8.3 (“Biodegradable adhesive based on a blend of Bioplastech mclPHA and SYNPOL polyester urethanes”). As an example, Fig.7 shows a compostable paper bag made of crafting paper and cellulose film window utilizing the 100% biobased adhesive (from SYNPOL biopolymers) for its assembling.
- The novel SYNPOL biopolymers prototypes will be patented in the future and therefore, no official publications have been produced within this demonstration task of the SYNPOL project.
Potential Impact:
Description of the potential impact
Waste is actually a pressing environmental, social and economic issue. Increasing consumption and a developing economy continue to generate large amounts of waste - with more effort required to reduce and prevent it. While waste was viewed as disposable in the past, today it is increasingly recognized as a resource; this is reflected in the waste management shift away from disposal towards recycling and recovery. Therefore, SYNPOL’s valorisation of anthropogenic derived wastes, as raw materials for the industrial production of polymers and chemicals, provided an effective response to the challenge of sustainable production of these materials, while in parallel handled the vast environmental and economic problem of wastes disposal.
Within the context of an ‘integrated operation’ there are multiple objectives satisfied in parallel at the benefit of environment, sustainability and bioeconomy. Therefore, the socioeconomic impact of SYNPOL project can be evaluated from different perspectives:
ϖ - Utilization/valorisation of the complex biowastes for the production of high-added value products like biopolymers
ϖ - Replacement of common chemically catalysed production processes for the transformation of syngas with eco-efficient microbial ones
ϖ - Reduction of the volume and polluting load of the complex biowastes
ϖ - Decrease of the waste treatment costs
ϖ - Establishment of the technological developments for the formation of renewable raw materials for the polymer industry contributing to its modernization, sustainability, consolidation and independence from the availability of fossil feedstocks
ϖ - Replacement of conventional polymer materials with biodegradable ones extending the range of polymer applications to food, biomedicine and pharmaceutics
ϖ - Production of novel biopolymer building blocks from renewable sources opening an increasing window-of-opportunity for bio-based products for the chemical industry and creating new markets to the manufacturers of pharmaceutical and nutritional care products
ϖ - Bridging the gap between the novel biomaterial production technologies and their industrial/commercial application and contribution to the faster satisfaction of the humanity need for renewable biopolymers that can be composted after use in contrast to conventional plastics
Enhanced environmental care and ecosystem protection both facilitating the safe and within the regulations discharge of wastes from colossal industries and contributing to the preservation of carbon balance in the atmosphere. This is due to the fact that the carbon sources used for the biotechnological production of biopolymers derived mainly from carbon dioxide that was previously fixed by plants. Hence, the release of CO2 by degradation of biopolymers is just the final step of natural mineralization.
Perhaps the greatest benefit generated by this project was traced with respect to the environment and the global climate through the reduced emission of fossil-carbon derived CO2. By the industrial implementation of the biochemical routes for the waste bioconversion to biopolymers and biochemicals, substantial reductions in greenhouse gas emissions could be realized. The more the produced technology is established and the proposed bio-based products replace the conventional existing ones, we could assume that the emission reduction can be of the order of 15-50%, which is a step closer to the satisfaction of global Kyoto target. Obviously, all these promises of the project with great impact on different sectors can only be realised provided the recognition and the adaptation of the produced technology by the industry, hence, the developments over the scaling-up capability of the produced technologies and the market value of the new products is the biggest challenge. The know-how gained from SYNPOL project can support any plans for large scale implementation of biopolymers and “green” chemicals.
To further appraise the project impact through the proposed cost- and eco-efficient production of bio-based polymeric materials we should contemplate the problems of the greenhouse effect and the global warming that the conventional polymers are highly linked with due to the utilization of constrained fossil resources for their production.
The main problem arising from incineration of plastics is the same as for energy recovery from fossil feedstocks: carbon that was fixed during millions of years and within this time was not part of the natural carbon cycle is converted to CO2, which can accumulate in the atmosphere, contributing to the mentioned climatic effects. In addition, incineration of plastics often generates toxic compounds. Besides, more and more waste of highly resistant plastics that are not incinerated is piled up every year. Recycling systems demand a certain degree of purity and a high sorting accuracy. In addition, the collection costs are fairly high, and recycling has a negative impact on the quality of the materials, such as an increase in brittleness of the recycled material.
In addition to these ecological considerations, the price of crude oil is unpredictably fluctuating, not least due to miscellaneous developments in the global political situation. This constitutes a factor of immense uncertainty especially for the highly petrol-dependent polymer industry. The technological developments produced by this project can contribute to overcome these problems and confront in parallel the increasingly posed question of how we will derive the materials we will need in the twenty-first century when the contingency of the depletion of fossil feedstock become one day a reality. In front of this contingency, the involved industrial branches (i.e. polymer and chemical industry), which are already aware of the necessity for promoting novel production techniques based on renewable resources, will turn to solutions also provided by the SYNPOL project.
The impact of the project can be also appraised from the perspective of the produced bio-based materials. It should be noted that the EU polymers and chemicals production sector is an industry in transition. This is a consequence of changing domestic demand and the pressures of globalization, both of which are driving the industry towards the manufacture of higher added value products and sustainable production. “Green” and thus biodegradable polymers and biochemicals from renewable resources is the technological response to the above challenge.
In addition to the project’s impact on the polymer industry, the contribution to a sustainable production of building blocks will strengthen the link of the biochemical industry with the pharmaceutical, nutritional and agricultural products industries creating an opportunity for the fine chemicals companies for growth and invigoration. These high added value biochemical products have the potential to become a significant part of EU’s economy in the future.
There is little doubt that the waste-based bioprocessing technology has great market potential both in replacement for current processes related to fossil resources, as well as in the creation of new markets through its sustainable, efficient and economically viable production of biopolymers and high value-added chemicals. Moreover, a successful research outcome in the production of these bio-based polymers and chemicals will enormously enhance the competitiveness and sustainability of the European biopolymer and biochemical industry and will generate new business opportunities. As “green” technology is making the industry more sustainable, it is expected that benefits will also be seen across a range of critical society-based areas representing other profound fields of economy.
The SYNPOL project responded to all these objectives with the technological advances described above. Precisely, in addition to cost reduction achieved by the efficient transformation of the low-cost complex biowastes in syngas as fermentation substrate, there are several lines of evidence supporting the feasibility of this objective, such as the genomic modification of the selected bacteria and the fermentation strategies properly designed for the selected bacteria. The synergistic action of the project approaches along all the routes together, (i.e. raw materials, microbial producers and operation strategies) generate the best outcome with respect to the low-cost production of biopolymers and bioconversion of wastes into high value-added products (Fig. 8).
To ensure the successful accomplishment of the project objectives, a multidisciplinary partnership has been established, consisting of academic and research scientists covering various disciplines and complementary scientific and technical areas (i.e. biotechnology, molecular biology, biochemistry, biosystems and bioprocess engineering, nanotechnology, chemical and process systems engineering, environmental engineering, etc.). These FP7 partnerships are also fruitful for future cooperation in further European projects, and indeed, some partners are actually working together again in other H2020 projects concerning the use of gases as feedstock or in related bioeconomy projects.
European dimension
The core idea of the SYNPOL project for a profitable utilization of selected complex biowastes for the production of bio-based products represents a strategic initiative for Europe in the area of biopolymers and high-added value biochemicals. The tactical decision on the complex biowastes, as raw materials, and the way to convert them into targeted products by fermentation of syngas reflects the needs, the capabilities and deficiencies as well of Europe, as a continent with low fossil-oil resources, high fossil-oil requirements though, abundant amount of the selected biowastes, legislations with intense environmental concern, political desire for imported-oil independence and capability to lead the world biopolymer and biochemical industry.
The various works for development of a novel and efficient industrial biowaste processing and valorisation technology, performed within the framework of SYNPOL project, successfully addressed community societal issues pertaining to the quality of life of European citizens by protecting the environment, climate, and increasing the energy resources via the use of innovative waste processing systems.
In order to be able to sustain the EU position, huge advances in the environmental control must be achieved. Such advances (i.e. CO2 mitigation and GHG reduction) would make it possible for the European members to successfully face the challenge of effective protection of the environment at affordable cost. In this sense, the SYNPOL project had a truly European dimension, as it served to protect and possibly enhance the European members’ position as the most ecologically sensitized.
Biobased products can replace 1:1 their chemically derived counterpart if the cost is competitive. Moreover, biobased products can bring new properties and functions to materials and chemical intermediates. This characteristic can enable a chain growth and development to new sectors as well as to already existing highly profitable ones such as the pharmaceutical and nutritional products industry, with the European representatives being of the pioneer ones world widely.
Community societal objectives
The current challenges in the area of renewable biodegradable polymers and biochemicals are to design, produce at a competitive cost and use industrial waste sources that satisfy the energy needs of the society and ensure an improved quality of life, whilst maintaining the capacity of the natural environment to renew itself. As a result of project exploitation, the industrial partners involved in the SYNPOL project benefit from the novel technological solutions and related markets that will improve their short and long-term profit. The research and academic centres scientifically benefited from the collaboration with each partner of the consortium exchanging know-how and creating background knowledge in a field that is anticipated to be at the edge of the technology. Finally, all partners had synergistically worked with common target the extroversion of the consortium as a whole, in order to make the scientific and technological developments known to the related industries and attract their investing interest for the commercial application of the produced technology. Innovation and intellectual property rights provide economic opportunities such as establishment of spin-off companies. The society at large will benefit from the innovations in different areas (e.g. energy, environment, and climate).
Additionally, the present project provided the core knowledge for potential exploitation of different industrial biowastes further promoting to the “integration” and link of heterogeneous industrial sectors and expanding their business opportunities. Moreover, it will have drastic impacts on all societal, economic, ethical, legislative and educational aspects of everyday life, since it supports innovative, fundamental research in the science and technology of the production of biobased materials leading to potential breakthroughs primarily to the directly involved industrial sector (i.e. biopolymer and biochemical industry) as well as to the indirectly involved one as recipient of these technological developments (e.g. pharmaceutical and food industry).
Main dissemination activities and the exploitation of results
Translation of basic foreground into popularised communications or executive summaries was handled by the dissemination work package of the project and freely distributed to all concerned stakeholders within and outside the EU (see summary of dissemination activities in the automatic generated table from the Participant Portal).
The scientific and technological expertise gained was disseminated through scientific publications, patents, presentations at conferences and press releases. Knowledge acquired was also used for advanced training purposes and as an input to further research and industrial applications. In order to promote academic and industrial cross-fertilization within the partners’ organizations and towards the ERA we have established a number of dissemination activities. In this sense it is worth mentioning that special attention will be devoted to the dissemination of the social benefits derived from the SYNPOL technology concerning to the transformation of different biowastes into recyclable bioplastics. We are aware of the enormous social, economical and technical problem involving the accumulation of wastes and of the great benefices that will provide the technology developed in this project.
To fulfil these goals the following dissemination activities have been carried out:
1. We created different Communication Materials (website, flyer & brochure) that was useful for improving not only partners’ communications but also the communication with other R&D initiatives and with potential partners interested in participating in or supporting the project activities:
- A specific internet website for the project (www.synpol.org). This tool was one of the elements to integrate the complete dissemination plan of the project. This internet web site was used to communicate the advantages of SYNPOL technology to scientific or other stakeholders at both European and international levels. The web page displayed links with other related and collaborating EU projects. Apart from being an effective way of advertising our research and main achievements, the website was used to promote events and publications or even to distribute key messages on the home page. The partners contributed scientific contents in order to ensure its continuous update and extension. The web page, which was updated more or less weekly, allowed and still allows the interested persons accessing the website to be informed on the project developments. Thus, the website supported a close co-operation between the consortium partners and helped raise public awareness.
- Information brochures and flyer on SYNPOL technology. A brochure and a flyer were produced presenting the SYNPOL project. It was intended to support the presentations at events and the individual meetings carried out and it served as a communication tool for potential mailings to target audiences. Flyer and brochure were project oriented, as both materials described the project, its objectives, the participating organisations, the expected results and the activities planned.
- SYNPOL updates were also published in the institutional or company web sites of nearly all beneficiaries. The CSIC website for example and the other available web sites have a great exposure to public and they constitute an excellent complementary dissemination mechanism apart from our own SYNPOL web site.
2. The SYNPOL web site also offered links to open the project to Social Networks. Therefore, we created SYNPOL accounts in the most important social networks such as Facebook and Twitter. The social networks provided us the opportunity to connect not only with the citizens but also with the companies and scientists interested in this technology.
3. Plenary lectures and workshops. We organized as satellite events of the annual General Assembly meetings (12, 24, 36 and 48 months) four specific workshops to invite technology users of SYNPOL achievements, including keynote lectures from partners as well as for specially invited speakers. Additionally, 3 further workshops (one in Spain [2014] and two in Switzerland [2014 & 2016]) on biotechnology issues were organized and carried out by SYNPOL beneficiaries.
4. The activities developed in the project were disseminated also by stimulating the presentation of results with poster and oral presentations at scientific international conferences, congresses, workshops, and meetings. The absolute scientific dissemination highlight for the SYNPOL project can be seen in the 15th edition of the International Symposium on Biopolymers 2016 (www.isbp2016.com) that was co-organized (and financially supported) by the SYNPOL project and that was held during the final week of the project from 26th to 29th of September 2016 in Madrid, Spain. The ISBP2016 was the final and main dissemination platform for the SYNPOL project and more than half of all SYNPOL consortium partners have presented their results and perspectives at this congress.
5. The publication of results in high impact scientific journals and books was also encouraged and successfully done by many SYNPOL partners (see automatically generated list of publications in the Participant Portal).
6. Theses; at the end of the project time (30th of September 2016) one PhD thesis can be announced as terminated. Title: “Strategies for utilizing biobased and recycled resources for polylactide plasticization”, Author: Xi Yang, Date of approval: 16/09/2016 at KTH - Royal Institute of Technology - School of Chemical Science & Engineering10044 Stockholm (Sweden). Further PhD theses will be defended late 2016 or in 2017 by doctoral candidates from the SYNPOL partner universities of Ulm (Germany), Münster (Germany), Manchester (UK), Amsterdam (The Netherlands), Strasbourg (France), HES-SO (Sion, Switzerland), and Dublin (Ireland).
7. In coordination with the press offices of the partners´ institutions we tried to transmit the results and advantages of SYNPOL technology to the journalists through radio interviews (2), videos (2) and specialized articles (5). A few press releases in different languages have been made and monitored as an indicator of dissemination of project results.
8. To improve the training of the personnel involved in the project, to stimulate and promote the transfer of knowledge and to increase the cooperation within the partners, we established 15 specific exchange/mobility activities for interchanging personnel between the academic institutions and the companies. This interaction favoured the flow of ideas across the partner laboratories.
9. Announcements of vacant research positions within the project at partner’s laboratories have been published on the website a few times.
10. General public open-doors activities. Some partners promoted participation of SYNPOL in activities to promote as “open days”, “science fairs” and “meet the scientist” presentations and and participated in workshops to train high-school teachers. These activities targeted to the public led to a better understanding of the efforts carried out by the project and the EU Commission to develop clean and sustainable technologies such as SYNPOL to efficiently recycle biowastes.
Dissemination & Exploitation outcomes of the SYNPOL project at a glance:
Patents: 2 (from academic partners, but there are more in the pipeline from the industrial partners)
Spin-off companies: 1 is planned by one industrial partner by the name of CEWAS, Consulting and Engineering technologies for Wastes (Seville, Spain)
Publications: 39 peer-reviewed scientific papers and a few more general project presentation papers (from all academic partners and one industrial partner; there are at least 3 more publications in the pipeline)
Workshops: 3 (2 in Sion, Switzerland and 1 in Madrid, Spain)
Annual SYNPOL courses: 4 (in Valencia, Oviedo, Murcia and Madrid, all Spain) with 50 – 75 participants from many European countries
Congresses: 1 (ISBP2016 from 26th to 29th of September 2016, thus in the final project week)
Open door activities: 4 (3 in Madrid & 1 in Sion, Switzerland)
Poster presentations on congresses: 16
Oral presentations on congresses: 29
PhD theses: 1 defended and thus terminated (7-8 ending soon)
List of Websites:
Website at www.synpol.org
Primary Coordinator Contact: Prof. Dr. José Luis García López (jlgarcia@cib.csic.es)
Coordinator Contact/Project manager: Dr. Oliver Drzyzga (drzyzga@cib.csic.es)
Both at: Centro de Investigaciones Biológicas (CIB) of the Agencia Estatal Consejo Superior de Investigaciones Científicas (CSIC), C/Ramiro de Maeztu 9, 28040 Madrid, Spain; Tel. +34 91 8373 112
