GRAIL e-NEWSLETTER #2 – July 2015

The GRAIL project is a 48-month Collaborative Project with funding by the European  commission under FP7 Programme for Knowledge Based Bio-Economy. The GRAIL project has been built with 15 partners from 9 different countries with the aim of finalising the solutions given previously to the valorization of glycerol and transform then in valuable products in a biorefinery approach.

Glycerol Biorefinery Approach for the Production of High Quality Products of Industrial Value

The global production and consumption of biodiesel is continually increasing, resulting in a stoichiometric increased generation of crude glycerol, due to its co-production in the transesterification process. As a consequence, a vast amount of raw glycerol is generated each year and its value on market is being reduced.
The point of becoming a “waste-stream” rather than a valuable “coproduct”. Glycerol prices fell, generating a bankruptcy of companies that produce glycerol chemically, reducing 10 times the price of glycerol in the market.

Concept of the project

GRAIL project is born aiming to produce a replicable methodology for using economic and scientific arguments to overcome the main scientific, technological and economical barriers to consider crude glycerol as a suitable feedstock for the production of economically value added products. To date, there is no real use for raw glycerol besides from calorific valorisation, which led to an accumulation and as storage or expend as waste cost for the biodiesel corporations. GRAIL, therefore, proposes a “green process” designed for the manufacture of various high value products and biofuels from glycerol side-streams. The GRAIL consortium is focused on the development of known and new types of applications using glycerol as the starting material and this project aims to develop a set of technologies for converting that waste glycerol from biodiesel into 1,3 propanediol, Fatty acid glycerol formal esters, PolyHydroxyAlkanoates (PHA), Hydrogen and Ethanol, Synthetic coatings, powder coating, resins, Secondary Glycerol Amines, Biobutanol, Trehalose, Cyanocobalamin (Vitamin B12), ß-carotene, Docosahexaenoic acid (DHA) and other products.

Project Activities Overview


The GRAIL project aims to convert crude glycerol occurring from the biodiesel process to valuable products in the fields of biofuels, green chemicals and food supplements. Within the activities of WP 1 the availability of crude glycerol is investigated and the requirements of this feedstock for the conversion processes are defined. The development of a purification step for the crude glycerol is discussed in order to meet these requirements. Furthermore, first plant layouts are developed and initial mass and energy balances are prepared for the conversion processes.

In order to determine the amount of crude glycerol available in an acceptable distance, the total amount of biodiesel plants in Europe was determined. The detailed status of the 347 European plants is presented in Figure 1.

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Regarding the operational plants, a capacity of 20.24 Mt/a was calculated, implying a theoretical crude glycerol potential of 2.02 Mt/a. By taking all 347 biodiesel plants into account, the theoretical European biodiesel and glycerol capacity could be determined to a value of 30.26 Mt/a and 3.03 Mt/a, respectively. Under consideration of the nationally evaluated degree of capacity utilisation, a technical crude glycerol potential of 828 000 t/a is realistic in Europe.

Currently, the approach and main results and outcomes of these considerations are published in an international journal and will be available soon.


The overall objective of WP2 concerns the industrial revalorization of glycerol into biofuels by development of biological processes using suitable microorganisms: Microbial Mixed Culture (MMC) and pure culture.

Lab-scale fed-batch experiments in non sterile conditions, without the control of the main parameters of fermentation, and using crude glycerol as the only carbon source, were carried out by ENEA.
All experiments pointed out that the increase in glycerol consumption was the critical parameter, since the fermentation of ITB glycerol by microbial community of GCL showed inhibition effects related to the chemical impurities of ITB glycerol (particularly to acetic acid) as well as to the metabolic products accumulated over time.

PUCV demonstrated the feasibility of selecting a MMC from an aerobic wastewater treatment plant. Batch and continuous fermentations were carried out for improving the production of ethanol and hydrogen. In batch system, the parameters studied were inoculum, inoculum pretreatment, addition of nutrients and substrate concentration. In continuous system, the parameters studied were inoculum, pH, hydraulic residence time (HRT) and substrate concentration.

Experiments for butanol production were carried out by STUBA using pure culture. Clostridium pasteurianum DSM 525 was chosen among four strains as the best butanol producer.
Batch fermentations with 99,5 % partially purified glycerol (Megara Resins) was very similar to experiments with pure glycerol (Figure 2).

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Clostridium pasteurianum DSM 525 was also suitable for cultivation in the continual system (Figure 3 Batch fermentations with C. pasteurianum DSM 525 inoculated with 5 % (v/v) of inoculum. Pure (left) and 99,5 % (right) glycerol was tested. Conditions: volume 1 dm3, 34°C, 200 rpm, pH 6 was kept by 2 M KOH.)

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Based on the obtained results, it was concluded that gene typed strain Clostridium butyricum DSM 10702 is metabolic deficient and therefore other Clostridium butyricum variants were screened for 1,3 propanediol productivities and yields. The fermentation media were optimized in order to achieve higher 1,3 propanediol yield and concentration. Addition of biotin is not necessary, the concentration of FeSO4.7H2O can be lowered to 0.01 g/L and its preparation does not need anaerobic conditions.
It is been concluded that the enrichment of suitable MMCs able to efficiently convert crude glycerol in butyric acid, by comparing different selection strategies and inocula. AAU has started working trying to adapt/enrich mixed microbial consortia to 2nd generation crude glycerol obtained from DAKA biodiesel in DK. Different adaptation strategies and different growth media are currently being tested.
Increased temperature improves the extraction of 1,3-PDO from fermentation broth into an ionic liquid phase, which means that the phase separation become faster (Figure 4).

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By the chemo-catalytic system the selective transformation of 1,3-PDO to C3 and C6 products has successfully been achieved. This system is recyclable.
On the other hand, analysis and characterization of alkyd resins and rosin esters, produced by using purified biodiesel glycerol, gave positive results with regard to the final properties of the targeted products.


During the first period of the GRAIL project within WP4, the main focus was put on establishing the biotechnological fermentation processes in lab scale, in order to obtain the high added value food compounds targeted in the project. Thus, in a first step the preparatory cultures of Blakeslea trispora to produce ß-carotene, Pythium irregulare to produce Eicosapentaenoic acid (EPA), Propionibacterium freudenreichii subspp. shermanii to produce cyancobalamin also known as vitamin B12, and finally Schizochytrium limacinum to produce docosahexaenoic acid (DHA), were done.
Once sufficient amounts of the microorganism were available, optimisations of the fermentation processes in lab-scale were performed, establishing as well the appropriate downstream processing protocols to separate bacteria biomass from the target food compounds. Overall, the results showed that crude glycerol could be used as a carbon source within the fermentation processes; by applying the appropriate conditions, sufficient amounts of the targeted compounds were obtained to allow the continuation of the studies to the up-scale phase.


The aim of WP5 is to demonstrate the viability, at industrial scale, of converting the by-product from the biodiesel production (glycerol) into new valuable feedstock, based on a new portfolio called BIO2.
Based on issued block diagrams (results of the first six months activities), the development of the Basic Design Engineering Package for selected processes has been started.
As first activity data from partners and literature (mainly for processes for which experimental data are not yet available) have been collected in order to go ahead with project development (especially Mass & Energy Balances to be performed by DBFZ/PI).
Preliminary identification of major bottlenecks in processes development at industrial scale has been started by PI and shared with all partners. PI is preparing a general document that will be used as basis for further discussions, also through dedicated meetings.


The overall objective of this activity is to control and support the glycerol-based products processing proposed in the project, taking into account environmental, economic and social aspects in order to foster sustainability. During the First Reporting Period, activities related to process design and optimization, as well as Life Cycle Analysis (LCA) and Life Cost Analysis (LCC) were performed. Initial actions have been conducted to GRAIL processes, mainly based on the evaluation and basic design for a multipurpose process for cost and energy efficient separation.
LCA and LCC methodology was determined including the scope and the system boundaries. Finally, an Environmental Technology Verification will be addressed for those technologies applied from producing glycerol-based products.


This GRAIL public website is a major dissemination tool intended to facilitate the spread of project’s information to different stakeholder groups. It is the core element of the external communication strategy of GRAIL project. The GRAIL website is also linked with some social media such as LinkedIn and Twitter, to improve the visibility of the project.

To this extend, the procedures i.e. the partners’ roles and responsibilities in updating/enriching the content, frequency of updates and improvements (when) and respective tools (how) will be developed. Thanks to this permanent update, the website has now a regular number of visitors each month, who are more than 6800 since the website real launch in April 2014 (more than 500 views a month).

GRAIL has also published two articles:

  • “Processi Innovativi partecipa al Progetto Grail” in the scientific journal Impiantistica Italiana (Italy, August 2014).
  • Comparison of different strategies for selection/adaptation of mixed microbial cultures able to ferment crude glycerol derived from second-generation biodiesel by Varrone, Heggeset, Le, Haugen, Markussen, Skiadas and Gavala in the scientific journal Biomed Research International (New York, in press by now)

The following information describes the main dissemination events of GRAIL during 2015:

  • The 2nd European life cycle assessment workshop: Workshop in Nice (France), led by Vertech (Organiser) and DBFZ (participant) on April 2015. It was focused for the scientific community.
  • Hydrogen transfer initiated dehydration of bio-renewable alcohols (UK Catalysis Conference): Presentation led by QUB in Loughborough (UK) on January 2015. It was also focused for the scientific community.
  • EU Project SENSE: Solutions for harmonized environmental sustainability in the European food chain: Organization of conference in Brussels (Belgium) led by BZN on January 2015 for several industries.
  • Combining Bio- and Chemo-catalysis for the Conversion of Bio-Renewable Alcohols by A.C. Marr (at the 3rd International Symposium on Green Chemistry) led by QUB on May 2015 in France.
  • EU FP7 GRAIL: Glycerol Biorefinery Approach for the Production of High Quality Products of Industrial Value by F. Lorenzini (presenting author), Y.-M. Wang, X. Liu, M. Rebros and A. C. Marr (at the 3rd International Symposium on Green Chemistry) led by QUB on May 2015 in France.
  • Comparison of different selection strategies for the bioconversion of crude glycerol derived from second-generation biodiesel by C. Varrone (presenting author), T.M. Bjerkan Heggeset, S. Balzer Le, G. Floriotis, T. Haugen, S. Markussen , I.V. Skiadas, H.N. Gavala (at the 11th International Conference on Renewable Resources and Biorefineries) led by AAU on June 2015 in York (UK).

For more information, please contact:

Institut Univ de Ciencia i Tecnologia, S.A.

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Main contact: Roberto Horcajada
Email: [email protected]

Please download the e-newsletter#2 here: #2 e-Newsletter_July_2015


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