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Progress on the fundamental research on microalgae

Optimization of lipid productivity

 

Two-stage batch cultivations with P. tricornutumhave been performed to study the effect of incident light intensity on lipid production. This showed that P. tricornutumis rather sensitive to a combination of nitrogen depletion and high light intensities, leading to rapid cell death at high incident light intensities. The highest lipid yield can be obtained at relatively low light intensity combined with nitrogen depletion. The knowledge obtained during these two stage experiments was used to design a one-step lipid production process.

 

Transcriptomics

 

We have successfully implemented methods and protocols for transcriptome profiling of N-depleted and N-repletedPhaeodactylumsamples from partner WU. Mapping of the transcriptome against the annotated genome sequence from P. tricornutumrevealed that 10213 genes out of 10391 annotated genes were expressed, indicating the sufficient coverage of transcriptome. We have analysed expression levels of genes by mapping transcriptome data against gene sequences from known lipid biosynthesis (KEGG, DiatomCyc) pathways. This approach enables the identification of differentially expressed genes underlying lipid biosynthesis in P. tricornutum. This platform is thus ready to perform detailed analysis on sample series

 

Proteomics

 

 

Several protein extraction procedures from intact Phaeodactylumcells have been tested and evaluated with the aim of obtaining a maximum of protein identifications plus an optimal quantitative reproducibility of the extraction efficiency. 3 grinding methods and 2 extractions buffers have been tested. In conclusion, grinding wet sample gives maximum identifications. The 2 buffers both give high yield, with some complementarity. We are ready to perform detailed quantitative analysis on sample series.

 

Metabolomics

 

 

Several protein extraction procedures from intact Phaeodactylumcells have been tested and evaluated with the aim of obtaining a maximum of protein identifications plus an optimal quantitative reproducibility of the extraction efficiency. 3 grinding methods and 2 extractions buffers have been tested. In conclusion, grinding wet sample gives maximum identifications. The 2 buffers both give high yield, with some complementarity. We are ready to perform detailed quantitative analysis on sample series.

 

 

 

 

Proof of principle of one-step process

 

 

A strategy for one-step lipid production in P. tricornutumwas designed and tested at lab scale (1.7L flat-panel photobioreactors). Simultaneous growth and lipid accumulation was reached in nitrogen limited turbidostats operated under day/night light cycles, thereby providing a proof of principle of one-step lipid production in P. tricornutum. The fraction of TAG increased from 7 %w/w in a nitrogen replete culture to 12 %w/w in a nitrogen limited culture. Optimization of this process is needed to further increase lipid content and productivity. These experiments will be coupled to transcriptome, proteome and metabolome analysis. The combination of continuous experiments with ~omics technologies will generate fundamental insight in the mechanisms underlying lipid accumulation in P. tricornutum, and will also give directions for the cultivation strategies that will be tested outdoors.

 

We have recently defined optimal two stage starvation protocols for maximal oil productivity with Nannochloropsis, including semi-continuous cultivation with limiting nitrogen content and batch cultivation with dilution of nutrient replete culture into nutrient deficient growth media. The impact of varying salinity on both approaches was tested. While the semi-continuous cultivation gives higher oil productivity at elevated salinity, the batch cultivation gives higher oil productivity if the starvation phase is performed at lower salinity. This approach was documented by metabolomics analysis indicating that high salinity demands high concentrations of free sugars as osmoticum, which are not required under lower salinity. Those findings are now being investigated outdoors in temperature controlled panel reactors.

 

 

 

FUEL4ME, September 2014: What's going on?

We just passed the firsts 18 months of the project. The last semester started with the General Assembly and the site visit in Florence and ended with the first report delivered to the commission. We entered in one the most challenging period of the project, in which, while in the laboratory the best culture conditions for optimal lipid productivity are being developed, outdoors, the 3 pilot plants were installed and the first results with the two selected microalgae were obtained. So the race, ….or more scientifically, the comparison for the best location and the best panel photobioreactor has begun, under the impartial judgment of LCA assessment! The large-scale plant is also being built and the first lipid-rich Nannochloropsisbiomass has been provided for the downstream processing.

 

Foto AG Florencia


International course Microalgae Process Design

International course
Microalgae Process Design - from cells to photobioreactors
11 - 18 July 2014 AlgaePARC, Wageningen, the Netherlands
Registration: http://www.vlaggraduateschool.nl/courses/algae-design.htm

This course provides the essential skills for designing optimal microalgae-based production processes, for both research and commercial purposes. Through lectures, digital cases and a photobioreactor practical, the participants will learn: 1) how to describe microalgal metabolism quantitatively, 2) how to apply basic design principles and set up mass/energy balances for photobioreactors, 3) how to cultivate microalgae in fully controlled photobioreactors, and 4) how to integrate all acquired knowledge into optimal production strategies for microalgae biomass or secondary metabolites.

The course is very popular and places are limited, so if you are interested, please subscribe soon!

Editorial e-Bulletin March 2014

This e-bulletin gives an update of the progress made so far by the partners of FUEL4ME. We are now one year into the project which started with a kick-off meeting in March 2013. The first experimental evaluations of microalgae performance in laboratory-scale photobioreactors are finished, analytical ~omics protocols have been developed and will be combined with detailed laboratory studies on optimization of continuous lipid production, which are currently running.

With Spring starting early this year in Europe, it is good to see that the development of the outdoor cultivation systems is on schedule: all systems in Italy, Israel, the Netherlands and Spain will be converting sunlight, seawater and CO2 into biomass and lipids this Spring!

Besides cultivation, the FUEL4ME partners have undertaken their first steps towards development of next generation fatty acid extraction protocols, and the development and description of the methodology for the sustainability assessment (economic, environmental and social assessment) is finished.

In the next years we hope to show you interesting results leading up to a sustainable process for continuous production of lipids from microalgae. We believe microalgae will become an important sustainable feedstock for production of commodities by innovative research. Through collaboration between research institutes, universities and industrial partners these innovations can be realized in a biobased economy.

FUEL4ME Work Package 1 leader
Dr. Packo Lamers
Wageningen UR - Bioprocess Engineering - AlgaePARC 

Sustainability assessment of integrated process

Work package 5 of FUEL4ME (Sustainability assessment of integrated processfocuses on the assessment of the sustainability including environmental, economic and social parameters of a continuous production and conversion process based on the work performed in WPs 1-4. The results will be used to guide the technical development and future implementation in the desired direction and to be able to determine economic feasibility and environmental sustainability.

Actually the development and description of the methodology for the sustainability assessment (economic, environmental and social assessment) is finished. The whole value chain of the FUEL4ME integrated process (see Figure 1), different algal concepts for the assessment (e.g. plant sizes, location, algae species), the reference system with conventional products, system boundaries and functional units have been defined. Within the sustainability assessment the whole value chain including cultivation of algal biomass, harvesting, transport to conversion centers, conversion to biofuels, disposal of wastes, transport to user, use in a final energy product of the FUEL4ME integrated process will be considered. 

 

Maria Hingsamer / Gerfried Jungmeier

 

Translation to outdoors and production

The Work Package 2 (Translation to outdoors and productiondedicated the activity of the last months to the detailed engineering definition for the Pilot (10 m2) and Demo (250 m2) plants at BGU, F&M, WU and BIT. A common basic design (flat panel reactors) was chosen for all the locations, but detailed engineering, based on the specific knowledge of each group, will results in four different systems, three of them made by flexible plastic film and one by rigid glass sheets.  F&M will adopt the Green Wall Panel design based on its patent (WO2011/013104), BGU will adopt the disposable reactor based on its patent (WO 2005/006838), WU will use a rigid glass panel and BIT will use the ProviAPT system.

In some of the locations the reactors are already in place, in others they are in course of realization. All the Pilot plants are expected  to be in operation in April 2014 and the Demo plant is expected to be fully operative in May-June 2014. The Demo plant will include two series of photobioreactors, a larger system for the production of biomass for the downstream process and a smaller system to obtain growth data over a longer period. 

Thanks to the previous work carried out by WP2 (Translation to outdoors and production) partners, together with WP1, the strains of Phaeodactylum and Nannochloropsis to be used in the four plants were identified.

Together with WP5 partners the list of inputs and outputs to be measured during the outdoor experiments for LCA evaluation and the procedures to be used for the measurement of the different parameters were defined.

Liliana Rodolfi

 

Fundamental research and enabling technologies


The effect of incident light intensity in two-stage batch cultures ofPhaeodactylum tricornutumwas evaluated. In the first stage,P. tricornutumgrew to a fixed biomass density in the presence of a nitrogen source. The second stage started when all nitrogen was consumed and the microalgae switched their metabolism towards accumulation of lipids. This work showed thatP. tricornutumis rather sensitive to a combination of nitrogen depletion and high light intensities, leading to rapid cell death at high incident light intensities. The highest lipid yield can be obtained at relatively low light intensity combined with nitrogen depletion.

Currently, experiments for continuous one-stage lipid production in nitrogen-limitedP. tricornutumcultures are ongoing. These experiments will be coupled to transcriptome, proteome and metabolome analyses, for which the basic analytical protocols have been set-up or are currently being finalized. The combination of these highly controlled photobioreactor experiments and the broad-scale ~ omics analyses will generate fundamental insight in the mechanisms underlying lipid accumulation inP. tricornutum, and will also give directions for the cultivation strategies that will be tested outdoors in Work Package 2 (Translation to outdoors and production).

Simultaneously, various two-stage and one-stage cultivation strategies are being evaluated forNannochloropsisin both indoor and outdoor experiments.

Packo Lamers