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					EPOBIO – Realising the economic potential of sustainable resources –
bioproducts from non-food crops


EPOBIO WORKSHOP 2: Products from Plants – from crops and forests to
zero-waste biorefineries


Eretria Village Hotel Resort and Conference Centre – Greece 15-17 May 2007




ABSTRACTS OF PRESENTATIONS
Biorefinery and Bioenergy in FP7

Alfredo Aguilar

Head of Unit Biotechnologies, European Commission – DG Research

Directorate Food, Agriculture and Biotechnology, B-1049 Brussels, SDME 08/42

Alfredo.aguilar-romanillos@ec.europa.eu

Renewable biological resources are the basis of a European knowledge based bio-economy (food, feed,
agriculture, forest based, fisheries, aquaculture, biochemistry, etc.) that today has an estimated annual turn-
over of more than €1500 million. The increasing demand for biological resources, both in quantity and
quality, can only be met through innovation and advancement of knowledge in the sustainable management,
production and use of these biological resources (micro-organism, plants and animals).

The target of the new FP7 - theme Biotechnology, Agriculture, Food, Forestry and Fishery - is to built a
European Knowledge Based Bio-Economy by bringing together science, industry and other stakeholders, to
exploit new and emerging research opportunities that address social, environmental and economic
challenges, production and sustainable use of renewable bio-resources.

The rationale behind this choice will be analysed and an overview about the topics that are included in the
next FP7 calls will be given.


Toward a sustainable bioeconomy: How can emerging bioproduction systems
promote environmental quality?

Robert P. Anex

Iowa State University

3202 NSRIC Building, Ames, IA, USA

panex@iastate.edu, http://www.biorenew.iastate.edu/

Mature technologies for converting biomass into fuels and chemicals mature are expected to utilize a high
degree of internal material and energy recovery. Although they make facilities more complicated and are
capital-intensive, refinements such as heat recovery and integration help optimize plant efficiency. A review
of developments in grain ethanol production technology shows that this trend is already well underway. As
resource constraints become more limiting, biomass utilization systems will be required not only to be energy
efficient, but ecologically efficient. Such systems will be optimized to provide a wider range of the ecological
functions that agricultural and natural lands currently provide, including nutrient cycling, carbon
sequestration, and the protection of soil and water resources. Conceived of in this way, advanced production
methods, such as cellulosic biofuel technologies, will be able to offer more than energetic, economic and
climate change mitigation benefits (Lynd et al. 1991, Farell et al. 2006). One can identify hints of these
trends as well in the maturing grain ethanol industry and the nascent cellulosic ethanol industry in the United
States. It is important to look for opportunities to design biorefineries not only for high economic return and
energy efficiency, but as integral parts of the agricultural and industrial ecosystems. Such “ecologically
intensive” systems of bioproduction hold the promise of promoting environmental quality rather than adding
additional stresses to soil, water, and air resources that are already under heavy pressure from agriculture
and industry.

Work supported by U.S. National Science Foundation Grant #CMS0424700
Farrell, A.E., R.J. Plevin, B.T. Turner, A.D. Jones, M. O’Hare, and D.M. Kammen. 2006. Ethanol can
contribute to energy and environmental goals. Science. 311: 506-508.
Lynd, L. R., J.H. Cushman, R.J. Nichols, and C. F. Wyman. 1991. Fuel ethanol from cellulosic biomass.
Science. 231: 1318-1323.


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New tools for plant breeding

Abdel Bendahmane

Unité de Recherche en Génomique Végétale (URGV)

URGV, 2, rue Gaston Crémieux CP 5708, 91057 Evry Cedex, FRANCE

http://www.evry.inra.fr/, bendahm@evry.inra.fr

National and international institutions have been engaged in large programmes aimed at improving the
nutritional or functional properties of the harvested plant for use in food, animal feed, or industrial products.
This far, these efforts have been mainly carried out through breeding. Over the last two decades, knowledge
of plant growth, development and the molecular composition of plant organs has increased tremendously.
The genes that control the function of the biological mechanisms involved are in many cases identified and
well characterised. Unfortunately, development of technologies to manipulate plant genomes has not
matched that progress. GM is the only current way to carry out this task but is currently rejected by the
consumer. Consequently, exploitation of the wealth of information available to scientists to modify output
traits in crops is still far from expectation.

TILLING (Targeting Induced Local Lesions IN Genomes; Colbet et al, 2001), offers an alternative way to
manipulate endogenous genes for functional evaluation and improvement of crops without GM. This target
gene modification system has additional merits. First, it can be automated in a HTP mode, which is an
absolute necessity to match the speed of candidate gene discovery. Second, it is an efficient way to isolate
an allelic series in a specific gene and consequently identify alleles with a higher potential agronomic value.
Third, it is very effective for identifying mutants in redundant genes, which is extremely difficult using
phenotypic screening as in classical breeding.

In URGV we have set a TILLING platform on different crop species. In this workshop I will review the
importance of TILLING in breeding using examples of TILLED genes that have impact on functional
properties of the harvested crop for use in food, animal feed, or industrial products.




The poplar genome – accessibility to woody species

Wout Boerjan

Flanders Institute for Biotechnology (VIB)

VIB Department of Plant Systems Biology, UGent, Technologiepark 927, 9052 Gent, Belgium

http://www.psb.ugent.be; woboe@psb.ugent.be

Poplar is one of the most intensively planted commercial forest tree species, mainly because of its fast
growth, its ease of vegetative propagation and the strong hybrid vigour of interspecific hybrids. For the same
reasons, and additionally because poplar can be easily genetically modified, poplar has become the model
of choice for molecular geneticists. This common interest has culminated in the sequencing of the poplar
genome, the first genome of a forest tree.

The increasing awareness that lignocellulosic biomass from fast-growing trees, such as hybrid poplar, holds
great promise to become an excellent renewable carbon-neutral raw material for conversion to bioethanol,
urges for accelerated genetic improvement of trees by smartly combining conventional and biotechnological
breeding tools. There is large potential in conventional breeding, because poplars, as all trees, are still
largely undomesticated. Genetic modification, on the other hand, can circumvent the long breeding cycles
and allows tailoring wood quality to the need of the industry far beyond the possibilities nature can achieve.

This paper will present an overview of the genetic improvements in poplar made so far and provides a
research agenda that addresses how information from the poplar genome sequence combined with

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knowledge on the genetics and molecular biology of poplar, can result in step-change improvements in the
quality of woody biomass to meet future bioenergy demands.


Pilate et al., Nature Biotechnol Vol. 20:607-612 (2002)

Boerjan Current Opinion Biotechnol. Vol. 16:159-166 (2005)

Tuskan et al., Science Vol. 313:1596-1604 (2006)




Global Vision – Zero Waste and Sustainability

Dianna Bowles

Centre for Novel Agricultural Products

CNAP Department of Biology, PO Box 373, University of York, York, UK. YO10 5YW

www.cnap.org,uk

Society is in transition: moving from a dependence on oil and fossil reserves to an economy that will be
based on biorenewables and the use of agricultural feedstocks for industrial production. The strategic vision
for long-term sustainability is under development. The move globally to replace the use of existing transport
fuels with biofuels is a clear indicator of this transition and the increasing global commitment to sustainable
development. However, the bioeconomy is far broader than the energy sector and offers major opportunities
for the production of chemicals and materials for many different sectors of industry. Ultimately, the aim will
be to use biorefineries in a way comparable to that in which oil refineries are currently used. This will enable
maximum outputs and value to be gained from the inputs of biorenewable feedstocks.

Feedstocks will increasingly be used from the agricultural, forestry and waste sectors as well as from marine
resources. The achievement of maximum utility from the biorenewable feedstocks is likely to necessitate
integrated supply chains. Stakeholders from the growers to the industrial users must be linked if efficient
production is to underpin effective use.

Realising this potential of a global bioeconomy will raise major issues of land-use, security and quality of the
food chain, and availability and use of water resources. This will require the application of sustainability
criteria to assess the new developments. In turn, this need will bring the research community from many
different disciplines together with the industrial and business sectors, to ensure a decision-making process
for new energy and non-energy products that reflects both the market and is acceptable to the policy-makers
and the public.




                                                       4
Sugarcane: A Crop with the Potential to Function as Both a Biofactory Producing
Industrial Chemicals and as a Large Scale Source of Carbohydrates for Future
Biorefineries

Brumbley, S.M.1,2,3, Purnell, M.P.1,2,3, Petrasovits, L.A.1,2,3, Anderson, D.A,1,2,3, McQualter, R.A.3, Chong,
B.F.2.3 Nielsen, L.K.1,2
1
Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, St Lucia 4072
Queensland Australia
2
 Cooperative Research Centre for Sugar Industry Innovation through Biotechnology, The John Hines Bldg,
The University of Queensland, St Lucia 4072 Queensland Australia
3
BSES Limited, 50 Meiers Road, Indooroopilly 4068 Queensland Australia

Sugarcane (Saccharum sp. hybrids) has the potential to be a key crop for biofactory production of industrial
chemicals. It is the second fastest growing tropical grass, produces a large biomass (90-250+
tonnes/hectare), partitions carbon into sucrose at up to 42% of the dry weight of the stalk, has a mobile pool
of hexose sugars through most of its life cycle, is vegetatively propagated, and can be harvested multiple
times before replanting. Sugarcane industries all ready have in place the infrastructure to haul the enormous
biomass from the farms to the sugar mills for processing and the mills generate their own energy.

The PHB biosynthesis enzymes of Ralstonia eutropha, PHAA, PHAB and PHAC, were targeted to plastids of
sugarcane variety Q117 (Petrasovits et al. 2007). Epifluorescence and electron microscopy of leaf and stem
sections from these lines revealed that PHB accumulated in plastids of all vital cell types except mesophyll
cells. The concentration of PHB in culm internodes of plastidic lines was substantially lower than in leaves.
Western blot analysis indicated that expression of the PHB biosynthesis proteins was not limiting in culm
internodes. Epifluorescence microscopy of culm-internode and node sections and electron microscopy of
node sections from plastidic lines showed that PHB accumulated in all vital cell types except photosynthetic
cortical cells in the rind. Six PHB-positive lines were further studied in a replicated glasshouse trial using a
randomized block design. These lines accumulated PHB in leaves to a maximum of 1.77% of dry-weight,
without incurring an agronomic penalty. There was a PHB concentration gradient from the top to the base of
the plant as well as a gradient from tips to the bases of individual leaves. The pattern of relative PHB
accumulation was the same in all lines with marked differences in absolute PHB concentration (Purnell et al.
2007). There were good correlations between PHB concentration and the abundance of the biosynthetic
enzymes, but not between the biosynthetic enzymes and their respective transcripts. The overall highest
producing line (TA4) produced PHB at ~2.5% of leaf dry weight. Although moderate PHB concentrations
were achieved in leaves, maximum recorded total-plant PHB yield was only 0.26% (11.9 g PHB in 4.60 kg
fresh-weight). Sugarcane was also evaluated as a production platform for p-hydroxybenzoic acid using two
different bacterial proteins that both provide one-enzyme pathways from a naturally occurring plant
intermediate (McQualter et al. 2005). The sugarcane line producing the highest levels accumulated a
glycosylated form of pHBA in the leaves at 7.5% of the dry weight and in the stems at 1.5% dry weight
(McQualter et al. 2005). In addition, sugarcane was transformed with a gene encoding sorbitol-6-phosphate
dehydrogenase to generate plants expressing Sorbitol in leaves and stems (Chong et al. 2007). In the leaf,
the mean sorbitol levels (across the entire leaf blade minus midrib) for six biological replicates was 12 % of
dry weight. In the stem, the mean sorbitol (transverse bore sample, i.e. rind-pith-rind) for six biological
replicates was 1 % of dry weight.

Chong, B.F., Bonnett, G.D., Glassop, D., O’Shea, M.G., Brumbley, S.M. (2007) Growth and metabolism in
sugarcane are altered by the creation of a new hexose-phosphate sink. Plant Biotechnology Journal 5:240-
253.

McQualter, R.B., Chong, B.F., Meyer, K., Van Dyk, D.E., O'Shea, M.G., Walton, N.J., Viitanen, P.V. and
Brumbley, S.M. (2005) Initial evaluation of sugarcane as a production platform for p-hydroxybenzoic acid.
Plant Biotechnology Journal 3:29-41.

Petrasovits, L.A., Purnell, M.P., Nielsen, L.K., and Brumbley, S.M. (2007) Polyhydroxybutyrate production in
transgenic sugarcane. Plant Biotechnology Journal 5:162-172.

Purnell, M.P., Petrasovits, L.A., Nielsen, L.K., and Brumbley, S.M. (2007) Spatio-temporal characterisation of
polyhydroxybutyrate accumulation in sugarcane Plant Biotechnology Journal. 5:173-184.


                                                        5
Challenges for Sustainable Feedstock Production

Dr Mike Bushell

Syngenta

Jealott’s Hill International Research Centre, Bracknell, Berkshire RG42 6EY, UK

mike.bushell@syngenta.com

The strategic development and realisation of biomanufacturing through biorefining is an increasingly
important area for the sustainable development of useful industrial products. It is already clear that very
many useful materials can be produced from plant derived biomass, although only a limited number will be
economically viable in the near term. Process Technology breakthroughs can be expected, but the
economic challenges will definitely mandate use of agronomic systems that deliver high yields of the plant
feedstocks.

The challenges facing agricultural production are immense. Simply to feed the 2050 world population will
require average yields 50% higher than today. Competition for the same high quality agricultural land to
grow biofuels as well as to produce chemical feedstocks and other products intensifies the challenge.
Maintaining high yields as well as promoting biodiversity, soil fertility and managing water resources are
issues that need to be tackled.

Success will require us to build on established methods of agronomy, crop protection and conservation
agriculture, but also to embrace modern methods of plant biotechnology to develop new varieties of Plants
for the Future.

Oil Crop Platforms for Industrial uses
Anders S. Carlsson

Swedish University of Agricultural Sciences

Department of Plant Breeding and Biotechnology, P.O.Box 44, SE-230 53 Alnarp, Sweden.

http://vfbt.vv.slu.se/, anders.carlsson@ltj.slu.se

Our society has become increasingly dependent on fossil fuels not just as an energy source for
transportation and heating but also for the provision of industrial feedstocks for a multitude of products that
we use in every aspect of our daily lives. Despite the tremendous positive impact that crude oil has had on
the advancement of human society, our current utilization of crude oil is unsustainable since this dominating
feedstock is a limited resource as well as a major cause of climate change.

Fossil fuels thus need to be replaced with alternative sustainable and environmentally friendly sources of
energy and industrial feedstocks such as plant oils. However, the successful development of oleochemical-
based products for global markets is critically dependent on the effectiveness and cost competitiveness of
the strategies chosen for the production of the industrial oils. In this respect it is evident that the choice of
robust crop platforms for the production of feedstock oils is a critical decision.

The latest report from the EPOBIO plant oil flagship titled “Oil crop platforms for industrial uses” addresses
the establishment of such industrial crop platforms for oil. Three crop platforms are considered. The first,
rapeseed, is already a major crop globally that is used primarily as a source of food and feed, but is
increasingly utilized as a source of biodiesel; the second, oat, is explored as a potential new oil crop platform
for Europe, since variations in existing germplasm suggest that starch to oil ratios may be further
manipulated to increase oil content; the third, crambe, is relatively undeveloped compared to rape and oat
but also holds significant potential for production of industrial oils, since it is a high yielding oil crop that is
excluded from the food chain.

Each of these crops was evaluated as a potential source of renewable industrial oils and fuels, and research
required for optimization of each crop platform will be described.

                                                         6
Policy Drivers Leading to the Bio-based Economy – EU Perspectives

David Clayton

EPOBIO

CNAP, Department of Biology (Area 15), PO Box 373, University of York, York, UK, YO15 5YW

www.epobio.net

The various policy drivers in place for the bioeconomy – biofuels, bioenergy and the non-energy bioeconomy
– will be considered in this presentation. The speaker will examine the risks that policies are intended to
address and consider the effect of those risks and policies on the development of the bioeconomy in Europe.
The impact of support for the agriculture sector and policies affecting other potential feedstocks will be
considered. Key policies for the successful delivery of the bioeconomy will be outlined.


Making Cellulosic Biofuel Feedstocks a Commercial Reality

Richard Flavell

Ceres-Inc

Thousand Oaks, California, USA

The US Government has set goal of generating 35 billion gallons of biofuel per year by 2017. To achieve this
will necessitate not only continued production of ethanol from corn starch but also the generation of ethanol,
or its equivalent, from cellulose. The latter is much more energetically efficient. To achieve the production of
a biofuel from cellulose needs very large sustainable supplies of plant feedstocks transported to biorefineries
where conversion of the cellulose to sugars and then fermentation to ethanol or thermochemical conversion
can be performed efficiently. At present, costs of the whole commercial chain need to be reduced but this is
happening as more people address the technical issues. The leading contenders to be the most suitable
feedstocks are the C4 grasses, including the perennials switchgrass, miscanthus and sugar/energycane.
More than 50% of the cost of the feedstock at the biorefinery gate is due to harvesting and transport. This
means that high density of plant material in the field is a crucial goal for germplasm selection, plant breeding
and production. The conversion efficiency of this material to a biofuel is also particularly important to
optimize. The shortage of time for optimized feedstock development to meet the political goals means that
the best germplasm needs to be urgently selected and then imaginative breeding programs adopted with all
the efficiencies that can be gained from molecular comparative genetics. Ceres has initiated programs to
develop and commercialize biomass energy crops with high conversion efficiencies using carefully selected
starting germplasm but also incorporating its leading gene-trait knowledge base and composition analytical
skills. Its strategy and progress to achieve the commercial objectives will be summarized.


Oil crops – biofuels and beyond

Professor Ian A. Graham

CNAP, University of York,

Department of Biology (Area 7) PO Box 373, York, YO10 5YW, UK

iag1@york.ac.uk,: www.cnap.org.uk

The seed oils of plants are structurally similar to long chain hydrocarbons derived from petroleum, and thus
represent excellent renewable resources. About 15-20% of the plant oils produced in crops (15-20 million
metric tons) are currently used in non-food applications. These oils are competitive alternatives to petroleum-
based products such as detergents, paints, plastics and lubricants. Seed oils are also increasingly used to
produce biodiesel with oilseed rape being the major crop serving this market in Europe. Demand for plant

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derived oil, particularly from the biodiesel market, is currently outstripping supply. Yield of oil per hectare of
crop is recognised as a major target for improvement in this sector.

Efforts to overcome the bottlenecks that prevent increasing the yield of existing and designer oils in crop
plants have to date been piece-meal and so the full potential of plant-derived oils have not been realised. In
this presentation potential bottlenecks that limit the accumulation of such oils will be highlighted and
strategies to overcome these bottlenecks will be discussed. Looking to the future the prospects for
development of new and improved oilcrops that compete alongside other biomass crops for the production of
biofuels will be considered.




Challenges of the conversion of lignocellulosic biomass into ethanol

Kevin A. Gray, PhD

Diversa Corporation

4955 Directors Pl, San Diego, California, 92121. USA

kgray@diversa.com, www.diversa.com

Commercial implementation of a biomass to liquid transportation fuel process has been hindered by a
number of challenges. First, biomass collection and transport have issues pertaining to cost and abundance.
Second, preprocessing or “pretreatment” of the raw material is required to make it more digestible by
enzymes and this process tends to be very capital intensive. Third, enzymatic saccharification of the
pretreated material is difficult and expensive due to the natural recalcitrance of plant cell walls to biological
attack. In addition the presence of non-carbohydrate molecules like lignin have an impact on digestibility.
Finally, conversion of biomass sugars to alcohol is made difficult due to the mixed nature of the sugar
streams. Diversa Corp. has pioneered the discovery of novel, highly active enzymes from the environment.
In addition we have developed technologies to optimize biomolecules for specific industrial purposes. This
presentation will cover the discovery of these enzymes, the development of specialized automation
technologies to evaluate enzyme activity on insoluble substrates and the performance of various enzyme
cocktails on various feedstocks.



Oil modification and oleochemicals

William Hitz, Anthony Kinney

DuPont Company

Experimental Station, Wilmington, Delaware USA 19880

William.d.hitz@usa.dupont.com

Temperate oilseed crops typically comprise five main fatty acids in the triacylglyceride that makes up the
seed oil. Tropical crops might add one or two more. Each crop is individualized by the proportion of these
fatty acids but they are all characterized by having multiple fatty acids. The modification to the standard
carbon backbone that gives rise to the different fatty acids is the number of double bonds present. The
number of double bonds controls physical properties of the vegetable oil and its chemical reactivity. The
exact properties of an oil that are imparted by double bond number make an oil useful in some applications
but undesirable for others and these varied properties can be opposing even within an oil use. A relatively
high number of double bonds (unsaturation) is a desired nutritional property but an undesirable characteristic
in food processing. For industrial uses that require chemical reactivity such as drying oils for inks and paints,
a moderately high degree of unsaturation is required while for fuel and lubricant uses the lowest degree of
unsaturation consistent with solids requirement is desired.



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Plants that were not chosen for domestication for food use as food crops produce a much wider variety of
fatty acids in storage lipids. In some cases those fatty acids have desirable industrial properties and might
make valuable chemical intermediates if they can be economically produced.

We have the ability to modify the fatty acid profile of temperate crop plants through traditional breeding and
through the application of genetic modifications that result in changes to the fatty acid synthesis pathway.
Conversion of these modified plants into economically viable crops must take into account all of the
conflicting use requirements described above in addition to remaining within limits of the type of fatty acid or
the amount of any one fatty acid accumulated that are imposed by the required physiological function of
lipids containing these fatty acids in the plant. Compromises are possible but these must also fit into a
commerce framework that functions efficiently with large scale commodity crops to provide cost effective raw
ingredients for both food and industrial uses. Smaller scale crops must have the demand and value to
support higher production cost since they do not readily fit into the large scale.




Breaking the Biological, Chemical and Engineering Barriers to Lignocellulosic
Biofuels

George W. Huber

University of Massachusetts-Amherst

Chemical Engineering Department, 686 N. Pleasant St,112 Goessmann Laboratory, Amherst,
MA 01003-9303

huber@ecs.umass.edu

Concerns about global warming and national security, combined with the diminishing supply and increased
cost of fossil fuels are causing our society to search for new sources of transportation fuels. In this respect
the only sustainable source of renewable carbon that could be used to produce liquid transportation fuels is
plant biomass. Currently cellulosic biomass is significantly cheaper than petroleum (at $15 per barrel of oil
energy equivalent) and abundant (have the energy content of 60 % of our domestic crude oil consumption).
However, the chief impediment to the utilization of our biomass resources is the lack of economical
processes for conversion of biomass resources into fuels. To develop these processes, it is necessary to
understand and overcome the key biological, chemical and engineering barriers, and develop the enabling
technologies that will allow us to efficiently use our biomass resources. A major 21st century goal for
academia, industry, and government should be the emergence of efficient and economical utilization of
biomass resources.

We will compare and discuss strategies for green gasoline, green diesel and green jet fuel production
(www.ecs.umass.edu/biofuels). These strategies include: selective thermal processing of cellulosic biomass,
utilization of petroleum refining technologies for biofuel production, aqueous-phase processing, and syn-gas
conversion. Recent advances in theoretical chemistry combined with new in-situ catalyst characterization
methods allow us to understand chemistry at a fundamentally new level. Combining fundamental chemical
understanding with new methods to synthesize nanostructured catalytic materials, the ability to design and
simulate complicated reaction networks, and the ability to perform conceptual design and optimization
problems allow us to engineer efficient and economical processes for biofuel production. While biology is
important in biofuel production, chemistry, chemical catalysis and engineering will be equally vital to make
lignocellulosic biofuels a practical reality.




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Biorefineries in China: policy, R&D capacity & resource

Dr Julia Knights

British Consulate-General Shanghai

Consul, Head of Science and Innovation section, Suite 301, Shanghai Centre,
1376 Nanjing Xi Lu, Shanghai 200040, China.

China’s R&D capacity will be discussed in relation to government policies at local and national level together
with China’s research efforts on biorefineries. Development of a domestic biofuels industry will be debated
in terms of feedstocks, climate, geography and food security.

For more information on the UK Foreign and Commonwealth Office’s Science and Innovation network in
China, please visit www.uk.cn/science



A Canadian Approach to Biorefining

Jerome Konecsni, President and CEO

Genome Prairie

101-111 Research Drive
Saskatoon, Saskatchewan
S7J 0R1
Canada

jkonecsni@genomeprairie.ca

The presentation provides an overview of Canada’s approach to biorefining. It represents a team effort as
information was put together by a team of Canadians representing national and regional research
organizations, the National Research Council and Agriculture and Agri-Food Canada and universities.

Canada’s strengths, funding agencies, research networks/activities and examples of national and regional
initiatives related to the development and commercialization of bioproducts are highlighted. Models of multi-
stakeholder ventures are presented that parallel the efforts of EPOBIO. International linkages are an integral
part of the Canadian approach and expanding these partnerships is an objective of the Canadian team that
will be in attendance at the workshop.

Canada has a diverse and well-resourced R&D infrastructure dedicated to the development of value added
products from raw materials produced on vast quantities of arable land. It represents a wealth of
opportunities for collaboration with European and American scientists and industry.




Is micro-algae a future bio-fuel?

Barrie Leay

Aquaflow Bionomic Corporation Limited

PO Box 949, Nelson, New Zealand

barrie@thepacific.net

Imagine if we could:

Grow a fuel without using land
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Grow a fuel without polluting the atmosphere
Grow a fuel without creating green house gases
Take a crop every day instead of yearly
Generate up to 300 times more per acre
Grow crops in both fresh and sea water


Lessons learnt from the bio-economy - policy in partnership with research

Wilfrid Legg

Organisation for Economic Cooperation and Development

2 rue André-Pascal, 75016 Paris, France

wilfrid.legg@oecd.org, www.oecd.org/agr

Governments in many countries are putting in place targets and policies to encourage the production of
biomass from agriculture, forestry and waste for energy and materials. This paper focuses in particular on
policies related to the agricultural sector. Policies to increase biomass have been driven by concerns to
increase energy security, reduce greenhouse gases and to widen the diversification of income sources for
primary producers. Studies to date in OECD and elsewhere suggest that in many countries agricultural feed
stocks for bio energy are not currently economically efficient without subsidies, and it is not clear if there will
be improvements in overall environmental performance, particularly when considered on the basis of a life-
cycle approach. However, the assessments depend on a number of key assumptions related to fossil based
energy prices, bio refinery costs, crop yields and prices, and land availability. Three key issues are stressed
in this paper: policies to promote bio energy are often running ahead of the underlying economic and
environmental research; there are secondary impacts on agricultural, food and land markets – as well as
environmental impacts not only on greenhouse gases but also on water, soil erosion and biodiversity - that
are not always taken into account; and there are many uncertainties concerning the development of future
costs and prices, in particular in relation to second generation bio-fuels. An important issue concerns the
coherence of policies related to energy, agriculture, environment and taxation. The paper points out that
there can be significant risks of misallocating resources through locking-in policies that can also lead to
unintended consequences, and that closer linkages between the research and policy communities would be
desirable.


Crop Platforms for Cell Wall Biorefining – Lignocellulose Feedstocks
Ralf Möller
Max-Planck-Institute of Molecular Plant Physiology
Wissenschaftspark Golm, Am Mühlenberg 1, 14476 Potsdam – Golm, Germany
moeller@mpimp-golm.mpg.de


Cell walls are the main components of plant biomass on Earth and represent a largely untapped renewable
resource for bio-based products. For the successful development of the bio-based economy it is important to
consider a range of crop platforms that can provide the feedstock for cell wall biorefineries. The EPOBIO
report “Crop Platforms for Cell Wall Biorefining – Lignocellulose Feedstocks”, analysed the strengths,
weaknesses, opportunities and threats (SWOT) of four sources of biomass of relevance to Member States of
the EU as case studies. These were poplar and willow, Miscanthus and wheat straw, which have been
chosen as representative of woody species, grass and a co-product from arable crop cultivation.

The analysis has shown that the science base of poplar and the fact that its genome sequence is known,
provides an excellent foundation for the development of woody species for biorefining. Together with willow,
short rotation coppicing of poplar offers many opportunities for the agricultural sector, providing initial
investment costs and the timeline to investment recovery are acceptable. Information on the synthesis and
organisation of cell walls in these species with targeted studies to define properties of direct relevance to
ease of hydrolysis would be highly beneficial as these crops are developed in the longer term. Miscanthus

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undoubtedly holds great promise as a bioenergy crop for the future, because of its high biomass yield and
the low agrochemical inputs needed. However, studies are only beginning to understand the molecular
features of the grass, its cell walls and its optimisation for large-scale commercial cultivation. The decision to
undertake the very considerable amount of R&D needed to bring Miscanthus up to speed must be a strategic
commitment to perennial grasses as an industrial crop platform in the EU. Agricultural co-products as
feedstocks for biorefining have the advantage of adding value to the main use of the crop. These feedstocks
are already available now; however, improvement of the functionality of these co-products for biorefining
whilst maintaining the high quality bred into the crop as a food feedstock over many generations may prove
problematic. It will be a strategic decision, in terms of development of new feedstocks for energy and
chemicals biorefining, whether to disadvantage use and yield of crops for food production.




The problem of the lignocellulosic bottleneck I: material properties, architecture,
and pretreatments

Dr Ralph P. Overend

National Biofuels LP., and formerly National Renewable Energy Laboratory

310 Crichton St, Apt 402, Ottawa, Canada K1M 1W5

Overend@attglobal.net

The lignocellulosic (lc) resource is geographically diverse, of a very large magnitude, and is very variable in
physical form. While cellulose, hemicellulose and lignin, the major polymer families in wood and straw are
the majority of the lc material, there can be significant amounts of mineral matter, and non-structural
materials often called extractives. The chemical composition of the lignin and hemicellulose is variable with
season, bulk material storage and microbial attack. The challenge and opportunity, is to isolate the
carbohydrate components for biochemical conversion in a cost effective manner, while adding value to the
remaining components that range from 25% - 40% of the lc feedstock. Traditionally the plant components
that have been used for fuel production are the storage compartments such as cereal grains, while the lc has
been used as combustible fuel.

The major barrier to biotechnical exploitation is that trees, straws and stalks are all structural materials –
assembled to resist mechanical stress and biological degradation. Deconstructing these materials ideally
should take place in the context of an understanding of their architecture. In the last 20 years and particularly
since the sequencing of Arabidopsis the exquisite mechanisms of cellular construction in plants are being
revealed. Contrary to the given wisdom from many years of studying pulping, it now appears that there is
genetic control over the production and placement of lignin in the plant cell wall. This and evidence from
genetically engineering plant tissues suggests that there may yet be a biological pathway towards
deconstructing the lc cell walls. In the meantime the strategies are a combination of the physical treatments
(attrition, heat), and chemical reactions via hydrolysis and solvolysis of weak chemical bonds which bind the
hemicellulose and lignin to the cellulose. The control of the chemistry of deconstruction is a challenge as the
reactivity of the bulk polymeric materials is far less than that of the carbohydrate oligomers and monomers
that are released during the solvolysis. As a consequence the yield of desired polymers and oligomers is
reduced by the production of degradation products from the dissolved polymers, some of which have been
demonstrated to be inhibitory in downstream biological processing. Industrially feasible pretreatments are
commercial today, but the material challenge remains and its resolution would enhance the feasibility of the
lignocellulosic resource as a major feedstock for biorefining.




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Jatropha in developing countries

Winfried Rijssenbeek

Fuels for Agricultural Communal Technologies (FACT)

Horsten, 1 5612 AX Eindhoven, The Netherlands.

rijssenbeek@rrenergy.nl

This presentation will discuss the actual agricultural data and potential of Jatropha curcas in developing
countries. It will present some of the current yield data, information on climate and soil fertility and pest and
disease factors. It will also give some clues on how to further the potential of J curcas.. This requires
research and development in the plant genetics as well as in the best agronomic practices. The potential of
J. curcas is quite high and can be delivered under good policy measures.



Public attitudes towards the industrial uses of plants: The EPOBIO survey
Giorgos Sakellaris and Maria Paschou

Institute of Biological Research and Biotechnology, National Hellenic Research Foundation,

48 Vas. Konstantinou Ave., 11635, Athens, Greece

www.eie.gr

It is generally recognised that public opinion is a main contributor to the social trajectories of novel
technologies. Following the recent technological developments in the field of industrial plant exploitation for
energy production and manufacture, the aim of this study was to identify public attitudes towards the projects
and products identified by EPOBIO, which are expected to build the bio-economy of the near future. For that
purpose, a survey with national representative samples of seven EU countries and an electronic survey were
carried out in the period October-November 2006. The findings provide a valuable tool for the development
of a proactive communication strategy and could assist policy makers and investors in understanding the
social parameters which could help secure public acceptance of the new technological projects.

The questionnaire which was used in the survey examined opinion on plant oils, biopolymers and the bio-
refinery alongside with attitudes towards special issues and background information. The survey findings
suggest the following ideas. Firstly, Europeans are prepared to welcome the introduction of the novel
products proposed and are in favour of giving the Flagship areas incentives to support development:
Secondly, with regard to the special issues, energy production by combustion of plant-made products and
the usage of food crops in industry are viewed as being useful, morally acceptable and not a risk for society
by most Europeans. In contrast, the European public is ambivalent towards genetic engineering. Thirdly,
regarding decision making, the existence of appropriate regulation would determine public approval of the
technological projects and process involved. While there is a lack in trust in politicians, most Europeans
would prefer the European level for decision making over their national Governments. Fourthly, the national
populations were clustered into two groups, following systematic within-group consistency and between-
group variation with regard to several social parameters. Spaniards, Germans and Swedes were found to be
more attentive to science and technology matters, more knowledgeable about the industrial uses of plants,
more willing to buy the proposed products and more supportive of the relevant issues compared to Italians,
Greeks and the French. Another difference between those two clusters is that the former would be primarily
motivated by environmental reasons to supporting the novel technologies, whilst the latter would be mainly
motivated by the reduced dependency on petroleum. Fifth, the socio-demographic profile of those who are
more likely to support the novel plant-made products is males, urban dwellers, highly educated and those
aged 35-54. Finally, from a communication perspective the top priorities recommended are the intensification
of media coverage, the improvement of the corporate profile of industries, the creation of opportunities for
public participation and the promotion of scientists’ view-points in public debates.




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Phytochemical Genomics for Manipulation of Plant Secondary Products

Kazuki Saito

Graduate School of Pharmaceutical Sciences, Chiba University, Inage-ku, Chiba 263-8522, Japan; RIKEN
Plant Science Center, Tsurumi-ku, Yokohama 230-0045, Japan

ksaito@faculty.chiba-u.jp, http://www.p.chiba-u.ac.jp/lab/idenshi/index-e.html, http://www.psc.riken.go.jp/

Since ancient times, plant secondary products have been used as medicines, pesticides, flavors, dyes and
other industrial materials. This is still true even in the modernized societies. New plant products potentially
leading to the innovative drugs are being discovered from plants. Often we are astonished by the fact how
much man enjoys the benefits of the huge chemical diversity of plants for those non-food materials. More
recently plant biotechnology based on phytochemical genomics offers new possibilities for developments of
more beneficial bio-products such as pharmaceuticals and their feasible production. In this seminar, I will
discuss on the elucidation of the genomic basis of chemical diversity of plant secondary products for rational
manipulation of their complex pathways, by exemplifying our recent case studies on Arabidopsis thaliana
and an exotic plant producing camptothecin, a clinically used anti-cancer compound.



Engineering novel platforms for terpene natural product biosynthesis
Michel Schalk1, Anthony Clark1, Shuiqin Wu2, R. Brandon Miles3, Robert Coates3, Jéròme Maury4,
Mohammad Ali Asadollahi4 & Joe Chappell2
1
Corporate R&D Division, Firmenich SA, Geneva, CH-1211 Switzerland.
2
Department of Plant & Soil Sciences, University of Kentucky, Lexington, KY 40546-0312, USA.
3
Department of Chemistry, University of Illinois, Urbana, IL 61801 USA.
4
 Center for Microbial Biotechnology (Biocentrum/DTU)
Søltofts plads, Building 223, Kgs Lyngby, 2800, Denmark.

Terpenes comprise a large and structurally diverse class of natural products that serve a vast array of
biological functions in nature from microorganisms to animals. In plants for example, they serve as insect
attractants, defense compounds against pathogenic microbes, or as herbivore repellents. Terpene molecules
have been of interest for thousands of years because of their flavor and fragrance properties and their
cosmetic, medicinal and anti-microbial effects. In the Flavor and Fragrance industry alone, the world annual
consumption of volatiles terpene molecules represents 1000s of tons. Terpenes, like most natural products,
are often biosynthesized in small amounts and as complex mixtures by plants and microbes and are thus in
limited supply and/or too expensive. In addition, because of the often-complex structures of these molecules
and because of the requirement of enantiomeric pure molecules, synthetic approaches are often too costly
or inefficient. Therefore, efforts to genetic engineer high yielding production platforms in microbes and plants
have been sought.

The strategies employed to construct biosynthetic production platforms for the production of terpene
molecules for flavor, fragrance and other industrial applications will be presented. This strategy implies the
isolation and characterization of key biosynthetic enzymes and the metabolic engineering of microorganisms
and plants to construct platforms for industrial production of volatile terpenes. The strategies and potential of
engineering plants, relying on the diversion of carbon flow from isopentenyl diphosphate (IPP)
biosynthesized in either the cytosolic or plastidic compartments of plant cells, will be discussed in details.
These developments provide new precedents for engineering natural product biosynthetic pathways into
plants, and specifically a means for generating high levels of terpenes that until now has not been available
for scientific investigation, industrial production, or therapeutic applications.




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Policy Drivers Leading to the Biobased Economy—US Perspectives
Judith B. St. John

United States Department of Agriculture,Agricultural Research Service

5601 Sunnyside Avenue, Beltsville, Maryland 20705-5139, United States of America

judy.stjohn@ars.usda.gov

In 2001, the President put forward his National Energy Policy with over 100 recommendations to increase
domestic energy supplies, encourage efficiency and conservation, invest in energy-related infrastructure,
and develop alternative and renewable sources of energy. Then in 2005 the President signed the first
comprehensive energy legislation in a decade. The Energy Policy Act mandated strengthening America’s
electrical infrastructure and reducing the country’s dependence on foreign sources of energy, increasing
conservation, and expanding the use of clean renewable energy. The next policy driver in the US was the
Advances Energy Initiative announced by the President in the 2006 State of the Union Address. This
Initiative, lead by the Department of Energy, set a national goal of replacing more than 75% of US oil imports
from the Middles East by 2025. The most recent driver is the Administration’s recommendation for Title VII
“Research and Related Matters” of the proposed 2007 Farm Bill. The recommendation calls for establishing
within the Department of Agriculture an Agricultural Bioenergy and Biobased Products Research Initiative
with$500 million over 10 years to advance fundamental scientific knowledge for the improved production of
renewable fuels and biobased products.




Agriculture: the essential underpinning for the bio-based economy

Hilkka Summa

European Commission, Directorate-General for Agriculture and Rural Development

Rue de la Loi 130, 1040 Brussels, Belgium

hilkka.summa@ec.europa.eu

Alternative uses of what the land produces have been at the center of attention since the European
Commission presented its Energy Package with ambitious targets for expanding renewable energy sources.
The European Council endorsed in March the proposal that the EU should set legally binding targets for 20
% of renewable energy and 10 % of transport biofuels by 2020 in overall EU energy consumption. The
energy package was adopted together with a Communication on policy options for limiting the global climate
change to 2° Celsius, emphasising the integration and interdependence of climate and energy policies.

The targets for renewable energy are seen as good news for European agriculture: they promise new outlets
and a positive development of demand and prices at a time when farmers are increasingly faced with
international competition. Biomass is the main source (65 %) of renewable energy in the EU, which means
that agriculture and forestry are the main contributors to a more secure and sustainable energy and climate
policy. Furthermore, expanded uses of agricultural biomass can create value-added production and support
economic fabrics of rural areas.

As renewable energy is promoted to achieve a more sustainable energy future for Europe, concerns have
been raised about the sustainability of increased production and use of biomass for energy. Questions are
asked about impacts on the agricultural environment, deforestation and biodiversity, as well as about
impacts on prices of food and feed and availability of bio-based materials for other non-energy uses. The
Nuremberg Declaration, which the German Presidency addressed to Council, emphasises the ambitious
targets for renewable energy while calling for realising the full potential of renewable resources – including
the expansion of their industrial utilisation. Networking between scientists, industry and policy-makers is
necessary for realising this potential, as there is still some way to go towards a full bio-based economy, not
least because of the relatively high costs of most bio-based products.


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The Common Agricultural Policy includes some specific support measures for the production of energy and
other non-food crops. In the development of a longer term vision of the CAP, the best possible integration of
energy and climate policy into the policy instruments will be continuously assessed, while eyes will be kept
open for the balance between the food, feed and non-food markets.




Crops for biopolymers and platform chemicals

Jan van Beilen

University of Lausannne

Département de Biologie Moléculaire Végétale, Le Biophore, Quartier Sorge, Université de Lausanne

CH-1015 Lausanne, Switzerland

vanbeilen@biotech.biol.ethz.ch

The second report of the biopolymer flagship for the EPOBIO project analyses the suitability of the three
crops, sugar beet, tobacco, and Miscanthus, for the production of platform chemicals and biopolymers. Most
applications in this theme are in an early stage of development, necessitating a longer lead-time to market
(10/15 to 20 years). The strengths and weaknesses of developing each of these three crops as future
industrial crop platforms turned out to be quite different.

Sugar beet has the highest biomass yield of all conventional crops, and was very profitable prior to the CAP-
reform. Industrial utility of sugar beet would be greatly enhanced if new breeding targets aimed at industrial
applications were undertaken. Beyond bioenergy, there are clear opportunities for using beet to produce
novel chemicals and biopolymers. However, social acceptability of transgenic beet for this purpose and the
problematic gene flow situation are likely to play a major determining role in decisions. Tobacco is a
conventional crop that offers interesting potential as an industrial crop. It has many strengths for high yield
production of designer compounds by GM and the possibility for development into a relatively high yielding
biomass crop, if breeding targets are changed. As a non-food crop with limited risks of outcrossing it stands
a good chance of being accepted by the general public in European countries. Miscanthus undoubtedly
holds great promise as a bioenergy crop for the mid- to long-term future. This promise can only be realised
once the grass has been optimised for large-scale commercial cultivation. Miscanthus offers potential for co-
production of added value products in parallel with biomass for biofuels, once genetic transformation has
been established.

Central issues are the future acceptance of GM-crops, especially in the case of sugar beet, fitting the new
crops in the existing supply chains, and integrating the crops in existing or future processing schemes.




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