A New Biorefinery Platform Intermediate

1. A New Biorefinery Platform Intermediate Principal Investigator: Doug Cameron Project Managers: Jim Millis, Paris Tsobanakis 2005 OBP Bi-Annual Peer Review Integrated Biorefinery Platform November 16, 2005

2. Overview • Scope of Work Plan • Cargill, in partnership with Codexis, is developing a fermentation route to a novel platform chemical, 3-hydroxypropionic acid (3-HP) • We are also collaborating with PNNL to develop novel catalytic process for the conversion of 3-HP to acrylic acid and acrylamide Microbial fermentation Chemical catalysis Acrylic Acid Acrylamide Glucose 3-HP

3. Overview • Award was for $6.09 million in matching DOE funds for a 3 year program (total $12.2 MM), representing a 50/50 cost share • Biotech/Codexis work started in May 2003; Chemical conversion/ PNNL work started in December, 2003 • The pace of the program was slowed after one year as commercial activities lagged technology development. • $0.60 MM received in FY03 (May to September) • $1.66 MM received in FY04 • $0.77 MM received in FY05 • Request for FY06 and beyond to be submitted by end of November • 50% complete • Completion of the program will require a one year extension

4. Overview • The program is currently in Development Phase • Barriers addressed • 3-HP is a novel platform chemical (identified as a Top Ten product) with the potential to impact both large, commodity markets and new, emerging markets • Technical barriers addressed: • Specific improvements in metabolic pathway engineering to yield highly productive, high yield fermentation technology • Novel product recovery and process integration • Design of new catalysts and processes (for derivatives of 3-HP)

5. Project Goals and Objectives Cargill, through collaboration with their partners Codexis and PNNL will develop a new bio-based platform technology to produce a portfolio of products based on 3-HP produced by the fermentation of carbohydrates. This project will deliver an organism and process for 3-HP production, and catalysts and complete process concepts suitable for piloting and scale-up for industrial production. Our current timeline fits well with the FY08 Solicitation schedule

6. Project Strategic Fit poly(hydroxypropionate) 1,3-propanediol EEP 3-hydroxypropionic acid acrylamide malonic acid acrylic acid

7. Project Strategic Fit 1994 = 100 Market dynamics have emerged to make 3-HP conversion to acrylic as an attractive alternative to propylene oxidation

8. Project Strategic Fit 57% Propylene Polypropylene • Despite a global market of 6.6 billion pounds for acrylic acid, propylene used for acrylic production is a small portion of propylene supply, with price driven principally by polypropylene • Growth requires one new world class plant every year 12% Acrylonitrile Adiponitrile ABS/SAN resins Acrylamide Nitrile rubber 10% Propylene oxide Polyurethane polyols Propylene glycols Glycol ethers 8% Cumene Phenol Acetone 7% Oxo Chemicals (butanol, 2-EH) 4% Acrylic acid 4% Isopropyl alcohol 4% Polygas chemicals (nonene, dodecene) 3% Other chemical uses

9. Project Strategic Fit • Sponsoring BU within Cargill is Industrial Bioproducts, a member of the Emerging Business Platform • This project enables a lower cost (and less price volatile) route to acrylic acid from renewable sugars. Joint analysis of production economics with an industry leader indicates a potential manufacturing cost advantage vs. propylene oxidation of >5¢ per pound for a Midwest plant. • Energy consumption would be reduced 61% compared to the petrochemical route. By 2020, the acrylic acid product alone would displace 7,200 barrels of oil/day with domestically produced corn. • The biorefinery route to acrylic acid would eliminate 600,000 lbs of SOx, 1 million lbs of NOx and 2.8 billion lbs of CO2 annually.

10. Project Approach Glucose NADH ATP 1 2 3 malonate semialdehyde L--alanine -alanine Pyruvate glutamate α-ketoglutarate α-ketoglutarate glutamate 4 NADH 1 – Pyruvate / alanine aminotransferase 2 – Alanine 2,3-aminomutase 3 – -alanine aminotransferase 4 – 3-HP dehydrogenase 3-HP

11. Project Collaboration Microbial catalysis/ fermentation Chemical catalysis Acrylic Acid Acrylamide Glucose 3HP Task Group 1 Task Group 2 Task Group 3 Codexis: Enzyme & pathway evolution Cargill: Host strain construction & fermentation process PNNL: Catalyst & process development Cargill: Process design, integration & economics

12. Project Tasks Task Groups • Biochemical pathway optimization • Host strain construction and fermentation process development • Catalyst screening and development, process development and process integration for the conversion of 3-HP to acrylic acid

13. Project Tasks • Random • Phylogenetic – semisynthetic • Targeted • Hit shuffling • ProSAR Relative improvement WT Rd4 Rd1 Rd2 Rd3 Rd6 Rd7 Rd5 Rd8 Parent Rounds of shuffling Task Group 1: Biochemical pathway optimization (Codexis) Targets • Improve performance of Enzyme 2 to meet productivity targets • Balance supporting pathway enzymes to maximize productivity and yield Milestones • Performance measured by fermentation productivity (see Task Group 2) Technical Risks • Enzyme 2 does not meet activity target Future plans • Continue optimization of Enzyme 2 • Perform pathway shuffling to optimize pathway performance

14. Project Tasks Task Group 2: Host strain construction and fermentation process development (Cargill) Targets • Host strain selected and pathway engineered into strain • Pathway constructed and 3-HP produced • Fermentation optimization to meet economic targets Milestones • Specific productivity targets communicated to DOE (confidential) Technical Risks • Targets for high productivity and titer are not met Future plans • Incorporate improved variants of Enzyme 2 into pathway • Improve yield and productivity of fermentation process

15. Project Tasks Task Group 3: Catalyst screening and development, process development and process integration for the conversion of 3-HP to acrylic acid (PNNL, Cargill) Targets • Catalysts screened and selected for the dehydration of 3-HP to acrylic acid • Catalyst & process conditions demonstrated near quantitative conversion and selectivity Milestones • Milestones met for pure 3-HP Technical Risks • Catalyst robustness/ effect of impurities Future plans • Demonstrate extended catalyst life • Demonstrate catalyst performance on 3-HP produced by fermentation • Process integration

16. Market & Customers • This project enables the first biobased route to acrylic acid, a commodity product with a global market approaching 7 billion pounds per year. • Meeting economic targets for acrylic will also enable other 3-HP derivatives. • 1,3-propanediol • malonic acid and others

17. Competitive Advantage • Cargill’s route to acrylic acid through 3-HP is advantaged over other technology options • Propane oxidation is unproven and does not meet DOE OBP objectives • Lactic acid dehydration pursued by many parties without success • Future may allow use of biomass feedstock for Cargill process, thus driving costs downward • Relative attractiveness of 3-HP to acrylic technology will depend on propylene price, but current forecast for high prices and volatility provides the basis for optimism

18. Progress and Accomplishments • Production of 3-HP from glucose demonstrated by a novel pathway • Catalyst discovered for 3-HP to acrylic acid process • Dehydration reactor designed to overcome equilibrium limitations

19. Future Work Development Stage Plans • Due to a reduced rate of spend in FY04 and FY05, Cargill requests an extension of one year to complete the program. New proposed date for completion is May ‘07. • Plans are to accelerate the spend over the next 18 months. • Interim milestones and commercial plans communicated to DOE in July 2005 review (confidential). • Our current timeline fits well with the FY08 Solicitation schedule