Enzyme Sugar-Ethanol Platform Project National Renewable Energy Laboratory Enzyme Sugar-Ethanol Platform Project Operated for the U.S. Department of Energy by Midwest Research Institute • Battelle • Bechtel

Project Goal Objective: Develop and demonstrate economical bioethanol technology based on enzymatic cellulose hydrolysis Feedstock Constraint: Develop the technology for an abundant biomass resource that can support production of at least 3 billion gallons of ethanol per year

Approach Select corn stover as feedstock Most abundant, concentrated domestic biomass resource Potential to leverage existing corn harvesting and processing (esp. to produce fuel ethanol) infrastructure and “bridge” industrial contacts Utilize low cost enzymes now being developed Genencor International and Novozymes Biotech Inc. are leading enzyme development work through cost-shared subcontracts from the USDOE. Lower cost enzymes are anticipated in 2003-2004. Demonstrate compelling process economics Validate improved process performance and identify potentially attractive commercialization scenarios.

Increasing Cost & Industrial Involvement Project Scope NREL-led Development Industry-led Commercialization Prelim. Studies Detailed Investigation Process Development Testing and Validation Commercial Launch Stage 1 Stage 2 Stage 3 Stage 4 Stage 5 Gov. & Univ. & Corp. R&D Industry-led deployment Increasing Cost & Industrial Involvement

Strategic Fit The project demonstrates enabling technology for a lignocellulose-based biorefinery The project focuses on the core steps needed to produce sugars, fractionated lignin, and ethanol Industry is focusing on the application of this technology to make new products

External Drivers or Showstoppers Price of Oil and Gasoline (Transportation Fuels) Global supply and demand issue Contingent on fuel standards and energy policy Price and Availability of Starch (Grain) Ethanol Existence of renewable fuel standard Markets for starch ethanol co-products Price and Availability of Corn Stover How much can be removed and what does it cost? What infrastructure needed for collection, storage and delivery? Are there alternative markets that will out compete ethanol? Environmental Regulations and Policies Greenhouse gas mitigation, carbon tax, etc.

Simplified Process Schematic Steam & Acid Enzyme Feed Handling Corn Stover Pretreatment Fermentation Hydrolyzate Broth Recycle Water Waste Water Recycle & Condensate Solids S/L Sep Liquor Conditioning Distillation & StillageTreatment Steam Waste Water Treatment Waste Water This block flow diagram represents the enzymatic biomass to ethanol process as we currently envision it. Corn stover is brought into the plant baled. The bales are then broken, material is washed and sized, and sent to pretreatment where hemicellulose is broken down into its component sugars. The hydrolysate must be conditioned prior to fermentation to eliminate potential fermentation enhibitors, which is where the first S/L separation step takes place. Enzyme is added and simulataneous saccharification and fermentation (SSF) takes place. The broth or “beer” that results is sent to product recovery steps where the ethanol product is concentrated via distillation and other products are treated and segregated. The lignin-rich residue that results is currently sent to the burner/boiler to generate steam and electricity for the plant. If enough residue exists, excess steam and electricity can be made and sold for a credit. Waste Water Biogas & Sludge Syrup Ethanol S/L Sep Cake Steam Utilities Burner/Boiler Turbogenerator Storage Electricity

Relative Cost Contribution by Area Capital Recovery Charge Raw Materials Process Electricity Grid Electricity Total Plant Electricity Fixed Costs Biomass Feedstock 33% Feed Handling 5% Pretreatment / Conditioning 18% SSCF 12% Cellulase 9% (after ~10x cost reduction) Distillation and Solids 10% Recovery Wastewater Treatment 4% 4% Boiler/Turbogenerator Utilities 4% Storage 1% (0.30) (0.20) (0.10) - 0.10 0.20 0.30 0.40

Process Economics Production costs dominated by Feedstock Enzymes - cellulases Capital equipment throughout the plant The focus of the project is to work closely with USDOE, ORNL, USDA, and others, to decrease these key cost factors.

Key Cost Reduction Strategies Minimize feedstock cost Work with ORNL, USDA, and others to reduce the cost of corn stover by developing policies and infrastructure for efficient collection, storage and delivery Minimize enzyme cost Exploit anticipated thermo-stability of lower cost enzymes being developed by Genencor and Novozymes to reduce enzyme and capital costs for process Reduce processing plant capital cost Demonstrate improved integrated process performance Use process engineering techno-economic models to explore potential benefits of co-location and co-products

Market Goals The project targets achieving a commercial production cost of $1.10 per gallon by 2010 This target is based on a combination of technical conversion process performance goals and market considerations The market for ethanol is driven by refinery demand for ethanol as a gasoline blend stock

Ethanol Value-Demand Curve Oak Ridge National Lab’s linear programming model for a generic oil refinery used to estimate ethanol value as a function of demand (usage) Results quantify how the value of ethanol decreases as more of it is used

Higher ethanol demand scenario Refiner Ethanol Demand Curve Reference conditions Higher ethanol demand scenario From G. Hadder (ORNL, 1999)

Demand Curve Findings At $1.10 per gallon, refiners can afford to use 1-5 billion gallons per year of ethanol, depending on the future price of petroleum This estimate does not include the effect of a tax incentives If the tax incentive continues at $0.50 per gallon ethanol, refiners can afford to use 10-11 billion gallons per year

Possible Process Scenario Feedstock Handling Pretreatment S/L Separation Corn Stover Steam & Acid Liquor Solids CO2 Enzyme Lime Ethanol Gypsum Steam Saccharification & Fermentation Conditioning This block flow diagram represents the enzymatic biomass to ethanol process as we currently envision it. Corn stover is brought into the plant baled. The bales are then broken, material is washed and sized, and sent to pretreatment where hemicellulose is broken down into its component sugars. The hydrolysate must be conditioned prior to fermentation to eliminate potential fermentation enhibitors, which is where the first S/L separation step takes place. Enzyme is added and simulataneous saccharification and fermentation (SSF) takes place. The broth or “beer” that results is sent to product recovery steps where the ethanol product is concentrated via distillation and other products are treated and segregated. The lignin-rich residue that results is currently sent to the burner/boiler to generate steam and electricity for the plant. If enough residue exists, excess steam and electricity can be made and sold for a credit. Wastewater Treatment Distillation & Ethanol Purification Burner/Boiler Turbogenerator Lignin Residue Steam Electricity

Feedstock – Corn Stover Model Parameter Value Feedstock Cost $35/dry ton Cellulose Fraction 37.1% Xylan Fraction 19.9% Arabinan Fraction 2.5% Mannan Fraction 1.3% Galactan Fraction 1.7% Lignin Fraction 20.7% * Composition is average of 5 stover pretreatment runs at NREL

Feedstock – Corn Stover Rationale for data: Feedstock Cost: Walsh, et.al. (ORNL) Demonstrated at Harlan, IA Feedstock Composition: Averaged stover data (NREL) Research underway to improve analysis methods and understand major sources of compositional variance Model Parameter Value Feedstock Cost $35/dry ton Cellulose Fraction 37.1% Xylan Fraction 19.9% Arabinan Fraction 2.5% Mannan Fraction 1.3% Galactan Fraction 1.7% Lignin Fraction 20.7%

Feedstock – Corn Stover Large Cost Impact Feedstock Cost Impact $1.50 $1.48 $50 / BDT $1.28 $0.65/gal $35 / BDT $1.00 $0.83 Minimum Ethanol Selling Price ($/gal etoh) $0 / BDT $0.50 $0.00 Process Case

Feedstock Handling Brings biomass into facility Prepares biomass for pretreatment Subcontract work to develop less expensive handling systems

Pretreatment - Example Converts hemicellulose to monomeric sugars Makes cellulose more susceptible to enzymatic hydrolysis Conditions: Technology Dilute Acid Reactor Solids Concentration 30 % Residence Time 2 min Acid Concentration 1.1 % Temperature 190 °C Reactor Metallurgy Incoloy 825-clad

Pretreatment - Example Converts hemicellulose to monomeric sugars Makes cellulose more susceptible to enzymatic hydrolysis Conditions: Technology Dilute Acid Reactor Solids Concentration 30 % Residence Time 2 min Acid Concentration 1.1 % Temperature 190 °C Reactor Metallurgy Incoloy 825-clad Rationale for Data: Corn stover steam gun expts Hot wash process expts Prior research on hardwood feedstocks

Pretreatment - Example Reactor Solids Cost Impact: Prehydrolysis Solids Concentration Sensitivity $1.50 $1.45 $1.40 Minimum Ethanol Selling Price ($/gal) $1.35 $1.30 $1.25 $1.20 10% 15% 20% 25% 30% 35% 40% Prehydrolysis Solids Concentration inside Reactor

Pretreatment - Example Xylose Yield Cost Impact: Xylose Yield Cost Impact $1.50 50% xylose $1.50 $0.27/gal 85% xylose $1.28 $1.23 95% xylose $1.00 Minimum Ethanol Selling Price ($/gal etoh) $0.50 $0.00 Process Case

Solid/Liquid Separation Separate pretreated solids from liquor Countercurrent hot water wash increases enzymatic digestibility and solubilizes recoverable lignin Conditions: Equipment Pressure Filter Separation Temp 135 °C Separation Pressure 5 atm Conditioning Overlime only Wash / Hydrolysate Ratio 0.58 kg/kg

Solid/Liquid Separation Separation of pretreatment solids from liquor Countercurrent hot water wash increases enzymatic digestibility and solubilizes recoverable lignin Rationale for Data: Lower acetylation of corn stover hemicellulose means IX not needed to reduce acetic acid levels Hot wash process expts Harris subcontract Working towards pilot scale demonstration at NREL Conditions: Equipment Pressure Filter Separation Temp 135 °C Separation Pressure 5 atm Conditioning Overlime only Wash / Hydrolysate Ratio 0.58 kg/kg

Solid/Liquid Separation Cost Impact: Conditioning Sensitivity $1.50 $1.45 $1.40 Minimum Ethanol Selling Price ($/gal etoh) $1.35 $0.08 / gal $1.30 $1.25 $1.20 OL only IX / OL Process Case

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Saccharification: Enzyme Source purchased Enzyme Cost $0.11/gal EtOH SHF vs. SSF Hybrid Temperature 65 °C Residence Time 1.5 days Cellulose to Glucose Yield 90%

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Saccharification: Enzyme Source purchased Enzyme Cost $0.11/gal EtOH SHF vs. SSF Hybrid Temperature 65 °C Residence Time 1.5 days Cellulose to Glucose Yield 90% Rationale for Data: Enzyme Cost is 10x-reduction from Glassner-Hettenhaus parameters 10x-reduction is goal of enzyme subcontracts Hybrid design advantageous for more thermotolerant enzyme system

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Fermentation: Residence Time 2 days Temperature 37 °C Nutrient Requirement 0.25% CSL 0.33 g/L DAP Effective Solids Conc. 20%

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Rationale for Data: Previous work based on conversion of hardwood hydrolyzates using Z. mobilis Nutrients Strain improvements 2nd Gen. ethanologen projects at NREL Literature search Fermentation: Residence Time 2 days Temperature 37 °C Nutrient Requirement 0.25% CSL 0.33 g/L DAP Effective Solids Conc. 20%

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Yields: Glucose to Ethanol Yield 92% Xylose to Ethanol Yield 85% Arabinose to Ethanol Yield Contamination Loss 5%

Saccharification & Fermentation Enzymatic hydrolysis of cellulose to glucose Microbial conversion of sugars to ethanol Rationale for Data: Initial work based on glucose and xylose cofermenting Z. mobilis Improved strains constructed with broader pentose and hexose substrate ranges rDNA yeast Ingram et al. constructs Yields: Glucose to Ethanol Yield 92% Xylose to Ethanol Yield 85% Arabinose to Ethanol Yield Contamination Loss 5%

Saccharification & Fermentation Enzyme Cost Impacts: Enzyme Cost Impact $1.07 / gal $2.24 $2.00 $1.01/gal $.50 / gal $1.67 $1.50 $0.11 / gal $1.28 Minimum Ethanol Selling Price ($/gal etoh) $1.23 $0.06 / gal $1.00 $0.50 $0.00 Process Case

Saccharification & Fermentation Cost Impacts: Fermentation Residence Time Cost Impact $1.50 $1.45 $1.40 Minimum Ethanol Selling Price ($/gal etoh) $1.35 $1.32 $1.30 7 days $0.07/gal $1.28 3.5 days $1.25 $1.25 1 day $1.20 Process Case

Saccharification & Fermentation Cost Impacts: Fermentation Yield Cost Impact $2.40 $2.10 70% Minimum Ethanol Selling Price ($/gal) $1.80 92% $1.50 95% $1.33 $1.28 $1.23 $1.20 glucose only add 85% xylose add 85% arabinose all other sugars 85%

Saccharification & Fermentation Cost Impact: Contamination 5%  7% equates to $0.02/gal increase Nutrient Cost $0.035/gal 89% CSL, 11% DAP

Distillation & Ethanol Purification Separation of ethanol and CO2 from “beer”

Wastewater Treatment Anaerobic and aerobic treatment Reduce Biochemical Oxygen Demand (BOD) Recycle water

Burner/Boiler/Turbogenerator Biomass boiler generates steam from lignin residue Excess electricity from generator sold to power grid ($0.04/kWh credit) High capital cost area

Current Status Completing Stage 2 Next step: Gate 3 review Compelling scenario identification Technology selection Stage 3 plan development Next step: Gate 3 review Planned for January 2002

Conversion-related Cost Reduction Opportunities Stage 2 technology selection focus Is a better pretreatment technology available? Higher yields, lower capital or operating costs Is a better fermentation strain available? Broader sugar utilization range, higher ethanol yields, better compatibility with enzyme Stage 3 technology improvement focus Are better cellulases available and how do they benefit integrated process economics?

Technology Selection Tiered screening approach being applied to ensure best options will be studied in Stage 3 1st screen: Efficacy 2nd screen: Readiness and availability Stage 2 focus: Pretreatment technology Fermentation strain

Co-location-related Cost Reduction Opportunities Better feedstock price Proximity to transportation Farmer cooperative Reduce capital cost Utilize existing utilities and processing infrastructure within site constraints

Cost Reduction Strategies, cont. Reduce conversion plant capital cost co-locate into a dry mill expansion co-locate with a coal-fired power plant co-locate with both a dry mill and power plant Reduce capital cost through better financing Loan guarantee? City/county/state/federal support or tax benefits?

Cost Reduction through Co-products New process case potentially enables “sugar platform” and “lignin platform” co-products Value-added co-products can increase upside of process commercialization and mitigate overall risk While we can explore the possibilities, development of prospective co-products must be led by industry!

Potential Bioethanol Co-products Hemicellulose Hydrolyzate (Xylose) Cellulose Hydrolyzate (Glucose) Cell Mass, Enzymes (Protein, etc.) Process Residue Liquids Pretreatment Hemicellulose Hydrolysis 1o Enzymatic Cellulose Hydrolysis 2o Enzymatic Hydrolysis & Fermentation Ethanol Recovery & Purification Biomass EtOH Soluble Lignin (Low/Medium MW Phenolics) Process Residue Solids Insoluble Lignin (High MW Phenolics)

Sugar & Lignin Platform Biorefinery Product(s) Recovered Lignin Purification & Drying of Lignin Product(s) Renewable Biomass Feedstock Catalyst Steam, Acid, Enzyme, etc.) Water Steam Unrecovered Lignin Sugar Product(s) Sugar-rich Hydrolyzate Feedstock Handling Concentration & Purification of Sugar Product(s) Biomass Fractionation Water Steam Recycle Water Waste Water Unrecovered Sugars Hydrolyzate & Residual Solids Fuel Ethanol Ethanol Production & Recovery Waste Water Treatment Make-up Water Waste Water Biogas & Sludge Residual Solids & Syrup Steam Unrecovered Lignin Steam Generation Power Production (Turbogenerator) Electricity Steam