Presentation of Bin Yang for the Workshop on Hydrolysis Route for Cellulosic Ethanol from Sugarcane. Apresentação de Bin Yang realizada no "Workshop on Hydrolysis Route for Cellulosic Ethanol from Sugarcane" Date / Data : February 10 - 11th 2009/ 10 e 11 de fevereiro de 2009 Place / Local: Unicamp, Campinas, Brazil Event Website / Website do evento: http://www.bioetanol.org.br/workshop1

1. Progress and Outlook for Low Cost
Pretreatment of Cellulosic Biomass for
Biological Production of Fuels and Chemicals
Bin Yang and Charles E. Wyman
Chemical and Environmental Engineering and
Center for Environmental Research and Technology (CE-CERT)
University of California
Workshop on Hydrolysis Route for Cellulosic Ethanol From Sugarcane
February 11, 2009
Campinas, Brazil

2. Sustainable Alternatives for
Transportation
Sustainable Primary Secondary Human
Resources Intermediates Intermediates Needs
Sunlight
Wind Biomass Organic Fuels
Ocean/
hydro
Transportation
Geothermal Hydrogen
Electricity
Nuclear Batteries
By Lee Lynd, Dartmouth
2

3. Reaction Pathways for Biomass
Conversion
High Temperature
Cellulosic Conversion: Catalytic
Pyrolysis, Conversion
Liquefaction, in
Supercritical, Gas Phase Oil Refining
Gasification
Reactions:
Cellulosic Catalytic Biofuels
Biomass Cracking, Biochemicals
Hydrotreating
Catalytic
Conversion
in
Low Temperature Aqueous Phase
Cellulosic Conversion:
Acid Hydrolysis
Enzymatic Hydrolysis
From George Huber, UMass
3

4. Alternative Fuel Mandates in US
From Energy Independence and Security Act of 2007
4

5. Biological Processing of
Cellulosic Biomass
 Biological processing of cellulosic biomass
to ethanol and other products offers the high
yields vital to economic success
 Biological processing can take advantage of
the continuing advances in biotechnology to
dramatically improve technology and
reduce costs
5

6. Historical and Projected Cellulosic Ethanol
700 Costs
Minimum Ethanol Selling Price (cents/gal)
600
Cost reductions
500 to date
400
Future goal
300
200
100
0
2009
2007
2008
2006
2010
2005
2004
2003
2012
2002
2001
2011
Enzyme Feedstock Conversion
NREL Modeled Cost
6

7. Key Processing Cost Elements
~9% of cost
Total ~39% of cost
Cellulase enzyme
~18% of cost
~33% of cost
Biomass Stage 2 Residual solids:
Stage 1
Enzymatic cellulose,
Pretreatment Solids: cellulose,
Chemicals hydrolysis hemicellulose,
hemicellulose, lignin
lignin ~12% of cost Dissolved sugars,
oligomers
Dissolved sugars,
oligomers, lignin Stage 3
Sugar
fermentation
7

8. Pretreatment
 Reduce biomass recalcitrance to
attack by enzymes
 High sugar yields are vital
8

9. Disruption of Cellulosic Biomass by
Pretreatment
Heat Cellulose
Lignin
Disruption
Hemicellulose

10. Importance of Pretreatment
 Although significant, feedstock costs are low
relative to petroleum
 In addition, feedstock costs are a very low fraction
of final costs compared to other commodity
products
 Pretreatment is the most costly process step:
 Low yields without pretreatment drive up all
other costs more than amount saved
 Conversely enhancing yields via improved
pretreatment would reduce all other unit costs
 Need to reduce pretreatment costs to be
competitive
10

11. Central Role and Pervasive Impact of Pretreatment
for Biological Processing
Enzyme
production
Harvesting,
Biomass Enzymatic Sugar
storage, Pretreatment
production hydrolysis fermentation
size reduction
Hydrolyzate Hydrolyzate Ethanol
conditioning fermentation recovery
Residue Waste
utilization treatment
11

12. Feedstocks Vs. Yields
Biomass Glucan Xylan Theoretical Potential
feedstock % % Ethanol Yield Real Ethanol
(gal/ton) Yield
(gal/ton)
Corn stover 36.1 21.4 105 89
Switchgrass 35.0 21.8 104 88
Sugarcane
bagasse 38.6 20.4 108 92
Poplar 43.8 14.9 107 91
Aspen wood 44.8 14.9 109 98
Miscanthus 46.0 19.8 120 102

13. Economic Impact of R&D-Driven Improvements
Increase hydrolysis yield 3% Overcoming the
13%
recalcitrance of
Halve cellulase loading
biomass
Eliminate pretreatment 22%
Consolidated bioprocessing 41%
(CBP)
Simultaneous C5 & C6 Use 6%
Improving
Increased fermentation 2% production of
yield targeted
Increased ethanol titer 11%
products
Increased ethanol titer following 6%
CBP
0% 10% 20% 30% 40% 50%
Error bars denote two Processing Cost Reduction
different base cases
From Nature Biotech. 2008
13

14. Key Features of CAFI Leading Pretreatments
for Corn Stover
Pretreatment Temperature, Reaction Chemical Percent Other notes
Acid system oC time, agent used chemical
minutes used
Dilute acid 160 20 Sulfuric 0.49 25% solids concentration during
run in batch tubes
acid
Flowthrough 200 24 none 0 Continuously flow just hot water at
10mL/min for 24minutes
Partial flow 200 24 none 0 Flow hot water at 10mL/min from
4-8 minutes, batch otherwise
pretreatment
Controlled 190 15 none 0 16% corn residue slurry in water
pH
AFEX 90 5 Anhydrous 100 62.5% solids in reactor
(60% moisture dry weight basis), 5
ammonia minutes at temperature
ARP 170 10 ammonia 15 Flow aqueous ammonia at 5
mL/min without presoaking
Lime 55 4 weeks lime 0.08 g Purged with air.
Base CaO/g
biomass
14

15. CAFI Feedstock: Corn Stover
From BioMass AgriProducts, Harlan IA and Kramer Farm, Wray, CO
Component Composition Ethanol yield
wt % gal/ton
Glucan 36.1 62.1
Xylan 21.4 37.7
Arabinan 3.5 6.2
Mannan 1.8 3.1
Galactan 2.5 4.3
Lignin 29.1
Protein nd
Acetyl 3.6
Ash 1.1
Uronic Acids nd
Extractives 3.6
Total maximum ethanol potential 113.3

16. Overall Yields for Corn Stover
at 15 FPU/g Glucan
Xylose yields* Glucose yields* Total sugars*
Pretreatment
Total Stage Total Combined
system Stage 1 Stage 2 Stage 2 Stage 1 Stage 2
xylose 1 glucose total
Maximum
37.7 37.7 37.7 62.3 62.3 62.3 100.0 100.0 100.0
possible
Increasing pH
Dilute acid 32.1/31.2 3.2 35.3/34.4 3.9 53.2 57.1 36.0/35.1 56.4 92.4/91.5
SO2 Steam
14.7/1.0 20.0 34.7/21.0 2.5/0.8 56.7 59.2/57.5 17.2/1.8 76.7 93.9/78.5
explosion
Flowthrough 36.3/1.7 0.6/0.5 36.9/2.2 4.5/4.4 55.2 59.7/59.6 40.8/6.1 55.8/55.7 96.6/61.8
Controlled
21.8/0.9 9.0 30.8/9.9 3.5/0.2 52.9 56.4/53.1 25.3/1.1 61.9 87.2/63.0
pH
AFEX 34.6/29.3 34.6/29.3 59.8 59.8 94.4/89.1 94.4/89.1
ARP 17.8/0 15.5 33.3/15.5 56.1 56.1 17.8/0 71.6 89.4/71.6
Lime 9.2/0.3 19.6 28.8/19.9 1.0/0.3 57.0 58.0/57.3 10.2/0.6 76.6 86.8/77.2
*Cumulative soluble sugars as total/monomers. Single number = just monomers.
16

17. CAFI Feedstock: Poplar
Feedstock: USDA-supplied hybrid poplar (Alexandria, MN)
 Debarked, chipped, and milled to
pass ¼ inch round screen
Component Composition Ethanol yield
wt % gal/ton
Glucan 43.8 75.4
Xylan 14.9 26.1
Arabinan 0.6 1.1
Mannan 3.9 6.8
Galactan 1.0 1.8
Lignin 29.1
Protein nd
Acetyl 3.6
Ash 1.1
Uronic Acids nd
Extractives 3.6
Total maximum ethanol potential 111.1

18. Sugar Yields for CAFI Standard
Poplar at 15 FPU/g Glucan
Xylose yields Glucose yields Total sugar monomers
Pretreatment
Total Total Combined
system Stage 1 Stage 2 Stage 1 Stage 2 Stage 1 Stage 2
xylose glucose total
Maximum
25.7 25.7 25.7 74.3 74.3 74.3 100 100 100
possible
SO2 Steam
Increasing pH
19.2/14.0 2.4 21.6/16.4 2.3 72.0 74.3 21.6/16.3 74.4 95.9/90.7
explosion
Dilute acid
16.1 2.4 18.5 17.7 46.6 64.3 33.8 49.0 82.8
(Sunds)
Controlled
21.2/1.0 8.8 30.0/9.8 1.4/0.1 42.3 43.7/42.4 22.6/1.1 51.1 73.7/52.2
pH
AFEX 0.0 13.4 13.4 0.0 39.4 39.4 0.0 52.8 52.8
AFEX with
76.9/55.
cellulase + 0.0 17.5/13.0 17.5/13.0 0.0 76.9/55.0 0.0 94.3/68.0 94.3/68.0
0
xylanase
ARP 9.6/0.0 8.2/8.0 17.7/8.0 0.4/0.0 36.3 36.6/36.3 10.0/0.0 44.5/44.3 54.5/44.3
74.4/72.
Lime 1.1/0.0 20.1/17.1 21.2/17.1 0.2/0.0 74.6/72.5 1.3/0.0 94.5/89.6 95.8/89.6
5
*Cumulative soluble sugars as total/monomers. Single number = just monomers.
18

19. Projected Costs Virtually the Same with Oligomer
1.75
Utilization (Black Bars) for Corn Stover
1.50
MESP, $/gal EtOH
1.25
1.00
Dilute Acid Hot Water AFEX ARP Lime
w/o Oligomer Credit w/ Oligomer Credit

20. Opportunities to Reduce Pretreatment Cost
 Need to reduce cost from the operation units:
 Energy use
 Costs of chemicals
 Containment costs
 Size reduction requirements
 Prefermentation conditioning
 Achieve high yields for multiple crops, sites, ages, harvest
times
 While increasing yields
 And limiting inhibitors to bioprocessing
 Advanced pretreatment processes will pay big dividends
 Key: understand pretreatment mechanisms and how to
improve yields
20

21. Effect of Flow Rate on Xylan Removal from
Corn Stover and Oat Spelt Xylan
100
Xylan/2mL/min
90
Xylan/25mL/min
Percent of potential total xylose, %
80
Xylan/0mL/min
70
Corn stover/25mL/min
60
50
Corn stover/2mL/min
40
30 Corn stover/0mL/min
20
10
0
0 2 4 6 8 10 12
Time, minutes
21

22. Yield of Xylan Oligomers and Total Xylan
Recovery in Hydrolysate
Flow rate Yield, %
Feedstock mL/min Total DP1 to Long Ratio of
xylan 30 chain shorter chain
recovery1 oligomer2
to longer
chain
oligomer
Corn stover 0 (Batch) 38.1 28.1 10.0 2.8
2 48.2 20.3 27.9 0.7
25 73.3 9.1 64.2 0.1
Oat spelt xylan 0 (Batch) 73.1 30.1 43.0 0.7
2 92.1 0.3 91.8 0.003
25 91.1 0.4 90.8 0.004
1. Total xylan recovery = yield of xylose in hydrolysate+ yield of oligomers in
hydrolysate (xylose equivalent);
2. Yield of long chain oligomer (DP>30) = total xylan recovery – yield of DP1∼30.
22

23. Effect of Xylan Removal on Digestibility of Corn
Stover for Batch and Flowthrough Reactors
100
90
80
Enzymatic digestibility,%
70
60
50
40
30
Uncatalyzed batch tube (160-220C, 5% solid loading)
Catalyzed batch tube (160-220C, 5% solid loading, 0.1% acid)
Uncatalyzed flowthrough (160-220C, flow rate of 2ml/min)
20 Uncatalyzed flowthrough (160-220C, flow rate of 7.5ml/min)
Uncatalyzed flowthrough (160-220C, flow rate of 25ml/min)
Catalyzed flowthrough (160-220C, flow rate of 2ml/min)
Catalyzed flowthrough (160-220C, flow rate of 7.5ml/min)
Catalyzed flowthrough (160-220C, flow rate of 25ml/min)
10
0 20 40 60 80 100
Xylan removal,%
23

24. Effect of Lignin Removal on Digestibility of Corn
Stover for Batch and Flowthrough Reactors
100
90
80
Enzymatic digestibility,%
70
60
50
40
30 Uncatalyzed batch tube (160-220C, 5% solid loading)
Catalyzed batch tube (160-220C, 5% solid loading, 0.1% acid)
Uncatalyzed flowthrough (160-220C, flow rate of 2ml/min)
Uncatalyzed flowthrough (160-220C, flow rate of 7.5ml/min)
20 Uncatalyzed flowthrough (160-220C, flow rate of 25ml/min)
Catalyzed flowthrough (160-220C, flow rate of 2ml/min)
Catalyzed flowthrough (160-220C, flow rate of 7.5ml/min)
Catalyzed flowthrough (160-220C, flow rate of 25ml/min)
10
0 10 20 30 40 50 60 70 80 90
Lignin removal,%
24

25. Role of Lignin in Pretreatment
 Historically divergent opinions on role of lignin versus
hemicellulose in access of enzymes to cellulose in
pretreated biomass
 Our results suggest that lignin must be disrupted to achieve
high enzymatic hydrolysis
 Hemicellulose removal serves as a marker of lignin disruption but
is not the cause of better digestion
 Even better results if remove lignin
 Lignin-xylan oligomers and their solubility could have a large
effect on the rates and yields of lignocellulosic biomass
pretreatment
25

26. Mission of UCR Ethanol Research
 Improve the understanding of biomass
fractionation, pretreatment, and cellulose
hydrolysis to support applications and advances
in biomass conversion technologies for
production of low cost commodity products
 Develop advanced technologies that will
dramatically reduce the cost of production
26

27. Current Research Topics
 Diesel fuel from biomass – DARPA
 Effect of different pretreatments on enzymatic hydrolysis
of poplar wood and switchgrass – US DOE
 Lead Consortium with Auburn, Michigan State, NREL, Purdue,
Texas A&M, U. British Columbia, and Genencor
 Pretreatment of cellulosic biomass for BioEnergy Science
Center (BESC), $25million/yr DOE Center
 Continuous hydrolysis and fermentation – USDA
 Continuous fermentations of pretreated biomass - NIST
 Fundamentals of biomass pretreatment – Mascoma
Corporation
 Evaluation of advanced plants – Mendel Biotechnology
 Enzyme inhibition by oligomers – Bourns College of
Engineering
27

28. 4
”
Example Experimental Systems
Pretreatment tubes Pretreatment reactor Flowthrough Reactor
Pretreatment steam gun HTP pretreatment system Continuous Fermentation
28

29. Biomass Refining Consortium for Applied
Fundamentals and Innovation (CAFI)
29

30. Agricultural and Industrial Advisory Board
CAFI DOE Project
Quang Nguyen, Abengoa Bioenergy Kendall Pye, Lignol
Jim Doncheck, Arkion Life Sciences Wei Huang, LS9
Gary Welch, Aventinerei Jim Flatt, Mascoma
Mohammed Moniruzzaman, BioEnergy Intl Farzaneh Teymouri, MBI
Paris Tsobanakis, Cargill James Zhang, Mendel
James Hettenhaus, CEA Richard Glass, NCGA
Steve Thomas, CERES James Jia, NorFalco Sales
Lyman Young, ChevronTexaco Joel Cherry, Novozymes
Mike Knauf, Codexis Mark Stowers, Poet
Julie Friend, DuPont Ron Reinsfelder, Shell
Jack Huttner, Genencor Paul Roessler, Synthetic Genomics
Don Johnson, GPC (Retired) Carmela Bailey, USDA
Jeff Gross, Hercules Don Riemenschneider, USDA
Peter Finamore, John Deere Kevin Gray, Verenium
Glen Austin, Lallemand Ethanol Technology Chundakkadu Krishna, Weyerhaeuser
30

31. The BESC Team: Recently Funded by
DOE for $125 Million Over 5 Years
Joint Institute for Biological Sciences
• Oak Ridge National Laboratory
• University of Georgia
• University of Tennessee
• National Renewable Energy
Laboratory
• Georgia Tech
Alternative Fuels User Facility • Samuel Roberts Noble Foundation
• Dartmouth
• ArborGeni
• Mascoma
• Verenium
• U California-Riverside
Complex Carbohydrate Research Center
• Cornell, Washington State, U
Minnesota, NCSU, Brookhaven
National Laboratory, Virginia Tech
31

32. BESC - A Highly Integrated Cutting-
Edge Research Team
32

33. Closing Thoughts
 Biology provides a powerful platform for low cost fuels
and chemicals from biomass
 Can benefit both crop production and conversion
systems
 The resistance of one biological system (cellulosic
biomass) to the other (biological conversion) requires a
pretreatment interface
 Advanced pretreatment systems are critical to enhancing
yields and lowering costs
 Not all pretreatments are equally effective on all
feedstocks
 Focus on 2 biologies - plants and biological conversion -
without integrating their interface – pretreatment – will not
significantly lower costs
33

34. Charles Wyman Bin Yang
Simone
Brethauer Jaclyn
DeMartini Mirvat
Ebrik
Heather
McKenzie Tim Redmond
Jian
Shi
Michael
Studer Taiying Zhang
Rajeev Kumar Qing Qing

35. Acknowledgments
 Ford Motor Company
 The BioEnergy Science Center, a U.S. Department of Energy Bioenergy
Research Center supported by the of Biological and Environmental
Research Office in the DOE Office of Science
 DARPA
 Mascoma Corporation
 Mendel Biotechnology
 National Institute of Standards and Technology, award number
60NANB1D0064
 USDA National Research Initiative Competitive Grants Program, contract
2008-35504-04596
 US Department of Energy Office of the Biomass Program, contract DE-
FG36-07GO17102
 The University of California at Riverside
 The University of Massachusetts, Amherst
 Numerous past and present students, coworkers, and partners who make
our research possible
35

36. Questions???
36