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Comparative Data for Enzymatic Digestion of Corn Stover and Poplar Wood after Pretreatment by Leading Technologies. Charles E. Wyman, Dartmouth College/University of California Rajeev Kumar, Dartmouth College Bruce E. Dale, Michigan State University

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slide1

Comparative Data for Enzymatic Digestion of Corn Stover and Poplar Wood after Pretreatment by Leading Technologies

Charles E. Wyman, Dartmouth College/University of California

Rajeev Kumar, Dartmouth College

Bruce E. Dale, Michigan State University

Richard T. Elander, National Renewable Energy Laboratory

Mark T. Holtzapple, Texas A&M University

Michael R. Ladisch, Purdue University

Y. Y. Lee, Auburn University

Mohammed Moniruzzaman1, Genencor International

John N. Saddler, University of British Columbia

1 – now with BioEnergy International, LLC

AIChE Annual Meeting

San Francisco, California

November 16, 2006

Biomass Refining CAFI

central role and pervasive impact of pretreatment for biological processing
Central Role and Pervasive Impact of Pretreatment for Biological Processing

Enzyme

production

Biomass

production

Harvesting,

storage,

size reduction

Pretreatment

Enzymatic

hydrolysis

Sugar

fermentation

Hydrolyzate

conditioning

Hydrolyzate

fermentation

Ethanol

recovery

Residue

utilization

Waste

treatment

Biomass Refining CAFI

key processing cost elements

33%

Biomass Feedstock

Capital Recovery

Charge

5%

Feed Handling

Grid Electricity

18%

Pretreatment / Conditioning

Raw Materials

SSCF

12%

Total Plant

Electricity

(after ~10x cost reduction)

9%

Cellulase

Process Elect.

Distillation and Solids

10%

Recovery

Fixed Costs

4%

Wastewater Treatment

Net 4%

Boiler/Turbogenerator

Utilities

4%

Storage

1%

(0.20)

(0.10)

-

0.10

0.20

0.30

0.40

Key Processing Cost Elements

Biomass Refining CAFI

importance of pretreatment
Importance of Pretreatment
  • Although significant, feedstock costs are low relative to petroleum: $40/dry ton ~ $13/bbl oil
  • In addition, feedstock costs are a very low fraction of final costs compared to other commodity products
  • Pretreatment is the most costly process step: the only process step more expensive than pretreatment is no pretreatment
    • 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

Biomass Refining CAFI

biomass refining consortium for applied fundamentals and innovation cafi
Biomass Refining Consortium for Applied Fundamentals and Innovation (CAFI)
  • Biomass Refining Consortium for Applied Fundamentals and Innovation (CAFI) organized in late 1999 and early 2000
  • Included top researchers in biomass hydrolysis from Auburn, Dartmouth, Michigan State, Purdue, NREL, Texas A&M, U. British Columbia, U. Sherbrooke
  • Mission:
    • Develop information and a fundamental understanding of biomass hydrolysis that will facilitate commercialization,
    • Accelerate the development of next generation technologies that dramatically reduce the cost of sugars from cellulosic biomass
    • Train future engineers, scientists, and managers.

Biomass Refining CAFI

slide6

DOE OBP Project: CAFI II

  • Started in April 2004 after completion of USDA IFAFS funded CAFI I on corn stover
  • Funded by DOE Office of the Biomass Program for $1.88 million through a joint competitive solicitation with USDA
  • Using identical analytical methods and feedstock sources to develop comparative data for corn stover and poplar
  • Determining in depth information on
    • Enzymatic hydrolysis of cellulose and hemicellulose in solids
    • Conditioning and fermentation of hydrolyzate liquids
    • Predictive models
  • Evaluating AFEX, ARP, controlled pH, dilute acid, lime, sulfur dioxide pretreatments
  • Genencor supplies commercial and advanced enzymes

Biomass Refining CAFI

slide7

DOE OBP Project: CAFI II

  • Started in April 2004 after completion of USDA IFAFS funded CAFI I on corn stover
  • Funded by DOE Office of the Biomass Program for $1.88 million through a joint competitive solicitation with USDA
  • Using identical analytical methods and feedstock sources to develop comparative data for corn stover and poplar
  • Determining in depth information on
    • Enzymatic hydrolysis of cellulose and hemicellulose in solids
    • Conditioning and fermentation of hydrolyzate liquids
    • Predictive models
  • Evaluating AFEX, ARP, controlled pH, dilute acid, lime, sulfur dioxide pretreatments
  • Genencor supplies commercial and advanced enzymes

Biomass Refining CAFI

enzymes used and experimental conditions
Enzymes Used and Experimental Conditions
  • Enzymes from Genencor International
  • Spezyme CP (59 FPU/ml, 123 mg protein/ml)
  • GC-220 (89 FPU/ml, 184 mg protein/ml)
  • β-glucosidase (32 mg protein/ml)
  • Multifect Xylanase (41 mg protein/ml)
  • Experimental conditions
  • 1-2% glucan in 100 ml flasks in citrate buffer plus antibiotics
  • Digestion time – 72 hrs
  • CBU : FPU ~ 2.0
  • Enzyme loadings – 3.0, 7.5,15,50, and 60 FPU/g

glucan prior to pretreatment

Biomass Refining CAFI

cafi corn stover
CAFI Corn Stover
  • Second pass harvested corn stover from Kramer farm (Wray, CO)
    • Collected using high rake setting to avoid soil pick-up
    • No washing
    • Milled to pass ¼ inch round screen
  • Performed like CAFI I material

for all

pretreatments

Biomass Refining CAFI

overall yields for corn stover at 15 fpu g glucan from cafi i

Increasing pH

Overall Yields for Corn Stover at 15 FPU/g Glucan from CAFI I

*Cumulative soluble sugars as total/monomers. Single number = just monomers.

Biomass Refining CAFI

slide11

CAFI II Standard Poplar

Biomass Refining CAFI

slide13

CAFI II Initial Poplar

  • Feedstock: USDA-supplied hybrid poplar (Arlington, WI)
    • Debarked, chipped, and milled to pass ¼ inch round screen
    • Not enough to meet needs

Biomass Refining CAFI

slide14

CAFI II Initial Poplar

  • Feedstock: USDA-supplied hybrid poplar (Arlington, WI)
    • Debarked, chipped, and milled to pass ¼ inch round screen
    • Not enough to meet needs

Biomass Refining CAFI

slide15

C - Cellulase

(31.3 mg/g glucan)

X - Xylanase

(3.1 mg/g glucan)

A - Additive

(0.35g/g glucan)

UT - Untreated

AFEX condition

24 h water soaked

1:1 (Poplar:NH3)

10 min. res. time

AFEX Optimization for High/Low Lignin Poplar

slide16

SO2 Overall Yields at 15 FPU/g of Glucan (148 hours hydrolysis)

*Cumulative soluble sugars as total/monomers. Single number = just monomers.

Biomass Refining CAFI

why is variability important
Why Is Variability Important?
  • Impacts yields for all but SO2 pretreatment
  • Assumption is that all hardwoods of same type behave similarly
  • Woody crops proponents plan to use standing forest to “store” wood, but harvest season may affect yields
  • Important to understand what causes this and whether it impacts other feedstocks such as herbaceous crops or agricultural residues

Biomass Refining CAFI

differences among poplar species
Differences Among Poplar Species*

* Based on information provided by Adam Wiese, USDA Rheinlander, WI

Biomass Refining CAFI

slide19
Getting to the Root Cause of Factors That Impact Digestion of Different Substrates with Different Pretreatments
  • Degree of polymerization – DP
  • Available reducing ends
  • Βeta-glucosidic bond accessibility
  • Crystallinity index
  • Other?

Biomass Refining CAFI

slide20
Getting to the Root Cause of Factors That Impact Digestion of Different Substrates with Different Pretreatments
  • Degree of polymerization – DP
  • Available reducing ends
  • Βeta-glucosidic bond accessibility
  • Crystallinity index
  • Other?

Initial data!

Biomass Refining CAFI

measuring dp for pretreated biomass
Measuring DP for Pretreated Biomass
  • Difficult to directly measure DP for complex substrates
  • Delignification improves measurements but could alter cellulose DP, leading to wrong conclusions
  • ASTM capillary viscometer method modified to estimate DPv for cellulose by correcting for hemicellulose*:

(DPv) = {(1.65 [η] – 116 y)/x}1.111

η = intrinsic viscosity

y = hemicellulose fraction

x = glucan fraction

  • However, no information is available on DP of corn stover for comparison

*Heiningen et al.(2004) Journal of Pulp and Paper Science 30(8):211-21

Biomass Refining CAFI

estimation of reducing ends for corn stover
Estimation of Reducing Ends for Corn Stover
  • Copper number is often used to reflect number of reducing ends for cellulose in pulp
  • Applied Braidy’s method

Copper Number = (0.064*t) /w for which

t = ceric ammonium sulfate solution (ml) consumed

w = dry weight of wood/biomass

  • In general higher DP = lower copper number and vice versa

Roy L. Whistler et al.(1963). Methods in carbohydrate chemistry-III.

Holtzapple, M. T. (1981) Ph.D Thesis, Deptt. of Chemical Engineering. University of Penn.

Biomass Refining CAFI

estimating glucosidic bond accessibility
Estimating β-Glucosidic Bond Accessibility
  • Measured protein adsorption:
    • At 40oC in 50 ml tubes with 0.5- 1 % glucan content
    • With SP-CP protein from 40mg to 800 mg/g glucan
  • Nitrogen content was determined after decanting unabsorbed protein and drying the substrate
  • Converted into mg protein adsorbed/gm cellulose substrate using established nitrogen factor
  • Then determined Langmuir isotherm parameters using nonlinear regression in Polymath per Lynd, et al.(2002) Microbiol. and Mol. Biol. Reviews. 66:506-577

Biomass Refining CAFI

initial digestion data for 7 5 fpu g enzyme loading vs substrate features
Initial Digestion Data for 7.5 FPU/g Enzyme Loading vs Substrate Features

Biomass Refining CAFI

closing thoughts
Closing Thoughts
  • The resistance of one biological system (cellulosic biomass) to the other (biological conversion) requires a pretreatment interface
  • All of the pretreatments studied were very effective in enhancing cellulose digestion for corn stover
  • Poplar wood presented a more challenging substrate for several pretreatments, particularly those at higher pH
  • The pretreatabilty of poplar changed with source
  • The cause of variations in poplar performance among pretreatments and biomass sources must be understood to identify paths to 1) better pretreatments and 2) improved biomass production

Biomass Refining CAFI

acknowledgments
Acknowledgments
  • US Department of Agriculture Initiative for Future Agricultural and Food Systems Program, Contract 00-52104-9663
  • US Department of Energy Office of the Biomass Program, Contract DE-FG36-04GO14017
  • Natural Resources Canada
  • Numerous collaborators including the CAFI Team members, students, and others who have been so cooperative
  • Ford Motor Company and University of California for sponsoring this trip

Biomass Refining CAFI

slide28

Questions???

Biomass Refining CAFI