1. Biomass Refining CAFI Overall Sugar Yields from Corn Stover via Thermochemical Hemicellulose Hydrolysis Followed by Enzymatic Hydrolysis Todd A. Lloyd and Charles E. Wyman Thayer School of Engineering Dartmouth College Hanover, New Hampshire 03755 AIChE Annual Meeting San Francisco, CA November 20, 2003 Biomass Refining CAFI

2. Objectives Determine pretreatment conditions that lead to highest overall sugar yields Develop comparitive information on a consistent basis Identify opportunities to lower production costs while maintaining high yields

3. Biomass Refining CAFI Corn Stover Composition NREL supplied corn stover to all project participants (source: BioMass AgriProducts, Harlan IA) Stover washed and dried in small commercial operation, knife milled to pass ¼ inch screen Glucan36.1 % Xylan21.4 % Arabinan3.5 % Mannan1.8 % Galactan2.5 % Lignin17.2 % Protein4.0 % Acetyl3.2 % Ash7.1 % Uronic Acid3.6 % Non-structural Sugars1.2 % Biomass Refining CAFI

4. Hydrolysis Enzyme Batch Pretreatment H 2 O or H 2 SO 4(aq) Corn Stover Overall Flow Diagram for Dilute Acid Pretreatment Fermentation Ethanol Pretreatment Sugars Residue Digestion Sugars Stage 1Stage 2

5. Biomass Refining CAFI Stage 1 – Summary of Pretreatment Conditions Steam Gun-no added acid –150-220 o C –~25% solids Dilute Acid –140-200 o C –0.22-1.0 % H 2 SO 4 –5-25% solids –Heated in sand bath Biomass Refining CAFI Batch Reaction Systems 1.NREL steam gun 2.½”o.d. batch tubes 3.1l stirred autoclave

6. Biomass Refining CAFI NREL Steam Gun Biomass Refining CAFI

7. Batch Tube Experimental Apparatus for Dilute Sulfuric Acid 4” Biomass Refining CAFI

8. Parr Reactor Experimental Apparatus for Dilute Sulfuric Acid Biomass Refining CAFI

9. Stage 2 – Enzymatic Digestion Spezyme from Genencor - used by all investigators NREL LAP-009 used to evaluate digestibility –60 FPU/g original glucan used to determine ultimate digestibility –15 FPU/g tests done on selected samples

10. Biomass Refining CAFI Severity Parameter For no acid addition applied approach used by Overend and Chornet CS = t*A n exp[(T-100)/14.75] CS = Combined Severity A = Added Acid, % n = Arbitrary exponent T = Reaction Temperature, o C Biomass Refining CAFI Modified equation including acid addition has the form: R o = t*exp[(T-100)/14.75] Provides useful tools to compare results from a broad range of conditions

11. Biomass Refining CAFI Combined Stage 1 and 2 Steam Gun Results - No Acid Biomass Refining CAFI

12. Stage 1 Pretreatment Yield for 0.49% H 2 SO 4 Addition in Batch Tubes Biomass Refining CAFI

13. Stage 2 Digestion Yield for 0.49% H 2 SO 4 Addition in Batch Tubes

14. Biomass Refining CAFI Combined Stage 1 and Stage 2 Yield for 0.49% H 2 SO 4 Addition in Batch Tubes

15. Biomass Refining CAFI Combined Stage 1 and Stage 2 Yield for 0.98% H 2 SO 4 Addition in Batch Tubes

16. Biomass Refining CAFI 84.2% glucan to glucose + 54.5% xylan to xylose conversion at 15FPU/g glucan 89.7% mass balance closure ( all solids + G + GO + X + XO) 92% theoretical ethanol yield from glucose + xylose Hydrolysis Enzyme @ 15 FPU/g of glucan, 48h Residual Solids Hydrolyzate Liquid Parr Reactor Pretreatment 1 wt% H 2 SO 4 Corn Stover 100 lb (dry basis) 21.4 lb xylan 0.2 lb gluco-oligomer (GO) 21.8lb xylose (X) 64.0 lb 36.1 lb glucan Fermentation 4.08 gal Ethanol 1.3 lb xylose (X) 30.9 lb glucose (G) 0.1 lb xylo-oligomer (XO) 3.0 lb glucose (G) 32.4 lb Treated Solids 2.1 lb xylan 33 lb glucan 2.29 gal ethanol 1.79 gal ethanol 140 o C Mass Balance for Dilute Acid Pretreatment with Digestion (15 FPU/ g of glucan) in Parr Reactor

17. Biomass Refining CAFI Summary of Batch Tube Dilute Acid Performance at 15 FPU/g Glucan Biomass Refining CAFI T, o CA, %t, minX Yield, %G Yield, % 1400.9840 Stage 1 - 85 Stage 2 - 8 Total - 93 Stage 1 - 8 Stage 2 - 82 Total - 90 1600.4920 Stage 1 - 85 Stage 2 - 9 Total - 94 Stage 1 - 6 Stage 2 - 85 Total - 91

18. Biomass Refining CAFI Understanding Role of Dilute Acid and Oligomer Release Why does acid enhance performance? What is the role of oligomers in sugar release? Biomass Refining CAFI

19. Understanding Role of Dilute Acid and Oligomer Release Why does acid enhance performance? What is the role of oligomers in sugar release? Biomass Refining CAFI

20. Predicting pH at Elevated Temperature (1) (2) (3) Biomass Refining CAFI (4) (Amount of Acid Neutralized) Assume reaction (1) goes to completion (Amount of Added Acid)

21. Biomass Refining CAFI *Marshall and Jones Predicting pH at Elevated Temperature

22. Biomass Refining CAFI Predicting pH at Elevated Temperature Similar to approach used by Bienkowski et al. to predict the sulfuric acid degradation of glucose at elevated temperature and of Springer and Harris for the hydrolysis of wood

23. Biomass Refining CAFI Predicted pH vs. Temperature Biomass Refining CAFI

24. Predicted pH vs. Temperature Biomass Refining CAFI

25. Predicted pH vs. Temperature Biomass Refining CAFI

26. Predicted pH vs. Temperature Biomass Refining CAFI

27. Understanding Role of Dilute Acid and Oligomer Release Why does acid enhance performance? What is the role of oligomers in sugar release? Biomass Refining CAFI

28. Typical Stage 1 Yield vs Time Data

29. Biomass Refining CAFI Models to Predict Hemicellulose Hydrolysis Abundance of literature data Most kinetic studies neglect oligomers Saeman model: H M D khkh kdkd HfHf M D HsHs kfkf ksks kdkd Biomass Refining CAFI Have also added consideration of two fractions of hemicellulose in biphasic reaction (Kobayashi et al.)

30. Biomass Refining CAFI Hemicellulose Hydrolysis Kinetics A few models in literature include oligomers These treat oligomers as only one or possibly two species In fact, we would expect a number of different species of oligomers with varying chain lengths Some inconsistencies among models HfHf O M HsHs kfkf ksks khkh D kdkd Biomass Refining CAFI

31. Hemicellulose Hydrolysis Kinetics A few models in literature include more than one species of oligomer and suggest that oligomers may react directly to degradation products without first forming xylose monomer Biomass Refining CAFI XO H M HsHs kfkf ksks khkh D kdkd XO L k Od Garrote et. al, 2000, Process Biochemistry, 36/571-78

32. Biomass Refining CAFI Structure of Hemicellulose p-Coumaric Acid Xylose Acetyl Arabinose Ferulic Acid Glucuronic Acid Fragmentation Products + H 2 O Biomass Refining CAFI

33. Polymer Degradation 1 N-mer dC N /dt = -k h (N-1)C N C N = C N0 exp[-k h (N-1) t] Assumes random bond scission and uniform rate constant 1 Simha, R., (1941), Journal of Applied Physics, 12:569-578 Biomass Refining CAFI

34. Generalized Form for j-mer Distribution dC j /dt = 2k h  C i - k h (j-1)C j i=j+1 N C j = C N0 [1 –  ] (j-1)  [ 2 + (N-j-1)  ] 1< j < N-1 Where  = 1 - exp[ -k h t ] @ t=0, C N =C N0 ; C j =0 Biomass Refining CAFI For reactive monomer:

35. Biomass Refining CAFI Depolymerization Model 140 o C, 0.49% Added Acid

36. Biomass Refining CAFI Modified Depolymerization Model Assumes a change in activation energy with conversion

37. Biomass Refining CAFI Modified Depolymerization Model 140 o C, 0.49% Added Acid Biomass Refining CAFI

38. Conclusions Water only hydrolysis with a steam gun produced a maximum yield of about 60% X+G with 60 FPU/g glucan. For dilute acid pretreatment conditions for >95% yield of glucan and xylan ranged from 140 o C with 0.98% added acid for 40 minutes to 180 o C with 0.49% added acid for 5 minutes Xylanase activity in stage 2 enhances xylose yields Neutralization of added acid can have a significant effect on pH Hydrolysis can be viewed as a depolymerization process Biomass Refining CAFI

39. Acknowledgements The United States Department of Agriculture Initiative for Future Agricultural and Food Systems Program through Contract 00-52104-9663 for funding our research The United States Department of Energy Office of the Biomass Program and the National Renewable Energy Laboratory Our partners from Auburn University, Michigan State, Purdue, and Texas A&M Universities and the National Renewable Energy Laboratory The National Institute of Standards and Technology for funds to purchase some of the equipment used in this research The Thayer School of Engineering at Dartmouth College Biomass Refining CAFI

40. Questions? Biomass Refining CAFI