1 Auburn UniversityBiomass Refining CAFI Corn stover Wood chip Bagasse Rice straw Sawdust Biomass Ethanol Fuel

1

Auburn UniversityBiomass Refining CAFI

Corn stover

Corn stover

Wood chip

Bagasse

Rice straw

Sawdust

Biomass

Ethanol F

uel

2


0

5

10

15

20

25

30

35

0 5 10 15 20 25 30

% Xylan

% L

ign

in

Comparison among various feedstock

Lose lot of sugarsIn liquid stream/Low pretreatment yield

(considering solid only)

Very hard to treatOptimum

Corn StoverLow Lignin Hybrid PoplarHigh Lignin Hybrid Poplar

(Courtesy: US-DOE website)http://www.eere.energy.gov/biomass/progs/search1.cgi

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Substrate Dependency on the Effect of Pretreatment by Aqueous Ammonia

Rajesh Gupta, Tae Hyun Kim and Y. Y. LeeDepartment of Chemical Engineering

Auburn University

4


Outline

• Difference in physical & chemical features of biomass responsible for different results in pretreatment.

• Effect of xylanase supplementation on ARP treated biomass.

• How the cellulosic part in biomass affected during different pretreatment.

5



Low lignin Poplar seems the best choice among three feed stocks.

Composition of Untreated Biomass

36%

21%17%

21%

14%

29%

18%

45% 44%

0%

10%

20%

30%

40%

50%

Glucan Xylan Lignin

Component

co

mp

os

itio

n

Corn Stover

Low Lignin Poplar

High Lignin Poplar

6


Corn Stover

Low Lignin PoplarHigh Lignin Poplar


7


Processes with Ammonia

SAA(Soaking in Aqueous Ammonia)

• Batch Reactor

• Low Temperature

• Long Pretreatment time (several hours)

ARP(Ammonia Recycle Percolation)

• Flowthrough Reactor

• High Temperature

• Short Pretreatment time (several min.)

8


Effect of SAA treatment

Treatment Conditions: 15% Ammonia, 1:6 S:L & 12 hrs soaking time

Temperature: 60'C

0

20

40

60

80

100

0 20 40 60 80

Time(hrs)

% G

luc

an

Dig

es

tib

ilit

y

Corn Stover

Enzyme Loading: Cellulase (15FPU/g glucan) + Cellobiase (30CBU/g glucan)

9


Temperature: 120°C

0%

20%

40%

60%

80%

100%

0 20 40 60 80

Time(hrs)

Glu

can

Dig

esti

bil

ity

High Lignin Poplar

Low Lignin Poplar

Temperature: 150°C

0%

20%

40%

60%

80%

100%

0 20 40 60 80

Time(hrs)

Glu

can

Dig

esti

bil

ity

High Lignin Poplar

Low Lignin Poplar


Treatment Conditions: 15% Ammonia, 1:10 S:L & 24hrs soaking time

Effective treatment temperature for SAA is much higher for Hybrid poplar

than Corn stover.


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In comparison to Corn stover, higher delignification / xylan removal is required in case

of High lignin Poplar for attaining the good digestibility.

Corn Stover

21

181717

9

6

0

5

10

15

20

25

Untreated Case-1 Case-2

% X

yla

n/L

ign

in

Xylan

Lignin

High Lignin Poplar

1413

11

29

18

15

0

5

10

15

20

25

30

35


% X

ylan

/Lig

nin

Xylan

Lignin

~15-20% Xylan removal~50% Delignification

Corn Stover72hrs Glucan Digstibility

17

80

90

0

20

40

60

80

100


% D

iges

tibili

ty

High Lignin Poplar72hrs Glucan Digestibility

4

29 31

0

20

40

60

80

100


% D

iges

tibili

ty


11


ARP (Ammonia Recycle Percolation) Treatment

Optimum chosen treatment condition:

Corn Stover:

Temperature : 170°C

Reaction time : 20min.

Ammonia Conc.: 15%

Liquid : Solid : 3.33 : 1

Hybrid Poplar:

Temperature : 185°C

Reaction time : 27.5min.

Ammonia Conc.: 15%

Liquid : Solid : 3.667 : 1

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Effect of ARP Treatment

Xylan Removal

21%

18%

14%

10%

14%

10%

0%

5%

10%

15%

20%

25%

Corn Stover Low Lignin Poplar High Lignin Poplar

Feed stock

Treated Biomass

Lignin Removal

17%

21%

29%

5%

15%

18%

0%

5%

10%

15%

20%

25%

30%

35%


Feed stock

Treated Biomass

72hrs Glucan Digestibility

63%

88%90%

0%

20%

40%

60%

80%

100%


(15FPU + 30 CBU)

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Effect of Reaction time on Hybrid Poplar (High Lignin Poplar) during ARP treatment

Component Removal

5%

10%

15%

20%

0 20 40 60 80 100

Reaction time(min)

Xyl

an/L

ign

in R

emai

nin

g

Lignin Remaining

Xylan Remaining

Digestibility (15FPU+30CBU)

20%

25%

30%

35%

40%

45%

50%

55%

0 20 40 60 80 100

Reaction time(min)

Glu

can

/Xyl

an

Dig

esti

bil

ity

Glucan Digestibility

Xylan Digestibility

XRD Plots

0

500

1000

1500

2000

2500

3000

3500

4000

10 15 20 25 30 35 402 θ

Inte

nsi

ty(a

.u.)

Untreated(CrI: 64.41)

27.5min (CrI: 75.45)

82.5min (CrI: 76.32)

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Effect of Ammonia flow on Hybrid Poplar (High Lignin Poplar) during ARP treatment

Digestibility seems more sensitive towards xylan removal than delignification.

Component Removal

5%

10%

15%

20%

0 2 4 6 8

Flow rate(ml/min)

Xyl

an/L

ign

in

Rem

ain

ing

Lignin Remaining

Xylan Remaining

Digestibility( 15FPU+30CBU)

20%

25%

30%

35%

40%

45%

50%

55%

0 2 4 6 8

Flow rate(ml/min)

Glu

can

/Xyl

an

Dig

esti

bil

ity


Xylan Digestibility

XRD Plots

0

500

1000

1500

2000

2500

3000

3500

10 15 20 25 30 35 40

2 θ

Inte

ns

ity

(a.u

.)


2ml/min (CrI: 75.45)

7.5ml/min (CrI: 70.43)

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Effect of Temperature on Hybrid Poplar (High Lignin Poplar) during ARP treatment

Component Removal

5%

10%

15%

20%

165 170 175 180 185 190 195 200

Temperature('C)

Xyl

an/L

ign

in

Rem

ain

ing

Lignin Remaining

Xylan Remaining

Digestibility( 15FPU+30CBU)

20%

25%

30%

35%

40%

45%

50%

55%

165 170 175 180 185 190 195 200

Temperature('C)

Glu

can

/Xyl

an

Dig

esti

bil

ity


Xylan Digestibility

XRD Plots

0

500

1000

1500

2000

2500

3000

3500

10 15 20 25 30 35 402 θ

Inte

nsi

ty(a

.u.)


175°C (CrI: 75.25%)

195°C(CrI: 73.70%)

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Effect of Temperature on Hybrid Poplar (Low Lignin Poplar) during ARP treatment

Component Removal

18%

16%

14%

21%

16%15%

0%

5%

10%

15%

20%

25%

Untreated 170'C 185'C

Xylan Remaining

Lignin Remaining


Increase of temperature from 170°C to 185°C does not affect xylan/lignin removal, but increases cellulase accessibility to cellulosic part in biomass

Digestibility

0%

74%

87%

0%

69%73%

0%

10%

20%

30%

40%

50%

60%

70%

80%

90%

100%

Untreated 170'C 185'C


72hrs Xylan Digestibility

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Effect of xylanase supplementation onARP treated Corn Stover

Enzyme Loading:C+β-G : Cellulase (15FPU/g glucan) + Cellobiase (30CBU/g glucan)C+ β-G +X : Cellulase (15FPU/g glucan) + Cellobiase (30CBU/g glucan) + Xylanase (31.3mg protein/g glucan)


0%

20%

40%

60%

80%

100%

0 10 20 30 40 50 60 70 80

Time(hr)

% G

luca

n E

quiv

alen

t

C+β-G C+β-G+X

Xylan Digestibility

0%

20%

40%

60%

80%

100%

0 10 20 30 40 50 60 70 80

Time(hr)%

Xyl

an E

qu

ival

ent

C+β-G C+β-G+X

Xylan DigestibilityGlucan Digestibility

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Effect of xylanase supplementation on Glucan Digestibilityof Hybrid Poplar

0%

20%

40%

60%

80%

100%

0 20 40 60 80

Time(hr)

% G

luca

n E

qu

ival

ent

C+β-G C+β-G+X

0%

20%

40%

60%

80%

100%

0 20 40 60 80

Time(hr)%

Glu

ca

n E

qu

iva

len

t

C+β-G C+β-G+X

High Lignin Poplar Low Lignin Poplar


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0%

20%

40%

60%

80%

100%

0 10 20 30 40 50 60 70 80

Time(hr)

% X

ylan

Equ

ival

ent

C+β-G C+β-G+X

0%

20%

40%

60%

80%

100%

0 10 20 30 40 50 60 70 80

Time(hr)

% X

ylan

Eq

uiv

alen

t

C+β-G C+β-G+X

High Lignin Poplar Low Lignin Poplar

Effect of xylanase supplementation on Xylan Digestibilityof Hybrid Poplar

Near 100% Glucan & xylan digestibility can be achieved for ARP treated Low lignin Poplar with xylanase supplementation.

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Effect of xylanase loading


0%

20%

40%

60%

80%

100%

0 10 20 30 40

Xylanase Loading(mg/g glucan)

Dige

stib

ility Low Lignin Poplar

High Lignin Poplar

72hrs Xylan Digestibility

0%

20%

40%

60%

80%

100%

0 10 20 30 40

Xylanase Loading(mg/g glucan)Di

gest

ibili

ty Low Lignin Poplar

High Lignin Poplar

Cellulase loading: 15FPU/g glucanβ-G loading : 30CBU/g glucan

In ARP treated Low lignin Poplar, there is some easily digestible xylan.

In High lignin Poplar, minimum amount of xylan removal is required for betteraccessibility of cellulase.

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Basis: 100g dry biomass

Enzyme LoadingWithout xylanase : Cellulase (15FPU/g glucan) + Cellobiase (30CBU/g glucan)With xylanase : Cellulase (15FPU/g glucan) + Cellobiase (30CBU/g glucan) + Xylanase (31.3mg protein/g glucan)

Yield from different ARP treated Biomass

* Processing of liquid stream is eliminated.* High fermentation efficiency due to absence of inhibitor/toxins

* There is a need to increase the xylanase activity in cellulase* Low pretreatment severity/Low pretreatment cost

GlucanXylan

Glucan Xylan

Sugar YieldWithout Xylanase With Xylanase Without Xylanase With Xylanase Without Xylanase With Xylanase

Glucan 31.38 32.88 29.42 43.23 19.96 28.45Xylan 7.51 8.89 8.99 12.90 4.91 7.05

Sugar Yield 84.72% 91.00% 66.24% 96.77% 49.30% 70.35%38.89 41.77 38.42 56.13 24.87 35.49

67.63% 72.64% 61.21% 89.43% 42.96% 61.31%


79.83% 92.42% 87.15%

Feed Stock36.1 44.91 43.821.4 17.85 14.09

After ARP treatment

35.6 44.5 40.8410.3 13.5 9.61

Overall Sugar YieldTotal Sugars(g)/100g biomass

After Enzymatic Hydrolysis

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Solid remaining after treatment

100% 99%92%

0%

20%

40%

60%

80%

100%

120%

Untreated Cotton ARP treated Cotton Sulfuric acid treatedCotton

Treatment Conditions :

ARP treatment: 15% NH3, 175°C & 20min Rx. Time (Flow through)

Acid treatment: 0.05% H2SO4, 175°C & 30min Rx Time (Batch)

How Cellulose is Affected in Pretreatment

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Enzyme loading: 15FPU Cellulase + 30CBU β-G


0%

20%

40%

60%

80%

0 25 50 75 100Time(hr)

% G

luc

an

Eq

uiv

ale

nt

Untreated Cotton

ARP treated Cotton

Sulfuric acid treatedCotton

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Absorbance with DNS reagent

0.06750.0165

0.857

0

0.2

0.4

0.6

0.8

1

Untreated Cotton ARP treated Cotton Sulfuric acid treated Cotton


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0

500

1000

1500

2000

2500

10 15 20 25 30 35 40

2θ

Inte

sit

yHow Cellulose is Affected in Pretreatment

Untreated Cotton(77.1)

ARP treated Cotton(83.4)

Acid treated Cotton(81.35)

XRD Plots

Lower Crystallinity More Endo glucanase action

Higher Digestibility

(a.u

.)

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• Any pretreatment reagent( Ammonia/Acid) first attacks the amorphous part of cellulose & increase the crystallinity.

• Ammonia, being a weak base, cannot hydrolyse the crystalline part of cellulose and removes very little cellulose.

• H2SO4, being a strong acid, hydrolyze the crystalline cellulose as well and that’s why more cellulose is lost into liquid.

AmmoniaDoes not affect Cellulosic part

in biomass

Better sugar yieldin pretreatment

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Summary

• Lignin/xylan content plays an important role in pretreatment of lignocellulosic biomass.

• Without xylanase supplementation, the overall sugar yield with ARP treatment is seen in the order: Corn Stover > Low-lignin poplar > High-lignin poplar (~68%) (~61%) (~43%)

• With xylanase supplementation, the overall sugar yield with ARP treatment is seen in the order:Low-lignin poplar> Corn Stover> High-lignin poplar

(~90%) (~73%) (~62%)

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Summary

• Without xylanase supplementation, a certain degree of xylan/lignin removal is necessary in pretreatment to achieve acceptable level of digestibility.

• With xylanase supplementation, high digestibility and overall yield can be attained without removing much xylan/lignin during the ARP treatment.

• Higher xylanase activity in cellulase will improve the overall yield and economics of cellulosic ethanol process drastically.

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Future Work

• To compare the economics of following two bio-ethanol production processes using ARP pretreatment:* Severe pretreatment conditions/Processing of

liquid stream/No xylanase supplementation * Mild pretreatment conditions/No sugars in liquid

stream/ xylanase supplementation• To optimize the ARP pretreatment conditions for

different feedstocks with xylanase supplementation.• To characterize the changes in biomass during ARP

pretreatment using FTIR.

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Acknowledgements US Department of Energy Office of the Biomass

Program, Contract DE-FG36-04GO14017

CAFI Team:

* University of California, Riverside* Michigan State* Purdue* Texas A&M* University of British Columbia* National Renewable Energy Laboratory

Genencor International

Documents

1 Auburn UniversityBiomass Refining CAFI Corn stover Wood chip Bagasse Rice straw Sawdust Biomass Ethanol Fuel