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1‐2: Characterization of Plant Cell Wall P ti C t ib ti t I d RProperties Contributing to Improved Reponses
to Alkaline Pretreatment and Hydrolysis
Feedstocks I – Genetics and Recalcitrance
27 April, 2015
37th SIM SBFCJacob Crowe Muyang Li Daniel WilliamsJacob Crowe, Muyang Li, Daniel Williams,
Guilong Yan, and David Hodge
www.glbrc.org
Cell Wall Diversity
Monocots Woody
Dicots
Herbaceous
DiversityImportant pimplications for cell wall deconstruction
SorghumSorghum((Sorghum bicolorSorghum bicolor))((Sorghum bicolorSorghum bicolor))
Corn/Maize Corn/Maize ((ZeaZea maysmays subspsubsp. . maysmays))
Switchgrass(Panicum virgatum)
Arabidopsis thaliana
Black Cottonwood (Populus trichocarpa)
Cell Wall Diversity
Monocots Woody
Dicots
Herbaceous
DiversityImportant G lp
implications for cell wall
Goals• Understanding interaction between
d l ll ll ideconstructionpretreatments and plant cell wall properties
SorghumSorghum((Sorghum bicolorSorghum bicolor))
• Identifying cell wall phenotypes with improved responses to pretreatment + enzymatic hydrolysis
((Sorghum bicolorSorghum bicolor))
Corn/Maize Corn/Maize ((ZeaZea maysmays subspsubsp. . maysmays))
Switchgrass(Panicum virgatum)
Arabidopsis thaliana
Black Cottonwood (Populus trichocarpa)
Cell Wall Properties Impacting Pretreatability, Ruminant Digestibility, and Cellulolytic Enzyme Hydrolyzability
Structural DifferencesGenotype Phenotype
– Relative abundance of cell types?
• Epidermisl hEnvironmental/agronomic • Sclerenchyma
• Vascular bundle zone cells• Pith parenchyma
C ll ll thi k
Environmental/agronomic– Harvest time/maturity
– N, water, environment,…– Cell wall thickness, composition, accessibility,…
Composition80
90
100
Unassigned
Ash
– Lignin, structural polysaccharides
pCA and FA30
40
50
60
70
Compo
sition
(wt %)
Ash
Water+EtOH ExtractivesAcetate
Lignin
Uronic Acids
Galactan
Mannan
Arabinan
MaizeSwitchgrass
– pCA and FA
– Acetyl 0
10
20
Corn stover Switchgrass
Arabinan
Xylan
Glucan
(Pioneer hybrid 36H56) (cv. Cave-in-Rock)
Goals
• Understanding impacts of lpretreatments on plant
cell wall properties
• Identifying phenotypes y g p ypthat respond well to cellulolytic enzymes or y ypretreatment/hydrolysis
Li et al., (2015). J Exp Bot. 66(14):4305‐4315
Overview of Work
1. Response of diverse cell types in sorghum to alkaline pretreatmentsorghum to alkaline pretreatment
2. Impact of cell wall properties in diverse poplar on alkaline anddiverse poplar on alkaline and alkaline‐oxidative pretreatment
3 Impact of xylan o‐acetylation on3. Impact of xylan o acetylation on cell wall nanoscale porosity in Arabidopsisp
4. Impact of alkaline pretreatments on cell wall water sorption in pmaize and switchgrass
Overview of Work
1. Response of diverse cell types in sorghum to alkaline pretreatmentsorghum to alkaline pretreatment
2. Impact of cell wall properties in diverse poplar on alkaline and
Work performed by Dr. Muyang Li and Dr. diverse poplar on alkaline and
alkaline‐oxidative pretreatment3 Impact of xylan o‐acetylation on
In collaboration with John Mullet, TAMU
Guilong Yan
3. Impact of xylan o acetylation on cell wall nanoscale porosity in Arabidopsis
See poster: M28 – Tissue fractionation, extraction and characterization of energy p
4. Impact of alkaline pretreatments on cell wall water sorption in
sorghum and the development of a counter‐current extraction and alkaline pretreatment for high‐titer mixed sugar production (Tonight!)p
maize and switchgrass
Sorghum Physical Fractionation, Pretreatment, and HydrolysisPretreatment, and Hydrolysis
Energy Energy SorghumSorghum
Goals of this Study : • Comparison of within‐plant
diff i ll ll ti SorghumSorghum
Sweet Sweet SorghumSorghum
differences in cell wall properties in two sorghum cultivars
• Identify responses to alkaline d h d l i
1 10
Grain SorghumGrain Sorghum
Image : Bill Rooney TexasImage : Bill Rooney Texas A&MA&M
pretreatment and hydrolysis TX08001/ES5200• Photoperiod‐sensitive “energy
0 70
0.80
0.90
1.00
1.10
Cell Wall
Fructose
Glucose
Sucrose
Image : Bill Rooney, Texas Image : Bill Rooney, Texas A&MA&MPhotoperiod sensitive energy sorghum” hybrid
• Delayed flowering Extended vegetative growth
0.30
0.40
0.50
0.60
0.70
uble Sug
ar / g
Increased biomass yields
Della• Commercial sweet sorghum cultivar
0.00
0.10
0.20
0.30
TX08001 Della
g So
luCommercial sweet sorghum cultivar• Mid‐season variety, matures early• Stalk height 10–11 ft.
Energy Sorghum Stem Anatomy
Image: Tesfamichael Kebrom and John Mullet, Texas A&M
Pith Parenchyma: 30%Internal “Fiber” (Vascular Bundles): 15%Rind “Fiber”: 50%Epidermis: 5%
Industrial Physical Fractionation of Grass Stems
• Depithing performed prior to chemical pulping of non‐wood f d t k ( b )feedstocks (sugarcane bagasse)
• Industrial depithingB d h illi i– Based on hammermilling‐screening
– Integration with inorganics removal from the biomasso t e b o ass
– 1500 dry tonnes/day moist depithing of sugarcane bagasse
• Potential for integration intoPotential for integration into cellulosic biofuels technologies?
Image Source: FMW, GmbH
Physical Fractionation of Sorghum Stemsof Sorghum Stems
TX08001 or Della
mmee
Bottom
Rind
Rind
Rind
Rind
Rind
Rind
MiddleTop
Bottom
Bottom
Middle
Middle
Top
Top
Ep. + Outer R
VB + Inne
r R
Pith
Ep. + Outer R
VB + In
ner R
Pith
Ep. + Outer R
VB + In
ner R
Pith
Internode Cross‐Section Epidermis + Outer Rind Vascular Bundles + Inner Rind Pith
Confocal Microscopy of Sorghum Stem Fractions
Outer rind/EpidermisOuter rind/Epidermis100 μm
SclerenchymaSclerenchyma cellscells, , collenchymacollenchyma cells, vascular bundlescells, vascular bundlesCollenchymaCollenchyma cellscells
Inner rind / Vascular bundles Inner rind / Vascular bundles 100 μm
Vessel cellsVessel cells Fiber cellsFiber cells
PithPith100 μm
LowLow‐‐lignin lignin parenchymaparenchyma cellscells
UnseparatedUnseparated bundle bundle sheath (fiber cells)sheath (fiber cells)HighHigh‐‐lignin parenchyma cellslignin parenchyma cells
Comparison of Della vs. TX08001: Yields
• Substantial differences in hydrolysis yields
• General trend for recalcitrance: Hydrolysis:CTec3: 15 mg protein/g glucan
Ep+OR > VB+IR > PithCTec3: 15 mg protein/g glucanHTec3: 7.5 mg protein/g glucan
Bottom pith samplesBottom pith samples were most completely“defibered”
Hydrolysis OnlyHydrolysis Only
Epidermis + Outer Rind
Vascular Bundles + Inner Rind
Pith
Della
Epidermis + Outer Rind
Vascular Bundles + Inner Rind
Pith
TX08001
Comparison of Della vs. TX08001: Yields G l t d f l it• General trends for recalcitrance:
Ep+OR > VB+IR, Pith Pretreatment: 0.10 g/g NaOH; 80°C; 1 hr
Top > Middle > Bottom Hydrolysis:CTec3: 15 mg protein /g glucanHTec3: 7.5 mg protein /g glucan
Pretreatment + Hydrolysis Pretreatment + Hydrolysis
Hydrolysis Only Hydrolysis Only
Epidermis + Outer Rind
Vascular Bundles + Inner Rind
Pith
Della
Epidermis + Outer Rind
Vascular Bundles + Inner Rind
Pith
TX08001
Comparison of Della vs. TX08001: Properties
• Diverse range of properties
• Comparable tissues have comparable compositions and
90
responses to hydrolysis and pretreatment + hydrolysis
60
70
80
90
tion
Acetyl (mg/g)
Xylan (%)
Glucan (%)
Lignin (%)
100%
120%
s (D
ella ) Hydrolysis only
Pretreatment + Hydrolysis
30
40
50
60
la Com
posit Lignin (%)
40%
60%
80%
rolysis Y
ield
0
10
20
30Del
0%
20%
40%
72‐hr H
ydr
00 20 40 60 80
TX08001 Composition
0% 20% 40% 60% 80% 100% 120%
72‐hr Hydrolysis Yields (TX08001)
Cell Wall Properties Contributing to Differences in Sorghum Stem Internode Fractionse e ces So g u Ste te ode act o s• Most properties correlated to each other, e.g.:
Xylan Acetyl, Xylan 1/LigninXylan Acetyl, Xylan 1/Lignin
• Lignin strongest single property predictor of hydrolysis yields in untreated and pretreated sorghum fractions
1.2R = ‐0.596p = 0.0090
yields in untreated and pretreated sorghum fractions
30 Ac:Xyl = 0.764 mol/mol
TX08001: Solid data pointsDella: Open data points
0 6
0.8
1
olysis Yields
R = 0 85220
25
ylan
(%)
Top Pith
0.2
0.4
0.6
72‐hr H
ydro
Hydrolysis onlyPretreatment
R = ‐0.852p = 7.3 x 10‐6
5
10
15Ce
ll Wall Xy
Ac:Xyl = 0.543 mol/mol
p
00 5 10 15 20 25 30
7
Cell Wall Lignin Content (%)
Pretreatment + Hydrolysis
0
5
0 2 4 6 8 10
Cell Wall Acetyl Content (%)
Overview of Work
1. Response of diverse cell types in sorghum to alkaline pretreatmentsorghum to alkaline pretreatment
2. Impact of cell wall properties in diverse poplar on alkaline anddiverse poplar on alkaline and alkaline‐oxidative pretreatment
3 Impact of xylan o‐acetylation on3. Impact of xylan o acetylation on cell wall nanoscale porosity in ArabidopsisIn collaboration with Wellington
Work performed by Dr. AdityaBhalla
p4. Impact of alkaline pretreatments
on cell wall water sorption in
Muchero and Gerry Tuskan, ORNL
pmaize and switchgrass
Cell Wall Property Diversity in Populus trichocarpa
23
25
27
ent (%)
in Populus trichocarpa• Wild‐type P. trichocarpa genotypes
isolated from geographically and
17
19
21
ignin Co
nteisolated from geographically and
environmentally diverse sites – Exhibit wide phenotypic diversity
25
30
m) Cu
150 1 2 3 4 5
Li
Lignin S:G Ratio• Subset selected for diversity in
S:G ratio and lignin content
15
20
nten
t (pp
m CuFeMn
• Substantial diversity in cell wall‐associated inorganics as well
• Goal: Identify cell wall properties
5
10Metal Con• Goal: Identify cell wall properties
contributing to recalcitrance for: – No pretreatment
0
5
140
102S
1637
S2H6
303S
874
166S
443S
297S
1637
H39
S16
6H10
2H 15S
564H
303H 39H 2S
304S
564S
443H
304H
105S
319S
103
121S
105H 15H
193S
319H
121H 77S
77H
297H
193H
Diverse poplar genotypes
– Mild NaOH pretreatment– Mild alkaline‐oxidative pretreatment
Correlating Properties to Yields: Alkaline Pretreatment
80
90
100
(%)
80
90
100
(%)
80
90
100
%)
Alkaline PretreatmentR = ‐0.349p = 0.04
40
50
60
70
Hydrolysis Y
ield (
40
50
60
70
Hydrolysis Y
ield (
40
50
60
70
Hydrolysis Y
ield (
NaOH Pretreatment
R = ‐0.697
0
10
20
30
40
Glucose H
0
10
20
30
40
Glucose H
10
20
30
40
Glucose H
Hydrolysis Only
R 0.697p = 2.3x10‐6
• Significant negative correlation for lignin content
015.0 17.5 20.0 22.5 25.0 27.5
Cell Wall Lignin Content (%)
00.0 1.0 2.0 3.0 4.0 5.0
Lignin S:G Ratio
00.0 10.0 20.0 30.0
Cell Wall Transition Metal Content (ppm)
• Significant negative correlation for lignin content
• S:G and metals are not significant either i di id ll i bi ti ith li i t tindividually or in combination with lignin content
Correlating Properties to Yields: Alkaline‐Oxidative Pretreatment
80
90
100
%) 80
90
100
%) 80
90
100
%)
Alkaline‐Oxidative PretreatmentAlkaline‐Oxidative Pretreatment
R = ‐0.446 R = 0 287R = 0.363
50
60
70
80
ydrolysis Y
ield (%
40
50
60
70
ydrolysis Y
ield (%
40
50
60
70
Hydrolysis Y
ield (
R = ‐0 697
p = 0.006R = 0.287p = 0.055
p = 0.029
10
20
30
40
Glucose Hy
10
20
30
40Glucose Hy
10
20
30
40
Glucose H
Hydrolysis Only
R = ‐0.697p = 2.3x10‐6
015.0 17.5 20.0 22.5 25.0 27.5
Cell Wall Lignin Content (%)
00.0 1.0 2.0 3.0 4.0 5.0
Lignin S:G Ratio
00.0 10.0 20.0 30.0
Cell Wall Transition Metal Content (ppm)
• All three properties significant (both individual and in combination with each other) for alkaline‐oxidative pretreatment
See talk: 5‐1 Effective copper‐catalyzed alkaline‐oxidative pretreatment of woody biomass.
• First identification of the impact of differences native cell wall‐associated metals on hydrolysis yields
(Tuesday, 8:00)
Summary of Correlations• Combination of 3 properties provides good prediction• Combination of 3 properties provides good prediction• Lignin contribution negatively correlated to yields for all• S:G ratio and cell wall positively correlated to yields for
0.9Hydrolysis Only
• S:G ratio and cell wall positively correlated to yields for alkaline‐oxidative pretreatment
90
) NaOH
0.45
0.6
0.75
ength
Hydrolysis OnlyAlkaline‐OxidativeNaOH Pretreatment
60
70
80
ysis Yield (%
)
Alkaline‐Oxidative Pretreatment
NaOHPretreatment
0
0.15
0.3
ameter Str
40
50
60
ose Hydroly Pretreatment
‐0.45
‐0.3
‐0.15 Lignin S:G Metals
lative Para
10
20
30
edicted Gluco
Hydrolysis OnlyLinear model:
‐0.9
‐0.75
‐0.6Rel
0
10
0 20 40 60 80 100
Pre
Actual Glucose Hydrolysis Yield (%)
y
Y = Xβ + ε
Overview of Work
1. Response of diverse cell types in sorghum to alkaline pretreatmentsorghum to alkaline pretreatment
2. Impact of cell wall properties in diverse poplar on alkaline and
In collaboration with Markus P l B k l E
Work performed by Jacob Crowe
diverse poplar on alkaline and alkaline‐oxidative pretreatment
3 Impact of xylan o‐acetylation on
Pauly, Berkeley‐Energy Biosciences Institute
3. Impact of xylan o acetylation on cell wall nanoscale porosity in Arabidopsisp
4. Impact of alkaline pretreatments on cell wall water sorption in pmaize and switchgrass
Water‐Cell Wall InteractionsWh l k t t ?• Why look at water?
Understand cell wall porosity and polysaccharide accessibility in the context of water swellingaccessibility in the context of water swelling
Improved water penetration into cell walls yields improved enzyme accessibility to polysaccharidesimproved enzyme accessibility to polysaccharides
• Methods used: DSC for water freezing– DSC for water freezing point depression
– Water Retention Value (WRV)
Williams and Hodge, (2014). Williams and Hodge, ( 0 4).Cellulose. 21(1):221‐235.
Impact of Xylan o‐Acetylation on Alkaline Pretreatment and Hydrolysis
• Identification of esk1/tbl29 as a xylano‐acetyl transferase in Arabidopsis
y y
(Xiong et al., 2013. Mol Plant. 6(4):1373‐1375)
– Reduced size, lower cell wall glucancontent, collapsed xylem cellscontent, collapsed xylem cells
– Reduction in xylan o‐acetylation• Substitutions on hemicelluloses (e.g.Ara, GlcA, Ac) are hypothesized to control non‐covalent cross‐linking between polysaccharides
Xiong et al., 2015. Mol Plant. DOI: 10.1016/j.molp.2015.02.013
between polysaccharides– Impacts cell wall rigidity‐porosity?
• Goal: Quantify differences in cellGoal: Quantify differences in cell wall‐associated water
Impact of Xylan o‐Acetylation on Alkaline Pretreatment and Hydrolysis
• Identification of esk1/tbl29 as a xylano‐acetyl transferase in Arabidopsis
y y
(Xiong et al., 2013. Mol Plant. 6(4):1373‐1375)
– Reduced size, lower cell wall glucancontent, collapsed xylem cellscontent, collapsed xylem cells
– Reduction in xylan o‐acetylation• Substitutions on hemicelluloses (e.g.Ara, GlcA, Ac) are hypothesized to control non‐covalent cross‐linking between polysaccharidesbetween polysaccharides– Impacts cell wall rigidity‐porosity?
• Goal: Quantify differences in cellGoal: Quantify differences in cell wall‐associated water
Water Properties in Xylan o‐Acetylation‐Deficient ArabidopsisMutants
• No quantifiable differences in water retention value (WRV)• Lower content of nanoscale pore‐constrained water
Deficient ArabidopsisMutants
Lower content of nanoscale pore constrained water– Less porous cell walls due to tighter association between xylanand cellulose?
mass)
H2O
/g biom
WRV
(g H
Overview of Work
1. Response of diverse cell types in sorghum to alkaline pretreatmentsorghum to alkaline pretreatment
2. Impact of cell wall properties in diverse poplar on alkaline anddiverse poplar on alkaline and alkaline‐oxidative pretreatment
3 Impact of xylan o‐acetylation onWork performed by Dr. Dan Williams3. Impact of xylan o acetylation on
cell wall nanoscale porosity in Arabidopsis
In collaboration with Dr. Rebecca Garlock(MSU‐GLBRC)p
4. Impact of alkaline pretreatments on cell wall water sorption in pmaize and switchgrass
Grass Responses to Alkaline PretreatmentsSwitchgrassSwitchgrass(cv. Cave‐In‐Rock)• Goals:
– Relate differences in cell ll ti t i ldwall properties to yields
• Composition• Water sorption
Corn stover
Water sorption
• Pretreatments:• Pretreatments:– AFEX pretreatment
• Varying NH3:H2O loading,Varying NH3:H2O loading, temperature
– Alkaline hydrogen id (AHP)peroxide (AHP)
• Varying H2O2 loading
Correlating Yields to Composition• Xylan and lignin strong predictors of hydrolysis yields following alkaline‐oxidative delignificationyields following alkaline oxidative delignification
100%
eld100%
120%
elds 100%
eld100%
120%
elds
Corn Stover: Solid data pointsSwitchgrass: Open data points
60%
80%
drolysis Yie
60%
80%
drolysis Yie
60%
80%
drolysis Yie
60%
80%
drolysis Yie
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
0%0.0 0.1 0.2 0.3 0.4
G
Cell Wall Xylan Content (g/g)
0%0.00 0.05 0.10 0.15 0.20 0.25 0.30
G
Cell Wall Lignin Content (g/g)
0%0.0 0.1 0.2 0.3 0.4
G
Cell Wall Xylan Content (g/g)
0%0.00 0.05 0.10 0.15 0.20 0.25 0.30
G
Cell Wall Lignin Content (g/g)
Correlating Yields to Composition• Xylan and lignin strong predictors of hydrolysis yields following alkaline‐oxidative delignificationyields following alkaline oxidative delignification
• No obvious trends for AFEX pretreatment
100%
eld100%
120%
elds 100%
120%
elds 100%
eld
Corn Stover: Solid data pointsSwitchgrass: Open data points
60%
80%
drolysis Yie
60%
80%
drolysis Yie
60%
80%
drolysis Yie
60%
80%
drolysis Yie
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
0%0.0 0.1 0.2 0.3 0.4
G
Cell Wall Xylan Content (g/g)
0%0.00 0.05 0.10 0.15 0.20 0.25 0.30
G
Cell Wall Lignin Content (g/g)
0%0.00 0.05 0.10 0.15 0.20 0.25 0.30
G
Cell Wall Lignin Content (g/g)
0%0.0 0.1 0.2 0.3 0.4
G
Cell Wall Xylan Content (g/g)
Correlating Yields to Composition• Xylan and lignin strong predictors of hydrolysis yields following alkaline‐oxidative delignificationyields following alkaline oxidative delignification
• No obvious trends for AFEX pretreatment
100%
eld100%
120%
elds
Corn Stover: Solid data pointsSwitchgrass: Open data points
60%
80%
drolysis Yie
60%
80%
drolysis Yie
20%
40%
Glucose Hyd
20%
40%
Glucose Hyd
0%0.0 0.1 0.2 0.3 0.4
G
Cell Wall Xylan Content (g/g)
0%0.00 0.05 0.10 0.15 0.20 0.25 0.30
G
Cell Wall Lignin Content (g/g)
Cell Wall‐Constrained Water in Delignified Grasses• Water freezing point depression by DSC for AHP‐g p p ydelignified corn stover and switchgrass
• Clear trend of increasing cell wall‐associated water with increasing lignin removal– Corresponds to increasing hydrolysis yields
Corn Stover SwitchgrassCorn Stover Switchgrass
Correlating Properties to Water Sorption• WRV strongly correlated to hydrolysis yields and lignin content following alkaline‐oxidative d li ifi tidelignification
0.30
0.35
(g/g)
100%
120%
Yield
0.30
0.35
(g/g)
100%
120%
Yield Corn Stover: Solid data points
Switchgrass: Open data points
0.20
0.25
n Co
nten
t (60%
80%
100%
Hydrolysis
0.20
0.25
n Co
nten
t (60%
80%
100%
Hydrolysis
0 05
0.10
0.15
Wall Lignin
20%
40%
r Glucose H
0 05
0.10
0.15
Wall Lignin
20%
40%
r Glucose H
0.00
0.05
1.0 1.5 2.0 2.5 3.0 3.5
Cell
WRV (g/g)
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
0.00
0.05
1.0 1.5 2.0 2.5 3.0 3.5
Cell
WRV (g/g)
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
Correlating Properties to Water Sorption• WRV strongly correlated to hydrolysis yields and lignin content following alkaline‐oxidative d li ifi tidelignification
• Different trends for hydrolysis yields following AFEX t t t d d t NH l di
0.30
0.35
(g/g)
100%
120%
Yield
100%
120%
Yield
0.30
0.35
(g/g) Corn Stover: Solid data points
Switchgrass: Open data points
AFEX pretreatment – dependent on NH3 loading
0.20
0.25
n Co
nten
t (60%
80%
100%
Hydrolysis
60%
80%
100%
Hydrolysis
0.20
0.25
n Co
nten
t (
0 05
0.10
0.15
Wall Lignin
20%
40%
r Glucose H
20%
40%
r Glucose H
0 05
0.10
0.15
Wall Lignin
0.00
0.05
1.0 1.5 2.0 2.5 3.0 3.5
Cell
WRV (g/g)
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
0.00
0.05
1.0 1.5 2.0 2.5 3.0 3.5
Cell
WRV (g/g)
Correlating Properties to Water Sorption• WRV strongly correlated to hydrolysis yields and lignin content following alkaline‐oxidative d li ifi tidelignification
• Different trends for hydrolysis yields following AFEX t t t d d t NH l di
100%
120%
Yield Corn Stover: Solid data points
Switchgrass: Open data points
AFEX pretreatment – dependent on NH3 loading
60%
80%
100%
Hydrolysis Y
20%
40%
r Glucose H
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
Correlating Properties to Water Sorption• WRV strongly correlated to hydrolysis yields and lignin content following alkaline‐oxidative d li ifi tidelignification
• Different trends for hydrolysis yields following AFEX t t t d d t NH l di
Summary Water swelling a good predictor of hydrolysis
100%
120%
Yield Corn Stover: Solid data points
Switchgrass: Open data points
AFEX pretreatment – dependent on NH3 loadingyields for AHP‐ and AFEX‐pretreated grasses
Water swelling not correlated to any property
60%
80%
100%
Hydrolysis Y Water swelling not correlated to any property
other than hydrolysis yields for AFEX‐pretreated grasses
20%
40%
r Glucose H pretreated grasses
0%1.0 1.5 2.0 2.5 3.0 3.5
72 h
WRV (g/g)
SummarySorghum• Substantial differences in cell composition
and response to pretreatment and hydrolysis• Lignin strongest predictor of yieldsLignin strongest predictor of yields
Poplar• Lignin, S:G ratio, transition metals show
strong correlations to hydrolysis yields for alkaline‐oxidative pretreatment
ArabidopsisArabidopsis• Low‐acetate Arabidopsis cell walls contain
less pore‐constrained water
Corn Stover / Switchgrass• Substantial differences in cell wall response
to AFEX vs alkaline‐oxidative delignificationto AFEX vs. alkaline oxidative delignification• Water sorption is strong predictor of yields
for all pretreatments/feedstocks
AcknowledgementsCollaborators
Research Group:Dr. Muyang Li, Dr. Dan Williams, Dr. Aditya Bhalla, Dr. Ryan Stoklosa,
Collaborators• John Mullet, Texas A&M• Markus Pauly, Berkeley• Wellington Muchero ORNLy , y ,
Jacob Crowe, Lisaura Maldonado, Thanaphong Phongpreecha, DhruvGambhir, Nick Ferringa, Henry Pan
• Wellington Muchero, ORNL• Shi‐You Ding, MSU• Rebecca Garlock, MSU• Eric Hegg, MSUc egg, SU
Funding:Funding:• DOE, BER DE‐FC02‐07ER64494• NSF CBET‐1336622
ThankThank You!You!Questions?Questions?