Optimizing enzyme cocktails and process conditions for production

Optimizing enzyme cocktails and
process conditions for production of
cellulosic ethanol
René Verwaal, Herman Pel, Marco Hensing
DSM Biotechnology Center, Alexander Fleminglaan 1, 2613 AX, Delft, the Netherlands
POET-DSM JV Project Liberty
First commercial scale cellulosic ethanol plant of
POET-DSM Joint Venture
Starting capacity 75 million liter ethanol per year from
corn residues
Shared infrastructure with existing grain ethanol plant
in Emmetsburg, Iowa
Local corn residues
Power for both plants
Grand Opening on
September 3, 2014
Overview large scale
bio-ethanol plant
Enzyme screening
Enzyme mix optimization
Recombinant produced single enzymes
DSM thermostable enzyme cocktail
for biofuel production
Development target: Increase enzyme performance
Enzyme cocktail improvement
• Enzyme screening
• Classical strain improvement
• Genomics tools
• Protein engineering
Experimental set-up
● Enzyme mix varied in 4 cellulases
● Statistical mixture design:
- 55 incubations
- same total protein dose
- variation in ranges:
CBH 0.10-0.60
● Conditions wheat straw assay:
- pH 4.5
- temperature 65°C
- time 20h
● Spiking to base mix
● Compose artificial mixes
Enzyme cocktail build around enzymes from thermophilic
filamentous fungus Rasamsonia emersonii
In 3 steps from corn
residues to ethanol
● Identify weak links
● Identify enzymes with
highest impact on cellulose
Optimized enzyme cocktail
Optimized cocktail
reduces dosage by >40%
Enzyme cocktail is active above 60°C
● High performance
in broad
temperature range
● Active at acidic pH
● Improved control
of contamination in
Natural selection of improved cocktails
Classical Strain Improvement (CSI)
CSI effects enzyme
cocktail composition
Strong changes in amount and
ratio of main cellulases
CSI and process optimization
Fast feedstock liquefaction
Fast viscosity reduction. 20% acid-pretreated wheat straw, 62ºC
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DSM thermostable
Cost savings!
• Fast liquefaction allows
high dry matter content
up to 25%
• CAPEX savings due to
lower required capacity
of process equipment
• Thermostability allows
for less energy use for
• OPEX savings due to
reduced energy costs
• Thermostability gives
improved control of
• Reduced antibiotic costs
• Increased revenues
because less sugars are