Conversion of Cellulose, Hemicellulose And

Conversion of cellulose, hemicellulose and lignin into platform molecules: biotechnological approach Anders Frölander Gudbrand Rødsrud EuroBioRef Summer school Borregaard Industries Ltd, Lecce, Italy Norway 18-24 September 2011 Outline

reduce CO2 footprint

Agricultural products Food Lignocellulose Feed Algae

Organic waste Plastics (Materials)

Metals & minerals Chemicals Green electricity • Hydropower Transport • Solar Power • Wind power Building materials Geo-thermal Mechanical power Nuclear power

Gas and petroleum Heat

Coal Outline

The worlds first sulfite ethanol plant The inventors of sulfite ethanol production

Gösta Ekström Hugo Wallin

• Experimentation with fermentation of spent sulfite liquor (SSL) started around 1903 Skutskär sulfite ethanol plant in • They soon found out they had to Sweden started operation 1909 neutralize with lime Source: Persson, Bertil. Sulfitsprit. Förhoppningar och besvikelser under 100 år. Bjästa : DAUS Tryck & Media, 2007. ISBN: 91 7542 258-1.

Sugar composition of spent sulfite liquor (SSL) from sulfite pulping

% of DM in % of DM in Monosaccharide SSL from SSL from Sulfite Eucalyptus Spruce cooking Arabinose (C5) 0,3 0,8 Xylose (C5) 21,9 5,3 Filtration Galactose (C6) 1,6 2,1 Rhamnose (C6) 0,6 0,2 Glucose (C6) 1,6 3,7 Mannose (C6) 1,0 14,6 1 Spruce SSL • 20,6% of DM is C6 sugars • 77% of sugars are C6 sugars

Eucalyptus SSL Fibre SSL • 22,1% of DM is C5 sugars • 82% of sugars are C5 sugars 33 sulfite ethanol plants in Sweden from 1909 until today

• First sulfite ethanol plant ever opened 1909 in Skutskär, Sweden • 33 plants have been in operation in Sweden • Only one in operation after 1983: Domsjö, capacity of 15 000 m3/y

Source: Persson, Bertil. Sulfitsprit. Förhoppningar och besvikelser under 100 år. Bjästa : DAUS Tryck & Media, 2007. ISBN: 91 7542 258-1.

17 sulfite ethanol plants in Finland 1927 - 1977

• Sulfite ethanol production was stopped in 1977

• The last sulfite mill in Finland stopped production in the early 1990’ies

Source: 1. Biorefining in the pulp and paper industry. Niemelä, Klaus. Flensburg : s.n., 2008. 5th European Biorefinery Symposium. 2. Kaukoranta, Antti. Sulfittispiriteollisuus Suomessa vuosina 1918-1978 (Eng:"Sulphite alcohol industry in Finland in 1918-1978"). s.l. : Paino Polar Oy, 1981. ISBN 951-9479-25-2. 3. Niemelä, Klaus. Private communication. s.l. : VTT TECHNICAL RESEARCH CENTRE OF FINLAND , 2010. Sulfite ethanol plants in Central Europe

• Attizholts (later Borregaard) in Switzerland – Production from 1912 to 2008 – Capacity 13 mill litres – Also produced yeast and yeast extracts • M-Real in Hallein in Austria – Sulfite ethanol production 1941 – 1988 – Capacity 6 mill litres – Evaluating to restart production in 2016 • Kirov only plant still in operation in Russia

Source: 1) Borregaard internal files 2) Conference Austria April 2011 3) IEA Report: Status of 2nd Generation Biofuels Demonstration Facilities in June 2010, A REPORT TO IEA BIOENERGY TASK 39 Sulfite ethanol plants in USA

• Georgia Pacific – Bellingham mill produced ethanol from 1976 – 2001 – Capacity 24 million liters

Source: 1) Katzen customer reference list (http://www.katzen.com/projects.html) 2) Borregaard internal files 3) Graf and Koehler, June 2000, OREGON CELLULOSE-ETHANOL STUDY, An evaluation of the potential for ethanol production in Oregon using cellulose-based feedstocks. Hydrolysis of wood for ethanol, SCP and furfural • Initially developed in Germany around 1900. Yields up to 190 L/mt dry wood • Used in the USA during World War I and II – Converted further to butadien for rubber during WW II • USSR 1935 – 1985: Construction of – 18 Ethanol plants, – 16 SCP yeast plants – 15 furfural/xylitol plants – Feedstock hardwood:softwood 6:4 • Technology: weak sulfuric acid (130 – 150°C), 1 or 2 step hydrolysis • None are profitable without subsidies

Sources: Wood hydrolysis industry in the Soviet Union and Russia: What can be learned from the history? Rabinovich, M.L. Helsinki, September 2009. The 2nd Nordic Wood Biorefinery Conference (NWBC-2009), 111-120. Wikipedia contributors. Cellulosic ethanol. Wikipedia, The Free Encyclopedia. March 2, 2011, 16:08 UTC. Available at: http://en.wikipedia.org/w/index.php?title=Cellulosic_ethanol&oldid=416750931. Accessed March 8, 2011.

USSR wood hydrolysis plants 1935 - Production of ethanol, SCP and furfural Borregaard – world’s largest producer of 2nd gen bioethanol

BRG capacity 20 mill litres of bioethanol pr year 1/3 as 99,5% and 2/3 as 96%

From hemicellulose from spruce in SSL (spent sulfite liquor)

Production started 1938

Yeast strain: Baker’s yeast, Saccharomyces cerevisiae Adapted to industrial SSL continuously since 1938 Comparison of CO2 footprint of ethanol produced in different ways

Source: 1. Brekke, A., Modahl, I.S. and Raadal, H.L. Konkurrentanalyser for cellulose, etanol, lignin og vanillin fra Borregaard (Eng: Competitive CO2 footprint analysis for cellulose, ethanol, lignin and vanillin from Borregaard). Fredrikstad : Ostforld Research, Des. 2008. Confidential report. Will be published. 2. Sutter, J. Life cycle inventories of petrochemical solvents. [red.] H.-J., Chudacoff, M., Hischier, R. Jungbluth, N., Osses, M. and Primas, A. Althaus. Life cycle inventories of chemicals. Final report ecoinvnet data v2.0. Duebendorf and St. Gallen : Swiss Centre for LCI, Empa - TSL, 2007, Vol. 8 / 22. 3. Jungbluth, N., Chudacoff, M., Dauriat, A., Dinkel, F., Doka, G., Faist Emmenegger, M., Gnansounou, E., Kljun, N., Speilmann, M., Stettler, C. and Sutter, J. Life cycle inventories of bioenergy. Final report ecoinvnet v2.0. Volume 17. . Duebendorf and Uster : Swiss Centre for LCI, ESU, 2007.

Sulfite ethanol production 2011 Outline

• Leading supplier of specialty cellulose • Global leader in lignin performance chemicals, 50%+ market share • Only producer of vanillin from lignocellulosics • Production of lignocellulosic bioethanol since 1938 • World’s most advanced biorefinery in operation Borregaard product tree

Prodction cont Production stopped Product tree from 2G bioethanol 1950 - 1980

n Outline

LIGNOCELLULOSICS contain:

Lignin LIGNIN Cellulose CELLULOSE Binder Hemicellulose Fiber 20- 30% 35 - 45%

HEMICELLULOSE Various sugars 25-30% Lignocellulosic biomass structure

Cellulose fibres for chemicals Width: μm Micro fibrillar cellulose Logs Length: mm Width: nm Meters, m

Length: μm - mm

Plant cells Width: μm - mm Length: mm

Planks M and cm

Polymer chains 10 – 100 Å Glucose monomers

A few Ångstrøm

Cellulose LIGNIN CELLULOSE Binder Fiber 30% 40%

HEMICELLULOSE Various sugars 25%

Cellulose – Long chains of ONE type of ”beads” (polymer of glucose) – Forming crystals - crystalline – Same chemical structure in every plant

Hemicellulose LIGNIN CELLULOSE Binder Fiber 30% 45%

HEMICELLULOSE Various sugars 25% Hemicellulose – Long branched sugar chains (polymer, polysaccharide) – Amorphous – Composition varies largely from species to species – C6 and/or C5 sugars

Lignin LIGNIN CELLULOSE Binder Fiber 30% 45%

HEMICELLULOSE Various sugars 25% Lignin – Branched long-chain molecule (polymer) made up of 3 types of HO monomers OH HO Carb. O – Amorphous (non-crystalline) Carb. HO OH H3CO O O OH OH – Composition varies from species to OCH OH CH 3O 3 H3CO O species HO OH H3CO OH HO OCH O OH 3 O HO O O – Is the binder in all plants gluing the OH HO cellulose fibres together HO O O OCH 3 OH O H3CO OCH 3 OH O H3CO HO OCH O 3 HO O

HO OH H3CO H3CO O O OH O

(Adler, 1977) H3CO OCH 3 OH O Composition of some lignocellulosic feedstocks Outline

Chemical and/or (Partly degraded) Marketable Natural polymers mechanical products Pre- processing treatment - Biocehmicals

Separation Liquefaction/ Fermentation CCS - Biomaterials hydrolysis Sugar - Enzymatic in solution - Weak acid Chemical - Proteins - Strong acid conversion

Pyrolysis Extraction, - Biofuels

BCD ” Bio-monomers ” Chemical &

Solvolysis Catalytic conversion

Purification Gasification Synthesis gas, Catalytic synthesis CO + H2 - Energy Refining, (CCS?)

Combustion Heat, energy CO2, CCS

Outline

1. Introduction 2. History of second generation bioethanol production 3. World’s most advanced biorefinery – history and learning points 4. Lignocellulosic biomass 5. Biorefinery options 6. The biochemical route (sugar plattform) 7. Pretreatment processes 8. Hydrolysis of cellulose 9. Anaerobic and aerobic fermentation 10. Lignin options 11. Hemicellulose/pentose options 12. Process integration & closing remarks Sugar plattform path ways Hydrolysis processes • Dissolving celluose and hemicellulose leaving hydrolysis lignins undissolved – Strong acid Hydrolysis – Weak acid Lignin (S)

– Enzymatic – Microbial Cellulose (L) Hemi- Cellulose (L) Pulping processes LIQUID SOLID • Dissolving lignin (and hemicellulose) leaving cellulose undissolved – Kraft – Soda

– Sulfite Lignin – Solvent (L) – Extrusion Cellulose (S)

Lignin quality depends strongly on process Hemi- and biomass source Cellulose (L) Hemicellulose/xylan form and quality LIQUID depends on process and biomass SOLID Outline

1. Introduction 2. History of second generation bioethanol production 3. World’s most advanced biorefinery – history and learning points 4. Lignocellulosic biomass 5. Biorefinery options 6. The biochemical route (sugar plattform) 7. Pretreatment processes 8. Hydrolysis of cellulose 9. Anaerobic and aerobic fermentation 10. Lignin options 11. Hemicellulose/pentose options 12. Process integration & closing remarks Pretreatment processes • Hydrolysis processes – Strong acid pretreatment (low temp, large consumption of acids, need regeneration of acids, low yields): Weyland, TNO, BlueFire – Weak acid pretreatment (high temp and pressure, creates large amounts of inhibitiors): SEKAB, Iogen – Steam explosion (followed by enzymatic hydrolysis, also combined with acids or SO2): Abengoa, Inbicon, BioGasol, University of Lund, Andritz

• Microbial (microbes doing the whole job of hydrolysis and fermentation) – Mascoma, Arbor Fuel etc. – Solid state fermentation

• Pulping processes – Kraft: evaluated by Innventia, most common commercial chemical pulping process – Soda: evaluated by Innventia, old pulping process – Sulfite: Borregaard, Wisconsin Uni. (SPORL), modified sulfite pulping processes – Solvent/Organosolv : Lignol, CIM-V – Extrusion: PureVision (autohydrolysis)

Formation of fermentation inhibitors High temp, water, acidic conditions

Xylose Furfural

Glucose HMF – Hydroxymethyl furfural

Acetic acid

Hemicellulose Steam explosion pretreatment

• http://www.youtube.com/watch?v=jpMAiyWoEFo

BALI™ – the holistic pretreatment process • The pretreatment and separation process used in EuroBioRef for lignocellulosics Supplying sugars in solution • A pretreatment process that enables production of valuable products out of all three main lignocellulosic components – Cellulose – Hemicellulose – Lignin • A pretreatment process that facilitates low cost hydrolysis of cellulose – Low enzyme consumption (lignin inhibition avoided) – Resirculation of enzymes (no adsorption to lignin)

Ethanol C6 Chemicals

BALI Ligno- Ethanol? Pretreatment and C5 Chemicals cellulose separation Yeast

Performance Lignin chemicals BALI™ in a nutshell BALI™ in a nutshell

pulp cellulases

hydrolysis fermentation BALI™ Step 1: pretreatment & separation

Pretreated and ”reactive” pulp

Bagasse or other biomass

Water soluble lignin Mass Balance of BALI™ pretreatment process Flexibility from two optional processes

Lignin (L) Cellulose (S) Bagasse BALI Acidic Hemi- Lignin Cellulose (L) LIQUID SOLID Hemicellulose PULP Cellulose

Lignin (L)

BALI Cellulose (S) LIQUID Alkaline Hemi- Cellulose (S) SOLID PULP BALI pilot plant

• Location: Borregaard Sarpsborg, Norway • Flexible feedstock • 1 metric ton dry matter/day • Start-up Q2 2012 • Budget: 130 MNOK

Outline

Hydrolysate = monosaccharides in solution Cellulose hydrolysis

xylanases, mannanases, hemicellulases

Cellobiohydrolase

Endoglucanase

b- glycosidase

Enzymatic hydrolysis of BALI™ cellulose Yield and Viscosity

4 → 6 h Enzyme hydrolysis of BALI pulp – better substrate than soda pulp Enzymes not inhibited by residual lignins

BALI cooks

Soda cooks 140-160 °C 120-180 min Enzymatic hydrolysis - carbohydrate conversion - dose response Accellerase DUET at 7% cellulose, 50°C, pH 5.0, 72h

120,00%

100,00%

80,00%

60,00%

Reference (hardwood pulp) 40,00% BALI bagasse A

BALI bagasse B

20,00% % total carbohydrate conversion carbohydrate total % 0,00% - 0,10 0,20 0,30 0,40 0,50 0,60 ml Accelerase DUET / g glucan Outline

C6H12O6 —> 2 CH3CH2OH + 2 CO2 glucose ethanol carbon dioxide

Mw (w%) 46 (51%) 44 (49%)

• Saccharomyces cereviciae (Baker’s yeast) – Only fermenting hexoses, not pentoses – Anaerobic fermentation for production of ethanol – Aerobic fermentation for production of yeast cells – GMOs for C5 fermentation

Aerob fermentation Reproduction and production of yeast/bacteria/chemicals Example of simple aerob fermentation to yeast from pentoses with added nutrients: Outline

• Low % of feedstock useful for ethanol production – Only approx. 40%- 45% of biomass can be converted to product • Low yield in several process steps – Theoretically maximum 51% yield of ethanol from C6 sugars – No industrial solution for fermenting C5 sugars to ethanol – Several process steps with 80%-95% yield create loss and sidestreams – Lignocellulosic biomass is recalcitrant to degradation – tough demands on pre-treatment and liquefaction/hydrolysis steps – Sidestreams impure – challenge to convert into valuable products

Properties of hydrolysis lignins • Low Mw – high polydispersity • Strongly condensed (high temp) • Very few ß-O-4 bonds left – mainly C-C bonds • Few –OH groups left • Generally low O content relative to other lignins • Water insoluble • Low reactivity - hard to modify chemically at a reasonable cost • Impurity level will be high – hard to separate – impure products – many side streams • NOT A GOOD STARTING POINT FOR CHEMICALS

IAR Reims G Rødsrud 8.9.2010 Borregaard BALI lignin is water soluble

BALI lignin is sulfonated and therefore highly water soluble at almost every pH

Major challenge is to make high quality lignin specialty chemicals

Extensive application tests have been conducted

Possible uses: dispersing agent, soil conditioner, antioxidant, emulsion stabilizer, crystal modifier for batteries, dust control, binding agent, etc. Lignosulfonate structure

- At least one SO3 for every four C9 units needed to be water soluble Properties of Lignosulfonates

MW 5,000 – 80,000 Da Polydispersity 6-8 Sulfonate groups 0.6-1.2 per monomer Organic sulfur 4-8% Solubility soluble in water at all pH insoluble in most organic solvents Color light to dark brown

Delivery powder or liquid form (40-50% DS)

Non-toxic: LD50 > 5 g/kg

Quality Softwood: good Hardwood: medium Annual plants: low

Intrinsic properties of lignosulfonates

In frequent use • Binder • Dispersing agent • Emulsifier • Complexing metal ions

Under exploration commercially • Corrosion reduction • Plant growth stimulation • Antioxidant

Not in commercial use • Flame retardant • Resins (old, not in use any more) • UV-absorption/UV-protectant • Protein precipitation (old, not in use any more)

BALI - Examples of possible product mixes

100% 5 Energy 90% 18 24 80% CO2

70% 20 Yeast 8 60% 5 Ethanol 16 50% 15 Lignin 40%

30% 46 20% 41

10%

0% Acidic Neutral

% of incoming biomass + added chemicals LS decreases viscosity in mortar and concrete

Flow table test Lignosulfonate - emulsifier and dispersing agent

stabilize emulsions disperse color pigments disperse pesticides

Future use: disperse carotenoids and fat soluble vitamins Lignosulfonate in lead acid batteries crystal growth modifier => better discharge/charge performance

Soil conditioner Oxidation of lignosulfonate to vanillin

Copper catalyst is recycled due to strict limitations on copper in effluent Outline

C6 sugars to ethanol (anaerobic) • Absence of fermentation inhibitors • High yields

C5 sugars to SCP – single cell proteins (aerobic) • No inhibitors • No toxic compunds • Interesting yeast strains identified and tested

C5 sugars to ethanol (anaerobic) • Hydrolysates under testing with many GMO microbes

Theoretical yield of ethanol from biomasses How much do we gain from using GMO yeasts? Outline

• Higher turnover

BUT

• Also additional manufacturing costs and capital cost

Will it be more profitable ?????? Turnover for ethanol production and biorefineries ROCE for a biorefinery compared to P&P

Sources: 1.CEPI. [Internett] http://www.cepi-sustainability.eu/uploads/graphs/CEPI_graph_18_3.eps. 2. [Internett] Poyry. http://www.poyry.com/linked/en/publications/FIC.pdf. 3. Orkla annual reports Environmental Impact

Future limit for advanced fuels in EU and US

BALI + CCS BALI

Sources: 1. Directive 2009/28/EC of 23 April 2009 On the promotion of the use of energy from renewable sources and …. 2. Modahl, I.S., 2011, Klimagasspotensialet ved komprimering, transport og lagring av biologisk CO2. Screening LCA. Confidential report by Ostfoldforskning for Borregaard. Funding

• EuroBioRef – Borregaard granted EUR 3.0 mill funding (2010 – 2013) Acknowledgement The research leading to these results has received funding from the European Union – BALI pretreatment & hydrolysis Seventh Framework Programme (FP7/2007-2013) under grant agreement n° 241718 EuroBioRef.

• Suprabio – Borregaard granted EUR 1.1 mill Acknowledgement funding (2010 – 2013) The research leading to these results has received funding from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreement n° – Microfibrillar cellulose 241640 SupraBio.

• Biomass2Products – B2P

– Borregaard granted 2,3 mill EUR from Acknowledgement The research leading to these results has the Norwegian Research Council received funding from the Norwegian Research (2009 – 2012) Council, the BIA programme, proj. no. 193217

• BALI·PILOT BALI • PILOT – Borregaard granted EUR 7,25 mill from Innovation Norway (2011-2012)

Thank you