Conversion of macroalgae to bioethanol and fish feed preliminary trials in the MAB3 project

Conversion of macroalgae to bioethanol and fish feed preliminary trials in the MAB3 project Anne-Belinda Bjerre, Senior Scientist PhD Danish Technological Institute 3rd Danish Macroalgae conference Grenå

Welcome to Danish Technological Institute

Division for Energy and Climate Center for Biomass & Biorefinery 4 locations in Denmark Several projects related to solid and liquid biofuels EU-projects / National projects / Industry Denmark Grenå Algae Center Denmark Aarhus Biomass Laboratory and Offices Taastrup Enzyme Laboratory and Offices Sdr. Stenderup Pilot plant and production facilities

Contents Background Processes on macroalgae Cultivation Harvest Conditioning Enzymatic hydrolysis Ethanol production Sustainability and economic feasibility

BACKGROUND the MAB3 project MacroAlgaeBiorefinery for 3G bioenergy and fish feed An integrated biorefinery concept to be developed for conversion of two species of macro algae Laminaria digitata and Saccharina latissima into energy carriers, together with a protein enriched fish feed derived as a residual from the energy conversion processes Financed: the Danish Strategic Research Council Total budget of 24 million DKK Project period: March 1 st, 2012 ---- March 1 st, 2016 12 Partners from Denmark, Germany, Ireland, Italy Education of 4 PhD and 2 postdoc Coordinator: Danish Technological Institute v/ Anne-Belinda Bjerre

MAB3 Partners Danish Technological Institute (Coordinator) (DK) Aarhus University (2 institutes) (DK) Technical University of Denmark (3 institutes) National University of Ireland, Galway (IRL) University of Hamburg (DE) University of Sienna (IT) Orbicon (DK) DONG Energy (DK) Aller Aqua (DK) VitaLys (DK) DanGrønt Products (DK) Affiliated partner: Novozymes (DK) [delivery of enzymes and participating in the advisory board]

MacroAlgaeBiorefinery

Which species are of relevance? Experiences from the North Atlantic Sea show promising results from Laminaria digitata Fingertang (a) Saccharina latissima Sukkertang (b) Palmaria palmata Søl (c) c a b

CULTIVATION MAB3 10 km of seeded lines Saccharina latissima Laminaria digitata Deployed in September 2012 Line mussel system Limfjorden, Denmark

HARVEST MAB3 Saccharina latissima Harvest time: May 2013 Growth period: 7-8 months Yield: 2 wet tons of S. latissima (2 km line) Harvest technology: line-mussel cultivation

HARVEST Laminaria digitata Natural population 300 kg August 2012 Adams et al., Bioresource Technology 102 (2011) 9976 9984

RAW MATERIAL CHARACTERISATON Species Harvesting Time Protein (%) Sulphated fucoidan (%) Mannitol (%) Glucose (%) Minerals (%) Xylose +Mannose (%) Residues (%) SUM (%) Laminaria digitata Laminaria digitata Saccharina latissima 2012.04. 14.6 6.8 3.7 7.7 31.8 1.2 5,5 71.3 2012.08. 3.9 3.5 6.4 56.9 8.1 0.5 2.7 82.1 2013.05. 20.1 4.3 5.1 5.8 35.4 0.6 8.6 79.9

CONDITIONING MAB3 Screw Pressing -Algae juice Drying Water content Water content 50 C (4 days): Laminaria 78% 75% Laminaria: 78% 10% Saccharina: 89% 9% Saccharina 89% 88%

PRETREATMENT AND ENZYMATIC HYDROLYSIS Laminaria digitata (harvested in August 2012) Wet Brown Seaweed Dried and Grinded Re-adjust DM by water Enzymatic Hydrolysis Wet Brown Seaweed Wetmilled by Disc mill Collect fibers Enzymatic Hydrolysis

PRETREATMENT AND ENZYMATIC HYDROLYSIS MAB3 Wet Brown Seaweed Wet-milled by Disc mill Collect fibers by centrifugation Enzymatic Hydrolysis 1.0 mm 0.2 mm (Picture: Dirk Manns DTU) Enzymatic hydrolysis (Picture: Annette Bruhn) Laminaria digitata (approx. 2m, DM 27%) (Picture: Dirk Manns DTU) Sprout-Bauer 12 Lab disc mill (disc distances 1.0 and 0.2mm) (Picture: Stinus Andersen DTU) ph 5.1, T 40 C, t 72h, 4% [S]/[V], CellicCTec2 (Novozymes): 5% [E]/[S] Alginate Lyase (EC 4.2.2.3 ): 0.25% [E]/[S] Disc distance 0.2 mm 1.0 mm Glucose [% dry fibers] 22.8 ± 0.9 32.7 ± 0.7

3. generations bioethanol Recovery of fucoidan Wet milling Fucoidan (value-added product) Enzymes Yeast 3rd Danish Macroalgae conference Grenå Fucoidan extract Polysacchari des, Protein Hydrolyses Hexose fermentation Destillation EtOH Protein

Fermentation to ethanol SSF

Protein Concentration (g/100 g material) 12 ETHANOL PRODUCTION MAB3 Exp ID Lam 1 Lam 2 Lam 3 Lam 4 Pretreatment Wet-milled by disc mill (0.2 mm) Wet-milled by disc mill (1 mm) Dried, grinded by pin-mill and screened (ø 1 mm) Dried, grinded by pin-mill and screened (ø 1 mm) Substrate DM (%, w/v) 4 4 5 10 Glucose in substrate (g/l) 9.12 13.1 28.5 56.9 10 8 6 4 2 0 BF 3,8 11,5 AF Enzyme loading CellicCtec2: 5 % v/w [E]/[S] Alginate Lyase (EC 4.2.2.3): 0.25 % v/w [E]/[S] CellicCtec2: 5 % v/w [E]/[S] Alginate Lyase (EC 4.2.2.3): 0.25 % v/w [E]/[S] Celluclast 1.5L: 40 U/g DM Alginate lyase (EC 4.2.2.3): 2 U/g DM Celluclast 1.5L: 40 U/g DM Alginate lyase (EC 4.2.2.3): 2 U/g DM Hydrolysis temperature ( C) Yeast inoculation conc. (g/l) Fermentation temperature ( C) Ethanol yield (% theoretical value), after 48 h fermentation 40 40 40 40 2 2 2 2 32 32 32 32 73 77 73 75 Ethanol concentration (g/l) 3.4 5.2 10.5 21.8

ECONOMIC POTENTIAL 100 kg wet algae biomass (Laminaria digitata) 2 kg Algae juice (dry matter) Conditioning 1 kg Value added products (Fucoidan, etc.) 7,4 kg Ethanol + 7,1 kg CO 2 1 kg Protein Pretreatment C6-Fermentation Separation of liquid and solid Input: construction and building materials, enzymes, energy and electricity, water MAB3 MacroalgaeBiorefinery Output CO2eq Other output Emission in air, water and soil Enzymatic prehydrolysis Bioethanol Proteins 2 kg Amino acids Residual sugars fermentation 3 kg fertilizer (inorganic salts and silicium) Residuals

ECONOMIC POTENTIAL MAB3 Price ( /kg) Weight Scenario Scenario Scenario Scenario (kg) 1 a 2 b 1 2 Wet algae 100.0 1.12 0.08 112 8 Cost Value added products (e.g. Fucoidan) 1 2.9 2.9 2.9 2.9 Ethanol 7.4 1 1 7.4 7.4 Protein 1.0 1.5 1.5 1.5 1.5 Income Amino acids 2.0 1 1 2 2 Fertilizers 3.0 0.35 0.35 1.05 1.05-97.15 6.85 Margin a Scenario 1: Price of macroalgae from Watson, L. and Dring, M., 2011. Business plan for the establishment of a seaweed hatchery & grow-out farm. Irish sea Fisheries Board, pp 41. B Scenario 2: Price of macroalgae from Michael Bo Rasmussen personal communication.

ENVIRONMENTAL SUSTAINABILITY ASSESSMENT MAB3 Input: cultivation system (lines, buoys, water, nutrients) GHG savings on the climate mitigation bank account Macroalgae cultivation and harvesting Output: CO2eq Other outputs: Emissions in air and water Algae Amounts (g/kg) Macroalgae carbon content 43-50 Macroalgae nitrogen content 3-5 CO 2 assimilation 158-182 Avoided N 2 O emission from N assimilation 4-8 Total CO 2 eq 1524-2519 According to IPCC guidelines

Conclusions Macroalgae cultivation has high potential for CO 2 saving providing water quality protection by assimilating excess nutrients Raw material price is essential for the overall feasibility Knowledge about biomass composition is important for the choice of biorefinery processes Ethanol and protein are realistic products from Laminaria digitata, however the algal biomass must be harvested at the right time in August/September due to the need of high C6-sugar contents

ACKNOWLEDGEMENT Thanks to the Danish Council for Strategic Research for financing the MAB3 project More information: www.mab3.dk Thanks to co-authors: Xiaoru Hou, Karin Svane Bech, Jonas Høeg Hansen, Annette Bruhn, Bodo Saake, Anne Meyer, Dirk Manns, Michael Bo Rasmussen, Mette Nielsen, Jens Kjerulf Petersen, Ditte Tørring, Peter Daugbjerg Jensen, Michele Seghetta, Simone Bastanoni, Marianne Thomsen

DTI CONTACTS: Anne-Belinda Bjerre (Senior Scientist, Senior Consultant): anbj@dti.dk Coordinator Conditioning, bioethanol and pretreatment Biorefinery Lars Steenberg Nikolajsen (Section leader, Senior Consultant): lsn@dti.dk Macroalgae cultivation and harvesting Conditioning and Pretreatment Karin Svane Bech (Consultant): ksb@dti.dk Newsletter and dissemination