May 3, Claudia Stamme, Andreas Scheidig, Klaudija Milos

Transcription

1 Enzymatic algae processing Claudia Stamme, Andreas Scheidig, Klaudija Milos

2 Direvo Industrial Biotech In 2008 Direvo Industrial Biotech was spun out of Direvo AG which pharmaceutical applications was sold to Bayer Health Care for about 210M 30 employees Optimization of proteins and strains via ultra-high Throughput Screening platform (fully automated wells/day, 1536 well-format) Protein & strain libraries u-hts Iterative optimization Fermentation New strategy developed in 2010, focusing on developing and marketing of biological solutions to the biomass conversion industry (e.g. production of aromatic chemicals from lignin, improving production efficiency of biofuels from cellulosic feedstocks, improving grain-to-ethanol processes, and developing novel algae extraction technologies) 2

3 Composition of algae cell walls Specific composition of cell wall in different classes of algae Polysaccharides: cellulose, hemicellulose, mannanes, xylans, alginic acid, sulfonated polysaccharides (agarose, carrageenan, porphyran, furcelleran and funoran) Glycoproteins Further cell wall compounds: silicon, sporopollenin, algaenan, calcium Proof of concept for enzymatic disruption of algae: Degradation of Haematococcus pluvialis (H. pluvialis) cysts for isolation of astaxanthin 3

4 H. pluvialis - Astaxanthin The green algae H. pluvialis is the major natural source of astaxanthin (AX) AX is a member of the family of carotenoids Natural AX is market as a lifestyle product (antioxidant, anti-inflammatory, ) and only natural astaxanthin has an approval by the FDA as a food additive Chemical produced AX is used in feed market (e.g. salmon) and approved as a food colorant, it is not approved for human nutraceuticals 4

5 Production of astaxanthin during formation of H. pluvialis cysts Astaxanthin is only produced in the cyst stage of H. pluvialis Trilaminar sheath (Algaenan) Source: DAMIANI et al., Phycologia (2006) AX accumulates in the cytosol of H. pluvialis in lipid globules in concentration of up to 4% and comes in a natural complex of other carotenoids Cysts are very stable by a trilaminar sheath with incorporated algaenan The extraction of astaxanthin via beating and supercritical CO 2 extraction out of the cyst stage is an expensive process 5

6 Enzymatic extraction of astaxanthin A biological solution to improve the extraction of substrates from microalgae The right enzyme mix is breaking or destabilizing the H. pluvialis cysts by degrading cell wall components w/o enzyme with enzyme A with enzyme B Absorption 474/485nm 1,2 1,0 0,8 0,6 0,4 0,2 0,0 Screening for optimal enzymes for degradation of cysts Milled cysts No enzyme Increased release of astaxanthin from algae cells 6

7 H. pluvialis cell disruption with enzymes Relative carotenoid yield [% ] 200% 150% 100% 50% 0% Carotenoid realease [% ] 214% 192% 100% untreated Enzyme 1 Enzyme 5 Carotenoid yield increased significantly Lab-scale trials: Degree of disrupted H. pluvialis cysts is higher Enzymes significantly increase the carotenoid yield from H. pluvialis 7

8 Pilot trials processing time Pilot trials at a commercial facility with standard bead mill Degree of cell disruption [%] Reduced milling time enzymatic process milling time [min] standard process without enzymes Higher degradation 8

9 Pilot trials extractable carotenoids Carotenoid yield [% of dry matter] Carotenoid concentration of dry biomass [% dry matter] 4,0% 3,8% 3,6% 3,4% 3,2% 3,45% 3,70% Carotenoid yield increased significantly 3,0% standard process enzymatic process 9

10 Application trials astaxanthin yield 4 standard process enzymatic process Astaxanthin content g / 100g No changes in astaxanthin quality or quantity (determined by HPLC analysis) 0 AX free AX ester AX total 10

11 Enzymatic algae processing Lab scale showed clear effect of enzymes on H. pluvialis cyst disruption Enzyme effects are confirmed in pilot scale Enzymatic process significantly decreases processing time Total carotenoid yield is increased HPLC analysis shows same distribution of different carotenoids No changes in free and esterified astaxanthin concentrations Industry scale trials are the next step to confirm the enzymatic effects Developing novel algae extraction technologies that allow the costefficient recovery of high-value metabolites and biochemicals 11

12 Contact information Claudia Stamme DIREVO Industrial Biotechnology GmbH Nattermannallee 1 D Köln (Cologne) Germany Phone Fax Claudia.Stamme@direvo.com Internet 12