Challenges in co-extraction of marine algal active ingredients: An exemple of co-extraction of alginates and fucoidans

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1 Challenges in co-extraction of marine algal active ingredients: An exemple of co-extraction of alginates and fucoidans Franck Hennequart, Director Research & Innovation, ALGAIA St Lô

2 Algaia portfolio Red Seaweed Extracts Carrageenans Sold under Carra trade name Produced in Chile at our strategic partner Gelymar Brown Seaweed Extracts Alginates Sold under Satialgine, AlgoGel and Cecalgum Produced at Algaia s French factory in Lannilis Tailor made solutions Customized Solutions Sold under Algogel tradename Produced in France R&D services with partners Algaia Services Biomass, analytical, bioactivity etc Provided from our R&D center in Saint Lô (France) Specialty Seaweed extracts Under Development with specific partners or through JDA To be produced at Algaia s French facility in Lannilis 2

3 Algaia at a glance Production facility at Lannilis Next to Brest 80,000 m2 land space 15,000 m2 occupied Over 40 years experience in Alginate production ISO 9001 since 1994 FSSC certification Lannilis R&D center at Saint Lô Next to Caen Saint Lô Paris 1000 m2 state-of-the-art R&D facility Extraction laboratory Analytical laboratory Application laboratory Pilot room (in development) Headquarter in Paris Strategic Alliance with Gelymar Located in Chile 4 th largest Carrageenan producer globally ISO 9001 FSSC certification Pharma cgmp production line 3

4 Algaia at a glance Laminaria digitata France, 2 nd processor country of seaweed in volumes in Europe (about 80,000 Ton/year) Algaia: 40,000 Tons of fresh seaweed/ year Laminaria hyperborea 4

5 Algaia at a glance Algogel Customized Brand Name Innovative Natural Ingredients Specialties Green Biorefinery Alginates globally Upon Cargill s acquisition :. Satialgine range. Algogel range. Cecalgum range Functional Systems Carrageenans in Europe Carra Brand Name Red Seaweed & other natural extracts Brown Seaweed Algaia s Services Biomass Potential Analytical R&D services Enhance functionalities Bioactivity Process development 5

6 Industries we serve 6

7 ALGAIA Services Analyses & Bioactivities Saccharides Total sugars Identification and quantification of polysaccharides Characterisation in the composition Study of the size range Fatty acid profile Omega 6 Omega 3 Phospholipids Antioxidant compounds Phenolic compounds Vitamins Pigments Proteins Total Proteins Amino Acids Pigment profile Chlorophylls Fucoxanthin Astaxanthin Phycocyanin. Genomic identification Microalgae strains Seaweeds. Ionic Profile Anions Cations Heavy metals Isotopic ICPMS 7

8 ALGAIA Services Analyses & Bioactivities Prebiotic Resistance to gastric enzymes Selective fermentation of intestinal bacteria strains Anti-cholesterol Influence on activity of selected genes involved in cholesterol absorption Influence on activity of selected genes involved in cholesterol refining Intestinal Absorption of cholestérol in vitro Anti-cancer Influence on selected genes involved in apoptosis Proliferation Adhesion Migration and invasion High flow screening Anti-microbial Antiviral Antibacterial Antifungal Anti-inflammatory Enzymatic assays Cytokines, interleukines. Cosmetic Anti-ageing Anti-UV Antioxidant Depigmentation Cicatrisation Plant biostimulation Germination Defense elicitors Plant nutrition / Stress Plant defense gene expression Cardiovascular Atherosclerosis Heart failure hypertension Obesity Diabetes 8

9 ALGAIA Services boost R&D and markets Culture/Wild stock of Macro & Micro algae Producers of algal extracts Users of algal extracts Active formulae Analyses of batch composition in production Study on batch variability Statistic studies on influence of key cultivation parameters Study on the valorisation of the production Support on potential markets of interest Support on wild biomass assessment and management Support in cultivation system setup Analyses of batch composition in extract production Study on batch variability Statistic studies on influence of key production parameters Tests on extract bioactivities Valorisation of by-products Studies on compatibility of extracts in formulated products Studies on products stability based on algal extracts Test on bioactivities of formulated products Identification of active algal extracts Development of novel algal extracts Get into sustainable commercial development as soon as possible! 9

10 Biorefinery / Cracking ALGAIA First Biorefinery studies Lessonia nigrescens Lessonia trabeculata Macrocystis pyrifera Dietary Fibers Oligo-elements Proteins Alginates Fucoidans Pigments Lipids Laminarans Iodine Polyphenols 10

11 Biorefinery / Cracking Brown Seaweeds Main Active Ingredients known Proportions in Dry Matter Sugars (carbohydrates) Alginates / Uronic ac. (total > 60%) Plant Biostimulant (liquids) > 30% Binding capacity, contact effects, root (?) Laminarans 1-20% Elicitors, source of glucose bacteria Fucoidans 1-20% Elicitors, binding capacity, Antioxidants Feed Applications Binders / Stabilisers Elicitor, Prebiotics Anti-microbial, Anti-inflammatory Food / dieatary Applications Binders / Stabilisers Elicitor, Prebiotics Anti-microbial Personal Care Binders / Stabilisers Cicatrisation unknown Skin Protection Fucoxanthins 1-4% Unknown Dietary fibers Dietary fibers Slimming Mannitol 1-20% Osmoregulant, antioxidant Ashes (Sea Minerals) 5-30% Oligo-element support, root physiology (?) Antioxidant Oligo-elements + Iodine Proteins / AA 2-8% Low amounts in liquids Marine Protein / Peptides Antioxidant, Cholesterol Oligo-elements + Iodine Marine Protein / Peptides Antioxidant Natural mineral salts Unknown Phlorotannins 2-15% Antioxidant Antioxidants Antioxidants Antioxidants Lipids (fatty acids) 2-5% Low amounts in liquids Marine lipids Marine lipids Oily solutions 11

12 Biorefinery / Cracking COLLOIDS Functional Systems Personal Care Ex: anti eczema Anti-inflammatory Feed Health (incl. aqua) Preventing shrimps diseases Preventing chicken diseases Antibiotic alternative Could be the most active component of seaweed extracts used as Biostimulants Pharma applications (multiple) Anti-microbial 12

13 Material & Methods Objective: Co-extraction proof of concept Raw material Seaweed Extraction Liquid Solid Cracking Fucoidan Others Alginate Others Alginate Fucoidan 2 to 3 products of interest 1 st yield & feasability evaluation Products quality characterisation 13

14 Experimental design Single extraction Co-extraction Raw material Raw material Seaweed Species Ca neutralisation Alginic acid protocol Na Alginate Liquid fraction (Fucoidan) Soft aqueous extraction (T C, H2O, Time h) Solid fraction (Alginate) Ca neutralisation X3 X3 Alginic acid protocol Na Alginate 14

15 Sampling overview Single extraction Co-extraction Frozen Dried Frozen Dried Protocole development + Evaluation 15 samples (alginate) 9 samples (alginate) Macrocystis pyrifera 6 samples (alginate) 27 samples (fucoïdans) cracking 12 samples (alginate) 45 samples (fucoïdans) Single extraction Co-extraction Frozen Dried Frozen Dried Protocole development + Evaluation 6 samples (alginate) 6 samples (alginate) Lessonia nigrescens 6 samples (alginate) 27 samples (fucoïdans) cracking 12 samples (alginate) 36 samples (fucoïdans) Total Samples studied >220 samples + FRESH SAMPLES 15

16 Macrocystis pyrifera Classical extraction versus Co-extraction Products yields Macrocystis pyrifera (frozen) Trial 1 Trial 2 Trial 3 Average Alginate yield, single extraction (% dry weight) Alginate yield, co-extraction (% dry weight) Fucoidan yield, co-extraction (% dry weight) Better extraction yields of Alginates with co-extraction? Fucoidan extraction yields quite low 16

17 Macrocystis pyrifera Classical extraction versus Co-extraction Positive effect on final product [color and yield] whitening = current challenge in alginate industry Significative quality difference = low viscosity Extract viscosity Single alginate extraction (cpo at 60rpm) Alginate co-extraction (cpo at 60rpm) Macrocystis pyrifera (frozen) Trial 1 Trial 2 Trial 3 Average

18 Macrocystis pyrifera Co-extraction at various conditions Experimental data Control 6h/X C 3h/2X C 1h/3X C 1h/5X C Alginate co-extraction yield (% MS) Fucoïdans co-extraction yield (% MS) Alginate viscosity (cpo) 1% at 25 C Average values based on triplicats Improvement of final alginate viscosity (low viscosity) Pre-extraction step at 2X C seems interesting Experimental conditions must be improved again regarding alginate polymerisation (MV) 18

19 Importance of process condition on yields EXPERIMENTAL PLAN SETUPS Controllable parameters (water, stirring intensity ) Studied parameters (time, temperature, ratio) Extraction Responses Extraction yields Extract composition Polysaccharide composition Uncontrollable parameters (seaweed wash, age of seaweed ) 19

20 Importance of process condition on yields Full factorial design: 2 n experiments (3 factors: 2 3 = 8 extractions) Effect calculation of extraction parameters: x a = ( Y1 Y2 Y3 Y4 + Y5 + Y6 + Y7 + Y8) 8 Extractions done in triplicate: calculation of confidence interval (t. σ s2 3 k ) Definition of influential parameters: effect ± confidence interval 0 PROPOSITION OF A MATHEMATICAL MODEL 20

21 Importance of process condition on yields EXPERIMENTAL PLAN SETUPS Effect of extraction time Ash content Phenolic compounds content Mannitol content 0.26± ± ±0.67 Fucoidan content Laminaran content -0.58± ±

22 Importance of process condition on yields EXPERIMENTAL PLAN SETUPS Effect of extraction temperature Ash content Phenolic compounds content Mannitol content -0.09± ± ±0.67 Fucoidan content Laminaran content -2.68± ±

23 Time Importance of process condition on yields EXPERIMENTAL PLAN SETUPS Effect of combined parameters Time Temperature Ratio 5.97% 5.16% % Fucoidan = 4.50% %A %B %C %AB 0.09AC 0.06%BC 4.50% 3.84% +1-1 Confidence 3.04% -1 interval at 0.95% calculated = 0.27% 4.50% 4.50% +0.66% % % Fucoidan content in extract expressed as a percentage of the total solid content (n=3) 0.35% % Fucoidan = 4.50% %A %B %AB 2.30% 1.51% 1.13% Time/temperature = +0.49% Time/ratio = -0.09% Temperature/ratio = -0.06% Temperature 3.05% 2.81% Ratio 23

24 IN SUMMARY Co-extraction of functional alginates and fucoidan is viable Extraction conditions affect yields and chemical functionality Temperature, H2O Ratio & Extraction time must be assessed All these elements need to be studied before industrial & market positionning to avoid business downturns 24

25 THANK YOU! In Algae we trust

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