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Biotechnology in the Food Industry - From a Fermentation Technology Supplier Perspective CLIB Forum 4th July 2013 Technology and Tradition - Working for the Process Industry since 1878 History: • In 1878 Heinrich Frings sr. founded a vinegar factory in Aachen. • Acetic acid fermentation at the scientific-engineering level is initiated by Heinrich Frings jr. • At the beginning of the 20th century Frings developed into a constructor of fermentation and other process systems. • Frings processes have evolved through several generations of technologies. Frings today: • Headquarters in Bonn, Germany • International constructor of plants for food technology‚ environment technology and biotechnology, • 80 employees (worldwide) • Largest project realized by Frings: 23 Mio. € Markets and Technologies - Actual Product Portfolio of FRINGS Key Technologies and System Solutions for the Fermentation Industry: Food Filtration Biotechnology Separation Environment Aeration Automation Others Mixing Fermenter Technologies - Sizes of FRINGS Fermenters Volumen Range: 500 mL – 500.000 L Lab Scale Fermenter PROREACT 5 P Pilot Fermenter PROREACT 75 P Large Scale Fermenters PROREACT 300 B Fermenter Technologies - Hygienic Standards FRINGS “Package Units” and Fermentation Groups for a Wide Range of Food Applications: Fermentation Groups with low hygienic standard (FRINGS HEJP Fermenter for Baker‘s Yeast) FRINGS Systems with CIP and SIP (PROREACT B Fermenter) Units for Recombinant Cells with highest standards (PROREACT G Fermenter) Industrial Bioprocesses - Block Diagram in Biotechnology Up-Stream Up-Stream CIP/SIP Raw Material-/ Mediapreparation Sterilisation Down-Stream Down-Stream Fermentation CIP/SIP Separation/Isolation Purification/Blending Confectioning/Packaging Product Waste Disposal Products made with Biotechnological Methods Biotechnology related with Food Industry - Product Clusters 1. Fermented beverages/Fermented liquid foods 2. Classical food additives ‚with E number‘ 3. Classical food additives ‚without E number‘ 4. Special yeast products 5. Technical enzymes 6. Vitamins 7. Biogenic nutraceuticals/special food additives 8. Algae biotechnology 9. (Bioenergy) 10.(Bioplastics production) 11.(Biopesticides) Biotechnology related with Food Industry - Product Clusters 1. Fermented beverages/Fermented liquid foods Beer, wine, fermented milk products, vinegar, other lactic acid fermentates, gluconic acid fermentates, soy sauce, kombucha, kwas, …. 2. Classical fermented food additives ‚with E number‘: MSG, citric acid, gluconic acid, xanthan, (carotinoids), (alginate), … 3. Classical food additives as ‚fermentates without E number‘: Fermentates as flavor enhancers which are produced without purification with natural microorganisms and which therefore, under certain conditions, do not require declaration under an E number. Biotechnology related with Food Industry - Product Clusters 4. Yeast products Baker‘s yeast, starter cultures, yeast extracts, ‚savoury‘ yeasts, ‚detoxin‘ yeast, recombinant yeasts for enzyme production, … 5. Technical Enzymes: Enzymes for dairy, baking, sugar industry, agro-enzymes, lipases, proteases, amylases, pectinases, phytase, asparaginase, … 6. Vitamins Riboflavin (B2), cyanocobalamin (B12), vitamin C, … Biotechnology related with Food Industry - Product Clusters 7. Biogenic nutraceuticals/special food additives: Amino acids and intermediates like adenosylmethionine, aspartame, antioxidants, omega 3/6 fatty acids, special sugars like fucosyllactose, exopolysaccharides, probiotics, algae products, … 8. (Bioenergy) 9. (Bioplastics production) 10. (Biopesticides) Other Purposes: Energy Integration, Packaging, Agriculture, Added value byproducts, … Aspects of Food Industry Regulations - Restrictions for Innovative Products Excerpt of Regulations for the Food Industry (in Germany): • • • • • • • • • • • • • • • Lebensmittel- und Futtermittel-Gesetzbuch LFGB Verordnung Lebensmittelhygiene 2004/852/EG Lebensmittel Kennzeichnungsverordnung LMKV Zusatzstoff-Zulassungsverordnung ZZulV Zusatzstoff-Verkehrsverordnung ZVerkV Verordnung Nahrungsergänzungsmittel 2009/1170/EG Richtlinie Nahrungsergänzungsmittel 2006/37/EG Richtlinie Nahrungsergänzungsmittel 2002/46/EG Verordnung Enzyme 2008/1332/EG Verordnung Lebensmittelzusatzstoffe 2008/1333/EG Verordnung Aromen 2008/1334/EG Verordnung GMO 2003/1829/EG Verordnung Biologische Produkte 2007/834/EG Novel Food Verordnung 1997/258/EG … Example 1 - Classic Food Biotechnology - Baker’s Yeast Production and other Products produced with Yeasts Baker’s Yeast - Starter for the Bread Baking Process General Process Chain: Flour Verunreinigungen Enzymes, Blends, Salts Water, (Baking Powder) Heat Heat Screening Mixing Kneading Leavening 1. Rising Proofing 2. Rising Baking Bread Baker‘s Yeast Water Yeast Production - General Process Chains for Different Products Molasses Chemicals Molasses preparation Molasses separation Lab Culture Saccharomyces cerevisiae Pure-/Seed Yeast I/II Yeast Separation Yeast Milk 4-6 oC Commercial Yeast Yeast Separation Yeast Filtration Yeast D27-D32 Autolysis Separation Evaporation Spray Drying Yeast Extracts Granulation Moving Bed Drying Dry Yeast Extruder Cutter Block Yeast Baker‘s Yeast Technology - Large Scale Bulk Production CFD for Aeration Design Typical Yeast Plant, FRINGS Product: Mix of fresh and dry yeast Production Capacity: 3.000 – 8.000 to/a Fermenter Volume: 240 m3 Design: static Mixing and Aeration System FRIBORATOR 24000, FRINGS Bonn Air Flow: 8.000 m3/h Power 315 kW Yeast Technologies - Use of Yeasts for Different Food Related Products Yeasts for the Production of Enzymes in the Food and Feed Industry Yeast Saccharomyces cerevisiae S. carlsbergensis Kluyveromyces lactis Kluyveromyces fragilis Pichia pastoris (GMO) Product Invertase, Galactosidase Invertase Lactase, Chymosin Lactase, Inulinase, Invertase Phytase Yeasts for the Production of Food Yeast Saccharomyces cerevisiae Zygosaccharomyces, Candida spec. Phaffia rodozyma Kluyveromyces lactis Saccharomyces carlsbergensis/cerevisiae andere Kluyveromyces marxianus Candida (Torula) utilis Saccharomyces spec. Product Baker’s Yeast Soy Sauce, Kombucha Carotinoids Whey Fermentation Beer, Wine Brewing Kefir, Raw Sausage, Savoury yeast (MSG, 5’GMP, 5’IMP) Example 2 - Classic Food Biotechnology - Vinegar Production and other Products Produced with AAB Vinegar Production - Wine and Bulk Vinegars Sugar-/Starchbased Raw Material Enzymes Wine Mashing Fermentation Filtration (Dilution) Wine Vinegar Pressing/Extraction (Saccharification) Mashing Alcohol fermentation Yeast separation Acetic Acid Bacteria Yeast Rectification Ethanol (Mashing) Fermentation Filtration Dilution Bulk Vinegar Slop Water Vinegar Production - Traditional and Modern Production Traditional European Vinegar Fermentation Traditional Chinese Solid State Vinegar Fermentation Modern Acetic Acid Vinegar Fermentation Plant, KÜHNE Germany Production Capacity 45.000 to/a Vinegar Production - Increase of Performance Data Acid concentrations achieved by Frings Acetators 25 24 23 22,3 22 acetic acid concentration [% w/v] 22 21 21 19,8 20 20,5 19 20 19 18 16,5 17 17 16 16,2 15 15,5 15 14,5 15 14,5 14 13,5 14 13 12,5 13 12 11,5 12 11 19 17,5 18 11 10 year Normal process DSHS process Historical Data of ACETATOR Processes SSHS process Normal process future 20 20 20 15 20 10 20 05 20 00 19 95 19 90 19 85 19 80 19 75 19 70 19 65 19 60 19 55 19 50 10 Vinegar Fermentation - Main Factors who have increased Performance Microbiology: • Evolutionary Screening of Acid Resistant Mutants • Development of Special Nutrients and Micronutrients Bioreactor Design: • New Methods for Fermenter Design higher kLa Processes: • Optimization of Repeated Fed Batch Processes • Gradients of Relevant State Variables (T, CS, …) Automation: • Highly Reproducible Automatic Charge and Discharge Control • Exact Flow Count of Transferred Liquids and Online Control of EtOH Expert Knowledge: • Operational Skills/Training Potential of Processes with AAB - for Different Industries Vinegar Technology -Wine and Fruit Vinegars -Rice and Cereals Vinegars -Spirit Vinegar -Food Ingredient -Pickled Vegetables -Ketchup and Sauces -Vinegar Drinks … Food and Beverage Biotechnology -Fermented Softdrinks -Food Ingredient -Vitamins -Special Sugars -Kombucha Similars -Thickeners … -Cosmetic Industry -Neutraceuticals -Pharmaceuticals -Polymers -Fine Chemicals -Fragrances -White Biotechnology -… Substrate Spectrum of AAB - Metabolism of Acetic Acid Bacteria Ethanol Acetaldehyde Acetate Ox-PP Glucose Ox-PP Gluconate Lactate Pyruvate Glucose Gluconate UQH2 2-Keto-gluconate 5-Keto-gluconate D-Sorbitol, Gluconate, Glycerol 2,5-Diketo-gluconate L-Sorbose, 5-Ketogluconate, DHA Respiratory chain D-Sorbitol L-Sorbose D-Sorbitol UQH2 Pyruvate Ethanol Acetaldehyde D-Fructose Ox-PP Acetate D-Fructose Unknown substrates Unknown products Flavin PQQ Unknown substrates Unknown products Unknown substrates Unknown products Processes with AAB - for Different Industrial Production Purposes Product Process Remarks Dihydroxyacetone (Tanning Agent) Fed Batch - Product Concentration > 200 g/L Erythrulose (Tanning Agent) Fed Batch - pH control - Low Temperature preferred Ascorbic Acid (Vitamin C) and other Acid Intermediates (L-Sorbose 2,5-Diketogluconate 2- and 5-Ketogluconates,..) Batch Fed Batch - Direct Route under Development Miglitol (Anti Diabetic Drug) Intermediates (6-Amino-L-Sorbose, Nprotected) Two Step Process: 1. Cell Growth on Sorbit 2. Disc Stack Separation 3. Whole Cell Batch Catalysis - High Integral Oxygen Demand Fruit Esters/Acids (Aroma Industry) Fed Batch - ATEX Requirements Gluconic Acid Bases (Food and Beverage Ingredient) Batch/ Sequential Batch - Isolates from Kombucha Acetic Acid (Vinegar, Glacial Acid) Batch Repeated Batch Repeated Fed Batch - Autosterile Process - Highly Sensible to Lack of Oxygen - Product Concentration > 200 g/L Scale Up/Down - AAB Fermenter for R&D Industrial AAB Vinegar ACETATOR Lab-/Pilot-Vinegar ACETATOR Industrial Sterile AAB PROREACT Mini-PROREACT Pilot-PROREACT Example 3 - Biorefinery and Food - Added Value Byproducts in 1st Generation Biorefinery Biorefinery - With Added Value Byproducts for Food and Feed Industry Wheat Grinding Mashing Wheat bran Gluten Gluten Separation Liquefaction Enzymes Separation Saccharification Enzymes Alcohol fermentation Yeast separation Compression Yeast Propagation Yeast Stillage Drying DDGS Rectification Dehydration Bioethanol CO2 Biorefineries - With Added Value Byproducts for Food and Feed Industry Plant erected in Kasachstan in 2006 by FRINGS Austria Biorefineries - With Added Value Byproducts for Food and Feed Industry Frings Biorefinery for the Production of Bioethanol, CO2, Glutenprotein, Yeast and DDGS with Integrated Water Management System Production Capacity Ethanol: 200.000 L/24 h Example 4 - Modern Food Biotechnology – Development of Enzyme Processes (in Germany) Enzyme Technology - Food Enzymes in Use or Under Development Food Enzymes Purposes, Improvements Bread and Bakery Products Amylase, Lipase, Protease, Hemicellulase, Xylanase, Glukoseoxidase, Hexoseoxydase, Lipoxygenase, Transglutaminase, Asparaginase, … Optimisation of dough, shorter leavening, baking process, freezing, pumping, acrylamid reduction, … Sweets Invertase, Lactase, Isomerase, … Sweetening, improvement of rheology, liquifaction of „pralines“, lactose hydrolysis Starch Amylase, Pullulanase, Xylanase, Glucooxidase, Isomerase, CGtase… Saccharification, sweetening, modified starch, fat simulation, dextrins, water binding, .. Dairy Lactase, Transglutaminase, Protease, … Lactose hydrolysis, aroma modification, fat simulation, texture modification, … Eggs Katalase, Glucoseoxidase, … Conservation, colour reduction, … Meat and Sausage Protease, Transglutaminase, … Enhancement of maturation, better texture, „meat glue“, … Proteins Protease Foam, viscosity, milk imitation, soy sauce, savoury, Flavor Lipase, Protease, Pektinase, Pektinesterase Artificial cheese, butter and meat flavor, citric flavor, colour extracts, … Noodles Lipase, Hemicullase, Xylanase Texture and colour improvement, increase of hydrophobicity, Enzyme Processes - Fermenter for Lab- and Pilot Enzyme Development PROREACT P Seed Train 10.000 L PROREACT P Multi + Seed Train 150 L Remark: Specific Design for Pichia pastoris fermentation with methanol as Inducer Enzyme Processes - Fermenter for Lab- and Pilot Enzyme Development PROREACT P for Enzyme Development at c-LEcta, Leipzig Thank You very much for Your Attention