Renewable Carbon-Feedstock to Industrial Chemicals: Producing Renewable Materials from Granular Starch

Transcription

1 S E 1 E 2 E 3 E 4 E n Continuous Biocatalytic Systems P Renewable Carbon-Feedstock to Industrial Chemicals: Producing Renewable Materials from Granular Starch Manoj Kumar, PhD Sr. Scientist, Research & Development Genencor International, Palo Alto California, USA 57 th Starch Convention Detmold, Germany April, 26 th, 2006

2 2 Production of Chemicals: O&NG Feed Stocks Impact of Oil & Natural Gas Prices on Chemicals Cost of O&NG ($$ Billion) Cost of O&NG Feed Stocks ($$ billion) % Feed stocks Cost of Products Year : $$ billion American Chemistry Council

3 3 Industrial Fermentations Feedstock: Predominantly use glucose as feed stock Process: Batch, fed-batch, or continuous Operate under substrate-limited and minimal by-products forming conditions Optimize fermentation time, yield and efficiency Glucose as a feed stock: enzymatic inhibition and/or catabolite repression, and/or growth of microorganisms Products: Proteins, enzymes, & chemicals

4 Products - Chemicals - Enzymes - Proteins 4 Industrial Fermentations Feedstock -Glucose Run with glucose-limitations Enzyme Inhibition Catabolite Repression Growth of Microorganisms

5 5 Granular Starch: Advantages over Glucose Cost: Lowest cost substrate: Corn Starch (5 cents/kg vs. 22 cents/kg glucose) Availability: Readily available from a corn wet milling and wheat starch processing plants Processing: Glucose limited vs. glucose excess Feasible separation of unhydrolyzed granular starch resulting in a high purity product Process simplification No capital cost investment

6 6 Granular Starch to Biochemicals Conversion of granular-starch to glucose: GSHE:Use of hydrolyzing enzymes to convert granular starch to glucose in a rate same as glucose feed rate required to achieve desired fermentative product Glucose limited and avoiding catabolite repression or enzyme inhibition Sugar production and processing cost eliminated Genencor WO/US Patent Application: WO 03/ published August 14, 2003; US published Oct. 30, 2003

7 7 Glucoamylase: Granular Starch Hydrolyzing Enzyme Activity A. niger Glyco-hydro-15 SBD Humicola Glyco-hydro-15 SBD SBD 167 R.oryzae GSHE 550 References: Ueda,et.al(1984),Microbiol.Sci.1,21-24; Hayashida,et.al.(1989),Agric. Biolo.Chem.53, ; Ashikari,et.al.(1986),Agric.Biol.Chem.50, ; :Allison et.al,(1992)curr.genet21,225-22; Balls et.al (1945)JBC.156,

8 8 Alpha Amylases Exhibiting Granular Starch Hydrolyzing Enzymes Activity Catalytic domain Linker SBD Examples: 1)Fungi-Aspergillus niger,yamazaki et.al (1951),Nippon Nogeikagaku Kaishi,24, Aspergillus Kawachi,Kaneko et.al,1996j.ferm & Bioeng.81, Rhizopus niveus,yamamoto et.al(1990)denpun Kagaku,37, ) Bacteria-Baciilus circulans Ho et.al,1992,j. Microbiol.& Biotechnol.2,56 Bacillus polymixa Sohn et.al,1992, J.Microbiol & Biotechnol.2,183

9 S p e c i a l t y C h e m i c a l s Amino Acids Sorbitol MSG Citric Ascorbic Gluconic Lactic Antibiotics 1,3 Propanediol Enzymes Bio-pharmaceuticals Process for Converting Starch to Biochemicals Water Liquefaction Saccharification Wheat 9 Current Process Corn Jet Cooker 108 C, 5-8min. Starch Tank 95 C, 90min. 60 C, hrs. Tapioca Purification Ion Exchange Carbon Glucose Starch Separation

10 S p e c i a l t y C h e m i c a l s Amino Acids Sorbitol MSG Citric Ascorbic Gluconic Lactic Antibiotics 1,3 Propanediol Enzymes Bio-pharmaceuticals Process for Converting Granular Starch to Biochemicals Water Wheat L o w E n e r g y P r o c e s s 10 L o w E n e r g y Process Starch Tank Corn Tapioca Glucose Starch Separation GSHE For Biochemicals

11 11 Industrial Chemicals: Whole Cell or Enzyme Bioreactor Sugars from Granular Starch [G] < Ki Industrial Chemicals * Organic Acids Gluconic, Ascorbic, Lactic, Succinic acid * Amino Acids Lysine, MSG * Antibiotics * Solvents 1,3-Propanediol, Butanol, Acetone, Glycerol

12 12 Industrial Chemical: Gluconic Acid Chemical Volume (MT/yr) Price ($$/Kg) Market (MM$/yr) Sodium Gluconate 60, Glucono-δ-lactone 15, Chemical Reporter

13 13 Gluconic Acid Applications Dairy Industry Prevent the deposition of milk stone Food Industries Baking, Tofu, Meat processing, dressing, soft-drinks, Pharmaceutical Amion fluid, Calcium and Iron supplementation therapy, Cleaning Aid Aluminum cans, bottles, and equipment Gentle metal Cleaning operations Non-corrosive Textile Prevent iron deposition Metallurgy Alkaline de-rusting operation Cement Additive to modify settling

14 14

15 15 Single Step From Granular Starch To Gluconic Acid Starch HTAA Sac.Enz. Sac.Enzymes. GO/CAT Slurry Tank Liquefaction Saccharification Gluconic Acid Starch To Gluconic GSHE Blend GO/CAT Granular Starch Gluconic Acid

16 16 In vitro Biocatalytic Conversion of Granular Starch to Gluconic Acid Granular starch hydrolyzing enzyme mix D-Glucose CHO HCOH HOCH HCOH HOCH CH 2 0H Glucose oxidase ½ O 2 Catalase D-Gluconic Acid COOH HCOH HOCH HCOH HOCH CH 2 0H

17 Why Enzyme based Gluconate Process 17 Glucose as a Substrate Supply Whole cell process - Maximum solids (sugars) 20% due to inhibition A patented (US ), but not practiced enzyme process % glucose possible, but 1) partially inhibits the enzymes 2) need non-economical enzyme dosage 3) a high oxygen demand process 4) high viscosity reduces oxygen transfer

18 18 Why Granular Starch/Enzyme based Gluconate Process Better than the current (A. niger) gluconic acid production process A given bioconversion rate is achieved with a significantly lower enzyme concentrations Significantly lower substrate cost of granular raw starch vs. D-glucose The excess (i. e., residual) feed stock is recoverable along with substantial amount of enzyme for renewed bioconversion Reduced substrate and/or product-based inhibition of enzymes Control over feed-stock concentration: A major process parameter

19 19 Advantages of In-Vitro Biocatalytic Process for Granular Starch Conversion to Gluconate Simple & Economical Prevents product inhibition of granular starch Hydrolyzing enzymes by concurrent conversion to gluconate Quantitative conversion feasible Elimination of byproduct formation Higher productivity Increases production capacity Higher Yield on carbon

20 20 In-vitro Bioconversion Process of Granular Starch to Gluconate Granular Starch Enzymes GRANULATOR Electrodialysis BIOREACTOR

21 21 Conversion of Granular Starch to Gluconate Products (g/l) Gluconate (g/l) Glucose (g/l) Time (hrs) using Glucose Oxidase, Catalase and a GSHE Blend GSHE enzyme blend has excellent kinetics for granular starch to sugar conversion Glucose oxidase and catalase enzymes dosage is drastically lower.

22 22 Conversion of Granular Corn Starch to Gluconate Products (g/l) Gluconate g/l Glucose (g/l) Hours using Glucose Oxidase, Catalase and a GSHE Blend; ph 5, 40 C Good Conversion of Granular Starch to Gluconate

23 23 Granular Wheat Starch to Gluconate Glucose and gluconate (g/l) gluconate (g/l) Glucose (g/l) Time (hr) using GSHE, Glucose Oxidase, and Catalase at ph 5 and 40C

24 24 Key Learning! Rate GA > Rate of G [G] ~ 0 g/l Enzymes Stable Improved Volumetric Productivity with Wheat granular starch vs. Corn granular starch Reaction to Completion: Optimization Work Enzymes inhibition by H2O2?

25 1,3-Propanediol Production from Glucose Glucose G-3-P Pyr TCA DHAP Gly-3-P Gly 3HPA Propanediol Propanediol

26 26 Fermentative Bioconversion of Granular Starch to 1,3-Propanediol Products (g/l) EFT (hrs ) [Gluc] [Gyl] [1,3ppd] at ph 6.65, 34C, 300rpm, using GSHE and genetically engineered E. coli

27 27 Fermentative Bioconversion of Granular Starch to Lactic Acid Products (g/l) [Gluc] [lactate] EFT (hrs) using GSHE and Lactobacillus sp. ph 6.4, 34C

28 Fermentative Bioconversion of Granular Starch to Succinic Acid Products (g/l) Succinate] [Glucose] EFT (hrs) using GSHE and an E. coli strain at ph 6.65, 34C, 300rpm

29 29 Summary Better Process Continuous supply of glucose is provided under controlled rate conditions Key glucose-limited bioconversion conditions Avoiding many of the metabolic regulations and cellular inhibitions Altered levels of catabolite repression and enzymatic inhibition Improved oxygen transfer for metabolic efficiency, Lower cost Reduced raw material cost Significant energy savings (production of glucose) Improved bioconversion efficiency results in increased yield Lowered overall manufacturing costs.

30 Acknowledgements Jay Shetty & Karl Sanford Gopal Chotani & Jeff Pucci Roopa Ghirnikar S E 1 E 2 E 3 E 4 E n Continuous Biocatalytic Systems P 30

31 31 Conclusion Development of a new technology platform of producing industrial chemicals such as gluconic acid, lactic acid, ascorbic acid intermediates, 1,3-propanediol, and amino acids by using biocatalytic systems that use a less expensive feed-stock such as granular starch to more efficiently produce industrial chemicals