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Biomanufacturing 2016

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Presentation by Dr. Sonia Wallman at Summer Fellows Forum 2016

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Biomanufacturing 2016

  1. 1. The Bioeconomy Bioproducts Development, Production and Analysis Education and Training Skilled Technicians Needed Sonia Wallman, PhD NBC2 and c3bc Biomanufacturing Consultant Bio-Link Summer Fellows Berkeley, CA June 8, 2016
  2. 2. History of Northeast Biomanufacturing Collaborative and Center Since 2003, working with biomanufacturing employers to define the skills, knowledge and abilities (SKAs) needed by technicians to develop, produce and analyze bioproducts. Starting with biopharmaceutical production and analysis in 2003, 2007 and 2013; with biofuels in 2013 and finishing up with industrial biotechnology in 2015 (and 2016), we will synthesized these inputs from local (and global) employers into a single set of SKAs to educate and train technicians to support the development and growth of the bioeconomy. The synthesis of these inputs on what biotechnicians need to know and do is driving the production of hands-on and online curriculum and instructional materials, including laboratory manuals, textbooks and virtual industrial modules to facilitate deep understanding of what is required by technicians to biomanufacture bioproducts, creating an awareness of the many career pathways in biomanufacturing.
  3. 3. Biofuels Workforce Summit May 23, 2013 in Honolulu, HI
  4. 4. Bioproducts Upstream Processes BIOPHARMACEUTICALS, INDUSTRIAL ENZYMES, GREEN CHEMICALS, BIOFUELS • DEFROST CRYOVIAL OF CELLS FROM WORKING CELL BANK • INOCULATE MEDIA WITH CRYOVIAL OF CELLS • SCALE-UP • HARVEST • CENTRIFUGATION • FILTER MEDIA (DEPTH FILTRATION AND STERILE FILTRATION) • HOLD BIODIESEL FEEDSTOCKS: WASTE AND AG • COLLECT OIL PRODUCTS SUCH AS WASTE FRY OIL AND PLANT OILS • TRANSPORT • STORE • HEAT • FILTER • HOLD
  5. 5. Bioproducts Downstream Processes BIOPHARMACEUTICALS, INDUSTRIAL ENZYMES, GREEN CHEMICALS, BIOENERGY • DEPTH FILTRATION • LIQUID COLUMN CHROMATOGRAPHY (ION EXCHANGE, AFFINITY, HYDROPHOBIC INTERACTION=HIC, SIZE EXCLUSION=GEL FILTRATION) • TANGENTIAL FLOW FILTRATION • STERILE FILTRATION • COLLECT BATCH • FORMULATE, FILL AND FINISH (for biopharmaceuticals) • DISTRIBUTE BIODIESEL FEEDSTOCKS: WASTE AND AG • TRANSESTERIFY OIL WITH ALKALINE CATALYST AND METHANOL INTO FAME (FATTY ACID METHYL ESTERS=BIODIESEL) • SEPARATE INTO FAME-MeOH AND GLYCEROL-MeOH LAYERS • DISTILL MeOH FROM BOTH LAYERS – RECYCLE MeOH • WATER WASH FAME LAYER AND FILTER OR DISTILL AT HIGH VACUUM • DISTRIBUTE BIODIESEL • SELL GLYCEROL CRUDE OR PURIFIED BY HIGH VACUUM DISTILLATION
  6. 6. Pacific Biodiesel (Big Island Biodiesel) 5M gallon/year Fry Oil to Biodiesel Facility Collected fry oil into heated tanks, oil strained, placed into funnel shaped tanks where sodium hydroxide cleaves glycerol from triglyceride and methanol is used to add a methyl group to the free fatty acids to produce FAME or fatty acid methyl esters or biodiesel. Glycerol to bottom of funnel and biodiesel is drawn off and dewatered. BIB follows this by distillation, producing crystal clear biodiesel.
  7. 7. Vessels and Pumps Vessels store/contain liquids, solids, or gasses involved in bioproducts production, such as bioreactors, tanks, drums, cylinders, and bins. Pumps are used to move liquids through the system, such as into or out of a vessel. There are many different types of pumps, but they all use either rotational or centrifugal force to induce flow in liquids. Tank holding Water For Injection (WFI) Peristaltic Pump
  8. 8. Compressors, Fans and Motors
  9. 9. Piping and Valves Piping is cylindrical tubes made of varying lengths and materials such as 316L stainless steel, plastics, and iron that carry materials (usually in liquid or gas form) from one location to another. Valves are devices that are attached to piping or equipment to control the flow of liquids or gases. There are a wide variety of valve types that can start/stop or throttle (open or close in increments) the flow. Valves are vital to process control. They can be remotely opened or closed by computers to control variables such as flow or level. Piping and valves are the most common pieces of equipment found in a biomanufacturing facility.
  10. 10. Piping & Instrumentation Diagrams P&ID A drawing or diagram which shows the piping of the process flow together with the installed equipment and instrumentation, including computer process controllers.
  11. 11. Biomanufacturing is Process Controlled Big Island Biodiesel 5M Gallon/Year
  12. 12. Quality Control Laboratory BIOPHARMACEUTICALS, INDUSTRIAL ENZYMES, GREEN CHEMICALS, BIOFUELS BIODIESEL TriState Biodiesel in Bridgeport, CT Built Using Recycled Equipment Lonza Biopharmaceuticals in Portsmouth, NH
  13. 13. Quality Control Testing BIOPHARMACEUTICALS, INDUSTRIAL ENZYMES, GREEN CHEMICALS, BIOFUELS BIODIESEL • Cell Count (growth rate) • Analyate Analysis (metabolism) • ELISA (protein ID, amount, and activity) • SDS PAGE (protein ID and amount) • TLC (thin layer chromatography) • HPLC plus detector (protein, lipid, carbohydrate IDs and amount of each) • Stability Testing • Genome Sequencing • PCR Identification
  14. 14. Scale-Up Using Aseptic Technique Shake Flask Inoculation using Biosafety Cabinet Class 100
  15. 15. Upstream Processing Count Cells
  16. 16. Scale-Up to Stainless Steel Biopharmaceutical Production Bioreactors
  17. 17. Inoculation of Microalgae Seed Bioreactor
  18. 18. Scale-Up of Microalgae to Open Pond Production Solar Bioreactors
  19. 19. Sludge Continuous Centrifuge Media Out Cells + Media In Harvest Protein Rich Media by Continuous Centrifugation
  20. 20. Harvest by Batch Centrifugation (1000 liters of Chlorella vulgaris culture) Process controlled centrifuge to separate cells (microalgae) from media Centrifuge drum and fins for separation of microalgae from media
  21. 21. Cellular paste is collected below the blades of the centrifuge Microalgae are harvested; media is reprocessed into useable water
  22. 22. Depth Filtration to Remove Cells
  23. 23. Depth Filtration Housings, Piping, Valves
  24. 24. Tangential Flow Filtration (TFF) for Separation and Concentration Using TFF with the right cut off filters, the biologic of interest can be separated from other thing For instance HSA has a molecular weight of 69KD. To make sure that the protein of interest is retained, a 10KD cut-off filter is used. After ultrafiltration, we can diafilter, adding the phosphate buffer at pH 7.1 that we will also use to equilibrate our affinity column to prepare it for affinity chromatography of HSA.
  25. 25. Tangential Flow Filtration Separates and Concentrates
  26. 26. TFF for Biopharmaceutical Protein Concentration or Buffer Exchange
  27. 27. TFF for Microalgae Concentration Photo taken at the ATP3 Large Scale Algal Cultivation, Harvesting and Downstream Processing Hands-On Training at AZCATi (Arizona Center for Algal Technology and Innovation) in Mesa, AZ From Open Pond To Centrifuge
  28. 28. Chromatography for Purification and Quality Control Chemistry of the Protein, Lipid or Carbohydrate of Interest Separation Science Used for Downstream Processes and for Quality Control Chemistry Monitoring of both Upstream and Downstream Processes Stationary Media and Moving Buffer System Result in Competition and Separation
  29. 29. Liquid Column Chromatography
  30. 30. Large Scale Chromatography Column Packed with Beads (Media) for Separation
  31. 31. Chromatography System Components • Chromatography Column and Media (Beads) • Peristaltic Pump, Pipes and Valves • Mixer for Buffers • (UV) Detector to identify the product of interest
  32. 32. Liquid Column Chromatography Process PURGE Air from Column using Equilibration Buffer PACK Column with Media (e.g. ion exchange, HIC, affinity or gel filtration media) EQUILIBRATE Column with Equilibration Buffer LOAD Column with Protein of Interest in Equilibration Buffer WASH Column with Equilibration Buffer ELUTE Protein of Interest from Column with Elution Buffer of high or low Salt or pH REGENERATE Column or Clean and Store (NaOH)
  33. 33. Chromatogram WASH=Flow Through ELUTE=Eluate
  34. 34. Skills, Knowledge and Attributes for Biomanufacturing Technicians Specific SKAs for Biomanufacturing Time Spent/Importance of Each Production and Processing 22 Operation and Control 7.6 Quality Control Analysis 5.6 Computers and Electronics 3.9 Operation Monitoring 3.2 Equipment Selection 2.3 Equipment Maintenance 1.3 Repairing 1.1 Engineering and Technology 0.7 Design 0.7 Systems Analysis 0.6 Technology Design 0.1 Installation 0.1
  35. 35. Lab Science, Medical Devices and Biomanufacturing Common Core Critical Work Functions • Maintain a Safe and Productive Work Environment • Provide Routine Facility Support • Perform Measurements/Tests/Assays • Comply with Applicable Regulations and Standards • Manage and Communicate Information • Perform Mathematical Manipulations
  36. 36. Online Biomanufacturing Resource www.biomanufacturing.org/ CHAPTER 1 INTRODUCTION ENGINEERING UNIT OVERVIEW CHAPTER 2 FACILITIES CHAPTER 3 METROLOGY CHAPTER 4 VALIDATION CHAPTER 5 ENVIRONMENTAL, HEALTH, AND SAFETY (EHS) CHAPTER 6 OPERATIONAL EXCELLENCE QUALITY UNIT OVERVIEW CHAPTER 7 QUALITY ASSURANCE CHAPTER 8 MICROBIOLOGICAL CONTROL CHAPTER 9 QUALITY CONTROL BIOCHEMISTRY PRODUCTION UNIT OVERVIEW CHAPTER 10 UPSTREAM PROCESSING CHAPTER 11 DOWNSTREAM PROCESSING CHAPTER 12 PROCESS DEVELOPMENT APPENDIX A MASTER GLOSSARY Michael Cicio - Today’s Biologics: From Bench Top to Bottle. Tags: Keynote Speeches, Bioman 2006
  37. 37. Virtual Downstream Processing Monoclonal antibody proteins (or mAbs) are the single largest class of recombinant biological drugs to date and represent about a third of the total biopharmaceutical market. The recent success of monoclonal antibodies for a wide range of disease therapies has led to the development of industrial production operations that manufacture pharmaceutical-grade mAbs both efficiently and safely. The following modules introduce a typical mAb bioprocessing workflow, detailing the equipment and processes used in biomanufacturing within a regulated environment. http://faculty.mc3.edu/downstreamprocessing/stor y.html
  38. 38. NBC2 Biofuels Curriculum – Textbook
  39. 39. NBC2 Biofuels Curriculum – Laboratory Manual Table of Contents ► LAB 1: INTRODUCTION TO BIOFUELS PRODUCTION ........................ 05 ► LAB 2: BIOFUEL ENZYME KIT ............................................................. 25 ► LAB 3: USING BACTERIA TO PRODUCE BIOHYDROGEN ................... 39 ► LAB 4: ESTABLISHING AND SCALING UP MICROALGAE CULTURE FOR BIOFUELS PRODUCTION SOP ........................................ 55 ► LAB 5: CONVERSION OF MICROALGAE LIPIDS TO FATTY ACID METHYL ESTERS (FAME) SOP.................................................. 59 ► LAB 6: MICROALGAE LIPID EXTRACTION AND SEPARATION BY THIN LAYER CHROMATOGRAPHY SOP.................................... 63 ► LAB 7: USING ALGAE TO PRODUCE BIODIESEL................................ 69 ► LAB 8: EXTRACTION OF ALGAL OIL .................................................. 83 ► LAB 9: PRODUCTION OF BIODIESEL BY THE FOLCH METHOD ......... 89 ► LAB 10: BIODIESEL ANALYSIS USING HPLC ........................................ 103 Biofuels Productio n and Anal ysis Laboratory Manual Primary Author: Elmar Schmid, Ph.D. Associate Professor, Biotechnology Department Miracosta College, Oceanside, California 2nd Editio n Apr il 2014 Part One
  40. 40. Career Pathway Data Collection: Job Titles of Some Program Graduates • Environmental Monitoring Technician • Laboratory Automation Scientist • Laboratory Technician • Manufacturing Associate I • Manufacturing Tech I • Manufacturing Tech 2 • Process Improvement Leader • Quality Assurance Associate • Quality Analyst • Quality Control Analyst • Quality Operations Product Release Coordinator l • Research Associate • Research Scientist • Research Technician • Senior Laboratory Technician • Senior Scientist • Technical Writer • Technical Writer/Scientist III

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