Chapter 5 Downstream Processing

1. Chapter 5Downstream Processing

2. Downstream processing, what and why Downstream processing is any treatment of culture broth after fermentation to concentrate and purify products.

3. Separation Processes

4. The challenge • Low product concentrations • Large number of impurities • Thermolabile bioproducts

5. An ideal bioseparation process should combine high throughput with high selectivity, and should ensure stability of product.

6. Fermentation Drug producing Happy growing Nutrients Oxygen Genetic modification Microbes Downstream Processing Smash the microbes Remove cells/debris Kill the microbes Concentrate and Purify the product Formulate product Market

7. Biological products

8. Distillation Separating components with different volatilities. Such as organic solvents , volatile flavors and aromas.

9. Distillation

11. Stages in Downstream Processing • Removal of insoluble's • Product Isolation • Product Purification • Product Polishing

12. Cell removal (filtration, centrifugation , • coagulation and flocculation) • 2. Primary isolation (adsorption, precipitation) • Removing components with properties significantly • different from those of the products • Large volume, relatively non selective

13. 3. Purification (chromatography, ultrafiltration) • Removing components with properties similar to • those of the products • Highly selective • 4.Product preparation (crystallization, drying)

14. Removal of insoluble's Capture of the product as a solute in a particulate-free liquid Example Separation of cells, cell debris or other particulate matter from fermentation broth containing an antibiotic.

15. Typical Operations Filtration A mechanical operation used for the separation of solids from fluids.

16. Type of filtration unit Plate and frame filter

17. Rotary-drum vacuum filter

18. Centrifugation • Use of the centrifugal force for the separation of mixtures • More-dense components migrate away from the axis of the centrifuge • Less-dense components migrate towards the axis

19. Centrifuges t t

20. Decanter centrifuge

21. Coagulation  and  Flocculation Coagulation: Forming semisolid lumps in a liquid. Flocculation: Forming woolly cloudlike aggregations.

22. Product Isolation • Reducing the volume of material to be handled and concentrating the product.

23. Precipitation • Formation of a solid in a solution during a chemical reaction. • Solid formed is called the precipitateand the liquid remaining above the solid is called the supernatant.

24. 1. Isoelectric Precipitation GA+ GA± GA– GA= pH 2 3.22 7.0 >12 pI

25. 2. Organic solvent precipitation By adding an organic solvent to an aqueous fermentation broth, the dielectric constant will decrease causing the solubility to decrease.

26. Example Xanthan gum is recovered from an aqueous fermented broth containing the gum by adding to the broth organic solvent.

27. 3. Insoluble salt precipitation Usually used in extraction of amino acid and organic acid.

28. e.g. • Citric acid extraction and purification process • Heating (destroy the microorganisms ) • Neutralizition (forming calcium citrate ) • Acidifying (liberate the citric acid from its salt )

29. Disadvantages • high chemical cost • relatively complex technology

30. Ion Exchange Resins • Resins • Organic or inorganic polymer used to exchange cations ( positive ions)or anions ( negative ions)from a solution phase • General Structure • Polymer backbone not involved in bonding • Functional group for complexing anion or cation

31. Organic Resin Groups Linkage group Cation exchange Chloride Anion exchange

32. e.g. Purification of Lactic Acid from Fermentation Broths by Ion-Exchange Resins First, a strong cation resin was used to reduce the broth pH and remove the cations present in the solution; Then the acidified broth was purified by an anion exchanger.

33. Membrane Separation Function : clarification and sterilization

34. Advantages • No phase change • Energy efficient • A higher quality product • Environmentally friendly • Easy to scale-up

35. Limitations • Long-term reliability not proven • Excessive pretreatment • Concentration polarization and membrane fouling • Replacement of membranes

36. Microbes + fat Lactose Protein Water Salts Nanofiltration (200 - 700 MW) Reverse Osmosis (<200 MW) Ultrafiltration (2000 - 50,000 MW) Microfiltration (100,000 MW) Membrane Fraction Technologies

37. Semi-permeable Membrane

38. Wine Filtration –Why? • Filtration Objectives/Purposes Include • Remove insoluble contaminants • Improve clarity, brightness • Extend shelf life

39. Electrodialysis

40. Product Polishing • Final processing steps which end with packaging of the product in a form that is stable, easily transportable and convenient • Crystallization, concentration and drying are typical unit operations

41. Crystallization • Formation of a crystalline phase from a parent phase, e.g. solution • One of the oldest and most important unit operations, e.g. extracting salt crystals from sea water

42. Definition of Crystal • A substance in which the constituent atoms, molecules, or ions are packed in a regularly ordered, repeating three-dimensional pattern. • The distinction between a crystal and an amorphous solid is that between order and disorder over large distances

44. Crystallization Process Generation of Supersaturation :Driving force Nucleation: Birth of Solid Phase Liquid Mixture Final Product Crystal Growth

45. Supersaturation refers to a state in which the liquid (solvent) contains more dissolved solids (solute) than can ordinarily be accomodated at that temperature.

46. Solubility & Supersolubility Diagram

48. Generation of supersaturation • Solvent evaporation • Solution cooling • Altering pH

49. Crystallization- by evaporation of saturated solution

50. Crystallization- by cooling saturated solution