PART 6

Question 1

General Approach

Answer 1

1. Separation of insoluble products or components.
2. Primary isolation or concentration and removal of water.
3. Purification and removal of contaminated chemicals.
4. Product preparation.

Question 2

Factors that impact difficulty and cost of recovery

Answer 2

• Product can be biomass, intracellular or extracellular component.
• Fragile or heat sensitive.
• Concentration or titer in the broth.
• Typically recovery and purification is more than 50% of total manufacturing costs

Question 3

Insoluble Products or Components

Answer 3

• Filtration
• Centrifugation
• Coagulation and Flocculation

Question 4

Most Cost-efiectve, most common in industrial biotechnology.

Answer 4

Filtration

Question 5

_______________: traditional. Penicillin mold.

Answer 5

Rotay Vacuum precoat filter

Question 6

Cross fow ultrafiltration: _______________

Answer 6

0.02-0.2 um bacterial separations

Question 7

Cross flow microporous filtration: ___________

Answer 7

0.2-2 um for yeast

Question 8

• Used to separate solids of size. 0.1 um to 100 um using centrifugal forces.
• Being replaced by microfiltration

Answer 8

Centrifugation

Question 9

Pretreatment to centrifugation, gravity settling, or filtration to improve separation.

Answer 9

Coagulation and Flocculation

Question 10

________: formation of small flocs of cells using coagulating agents electrolytes.

Answer 10

Coagulation

Question 11

________: formation of agglomeration of flocs into settleable particles using flocculating agents, polyelectrolytes, or
CaCl2

Answer 11

Flocculation

Question 12

Used wastewater treatment processes to improve clarification.

Answer 12

Coagulation and Flocculation

Question 13

Cell Disruption - Intracellular Products
Mechanical Methods

Answer 13

• Sonication
• Bead beating
• Pressing

Question 14

Cell Disruption - Intracellular Products
Non-Mechanical methods

Answer 14

• Osmotic shock
• Freeze-thaw
• Enzymatic

Question 15

_______: disrupts cell membrane. Mostly used at the laboratory scale.

Answer 15

Ultrasound

Question 16

_________: extrude cell paste at high pressure.

Answer 16

Pressing

Question 17

__________: grind cells with glass, metal beads.

Answer 17

Bead beating

Question 18

________ is a problem with all of these methods.

Answer 18

Heat dissipation

Question 19

____________: salt differences to cause the membrane to rupture. Common.

Answer 19

Osmotic Shock

Question 20

________: Causes cell membrane to rupture. Common.

Answer 20

Freeze-thaw

Question 21

_________: Lysozyme attacks the cell wall.

Answer 21

Enzymatic

Question 22

Separation of Soluble Products

Answer 22

• Liquid-liquid extraction
• Aqueous two phase extraction
• Precipitation
• Adsorption
• Dialysis
• Reverse osmosis
• Ultrafiltration and microfiltration
• Cross-flow filtration and microfiltration
• Chromatography
• Electrophoresis
• Electrodialysis

Question 23

Separate inhibitory fermentation products from broth.
Based on the solubility difference for the compound between the phases.

Answer 23

Liquid-Liquid Extraction

Question 24

Solubility reduction at low temperatures (less than -5°C) by adding organic solvents

Answer 24

Precipitation

Question 25

_____________ - inorganic salts (NH4)2SO4 at high ionic strength

Answer 25

Salting out

Question 26

Removal of solutes from aqueous phase onto a solid phase. Chromatography is based on _____________.

Answer 26

Adsorption

Question 27

Membrane separation used to remove low molecular weight solutes. Used to remove salts from protein solutions.

Answer 27

Dialysis

Question 28

Transport occurs due to a concentration gradient driving force.

Answer 28

Dialysis

Question 29

In ______________, pressure is applied to the salt phase, causing water to move against a concentration gradient. Salt phase becomes more concentrated.

Answer 29

Reverse Osmosis

Question 30

Transport of water molecules from a high to a low concentration of pure water to salt water.

Answer 30

Osmosis

Question 31

Pressure driven molecular sieve to separate molecules of different size.

Answer 31

Ultrafiltration and Microfiltration

Question 32

______________: retained components accumulate on the filter. Gel layer formed on the filter.

Answer 32

Dead end filtration

Question 33

______________ : retained components flow tangentially across the filter

Answer 33

Cross flow filtration

Question 34

Separates mixtures into components by passing the mixture through a bed of adsorbent particles.

Answer 34

Chromatography

Question 35

Solutes travel at different speeds through the column resulting in the separation of the solutes.

Answer 35

Chromatography

Question 36

Highly specific interaction between a ligand on the particle and a component in the mixture. Often based on antibodies.

Answer 36

Affinity Chromatography

Question 37

Separation of molecules based on size and charge in an electric field.

Answer 37

Electrophorensis

Question 38

Membrane separation to separate charged molecules from a solution.

Answer 38

Electrodialysis

Question 39

Finishing Steps

Answer 39

* Crystallization * Drying