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Chapter 5
Downstream Processing
Downstream processing, what and why
Downstream processing is any treatment
of culture broth after fermentation to
concentrate and purify products.
2
Separation Processes
The challenge
•Low product concentrations
•Large number of impurities
•Thermolabile bioproducts
An ideal bioseparation process
should combine high throughput
with high selectivity, and should
ensure stability of product.
Nutrients
Oxygen
Genetic modification
Microbes
Fermentation
Happy growing
Drug producing
Downstream Processing
Remove cells/debris
Concentrate and
Purify the product
Smash the microbes
Formulate
product
Kill the microbes
Market
Biological products
Product
Nature of bioseparation required
Alcoholic beverages:
Beer, wine, spirits
Clarification, distillation
Organic acids:
Acetic acid, citric acid
Precipitation, filtration, adsorption
Vitamins:
Vitamin C, vitamin B12
Precipitation, filtration, adsorption
Amino acids:
Lysine, glycine,
Precipitation, filtration, adsorption
Antibiotics:
Penicillins
Precipitation, filtration, adsorption
Enzyme,Polysaccharides
Filtration, precipitation, centrifugation,
adsorption, chromatography
Distillation
Separating components with different
volatilities. Such as organic solvents ,
volatile flavors and aromas.
Distillation
Stages in Downstream Processing
 Removal of insoluble's
 Product Isolation
 Product Purification
 Product Polishing
1. 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
3. Purification (chromatography, ultrafiltration)
Removing components with properties similar to
those of the products
Highly selective
4. Product preparation (crystallization, drying)
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.
Typical Operations
Filtration
A mechanical operation used for the
separation of solids from fluids.
Type of filtration unit
Plate and frame filter
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Rotary-drum vacuum filter
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
Centrifuges
t
t
Decanter centrifuge
Coagulation and Flocculation
Coagulation:
Forming semisolid lumps in a liquid.
Flocculation:
Forming woolly cloudlike aggregations.
Product Isolation
Reducing the volume of material to be
handled and concentrating the product.
Precipitation
Formation of a solid in a solution during a
chemical reaction.
Solid formed is called the precipitate and
the liquid remaining above the solid is
called the supernatant.
1. Isoelectric Precipitation
pH
GA+
GA±
GA–
GA=
2
3.22
7.0
>12
pI
2. Organic solvent precipitation
By adding an organic solvent to an aqueous
fermentation broth, the dielectric constant will
decrease causing the solubility to decrease.
Example
Xanthan gum is recovered from an aqueous
fermented broth containing the gum by
adding to the broth organic solvent.
3. Insoluble salt precipitation
Usually used in extraction of
amino acid and organic acid.
e.g.
Citric acid extraction and purification process
 Heating (destroy the microorganisms )
 Neutralizition (forming calcium citrate )
 Acidifying (liberate the citric acid from its salt )
Disadvantages
high chemical cost
relatively complex technology
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
30
Organic Resin Groups
SO3 H
Linkage group
CH2 Cl
Chloride
Cation exchange
CH2 N(CH3 )3 Cl
Anion exchange
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.
Membrane Separation
Function : clarification and sterilization
Advantages
 No phase change
 Energy efficient
 A higher quality product
 Environmentally friendly
 Easy to scale-up
Limitations
 Long-term reliability not proven
 Excessive pretreatment
 Concentration polarization and membrane fouling
 Replacement of membranes
Membrane Fraction Technologies
Nanofiltration
(200 - 700 MW)
Reverse Osmosis
(<200 MW)
Ultrafiltration
(2000 - 50,000 MW)
Microfiltration
(100,000 MW)
Semi-permeable
Membrane
Wine Filtration –Why?
Filtration Objectives/Purposes Include
Remove insoluble contaminants
Improve clarity, brightness
 Extend shelf life
Electrodialysis
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
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
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
Crystallization Process
Liquid
Mixture
Generation of
Supersaturation
:Driving force
Final Product
Nucleation:
Birth of
Solid Phase
Crystal Growth
Supersaturation refers to a state in
which the liquid (solvent) contains
more dissolved solids (solute) than
can ordinarily be accomodated at
that temperature.
Solubility & Supersolubility Diagram
Generation of supersaturation
 Solvent evaporation
 Solution cooling
 Altering pH
Crystallization
- by evaporation of saturated solution
Crystallization
- by cooling saturated solution
Concentration
 Filtration
 Centrifugation
 Reduced pressure concentration
Drying
Drying involves the transfer of heat to the
wet material and removal of the moisture as
water vapor.
Usually, this must be performed in such a
way as to retain the biological activity of the
product.
Drying Methods
（1） Atmospheric drying
（2）Vacuum drying (Decompression )
（3） Freeze-drying (lyophilization)
（4） Spray drying
lyophilization
Freezing the material
Reducing the surrounding pressure and
adding enough heat to allow the frozen
water in the material to sublime directly
from the solid phase to gas.
Review Questions
1. Show the importance and main procedures of
the downstream processing of fermentation. State
the principles and the characteristics of every
separation methods.
2. Tell the difference between freeze drying and
vacuum drying.