This document summarizes key techniques used in bioprocessing, including media formulation, sterilization methods, inoculum development, fermentation monitoring, product recovery techniques like centrifugation and chromatography, and final processing steps like drying and packaging. The growth medium is designed to support microbial growth and provide nutrients. Sterilization kills microbes using heat, chemicals, or radiation. Inoculum development prepares cultures for large-scale growth. Fermentation is monitored to optimize conditions. Recovery purifies and isolates the desired product. Drying stabilizes products for storage and transportation.

1. Techniques used in
Presented by
Bilqees Fatima Ph.D.
Presented to
Dr. Muhammad Mohsin Javed
INSTITUTE OF INDUSTRIAL BIOTECHNOLOGY
G.C. UNIVERSITY Lahore
PAKISTAN

2. Media
formulation
Synthesis
Product
recovery
Final
polishing
Organism
Waste
Market
Raw
material
Is the use of biological materials (organisms, cells,
organelles, enzymes) to carry out a process for
commercial, medical or scientific reasons.

3. A growth medium or culture medium is a liquid or gel
designed to support the growth of organisms or cells
A bioprocess medium design principles take care of the
following in general
Source(s) of energy
Source(s) of carbon
Source(s) of nitrogen
Source(s) of minerals
Source(s) of growth factors
Source(s) of specific nutrient needs
Source(s) of oxygen environment
Raw material is used for the formulation of growth medium

4. Weigh out Heat to dissolve
Raw
Material
Sugarcane Bagasse
Treated Sugar-cane
Bagasse
Chemical sources
Mix in buffer
Biological sources

5. Methods of Sterilization
Chemicals (phenols, alcohols, aldehyde, helogens,
detergents, ethylene oxide, heavy metals)
Radiations (UV, γ-rays)
Filtration and
Heat
Is the process by which all living cells, viable spores, viruses
and viroids are either destroyed or removed from an object
or habitat.

6. Required high temperature and longer
exposure time
Hot air oven (160 C for 60 min)
Infra red conveyer (180 C for 7 min)
(Metal instruments and glass syringes)
1. DRY HEAT
Incineration
Hot air ovens

7. 2. MOIST HEAT
Autoclave
Sealed chamber that furnished
pressurized steam for sterilization
Sterilizes
Instruments
Syringes
Needles
Other materials
Steam
pressure
(lb/inch2)
Temperature
(°C)
Holding time
(min)
15
20
30
121
126
134
15
12
3
Autoclave

8. The preparation of a population of organisms from a
dormant stock culture to an active state of growth that is
suitable for inoculation in the final production stage is
called inoculum development
Organism,
cells
The process of inoculum development

9. Sample Analysis
•pH
•DO
•Sugar
•Ammonia
•Phosphate
•Sulphate
•Products
•Precursors
•Contamination
Pressure probe
Level probe
pH probe
Temp. probe
DO probe
Antifoam
Acid/Base
Cooling
Air/agitation
Sugar/Oil
feed

10. Foam separation
Filtration
Centrifugation
Cell disruption
Fractional
Precipitation
Dialysis
Ultrafiltration
Electrophoresis
Distillation
Reverse osmosis
Diafiltration
Precipitation
Gel Filtration
Aqueous two phase
extraction
Sterile filtration
Crystallization
Lyophilization
Drying Packaging
Product recovery encompasses all processes following the
fermentation by which the desired product is Isolated,
Purified and Formulated for different end uses.

11. It may be possible to make some
materials surface active by the
application of surfactants such as
long chain fatty acids
amines and
quaternary ammonium
compounds
Depends upon differences in
surface activity materials
Collectors
Colligends

12. to obtain some products or to remove some impurities
directly from the broth
use the technique after a crude cell lysate has been
obtained.
Typical agents used in precipitation render the
compound of interest insoluble, are include:
• Acids and bases
• Salts
• Organic solvents
• Non-ionic polymer
• Polyelectrolytes
• Affinity precipitatants

13. Microorganisms and other similar
sized particles can be removed from
a broth
Vg = d2g (P-L) /18 
Vc = d2r (P-L) /18 
1. Basket bowl centrifuge
2. Tubular bowl centrifuge
3. Solid bowl centrifuge
4. Disc bowl centrifuge
5. Multichamber centrifuge
Types of centrifuges in their manner of liquid solid discharge
Stoke’s law

14. to release cellular contents a number of
methods for cell disintegration have been
developed
i. Detergents
ii. Osmotic shock
iii. Alkali treatment
iv. Enzyme treatment
Chemical Methods
Physical Methods

15. This method is used
while separating two or
more immiscible liquids
with different densities.
Mixture is taken in a
separating funnel.
Allowed the mixture to
stand for some time.
This separates the
liquids into layers.

16. It involves the treatment of organic solvent with aqueous solvent
containing desired solute
The solute which is preferential soluble in aqueous solvent, more
solute would be present in aqueous solvent at equilibrium
However, the use of organic solvents has limited application in
the bioprocessing of sensitive bioproduct.

17.  Phase separation
occurs when
hydrophilic polymers
are added to an
aqueous solution, and
concentration exceed a
certain value, two
immiscible aqueous
phases are formed.
System Example
Non-ionic polymer
/non-ionic
polymer /water
Polyethylene glycol
/ Dextran
Polyelectrolyte
/non-ionic
polymer /water
Sodium
carboxymethyle
cellulose
/polyethylene glycol
Polyelectrolyte
/Polyelectrolyte/
water
Sodium Dextran
sulphate
/sodium carboxymethyl
cellulose
Polymer
/low molecular
weight
component/wate
r
Dextran
/propyl alcohol
A large variety of natural and
synthetic hydrophilic
polymers are used today

19. Dialysis Ultrafiltration Nanofiltration
•Separation affected by the use of thin, selective, semi-
permeable barriers
• Ceramic and metal filters also perform similar task
•Dialysis membrane is usually
made of cellulose acetate
•Pore size = 1-20 nm in
diameter
low-pressure (10-100psi)
cross-flow membrane process
for separating colloidal and
suspended particles in the
range of 0.05-10 microns.
Reverse
Osmosis
Microfiltration

20. Ultrafiltration is a
selective fractionation
process utilizing
pressures up to 145
psi (10 bar).
It concentrates
suspended solids and
solutes of molecular
weight greater than
1,000.
UF is widely used in
– fractionation of milk and
whey,
– protein fractionation.

21.  Reverse osmosis is a high-
pressure, energy-efficient technique
for dewatering process streams,
concentrating low-molecular-weight
substances in solution, or purifying
wastewater.
Pressure requirement
•2–17 bar (30–250 psi) for
fresh and brackish water
•40–70 bar (600–1000 psi)
for seawater

22. Nanofiltration is a special
process selected when RO
and UF are not the ideal
choice for separation.
NF can perform separation
applications that are not
otherwise economically
feasible, such as
demineralization, color
removal, and desalination.
Nanofiltration removes most
of the organic molecules,
nearly all viruses, most of
the natural organic matter,
polyvalent ions and a range
of salts.

23. Membrane processes are not fundamentally
different, except in terms of size of molecules they
retains

24. Mixer
Buffer Select
Column
UV Detector
Injector Valve
Conductivity Meter
Peristaltic
Pump

25. Creates a gentle
squeezing action to
move fluid through
flexible tubing
system of
chromatographic
apparatus.

27. Ion Exchange Chromatography relies on charge-charge interactions
between the protein of interest and charges on a resin (bead).
• Once the protein of interest is
bound, the column is washed
with equilibration buffer to
remove unattached entities.
• Then the bound protein of
interest is eluted off using an
elution buffer of increasing ionic
strength or of a different pH.
• Either weakens the
attachment of the protein of
interest to the bead and the
protein of interest is bumped off
and eluted from the resin.

28. Affinity chromatography
separates the protein of
interest on the basis of a
reversible interaction
between it and its antibody
coupled to a
chromatography bead (here
labeled antigen) .
To elute the protein of
interest from the affinity
beads, the interaction can
be reversed by changing
the pH or ionic strength.

29. Adsorption
takes place in
high salt and
elution in low
salt
concentrations.

30. Crystallization is a process where solid particles of specified size and
shape are formed from a homogeneous phase.
Crystallization process consists of two steps
– Nucleation
– Crystal growth
Super saturated state consists of three zones
– Metastable zone
– Intermediate zone
– Labile zone
Crystallization may be initiated by
– Homogenous nucleation
– Hetrogenous nucleation

31. Drying
Drum dryer Spray dryer
• Drying removes the solvent, mainly water, from the
desired product and in most cases stabilizes the product
making it amenable for storing, packaging or formulation.
• Drying takes place by movement of water vapour from the
saturated surface through to the stagnant air film in to the
main stream of drying air.

32. • used for more temperature
stable products
• solid is in contact with
the heating surface for 6-
15 seconds
 heat transfer
coefficients are generally
between 1-2 kWm-2 K-1

33. •Most widely used for drying
of biological materials when
the starting material is in the
form of a liquid or paste.
•The droplets then fall into a
spiral stream of hot gas at
150-250°C.
•The high surface area:
volume ratio of the droplets
results in a rapid rate of
evaporation and complete
drying in a few seconds.

34. • Pressure gradient 0.1-2.0 torr
• A heating system to provide
latent heat of sublimation
The latent heat of sublimation of
ice is 2838 kJ/kg
In a typical phase diagram, the
boundary between gas and liquid
runs from the triple point to the
critical point. Freeze-drying (blue
arrow) brings the system around
the triple point, avoiding the direct
liquid-gas transition seen in
ordinary drying (green arrow).
•Foodstuffs, pharmaceuticals and
biological materials which are heat
sensitive even to low or moderate
temperatures are freeze-dried.
•The method involves freezing of
the material by exposure to cold air
followed by sublimation of ice in
vacuum from the frozen state to
produce a dried product.

35. Product viewable single
shelf freeze-dryer. Production freeze-drier.
Development freeze-dryer
Benchtop manifold
freeze-drier
Unloading trays of freeze-dried material from a
small cabinet-type freeze-drier

36. Freeze-dried coffee, a form of instant coffee