Centrifugation

1. Centrifugation

2. What is centrifugation?
“Centrifugation is the process where a mixture
is separated through spinning by using a
centrifuge.”

3. Centrifuge
“A centrifuge is a laboratory device that is used
for the separation of fluids based on density.”

4. Principle of centrifugation
 The centrifuge works on
sedimentation.
 Spinning sample at a high
speed
 The component of a
mixture are subjected to
centrifugal force
 Dense particle migrate
away from axis of rotation
and lighter ones toward it

5. Centrifugal motion
• Centrifugal acceleration
= rω2
• ω is the angular velocity
in rad/s
• r is the radius of
rotation
• Centrifugal force = mrω2
• m is the mass of the
particle

6.  Centrifugation method is utilized to separate the cellular debris from
the released protein.
 Example-a key role in many industrial processes, including the
production of insulin, is to separate liquid phases and solids from
each other.
 It depends on particles size, density difference between the cells and
the broth and broth viscosity.
 It is based on the behavior of particles in an applied centrifugal
field.
Principle of centrifugation

7. Pellet
Supernatant
Principle of centrifugation

8. • The principle component of a centrifuge is
rotor, which is a moving part.
• Usually rotors are fixed but they can use
multiple rotors.
• Rotors have lid to prevent samples from flying
out
Component of centrifuge
Rotor

9. Parameter of centrifuge run
The parameters for each
run include
• Duration of the spin
• The temperature
• The magnitude of spin
in term of speed

10. • Spin speed is measured as RPM.
“Revolutions per minute (RPM) is a measure
of the frequency of rotation, specifically the
number of rotations around a fixed axis in one
minute.”
• Its not the speed of the centrifuge that causes
the particle to separate from a mixture, but
the force acting on the particle
Parameter of centrifuge run

11. Parameter of centrifuge run
• The force acting on the
particle is related to the
radius of the centrifuge
rotor
• As the centrifuge rotor
have different radius
sizes, so different forces
can be applied at the
same speed.

12. Components of a centrifuge
• Centrifugal strength can also be quantified as
relative centrifugal force (RCF) generally
presented as earth’s gravitational acceleration.
• RCF can be related to RPM and be converted
into RPM as well.
• RPM is mostly used.
Parameter of centrifuge run

13. Types of small scale centrifuges
Centrifuges
Small Bench
Centrifuges
Large Capacity
Refrigerated
Centrifuges
High Speed
Refrigerated
Centrifuges
Ultra
Centrifuges
Preparative Analytical

14. Small Bench Centrifuges
• Collect small amount of
material that rapidly
sediment like yeast
cells, erythrocytes etc.
• They have maximum
relative centrifugal field
of 3000-7000 g

15. Large Capacity Refrigerated
Centrifuges
• Refrigerated rotor
chamber
• Capacity to change rotor
chambers for varying size.
• Spin speed upto 6500 g
and sediment rapidly like
erythrocytes, yeast cell,
nuclei and chloroplast.

16. High Speed Refrigerated Centrifuges
• They can generate
speed of about 60000g
• Collect micro-organism,
cellular debris, larger
cellular organelles and
proteins precipitated by
ammonium sulphate.

17. Ultra Centrifuges
Preparative
• Spin speed 600000g
• chamber is refrigerated,
sealed and evacuated.
• It is employed for separation
of macromolecules/ligand
separation of various
lipoprotein and amino acid
analysis.

18. Analytical
• It is capable of operating at
500000 g.
Ultra Centrifuges

19. Industrial centrifugation
• Centrifugation and membrane filtration are
the only techniques used for large-scale cell
harvesting.
• Centrifugation is very efficient for harvesting
yeast.
• Centrifugation can be used to recover solids
from slurries, to clarify liquids, or to clarify
solids.

20. Tubular-Bowl centrifuge
• Tubular-bowl centrifuges are sedimentation
centrifuges. They can be used to separate
both solid/liquid and liquid/liquid mixture
• High speed
• length-diameter ratio 4:8
• 15000 r.p.m
• Foaming of liquids may occur in it.

21. Working
• Feed added from the bottom of the spinning
bowl
• A distributor and baffle assembly accelerates the
incoming liquid to rotor speed.
• Then, a baffle separates the feed into its
components
• The outer layer, which consists of the heavier
components, becomes concentrated against the
wall, while the inner layer, which consists of the
lighter components, floats on top.
• Each layer then travels up the side of the bowl as
an annulus . Liquid layers are discharged from the
top of the centrifuge. Solid buildups remain in the
bowl and are recovered manually.

23. 1. Clearing of fats and waxes
2. Fractionation of blood
3. Recovery of viruses
4. Very commonly used in food,
5. Purification of lubricating industrial
6. Pharmaceutical industries
Usage

24. Disc-stack bowl centrifuge
• Disk centrifuges are the
most commonly used
centrifuges in industry
• Baffle into bowl
• Length- diameter ratio
and operational speed
is lower than tubular
type
• Manner of Discharge
according to size

25. Working
This type is common in bioprocess. The developed forces is 5000-
15000 G with minimal density difference between solid and liquid is
0.01-0.03 kg/m3.
• Large particles have higher settling velocities than small particles
• Cellular debris ends up at the outer edge of the bowl
• Soluble intracellular material passes through with the clarified
liquid
• The minimum particle diameter is 5 µm.
• The discs split the stream into a large number of very thin layers
thereby improving separation.

26. •Solids flow downwards on bottom face of disc.
•Liquid flows upwards on top face of disc.
•The close packing of the discs assists rapid sedimentation
and the solids then slides to the edge of the bowl.
•Smaller in size compared with a bowl without discs for
given throughput.
•Requires in-situ steam sterilization and the discs
arrangement makes this type of centrifuge laborious to
clean.
•Feed rate 45-1800 dm³ min¯¹.

27. Usage
• used in the brewing
industry.
• refine oil and fat.
• as cream separation
and the concentration
of butterfat milk

29. Bowl Basket Centrifuge
Useful for separating mould mycelia or crystalline
compounds.
• Perforated & Lined with a filter bag of nylon, cotton.
• Normally operated at speed of up to 4000 rpm for feed rate
50-300 dm³ min¯¹.
• Continuous feed is used & when the basket is filled with the
filter cake it is possible to wash the cake before removing it.
• It may be considered to be a centrifugal filter and holding
capacity is 50-300 dm³

30. Bowl Basket Centrifuge

31. Merits and demerits
• The clarification efficiency of centrifugation process is affected by
harvest parameters such as centrifuge feed rate, G-force, bowl
geometry, operating pressure, discharge frequency.
• total cell density and culture viability during the culture process and
harvest will also affect separation performance.
• Particles of submicron size cannot be removed in the centrifuge,
thus increases the burden on subsequent depth filtration.
• Disk stack continuous centrifuge removes cell debris from viable
cells and liquid phase but some cells gets disrupted during process
especially feedstock with low viability culture fluid

32. Applications
• Centrifuges are basically employed for separation of whole large cells from
heterogeneous cell mixture.
• Can be used to separate viable cells from cell debris using disc stack
centrifuge.
• clarification (removal of solid impurities from milk prior to pasteurization)
• skimming (separation of cream from skim milk)
• standardizing
• whey separation (separation of whey cream (fat) from whey)
• Collection of starch
• The clarification of process liquor e.g. insulin
• Removal of bacteria in bacterial enzyme preparation
• Removal of dirt and water from oil. e.g. olive oil
• Recovery of lanolin in a the wool scouring process

33. • bactofuge treatment (separation of bacteria from milk)
• butter oil purification (separation of serum phase from
anhydrous milk fat).
• Disc-stack centrifuges used by some companies in
the oil sands industry to separate small amounts of
water and solids from bitumen.
• Large industrial centrifuges are commonly used
in water and wastewater treatment to dry sludges. The
resulting dry product is often termed cake, and the
water leaving a centrifuge after most of the solids have
been removed is called centrate.
• Centrifuge are also used in whole blood separation into
different components.
Applications

34. Other Industrial centrifuges
Tubular bowl Chamber Disc

35. • Bacteria
– Small cell size
– Resilient
• Yeast cells
– Large cells
– Resilient
• Filamentous fungi
– Mycelial
– Resilient
• Cultured animal cells
– Large cells
– High speed required
– Low cell damage
– Lower speed required
– Low cell damage
– Lower speed required
– High water retention in pellet
– Very susceptible to damage
Centrifugation properties of different cell types