Presented By: Seemab Javed
Zunaira Irfan
Centrifugation is a process used to
separate or concentrate materials
suspended in a liquid medium.
theoretical basis
The of this technique is the
effect of gravity on particles (including
macromolecules) in suspension. Two
particles of different masses will settle in
a tube at different rates in response to
gravity.
Centrifugation is a process which involves
use of the centrifugal force for the
the
sedimentation of heterogeneous mixtures with a
centrifuge, used in industry and in laboratory
settings.
This process is used to separate two immiscible
liquids.
More-dense components of the mixture migrate
away from the axis of the centrifuge, while less-
dense components of the mixture migrate
towards the axis.
A centrifuge is a device for separating
particles from a solution according to their
size, shape, density, viscosity of the medium
and rotor speed.
In a solution, particles whose density is
higher than that of the solvent sink
(sediment), and particles that are lighter
than it float to the top.
The greater the difference in density, the
faster they move. If there is no difference in
density (isopyknic conditions), the particles
stay steady.
To take advantage of even tiny differences in
density to separate various particles in a solution,
gravity can be replaced with the much more
powerful “centrifugal force” provided by a
centrifuge.
A centrifuge is a piece of equipment that puts
an object in rotation around a fixed axis (spins it
in a circle), applying a potentially strong force
perpendicular to the axis of spin (outward).
The centrifuge works using the sedimentation
principle, where the centripetal
acceleration causes denser substances and
particles move outward in the radial
to direction.
At the same time, objects that are less dense
are displaced and move to the center. In a
laboratory centrifuge that uses sample tubes,
the radial acceleration causes denser particles to
settle to the bottom of the tube, while low-
density substances rise to the top.
It consist of two components, an electric
motor to spin the sample and a rotor to hold
tubes.
LOW SPEED CENTRIFUGE
HIGH SPEED CENTRIFUGE
ULTRA CENTRIFUGE
Most laboratories have a standard low-speed
centrifuge used for routine sedimentation of
heavy particles.
The low speed centrifuge has a maximum speed
of 4000-5000rpm.
These instruments usually operate at
temperatures with room no means of
control. temperature
Two types of rotors are used in it, fixed angle
and swinging bucket.
It is used for sedimentation of red blood cells
until the particles are tightly packed into a
pellet and supernatant is separated by
decantation.
High centrifuges are used in more
speed
sophisticated biochemical applications,
higher speeds and temperature control of the
rotor chamber are essential.
The operator of this instrument can carefully
control speed and temperature which is
required for sensitive biological samples.
Three types of rotors are available for high
speed centrifugation-fixed angle, swinging
bucket, vertical rotors
It is the most sophisticated instrument
Intense heat is generated due to high speed
thus the spinning chambers must be
refrigerated and kept at high vacuum.
It is used for both preparative work and
analytical work.
Types of
Centrifugation
Techniques
Density
Differential Ultra
gradient
centrifugation centrifugation
centrifugation
It allow separation of many or all components in
a mixture and allows for measurement to be
made
There are two forms of Density gradient
centrifugation :
Rate zonal centrifugation
Isopycnic or sedimentation
equilibrium
centrifugation
In Rate zonal centrifugation the solution
have a density gradient. The sample has a
density i.e. greater than all the layers in
the solution.
The sample is applied in a thin zone at
the top of the centrifuge tube on a
density gradient. Under centrifugal force,
the particles will begin sedimenting
through the gradient.
The particles will begin sedimenting in
separate zones according to their size
shape and density.
In this type of centrifugation , the solution
contains a greater range of densities.
The density gradient contains the whole range of
densities of the particles in the sample.
Each particle will sediment only to the position
in the centrifuge tube at which the gradient
density is equal to its own density.
In Isopycnic centrifugation separation of
particles occurs into zones on the basis of
their density differences, independent of
time.
Differential centrifugation is a common
procedure in microbiology and cytology used
to separate certain organelles from whole
cells for further analysis of specific parts of
cells.
In the process, a tissue
first homogenized to break sample the is
membranes and mix up the cell contents. cell
The homogenate is then subjected to
repeated centrifugations, each time
removing the pellet and increasing the
centrifugal force.
Svedberg coined the term “ultracentrifuge". He
was colloid chemist.
He used the ultracentrifuge to determine the
MW and subunit structure of hemoglobin ,
studies which changed the ideas concerning the
structure of proteins.
The first commercial ultracentrifuge was
produced in 1940 by SPINCO.
An important tool in biochemical research is
the centrifuge, which through rapid spinning
imposes high centrifugal forces on suspended
particles, or even molecules in solution, and
causes separations of such matter on the
basis ULTRACENTRIFUGATION of differences
in weight.
Example:
Red cells may be separated from plasma of
blood, nuclei from mitochondria in cell
homogenates, and one protein from another
in complex mixtures
COMPARISON
Separation of
solid
substances
from highly
concentrated
suspensions
Separation of
Separation
Heavy
of oily
particles and
suspensions Centrifugation large sized
with low
grains by
SS
cycloning
contents
Separation
of Oily
concentrated
sludge
Separating chalk powder from water
Removing fat from milk to produce skimmed
milk
Separating textiles
Removing water from lettuce after washing it
in a salad spinner
Separating particles from an air-
flow using cyclonic separation
The clarification and stabilization of wine
Separation of water particles from clothes
while spin-drying in washing machines
Separation of urine components and blood
components in forensic and research
laboratory
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