Theory of Centrifugation

In centrifugation, centrifugal force is used as the driving force for the

separation of particles. Centrifugal force is replacing a gravitational
force which is responsible for the sedimentation of two particles. Thus
centrifugation is useful when the ordinary filtration methods do not
apply to the separation of particles. When two particles having different
sizes but same densities are suspended in any liquid medium then they
may not be able to separate by a simple filtration method. In such kind
of cases, centrifugation method is useful.

The particle size above 5-micrometre sediment at the bottom with the
help of gravity but the particles having a size less than 5 micrometres,
start Brownian motion and do not sediment because of the gravity that’s
why they require the centrifugal force to separate properly. The
centrifugal force causes the denser dense particle direction of the radical
where writer practical moves to the centre. The ratio of the force acting
on radical the direction to the gravitational force is the centrifugal
effect.

Process of Centrifugation

1) The centrifuge consists of a container in which a mixture of solid and

liquid or two liquids is placed. Then this container is rotated at very
high speed. When this container is rotated at high-speed mixture
separated into its constituent parts by the action of centrifugal force on
their density.

2) The solid or liquid of higher specific gravity is thrown out word with
greater force.

Types of Centrifugation Techniques

1) Density gradient centrifugation
It allows separation of many or all components in a mixture and allows
for measurement also. There are 2 forms of density gradient
centrifugation one is rate zonal centrifugation and the second is I saw
pyknic or sedimentation equilibrium centrifugation.

A)Rate zonal centrifugation – In rate zonal centrifugation the solution

has a density gradient. The sample has a density therefore greater than
all the layers 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 segmenting through the gradient. The particles will
begin segmenting in separate zones according to their size, shape and
density.

B) Isopycnic or sedimentation equilibrium centrifugation – In this

type of centrifugation, the solution contains a greater range of densities.
The density gradient contains the whole range of densities of the
particle in the sample pool stop each particle with sediment only to the
position in the centrifuge tube at which the gradient density is equal to
the phone density. In sedimentation centrifugation separation of
particles occur into the zone based on their density difference,
independent of time.

2) Differential Centrifugation

Differential centrifugation is a common procedure in microbiology and

cytology useful to separate certain organelles for further analysis of
specific parts of cells. In the process, a tissue temple is first
homogenised generalized to break the cell membranes and mix up the
cell contents. The homogenate is then subjected to repeated
centrifugation, each time remove in the palate and increasing the
centrifugal force.

3) Ultracentrifugation
Svedberg coined the term “ ultracentrifugation”.He was a chemist. He
used the ultra fuse to determine the MW and subunit structure of emo
globin. The first commercial ultra fuse was produced in 1940. An
important tool in biochemical research is the centrifuge, which through
rapid spinning imposes hi centrifugal forces on suspended particles, or
even molecules in solution, and causes separation of such matter on the
basis ultracentrifugation of differences in weight. For example, red cells
may be separated from the plasma of blood, nuclei from mitochondria
and cell homogenate, and one protein from another in complex
mixtures.