What is centrifugation? Basic principle of centrifugation Interacting Forces

1. Lecture no : 1
BioMedical Instrumentation-II
Mr. Muhammad Moazzam
Lecturer(Biomedical Department)
University of the Lahore

2. Contents
• What is
centrifugation?
• Basic principle of
centrifugation
• Interacting Forces

3. Centrifugation Method
Centrifugation method is a process of
separating different components or the
suspended particles from the homogenous
solution, by the practice of centrifugal
force. It separates particles based on
their physical properties like size, shape,
density etc. and chemical properties like
molecular weight and viscosity.

4. Centrifugation Method
A natural gravitational force can also
concentrate particles but in an indefinite
time so to increase its efficiency, a
centrifugal force function to separate
particles by their sedimentation property.
The rate of sedimentation increases and
decreases as per the increase in
gravitational force and viscosity of the
medium respectively.

5. History
•The first analytical ultracentrifuge was
developed by Svedberg in 1920

6. What is centrifugation?
• A technique used to accelerate the sedimentation process
by applying centrifugal force
• Different types of centrifugation techniques are used to
separate suspended particles from a liquid in which the
particles are not soluble
• Liquids of different densities may also be separated
• Applications of centrifugation: separation of serum or
plasma from RBCs, sedimentation of cells and viruses,
separation of subcellular organelles, and isolation of
macromolecules such as DNA, RNA, proteins, or lipids

7. Basic principle of
centrifugation
• A centrifuge uses centrifugal force (g-force) to
isolate suspended particles from their
surrounding medium
• When a suspension is rotated at a certain speed
or revolutions per minute (RPM), centrifugal
force causes the particles to move radially
away from the axis of rotation
• The force on the particles (compared to gravity)
is called Relative Centrifugal Force (RCF)

8. BASIC PRINCIPLES
OF
SEDIMENTATION
• When designing a centrifugation protocol, it is
important to keep in mind that:
– the denser a biological structure is, the faster it
sediments in a centrifugal field,
– The more massive a biological particle is, the faster
it moves in a centrifugal field,
– the denser the biological buffer system is, the slower
the particle will move in a centrifugal field,

9. BASIC PRINCIPLES
OF
SEDIMENTATION
• The greater the frictional coefficient is, the slower
a particle will move
• The greater the centrifugal force is, the faster the
particle sediments
• The sedimentation rate of a given particle will be
zero when the density of the particle and the
medium are equal

10. Which factors have an influence
on centrifugation :
•Density of both samples and solution
•Temperature/viscosity
•Distance of particles displacement
•Rotation speed

11. Types of centrifugation
Based on the type of separation
molecules, there are two significant types
of centrifugation:
•Preparative centrifugation: It is a
technique concentrating one component
from the other.
•Analytical centrifugation: It is a
method that measures physical, chemical
and hydrodynamic properties of
suspended molecules.

12. Principle of Centrifugation
Method
•A centrifugation method depends on the
gravitational movement of suspended
particles, which cause deposition of high-
density particles towards the bottom.
• It is important to note that higher is the
difference between the densities of particles,
easy will be the separation. If the density
difference is low, then the separation becomes
tough, and in such case, gravitational force
must be accelerated.

13. Principle of Centrifugation Method
•Therefore, a centrifuge accelerates the
rate of sedimentation by increasing
the relative centrifugal force (RCF)
measures by the rotations per minute
(rpm). If RCF is equal to 100X g, then it
is equivalent to the centrifugal force that
is a hundred times greater than the force
of gravity.

14. Centrifugal field
•A centrifugal force is a kind of force that causes
circular motion of the particles and can express
as:
Centrifugal force(CF) = ω2r
•Where,
ω= Angular velocity in radians per second
r= Radius of a particle from the axis of
rotation
ω2r= Centrifugal acceleration

15. Example
•What is the applied centrifugal field
at a point equivalent to 5 cm from the centre
of rotation and an angular velocity of 3000
rad/s?

16. Solution
•The centrifugal field, G, at a point 5
cm from the Centre of rotation may be
calculated using the equation
•Centrifugal force(CF) = ω2r
=(3000)2x5 cm/s2
=4.5x107 cm/s2

17. Centrifugal field
• In terms of gravitation force,
the equation will represent as,
• Gravitational force (G)= mg
• Where g= Gravitational constant

18. Interacting Forces
•Other than, centrifugal force, there are some other
interacting forces, namely buoyant and frictional
force.

19. Buoyant force
Buoyant force
•A force with which the particles must displace the
liquid media into which they sediment. The
opposing buoyant force is smaller for a dense
particle and can express as:
Buoyant force (Fb)= ρgV
•Where ρ= Fluid density
g= Gravitational or centrifugal acceleration
V= Volume of fluid displaced

20. Frictional force
Frictional force
•A force generated by the particles as they
migrate through the solution.
• Frictional force (Ff )= fv
•Where f= Frictional coefficient
v= Sedimentation velocity

21. Questions?