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centrifugation principle (1)

Centrifugation is a technique used to separate components of a mixture based on size, shape, and density by applying centrifugal force, which is crucial in biochemistry for isolating biological molecules and organelles. The process involves spinning a mixture to cause denser particles to settle at the bottom, forming a pellet, while lighter particles remain in the supernatant. Various types of centrifugation exist, including analytical, density gradient, and differential centrifugation, each serving specific applications in research and industry.

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Karthikaiselvi Lakshmanan
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14 views12 pages

centrifugation principle (1)

Centrifugation is a technique used to separate components of a mixture based on size, shape, and density by applying centrifugal force, which is crucial in biochemistry for isolating biological molecules and organelles. The process involves spinning a mixture to cause denser particles to settle at the bottom, forming a pellet, while lighter particles remain in the supernatant. Various types of centrifugation exist, including analytical, density gradient, and differential centrifugation, each serving specific applications in research and industry.

Uploaded by

Karthikaiselvi Lakshmanan
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© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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CENTRIFUGATION

PRINCIPLES
RITHIN C M
RA2432016010004
M.SC.BIOCHEMISTRY
INTRODUCTION
Definition:
Centrifugation is a technique used to separate components
of a mixture based on their size, shape, and density by
applying centrifugal force.

Importance in Biochemistry:
• Separation of biological molecules (e.g., proteins, DNA,
RNA).
• Isolation of organelles, cells, and sub-cellular components.
BASIC PRINCIPLES OF
CENTRIFUGATION
• The centrifuge uses the sedimentation principle.
• The centrifugation technique works on the principle of
separating particles suspended in liquid media using a
centrifugal field. These are placed in tubes or bottles in the
centrifuge's rotor.
• Sedimentation is the process by which suspended material
settles out of fluids due to gravity. The suspended material
could be clay or powder particles.
• Particles larger than 5 micrometers are separated by a simple
filtration process, whereas particles smaller than 5
micrometers do not sediment under gravity. The central force
can be used to separate those particles.
BASIC PRINCIPLE OF
CENTRIFUGATION
In biochemistry, centrifugation works by spinning a mixture at high speeds, causing
the denser particles to move to the bottom of the tube. This process, called
sedimentation, separates the mixture based on the size and density of its
components. The heavier particles form a solid layer, called a pellet, at the bottom,
while the lighter particles stay in the liquid above, known as the supernatant.
• The basic physics on which the centrifuge works is gravity and generation of the
centrifugal force to sediment different fractions.
• Rate of sedimentation depends on applied centrifugal field (G) being directed
radially outwards.
G depends on
1. Angular velocity (w in radians / sec).
2. Radial distance (r in cms) of particle from axis of rotation
G = w²r
TYPES OF CENTRIFUGATION
There are eight types of Centrifugation:
1.Analytical Centrifugation
Analytical centrifugation is a separation method where the particles in
a sample are separated on the basis of their density and the
centrifugal force they experience. Analytical ultracentrifugation (AUC)
is a versatile and robust method for the quantitative analysis of
macromolecules in solution.

2.Density gradient centrifugation


Density gradient centrifugation is the separation of molecules where
the separation is based on the density of the molecules as they pass
through a density gradient under a centrifugal force.
TYPES OF CENTRIFUGATION
3.Differential centrifugation
Differential centrifugation is a type of centrifugation process in which components
are separately settled down a centrifuge tube by applying a series of increasing
centrifugal force.

4.Isopycnic centrifugation
Isopycnic centrifugation is a type of centrifugation where the particles in a sample
are separated on the basis of their densities as centrifugal force is applied to the
sample.

5.Rate-zonal density gradient centrifugation/ Moving Zone


Centrifugation
Rate-zonal density gradient centrifugation is a type of centrifugation that
separates particles on the basis of their shape as size and works on the same
principle of density gradient centrifugation but works in a different way. It is also
TYPES OF CENTRIFUGATION
6.Differential velocity (Moving Boundary) centrifugation
Differential velocity centrifugation is a type of centrifugation process in which
components are separately settled down a centrifuge tube by applying a
series of increasing velocities.

7.Equilibrium density gradient centrifugation


Equilibrium density gradient centrifugation is a modified and specialized form
of density gradient centrifugation.

8.Sucrose gradient centrifugation


Sucrose gradient centrifugation is a type of density gradient centrifugation
where the density gradient is formed of sucrose by changing the
concentration of sucrose.
CENTRIFUGATION DIAGRAM
KEY PARAMETERS OF
CENTRIFUGATION
RPM (Revolutions Per Minute):
Speed of rotor.

RCF (Relative Centrifugal Force):


Formula: RCF=1.118*r*(RPM/1000)^2

Time:
Duration of centrifugation.

Temperature:
Critical for sensitive biological samples.
APPLICATIONS OF
CENTRIFUGATION
• Production of bulk drugs.
• Production of biological products.
• Evaluation of suspensions and emulsion.
• Determination of molecular weight of collides.
• Removing fat from milk to produce skimmed milk.
• The clarification and stabilization of the wine.
• Biopharmaceutical analysis of drugs.
• Use in water treatment.
• Separating particles from an airflow using cyclonic separation.
SAFETY PRECAUTIONS OF
CENTRIFUGATION
• Balance tubes carefully.
• Use sealed tubes for hazardous materials.
• Do not open the lid while the rotor is spinning.
• Wear appropriate PPE (gloves, goggles, lab coat).
THANK YOU…

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