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Centrifugation

Use of centrifugation in food analysis
by

Richard Marshall

on 2 February 2010

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Transcript of Centrifugation

Centrifugation
Ultracentrifuge
Benchtop centrifuge
Types
Preparative
Analytical
to 100,000rpm
to ~10000rpm
Essential safety
Cups & rotors must be carefully maintained
Inspect for damage (don't use)
Balance across centre of rotation
Fill tubes/bottles to approx. same level
Keep rotors, buckets, tube caps clean
Don't exceed designated speed
Record hours/speed of HS rotors
Why?
Spillages can cause corrosion that weakens the rotor
Rotors operate just below elastic limit
Exceeding this will cause permanent damage
Rotor explosion
Loss of samples (contamination?)
Damage to centrifuge
Injury to personnel
Uses
Separation of solids (pellet) from soluble fraction (supernatent)
Extraction of subcellular components
Measurement of molecular size
Theory
As particle sediments, experiences upthrust as it displaces solution below
This depends on:
Size of particle (related to square of radius)
Proportional to difference in density between particle and medium
Decreases as viscosity of medium decreases
Sedimentation rate decreases as frictional coefficient increases
Rate depends on applied centrifugal field
Calculating G force
But angular velocity is not how we usually measure rotational speed
Use rpm and compare G to multiples of Earth's gravity
Relative Centrifugal Force (RCF)
r = radius of rotation
Sedimentation coefficient, S
how fast a particle sediments in centrifugal field
Density gradient centrifugation
Separation of particles of different sizes
Examples of types of gradient materials:
Sucrose (high viscosity)
Ficoll - polysaccharide
Percoll - modified colloidal silica in solution
CsCl (caesium chloride)
Ludox - colloidal silica
Rate zonal - fractions separated on basis of size or sedimentation rate

Isopycnic - fractions separate on basis of bouyant density
Reach equilibrium where no further sedimentation occurs
http://vimeo.com/1177927
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