CHM 256 7a
CHM 256 7a
CHM 256 7a
BASIC
CHROMATOGRAPHY
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Learning Outcomes
At the end of the course, student should be able to explain:
Principle of chromatography
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PRINCIPLE OF CHROMATOGRAPHY
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For samples containing unknown compounds, the components must be
separated from each other so that each individual component can be identified
by other analytical methods.
3 Measurement techniques used for chemical analysis are usually specific for a
single chemical species.
5 Example of physical properties that are useful for separations are density and
size.
✓ All types of chromatography contain a stationary phase (solid or liquid) to adsorb or absorb the mixture being
separated and a mobile phase (liquid or gas) which passes over the stationary phase and competes with it for the
constituents of the mixture.
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PRINCIPLE OF CHROMATOGRAPHY
Competition
Sample is
between the
dissolved in a
sample molecules
solvent and
adsorbed by the
washed through a
stationary phase
stationary phase
and dissolved by
by a mobile phase
the eluent results
called eluent. Components in the mixture separate out
in separation.
as the mobile phase moves through
Substances that
Speed of the
the stationary phase.
are more soluble
substance passes
in the eluent and
through depends
less adsorbed by
on the nature of
the stationary
the stationary
phase move faster
phase and the
through the
eluent.
apparatus.
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Principles of chromatographic separations
Sample Mobile phase
Component A
Stationary
Component B
phase
Component C
• A small volume of sample is placed at the top of the column (filled with stationary phase and
solvent).
• Sample is eluted with mobile phase.
• The individual components interact with the stationary and mobile phase to different degrees.
• Component A interact more strongly to the stationary phase than component B and component C.
• Then, as the mixture travels down the column, component A will be retarded with respect to the
component B and C.
• In time, they will separate into bands, and be eluted at different times.
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• Retention time
Time taken for a component in the sample to pass through the column.
• Chromatogram
Visual output of the chromatograph. Different peaks or patterns on the
chromatogram correspond to different components of the separated mixture.
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Types of chromatography on the basis of interaction
of the analyte with stationary phase
Adsorption Partition
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In the adsorption process, 1. ADSORPTION
CHROMATOGRAPHY
the solute molecules are
The separation of solute
held on or attached on
depends on the different
the surface of the
in polarity.
stationary phase which is
solid. o Definition: A
chromatography where
the analytes
(components) are
separated based on their
Because of the different different degree of
degree of intermolecular
attraction, some adsorption (stick) on a
components of the surface of solid
mixture will be more stationary phase.
attracted or adsorbed to
the stationary phase thus
separated.
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1
o Substances that
are strongly 3
adsorbed move
through the o The stationary
stationary phase phase is a polar
much more slowly solid, usually finely 4
than those that are divided silica and
weakly adsorbed. alumina and the oThe separation of
solutes are polar the solutes depends
molecules. on the difference in
their polarity. The
more polar solutes oMobile phase may
are more readily be a liquid (LSC) or a
adsorbed than the gas (GSC).
less polar solutes.
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ADSORPTION
CHROMATOGRAPHY
Stationary
phase: solid
Mobile
phase: liquid
solute
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o Example:
1. Thin-layer chromatography (TLC)
- stationary phase: plane (a solid supported on
an inert plate)
- mobile phase: liquid (solvent)
2. Column chromatography
- Stationary phase: silica gel (solid)
- Mobile phase: solvent (liquid)
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2) PARTITION CHROMATOGRAPHY
PARTITION CHROMATOGRAPHY
Mobile phase:
liquid
Stationary solute
phase: liquid
supported on
solid
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❖ Examples:
i. High performance liquid chromatography (HPLC)
ii. Paper chromatography
iii. Gas chromatography (GC)
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TYPES OF CHROMATOGRAPHY AND TECHNIQUES
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PLANAR CHROMATOGRAPHY
COLUMN CHROMATOGRAPHY (CC)
TYPES OF CHROMATOGRAPHY
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COLUMN AND PLANAR CHROMATOGRAPHY
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TRY THIS !!!
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