Chromatography is a separation technique based on partitioning of analytes between a stationary and mobile phase. There are different types of chromatography based on whether the mobile phase is a gas, liquid, or supercritical fluid, and the stationary phase is either a solid or liquid. Chromatographic techniques include gas chromatography, liquid chromatography, ion exchange chromatography, size exclusion chromatography, and thin layer chromatography. These techniques separate analytes based on principles such as adsorption to a solid stationary phase, partitioning between immiscible liquid phases, differences in ionic interactions, molecular size exclusion in a gel, or differing migration speeds in an eluent up a plate.

1. Chromatographic methods
Types and classification of chromatography
Introduction to Chromatographic separations
Theories of chromatographic separations
Instrumentation
Qualitative and Quantitative analysis
HPLC theory
Ref:Principles of Instrumental Analysis: Skoog

2. Chromatography
 Separation technique- based on sample partitioning
between a mobile phase and stationary phase.
 Mobile phase- is the moving phase- it moves over
the stationary phase carrying the analyte along with
it. It can be gas, liquid or supercritical fluid.
 Stationary phase- is fixed in a column or on a planar
surface. Can be a liquid or solid.
 Solid stationary phase- Adsorption.
 Liquid stationary phase- Partition.

3. Chromatography
Gas (mobile phase) Liquid (mobile phase) SFC
GLC GBP GSC Column Planar
LLC LBP LSC IEC Size
Exclusion
TLC PC

4. Adsorption:
X
A
X – active site
A-mobile phase , gas or liquid
A adsorbs to surface X, and desorb from the surface resulting
in separation
Stationary phase
solid
Mobile phase
Gas or liquid

5. Partition:
A
Stationary phase
Liquid
Mobile
phase
Dissolution of A in liquid stationary phase

6. Supercritical fluid
chromatography
 This is a hybrid of gas and liquid chromatography,
combining best features of each.
 A supercritical fluid SCF is formed when a
substance is heated above its critical temperature,
which is the temperature at which a substance
cannot be condensed into its liquid state through
application of pressure.
 CO2 becomes a SCF at 31oC
 Used to analyse nonvolatile and thermally unstable
compounds

7. Ion exchange
 Ion exchange resins serve as stationary
phase in Ion chromatography
 Technique in which ions of the same charge
are separated by elution from a column
packed with finely divided resin.
+
B-
A-
Solid
phase

8.  A- displaces B- if its interaction with the
stationary phase is greater than that of B-
 xRN(CH3)3
+OH- + Ax- RN (CH3)3
+
xAx- + xOH-
 Addition of a dilute base reverse the above
reaction, releasing anions
 Partition ratios for various anions differ from
each other- fractionation occurs

9. Size exclusion
S
L
S-Small
L-Large

10.  Molecules are separated according to
molecular weight and size
 Molecules are trapped in pores according to
size
 Large molecules are not retained and elute
first
 Small molecules are retained in pores of
packaging and eluted last
 Gel filtration- based on hydrophilic packing
used for aqueous mobile phase
 Gel permeation- based on hydrophobic
packing used for separation of non polar
organic solvents

11. Thin layer chromatography
Solvent front
Original line
Sample at time x
dR
dm
RF = dR Retardation factor
dm