CHROMATOGRAPHY

B Y: Y L A I N E
What is Chromatography?
Chromatography is an important
biophysical technique that enables the
separation, identifi cation, and
purifi cation of the components of a
mixture for qualitative and quantitative
analysis.
Chromatography Phases

Stationary Phase Mobile Phase:

This is a fl uid that moves
This is typically a solid over the stationary phase,
or a liquid supported on carrying the components of
the mixture with it. The
a solid. The mixture to
components interact diff erently
be separated is applied to with the mobile and stationary
the stationary phase. phases, causing them to move
at diff erent rates.
Different Types of Chromatography

Gas Chromatography Liquid

(GC) Chromatography (LC)

Is an analytical technique Separates molecules in a

applicable to gas, liquid, and liquid mobile phase using a
solid stationary phase. Liquid
solid samples (components
chromatography can be used
that are vaporized by heat). If
for analytical or preparative
a mixture of compounds is
applications.
analyzed using GC system, each
compound can be separated and
quantifi ed.
Different Types of Chromatography

Column Chromatography Ion-exchange

Chromatography
This technique is used for the
purifi cation of biomolecules. On a Ion- exchange
column (stationar y phase) fi rstly the chromatography is based on
sample to be separated, then wash electrostatic interactions
buff er (mobile phase) are applied between charged protein
Their fl ow through inside column
groups, and solid support
material placed on a fi berglass
material (matrix).
suppor t is ensured. The samples are
accumulated at the bottom of the
device in a tme-, and volume-
dependent manner
 Different Types of Chromatography
Gel- permeation
(molecular sieve)
chromatography
The basic principle of this
method is to use dextran
containing materials to
separate macromolecules
based on their
diff erences in molecular
sizes .
Affi nity High-prssure liquid
Chromatography chromatography
(HPLC)
This chromatography
technique is used for Using this
the purifi cation of chromatography
enzymes, hormones, technique is possible
to per form
antibodies, nucleic
structural, and
acids, and specifi c
functional analysis,
proteins.
and purifi cation of
many molecules
within a shor t time.
 Different Types of Chromatography

Paper Thin-layer Gas chromatography

Chromatography
Chromatography In this method stationar y
Thin-layer phase is a column which
In paper
chromatography is a is placed in the device,
chromatography
“solid-liquid adsorption” and contains a liquid
support material chromatography. In this stationary phase which
consists of a layer of method stationary phase is adsorbed onto the
cellulose highly is a solid adsorbent surface of an inert solid.
saturated with water. substance coated on Gas chromatography is a
glass plates. “gas-liquid”
chromatography.
 Different Types of Chromatography

Dye- ligand Hydrophobic interaction Pseudoaffi nity

Chromatography chromatography (HIC) chromatography
In this method the
Development of this Some compounds as
adsorbents prepared as
technique was based anthraquinone dyes, and
column material for the
on the demonstration azo-dyes can be used as
ligand binding in affi nity
of the ability of many chromatography are ligands because of their
enzymes to bind used. affi nity especially for
purine nucleotides dehydrogenases,
for Cibacron Blue F3GA kinases, transferases,
dye. and reductases.
 Quiz

1-3. C h r o m a t o g ra p hy i s a n ( M I P R O N T N AT ) b i o p hy s i c a l t e c h n i q u e t h a t e n a b l e s t h e ( I ET S N R A A P O ) ,
i d e n t i fi c a t i o n , a n d ( R N O F I C I A P I U T ) o f t h e c o m p o n e n t s o f a m i x t u r e f o r q u a l i t a t i ve a n d q u a n t i t a t i ve
analysis.

4 - 5 . G i ve t h e 2 P H A S E S o f C h r o m a t o g ra p hy.

6 - 8 . G i ve a t l e a s t 3 T Y P E S o f C h r o m a t o g ra p hy.

9 . I n _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ _ s u p p o r t m a t e r i a l c o n s i s t s o f a l a ye r o f c e l l u l o s e h i g h l y s a t u ra t e d w i t h
wa t e r.

A. Pa p e r C h r o m a t o g ra p hy B . A ffi n i t y C h r o m a t o g ra p hy C . Io n - e x c h a n g e C h r o m a t o g ra p hy

1 0 . W h a t t y p e o f C h r o m a t o g ra p hy i s p r e s e n t e d i n t h e p i c t u r e ?

A . C o l u m n C h r o m a t o g ra p hy B. Mobile Phase C . T h i n L a ye r C h r o m a t o g ra p hy
 Quiz

1-3. C h r o m a t o g ra p hy i s a n ( I M P O RTA N T ) b i o p hy s i c a l t e c h n i q u e t h a t e n a b l e s t h e ( S E PA R AT I O N ) ,
i d e n t i fi c a t i o n , a n d ( P U R I F I C AT I O N ) o f t h e c o m p o n e n t s o f a m i x t u r e f o r q u a l i t a t i v e a n d q u a n t i t a t i ve
analysis.

4-5. Stationary Phase, Mobile Phase

6 - 8 . A ny 3 t y p e s o f C h r o m a t o g ra p hy ( W R O N G S P E L L I N G W R O N G ) ; )

9. A

10. A