Introduction Definition History Types of chromatography Principle of column chromatography Types of column chromatography Process of column chromatography Requirement Procedure Precautions Applications Advantage of Column chromatography Disadvantage of Column chromatography Conclusion References

1. COLUMN CHROMATOGRAPHY
By
KAUSHAL KUMAR SAHU
Assistant Professor (Ad Hoc)
Department of Biotechnology
Govt. Digvijay Autonomous P. G. College
Raj-Nandgaon ( C. G. )

2. Synopsis
Introduction
Definition
History
Types of chromatography
Principle of column chromatography
Types of column chromatography
Process of column chromatography
Requirement
Procedure
Precautions
Applications
Advantage of Column chromatography
Disadvantage of Column chromatography
Conclusion
References

3. Introduction
• It is a broad range of physical methods used to
separate and or to analyze complex mixtures
•A mixture of molecules is separated based on
each molecules degree of affinity for the mobile
and stationary phase.
• Column chromatography is the actual or
basically a type of adsorption chromatography
techniques

4. Definition
•Column chromatography is one of the most useful
methods for the separation and purification of both
solids and liquids.
•This is a solid - liquid technique.
History
•American chemist D.T. Day in 1900.
•Invented in 1906 by the Russian botanist Mikhail
Tsvet.

5. Types of chromatography
•Gas chromatography
•Liquid chromatography
•High performance Liquid chromatography
•Ion exchange chromatography
•Column chromatography
•Thin layer chromatography
•Paper chromatography.

6. Principle of Column Chromatography
•When a mixture of mobile phase and sample to be
separated are introduced from top of the column.
•The individual components of mixture move with
different rates.
•Those with lower affinity and adsorption to
stationary phase move faster and eluted out first
while those with greater adsorption affinity.
•slower and get eluted out last.

7. Two phase present in column
chromatography
Stationary phase
Mobile phase
•The stationary phase= It is a solid.
•The most common stationary phase is silica gel.
•An important ratio between the stationary phase
weight and the dry weight of the analyte mixture.

8. •The mobile phase
•It is either a pure solvent or a mixture of different
solvents.
•It is liquid form.
•There is an optimum flow rate for each particular
separation

9. Types of column chromatography
•Adsorption chromatography
•Partition chromatography
•Ion exchange chromatography
•Gel chromatography
Adsorption Chromatography
•It is one of the oldest types of chromatography.
•Utilizes a mobile liquid or gaseous phase.

10. •The equilibriation between the mobile and
stationary phase accounts for the separation of
different solutes

11. Partition Chromatography
•This form of chromatography is based on a thin
film formed.
•Solute equilibriates between the mobile phase
and the stationary liquid.

12. Ion Exchange Chromatography
•In this type of chromatography, the use of a
resin.
•resin is the stationary solid phase
•covalently attach anions or cations onto it.
•To the opposite charge in the mobile liquid
phase are attracted to the resin by
electrostatic forces

14. Molecular Exclusion Chromatography
• Also known as gel permeation or gel filtration
•An attractive interaction between the stationary
phase and solute

15. •The liquid or gaseous phase passes through a
porous gel which separates the molecules
according to its size.
Process of column chromatography
Requirements
•Cylindrical column
•Adsorbent
•Solvent
•Cotton wool

16. Procedure
•The stationary phase material is packed
sufficiently in the column with a cotton pad at the
bottom.
•The extract material to be separated is placed on
the top of packed stationary phase with a second
cotton pad in between.
•The mobile phase is poured into the column over
the sample. A collecting beaker to collect the
elute.

17. Precautions:
•Keep the column in a clean and dust free
place.
•Do not disturb the column till the
separation is complete.
•Avoid gaps within the stationary phase
packing

18. Applications
•To separate or purify natural compound mixtures
like alkaloids, glycosides.
•Column chromatography is best suited to separate
active principle from plant materials.
Advantage s of Column Chromatography
•Any type and quantity of mixture can be separated
•Wider choice of Mobile Phase
•Automation is possible

19. Disadvantages of Column
Chromatography
•Time consuming
•More amount of Mobile Phase are required
•Automation makes the techniques more
complicated & expensive.

20. Conclusion
•Chromatography is a very special separation
process for a multitude of reasons.
•It can separate complex mixtures with great
precision.
•The main advantage of column chromatography
is the relatively low cost.
•Disposability of the stationary phase used in the
process.

21. Reference
•Principle of biochemistry5th edition-
David L.Nelson Michael M.Cox.
•www.studymode.com(at10 pm 9 sep)
•http://en.wikipedia.org