The document outlines the principles and applications of chromatography, particularly focusing on column chromatography in pharmaceutical analysis. It discusses various types of chromatography, their mechanisms of separation, and their uses in chemistry, medicine, and everyday applications. Additionally, it highlights the advantages and disadvantages of column chromatography as a method for separating, purifying, and analyzing components in mixtures.

1. COLUMN CHROMATOGRAPHY
Course Title: Pharmaceutical Analysis And Quality Control
Course Code: BPH-311
Prepared By:
All An Shariah Brishti
Batch: 13th (B)
Department of pharmacy
Daffodil international university
Submission date: 6th March, 2017

2. Chromatography:
 Chromatography is usually introduced as a technique for separating,
identifying and purifying the components in a mixture. [1]
Types Of Chromatography:
 There are different kinds of chromatographic techniques and these are
classified according to the shape of bed, physical state of mobile phase,
separation mechanisms.
The commonly used chromatographic techniques are tabulated below: [2]
Technique Stationary phase Mobile
phase
Basis of separation
Paper
chromatography
solid (cellulose) liquid polarity of
molecules
Thin layer
chromatography
(TLC)
solid (silica or
alumina)
liquid polarity of
molecules
Liquid column
chromatography
solid (silica or
alumina)
liquid polarity of
molecules
Size exclusion
chromatography
solid (microporous
beads of silica)
liquid size of molecules
Ion-exchange
chromatography
solid (cationic or
anionic resin)
liquid ionic charge of the
molecules
Affinity
chromatography
solid (agarose or
porous glass beads on
to which are
immobilized
molecules like
enzymes and
antibodies)
liquid binding affinity of
the analyte molecule
to the molecule
immobilized on the
stationary phase
Gas
chromatography
liquid or solid support gas (inert
gas like
argon or
helium)
boiling point of the
molecules

3. Uses Of Chromatography:
Uses of Chromatography in Chemistry:
 Chromatography is used to figure out the relation ofdifferent mixtures with
one another.
 It is very effective technique to test the purity of the sample.
 The amount of mixture present in a sample can be calculated by using
chromatography.
 Thin layer chromatography is used to check and remove Polychlorinated
biphenyls, pesticides and insecticides in ground water and fish
contaminated by these.
Uses of Chromatography in Medicine:
 In pharmaceutical companies large amount of pure chemicals for making
further medicines is prepared by using chromatography.
 Paper chromatography is used to separate the various inks or dyes from the
mixture.
 Presence of alcohol or some other drugs in blood or urine are detected by
using gas chromatography.
 In pharmacy chromatography is very important to analyse whether correct
medicine is manufactured or not.
 Paper chromatography is particularly used to check the quality of food by
examining different vitamins, preservatives, amino acids and proteins.
Uses of Chromatography in Everyday:
 Paper chromatography and Gas chromatography are employed in finger
print, DNA RNA analysis.
 It helps in checking the level of pesticides, fungicides and contaminants
in the food and drinking water.
 It is also used to check the level of adulterants in the manufactured food
 It is applied in forensic science for investigation.
 Gas chromatography is used in the manufacture and separation of
essential oils. [3]

4. Column Chromatography:
 Chromatography is the most widely accepted and important technique in
the organic chemistry for the separation, purification, identification and
characterization of components of a mixture.
Types Of Column Chromatography:
The four different types of column chromatography are as following:
 Adsorption chromatography----the components of the mixture are made
to adsorb on the surface of the adsorbent.
 Partition chromatography----the components are differentiated between
the mobile and stationary phase. In partition chromatography the mobile
phase and stationary phase are liquid.
 Gel chromatography----the column is packed with a gel, through which
separation takes place. The stationary phase is solvent held in the
interstices of a solvent.
 Ion exchange chromatography----in which the stationary phase is ion
exchange resin.[4]
Principle:
Adsorption is the principle of separation and here a solid stationary phase and a
liquid mobile phase is used. When a mixture of different components which are
dissolved in the mobile phase is introduced in the column. The different
components moves with different rates depending upon their different relative
affinities.
 The lesser affinity compound which is towards the stationary phase moves
faster and it is eluted first.
 The greater affinity compound which is towards the stationary phase
moves later and it is eluted later.
Thus in this way depending upon their relative affinities thre separation is done.
The rate of movement of the compound(R) is given by the following formula:
R=rate of movement of a component/rate of movement of mobile phase or
distance travelled by the solute/distance travelled by the solvent
P=partition coefficient

5. Am=average cross section of mobile phase
As=average cross section of stationary phase
If the mobile phase is liquid then
R=Am/Am+P.As [5]
Some Terminology: [2]
Term Definition
Mobile phase or
carrier
solvent moving through the column
Stationary phase or
adsorbent
substancethat stays fixed inside the column
Eluent fluid entering the column
Eluate fluid exiting the column (that is collected in flasks)
Elution the process ofwashing out a compound through a
column using a suitable solvent
Analyte mixture whose individual components have to be
separated and analyzed
Instrumentation:
 Stationary phase
 Mobile phase
 Column characteristic
 Preparation of column
 Introduction of the sample
 Development technique
 Detection of components
 Recovery of components

6. Application Of Column Chromatography:
 Column chromatography is best suited to separate active principle from
plant materials. Since plants contain many ingredients like alkaloids,
resins, glycosides, tannins, flavonoids and other bio-molecules, the
individual constituents are to be separated.
 In separation of compounds after organic synthesis to obtain desired
molecule.
 To separate or purify natural compound mixtures like alkaloids,
glycosides.[6]
 Separation of different mixture of compounds.
 Removal of impurities.
 Isolation of active constituents.
 Isolation of metabolite from biological fluids.
 Estimation of drugs in formulations.[7]
Advantages Of Column Chromatography:
 Any type of mixture can be separated.
 Any quantity of the mixture can be separated.
 Wider choice of mobile phase.
 Automation is possible.
 In preprarative type the sample can be separated and reused.
Disadvantages Of Column Chromatography:
 Time consuming method.
 More amount of solvents are required which are expensive.
 Automation technique makes complication.[8]

7. 1. MCT5 Chromatography.pdf
http://media.rsc.org/Modern%20chemical%20techniques/MCT5%20Chromatography.pdf
2. Principles of chromatography (article) | Khan Academy
https://www.khanacademy.org/test-prep/mcat/chemical-processes/separations-
purifications/a/principles-of-chromatography
3. Uses of Chromatography | Chemistry@TutorVista.com
http://chemistry.tutorvista.com/analytical-chemistry/uses-of-chromatography.html
4. Column Chromatography | Chemistry@TutorVista.com
http://chemistry.tutorvista.com/analytical-chemistry/column-chromatography.html
5. COLUMN CHROMATOGRAPHY Pharmaceutical Information by Sandhya Sravya malla
http://www.pharmainfo.net/msandhyasravya/blog/column-chromatography
6. Column Chromatography| Principle, Procedure & Applications
http://www.studyread.com/column-chromatography
7. COLUMN CHROMATOGRAPHY Pharmaceutical Information by Sandhya Sravya malla
http://www.pharmainfo.net/msandhyasravya/blog/column-chromatography
8. Column chromatography
https://www.slideshare.net/krakeshguptha/column-chromatography-26966949