The Power Point Presentation includes The Paper Chromatography, Its Types and Method with applications. These Slides may be helpful for master of science students. The Syllabus for the slides was prepared by following as KSV, Gandhinagar. Paper Code is CH-AC-302, Unit-01

1. ANALYTICAL CHEMISTRY-4
By-Deepak Sharma
HVHP
Institute of P.G
Studies and Research
LECTURE – 05 PAPER CHROMATOGRAPHY
For M.Sc.
Analytical
Chemistry
Assistant Professor (Department of Chemistry)
M.Sc. (Analytical Chemistry), GATE, GSET, Research Scholar at HNGU

2.  Lecture Outline
Deepak Sharma
• Paper Chromatography Introduction
• Principle
• Type
• Experiment
• Applications

3.  Paper Chromatography
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• It is a method of separating components of a mixture by differential
movement through a two-phase system: the mobile phase and the
secondary phase.
• This movement is effected by the flow of a liquid or a gas (M.P) which
percolates through an adsorbent (S.P).
• Paper chromatography was first introduced by the German scientist,
Christian Friedrich Schonbein in 1865.
• It is a type of a planar chromatography.
• It is the simplest and widely used type of chromatography procedures
which runs on a specialized paper.

4.  Principle
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• Two principles of Paper Chromatography:
• Adsorption Chromatography- Paper impregnated with silica or
alumina acts as the adsorbent (S.P) and solvent as the mobile phase.
• Partition Chromatography- The moisture or water present in the
pores of the cellulose fibers present in the filter paper acts as the
stationary phase and another solvent as the mobile phase.
• In General,
Paper Chromatography = Paper Partition Chromatography

5. Deepak Sharma
 Principle Separation
• Cellulose layers in filter paper contains moisture which acts as
stationary phase & organic solvents/buffers are used as mobile phase
The principle of separation is mainly
Partition rather
than Adsorption

6. Deepak Sharma
• Ascending chromatography
• Descending chromatography
• Ascending- descending mode
• Radial mode/Circular Chromatography
• Two Dimension Chromatography
 Types of Chromatography

7. Deepak Sharma
 Ascending chromatography:
• As the name indicates, the chromatogram ascends. Here the
development of paper occurs due the solvent movement or travel in
upward direction on the paper.

8.  Descending chromatography
• Here the development of paper occurs due to solvent travel
downwards on the paper.
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9.  Ascending- descending mode
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• Here solvent first travels upwards and then down wards on the paper.

10. Deepak Sharma
• Here the solvent travels from center(mid point) towards periphery of
Circular chromatography paper.
 Radial mode/Circular Chromatography

11. Deepak Sharma
• Here the chromatogram
development occurs in
two directions at right
angles.
 Two Dimension Chromatography

12. Experiment
Paper
Chromatography
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13. Deepak Sharma
• Stationary Phase & Papers used
• Application of sample
• Mobile phase
• Development technique
• Detecting or Visualizing agents
 Requirements

14.  Stationary Phase
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• Whatman filter papers of different grades like No.1, No.2,
No.3, No.4, No.20, No.40, No.42 etc. are used.
• In general this paper contains 98-99% of α-cellulose, 0.3–1%
β–cellulose
• Factors that governs the choice of paper:
» Nature of Sample and solvents used.
» Based on Quantitative or Qualitative analysis.
» Based on thickness of the paper.

15.  Preparing the Paper Strips
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• Cut the filter paper into 5 x 4
measurement.
• Draw a line 0.5 cm above the bottom
edge of the strip with the pencil.
• Label each strip with its corresponding
solution.
• Place a spot from each pen on your
starting line.

16.  Mobile Phase
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• Pure solvents, buffer solutions or mixture of solvents
• Examples- Hydrophilic mobile phase
1. Isopropanol: ammonia:water → 9:1:2
2. Methanol : water → 4:1
3. N-butanol : glacial acetic acid : water → 4:1:5
• Hydrophobic mobile phases
1. dimethyl ether: cyclohexane
2. kerosene : 70% isopropanol

17.  Preparation of the solution
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• Choice of suitable solvent for making solution is very important.
• Pure solutions can be applied direct on the paper but solids are
always dissolved in small quantity of a suitable solvent.
• Biological tissues are treated with suitable solvents and their
extracts obtained.
• Proteins can be precipitated with alcohol and salts can be
removed by treatment with ion exchange resin.

18.  Application of Sample
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• The sample to be applied is dissolved in
the mobile phase and applied as a small
spot on the origin line, using capillary
tube or micropipette. very low
concentration is used to avoid larger
zone
• The spot is dried on the filter paper and
is placed in developing chamber.

19.  Developing the Chromatograms
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• Place the strips in the beakers.
• Make sure the solution does not come
above your start line.
• Keep the beakers covered.
• Let strips develop until the ascending
solution front is about 2 cm from the top
of the strip.
• Remove the strips and let them dry.

20.  Calculate the Rf value and make
conclusions.
Deepak Sharma
distance traveled by component from application point
distance traveled by solvent from application point
Rf =

21.  Application
• Separation of mixtures of drugs
• Separation of carbohydrates, vitamins, antibiotics, proteins,
etc.
• Identification of drugs
• Identification of impurities
• Analysis of metabolites of drugs in blood , urine ….
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22. Deepak Sharma
https://www.linkedin.com
/in/deepak-sharma-
a1399688
Name of Channel
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