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Paper Chromatography
What Is Paper Chromatography?
Chromatography technique that uses paper sheets or strips as the adsorbent being the
stationary phase through which a solution is made to pass is called paper chromatography.
It is an inexpensive method of separating dissolved chemical substances by their different
migration rates across the sheets of paper. It is a powerful analytical tool that uses very small
quantities of material. Paper chromatography was discovered by Synge and Martin in the year
Paper Chromatography Principle
The principle involved can be partition chromatography or adsorption chromatography.
Partition chromatography because the substances are partitioned or distributed
between liquid phases. The two phases are water held in pores of the filter paper and
the other phase is a mobile phase which passes through the paper. When the mobile
phase moves, the separation of the mixture takes place. The compounds in the mixture
separate themselves based on the differences in their affinity towards stationary and
mobile phase solvents under the capillary action of pores in the paper.
Adsorption chromatography between solid and liquid phases, wherein the solid surface
of the paper is the stationary phase and the liquid phase is the mobile phase.
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Rf value
Distance of the center of spot from the starting point or the distance travelled by the
sample or analyte divided by Distance of solvent front from starting point or the distance
travelled by the solvent front in chromatography.
Retention Factor Chromatography
Rf value or Retention factor is the difference in rate of
movement of the components in chromatography is caused by various factors.
Retention Factor depends on number of factors
Nature of Adsorbent: Different adsorbents will give different Rf value for same solvent.
The Mobile phase: The purity of solvents and quantity of solvent mixed also affect Rf
Activity: Temperature and storage conditions of adsorbent also affects Rf value.
Thickness of layer: The layers may be of higher or lower thickness which also varies Rf
Equilibrium: Equilibrium of chamber used for development is more important in TLC
than in paper chromatography and hence saturation of atmosphere with the solvent
vapor is important.
The Temperature: Generally constant temperature must be maintained to carry out
separation in order to avoid changes in solvent composition.
Loading: Loading of sample must be small and appropriate about 10micro gram. If
loading is more the spreading of spot and tailing may occur.
Dipping Zone: Distance of starting point from the solvent surface is also a important
Chromatographic Technique: Rf value is also affected by different techniques applied
for chromatography.
These are some of the possible factors which affects Retention Factor (Rf) Value.
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Paper Chromatography Procedure
Below we have explained the procedure to conduct Paper Chromatography Experiment.
Selecting a suitable type of development: It is decided based on the complexity of the
solvent, paper, mixture, etc. Usually, ascending type or radial paper chromatography is
used as they are easy to perform. Also, it is easy to handle, the chromatogram obtained
is faster and the process is less time-consuming.
Selecting a suitable filter paper: Selection of filter paper is done based on the size of the
pores and the sample quality.
Prepare the sample: Sample preparation includes the dissolution of the sample in a
suitable solvent (inert with the sample under analysis) used in making the mobile phase.
Spot the sample on the paper: Samples should be spotted at a proper position on the
paper by using a capillary tube.
Chromatogram development: Chromatogram development is spotted by immersing the
paper in the mobile phase. Due to the capillary action of paper, the mobile phase moves
over the sample on the paper.
Paper drying and compound detection: Once the chromatogram is developed, the
paper is dried using an air drier. Also, detecting solution can be sprayed on the
chromatogram developed paper and dried to identify the sample chromatogram spots.
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