Principle of Separation
Partition or distribution of components of the sample between a pair of liquid phases is the underlying principle of chromatographic separations. In paper chromatography separations the component is distributed between the water held between the pores of filter paper and the liquid phase which travels along the filter paper. Separation results from the differences in affinities towards water and mobile phase when travelling under capillary action between the pores of the filter paper.
Separation of the sample components results from a combination of different factors:
Partitioning of the solute molecules between the stationary liquid phase and the developing liquid is governed by differences in solubility of solutes in the two phases which in turn is dependent on differences in polarities between the molecules.
Propelling force tends to pull the substance in the direction of flow of developing solvent. This force depends upon the flow rate and solubility of the substance in the developing solvent.
Retarding force tends to prevent the substance from moving ahead with the mobile phase solvent. This depends on the partitioning of the solute between the mobile and stationary phases
Quantitatively the separation is expressed in terms of retardation factor (Rf) where
Rf = the distance travelled by the solute/ distance travelled by the solvent front
Rf is a unit less quantity. A value of 0 implies that the solute has not moved at all from the application spot and on the other hand if Rf is 1 it means that the solute has no affinity for the stationary phase and has moved the entire distance along with the solvent front.
Rf values provide a fair confirmation on identity of two solutes that have moved the same distance and are of same colour. It cannot be deduced that such compounds are always same as it is quite possible for two compounds to have same Rf value and colour.
In the next article we shall look into the different types of paper chromatography which can help resolve the minute differences between similar eluting compounds.