Presentation onPaper Chromatography

Principle and Rf Value

Presented to:-Dr. B.M.Gurupadayya,Dept.of Pharmaceutical Chemistry,JSSCP, Mysore.

Presented by:-Tekuri Manoj Kumar,

1st M.Pharm(IP).

Overview:-

• Introduction.• Definition of paper chromatography.• Principle involved in paper chromatography.• Types of paper chromatography.• Advantages of paper chromatography.• Rf value.

• Factors which affect Rf value.• References.

Introduction:-

• All the credit for the present status of paper chromatography goes to Martin and his co-workers viz. Consden, Gorden and Synge.

• Martin and Synge (1941) developed forty-plate complicated apparatus for the separation of acetylated amino acids.

• Consden, Gorden and Martin (1944) replaced silica gel with a filter paper by using n-butanol as a mobile phase.

• It was originally used to separate organic mixtures, but it was quickly adopted for separation of inorganic ions by various workers.

Definition:-

Paper chromatography is the technique in which the separation of an unknown substance is mainly carried out by the flow of solvents on the specially designed chromatographic paper.

In this case the solvent goes up by capillary action.

The separation is effected by differential migration of the substance due to difference in distribution coefficients.

Theoretical principle:-largely by partition coefficient

phenomenon. • separation is accomplished by

successive equilibrations of sample between two phases; one of which moves over the other.

• The stationary phase is made up of the solvent held by the paper and mobile phase is the irrigating eluent.

• Both these phases are in contact over a very large interface on filter paper.

• The water absorbed in amorphous regions of cellulose is distinct from the bulk of mobile water.

Types of paper chromatography:-

Depending upon the migration forces that are involved in separation, paper chromatography is classified into two types.

They are:-i. Paper partition

chromatography.ii. Paper adsorption

chromatography.

Paper partition chromatography:-In standard method of analysis where the paper is utilized as a support with one solvent as mobile phase and other as a stationary phase. The migration of substances is due to differences in partition coefficients.

Paper adsorption chromatography:-In some cases the paper to be used is coated or impregnated with adsorbents like silica gel or aluminia, is used as an adsorbent and the solvent is allowed to flow over the unknown components. Thus, migration of substances is due to the difference in adsorptive powers of substances to be separated. However, this technique is not much used and is of academic interest.

Advantages over other methods:-

i. The equipment is very simple and is easily available.

ii. It has high efficiency of separation.iii. Separation can be effected on macro,

micro and semi-micro scale.iv. Closely related homologues, isotopes,

isomers and very labile and reactive substances can be separated readily and satisfactorily.

Rf VALUE:-

• In paper chromatography the results are represented by Rf value which represent the movement or migration of solute relative to the solvent front. This is indicating position of migrated spots on chromatogram.

• The Rf value is calculated as:-

• Rf is a function of partition coefficients and is a constant for a given substance, provided the conditions of chromatographic system are kept constant.

• For greater reliability, reference standards are used in parallel runs or as internal standards.

• Sometimes the solvent front runs off the paper. Then the position of individual spot is measured relative to position of standard substances.

Rf value is always less than unity but Rx can be greater than 1.

Rf values of two different compounds are different as other physico chemical constants.

factors which affect Rf value are:-

• The solvent system and its composition.• Temperature.• The quality of the paper.• Distance through which the solvent

runs.• The quality and nature of solvents used.• The direction of the fibres of the paper.• The method of development.

References:-

A text book of Pharmaceutical analysis by Dr. A.V.Kasture.Volume – II, page no-31.

….Thank You