DI DIMENTIONAL PAPER CHROMATOGRAPHY

TWO DIMENSIONAL PAPER CHROMATE GRAPHY:

In uni dimensional paper chromatography it is not easy to separate between spots of equal Rf values. The decree of closely separated (or even overlapping) spots is improved considerably using two dimensional chromatography. In this two dimensional chromatography procedure re-chromatographing of the separated mixture at 90 degrees angles to the first direction of development is processed. A solvent is used for the second development.

paper-chromatography-two-dimentional-

In this method, the spot is employed near one corner area of a rectangular piece of Chromatography paper. Ascending Chromatography is perform in one direction with the solvent passing the usual method. The mixture separates in the vertical (upright)plane. In one dimensional Chromatography, the components A and B are not well separated .The paper is then dried out, all the solvent is removed and the paper is turned through 90°. The developed components of the Chromatogram lie in the horizontal(parallel) plane. The Chromatography is carried out in the second way with a other solvent in which the components of the mixture have different rf  value from those observed in first solvent. The second development remove the overlapping of the spots

PAPER CHROMATOGRAPHY-TYPES

TYPES OF PAPER CHROMATOGRAPHY

Based on the moving direction of mobile phase the paper chromatography is classified into following types:

(I) ASCENDING PAPER CHROMATOGRAPHY

(II) DESCENDING PAPER CHROMATOGRAPHY

(III) HORIZONTAL PAPER CHROMATOGRAPHY OR RADIAL PAPER CHROMATOGRAPHY

 

i) Ascending paper chromatography: In this process, solvent is present in a dish in the base. The liquid moves toward upside of the paper by capillary movement against to gravitational pull.

Ascending Paper Chromatography Procedure: a filter paper is taken and a pencil line is drawn along the width of the paper above  5 cm from one end. This is called base line.

ASCENDING-PAPER-CHROMATOGRAPHY

 Then the solution mixture is employed on the center of the base line very densely. The filter paper is suspended in a glass tank containing the developing solvent so that the base line dips in the solvent to a depth of 2 cm. Due to capillary action the solvent moves up. The movement is fast in the beginning and it progressively slows down after few hours and lastly it stops. This is the solvent front.

The movement stops totally due to  a balance is formed between the capillary strength and downward gravitational pull.

The solvent front is marked with a pencil. Paper is removed from the tank without delay and dried. The paper now having different spots is called chromatogram. If the paper is placed in the tank for more time, the spots may diffuse on the developed Chromatogram.

Ascending chromatography Limitations: this procedure is not suitable for slow moving solvents. That means those have low RF values are not suitable for this method.

 

 (II) DESCENDING PAPER CHROMATOGRAPHY: in this technique the solvent moves towards downside:

In this technique, the paper is dipped into the solvent (present in trough) near the top of the tank and a heavy glass rod maintains the paper in its alignment. The paper weighs on a other glass rod positioned horizontally parallel to the trough. The mixture of separation (dissolved in the solvent) is put on the paper at an outside spot (outside the trough) in a way that the solvent creates an uniformly moving front" prior to when it reaches the spot. In order to get even dripping and uniform movement, the base of the paper is cutting into a fitting edge.

Descending-Paper-Chromatography

DESCENDING CHROMATOGRAPHY LIMITATIONS: Calculation of Rf value is not possible in this process. So standards are run simultaneously and the distances moved by various constituents of the mixture are in comparison.

(III) RADIAL CHROMATOGRAPHY OR HORIZONTAL CHROMATOGRAPHY: This technique is suitable for fast isolations. In this technique, the spot of the mixture is employed at the centre of a round paper held in the flat surface. The solvent in fed at the centre. As the solvent spreads by the usual capillary force the components of the mixture are separated by radial development in the form of concentric arcs of circles.

HORIZONTA-CHROMATOGRAPHY-RADIAL  

HORIZONTA-CHROMATOGRAPHY-RADIAL 1