Paper chromatography is a type of chromatography used for the separation and identification of chemical substances. The stationary phase in paper chromatography is a sheet of cellulose paper of appropriate thickness and texture, which may often be impregnated with a solvent that is immiscible with the mobile phase. Chromatographic separation on paper is much slower than the plates typically used in thin-layer chromatography and is not as versatile as compared with TLC because the degree of variation of the stationary phase is very much limited. However, paper chromatography is still a valuable and very effective method for separations.
Principle of paper chromatography:
The solvent mixture rises by capillary action through the stationary phase. The concentrated sample applies at the bottom of the filter paper in the form of a minute spot. As the liquid mobile phase moves up onto the filter paper, the spotted sample mixture gradually travels along with the mobile phase. This ultimately results in the separation of the solutes. The solutes in the mixture of samples will be separated as per their ability to solubility. In other words, it is polarity like open column chromatography. The molecules have less polarity will travel faster, while more polar molecules are travel more slowly. Every separated spot has its own Rf value. The retention factor (Rf value) signifies the ratio amongst the migration distance of a solute and the migration distance of the solvent front.
Different types or modes of paper chromatography:
There are five types of paper chromatography available based on the development of chromatograms.
1. Ascending chromatography
2. Descending chromatography
3. Ascending- descending mode
4. Radial mode
5. Two-dimensional chromatography
In this article, we are discussing ascending paper chromatography and descending paper chromatography.
Ascending paper chromatography:
Ascending chromatography is a separation method of paper chromatography in that the solvent travels up the stationary phase against gravity. The development of the chromatogram is caused by the upward movement of the mobile phase on paper. The bottom of the paper immersed in the mobile phase, but the spots remain just above the mobile phase. The analytes are separated because each molecule has a different affinity for the stationary phase.
Principle of ascending chromatography:
In ascending paper chromatography, the mobile phase moves upwards by the capillary action, opposite to gravity, and compound are separated by using the principle of partition chromatography or adsorption chromatography.
Procedure of ascending paper chromatography:
Application of the Sample:
- Using a pencil marked the starting point on the paper.
- A spotter, like a thin capillary tube, is used to spot on paper; it can be a very small dot of the sample mixture. Dry the sample spot.
- The paper is put in a closed container like a beaker or chamber that contains a small amount of the mobile phase. The end of the paper is below the solvent level, and the starting point shown on the paper is above the solvent.
- The mixture of solvents will gradually rise by capillary action. Compounds will also rise-up, although the rate is slower than in the mobile phase.
- Analytes that have high solubility in the mobile phase and low solubility in the stationary phase are quickly moving up.
- After the solvent front has about reached the top of the stationary phase the solvent front is marked and the paper is placed aside to dry.
Detection:
- We can observe the position of each analyte on the paper if they are different in colors. However, when the analyte is colorless, then the several techniques are used to detect each analyte. For example, reactions with color-producing reagents: by spraying amino acid with ninhydrin reagent can produce visible violet spots. Or. If the sample has fluorescence or absorbs ultraviolet light, it can be viewed under ultraviolet light by placing it in a UV cabinet.
Calculating the retardation factor (Rf value):
- Calculate the distances traveled by the mobile phase and each analyte. The retention factor is useful to compare results with results from another.
Descending paper chromatography:
Descending chromatography is a type of paper chromatography in which the chromatogram is developed by a solvent that travels downstream of the paper. The mobile phase flows over the stationary phase and it moves by gravity. Descending chromatography is used in both thin layer chromatography and paper chromatography.
Principle of descending chromatography:
Partition chromatography or adsorption chromatography is the basic principle of descending chromatography. It works on the principle of adsorption chromatography for and chromatography for the partition.
Procedure of descending paper chromatography:
- The system of descending chromatography is the contrast of ascending chromatography. This is a considerably more complex setup than ascending chromatography.
- The tank of the mobile phase is positioned on top of the system. The chamber wants to saturate before separating the components.
- The spotting of the sample in descending chromatography is similar to that in ascending chromatography. After spotting the compounds, adsorption paper is attached to the top of the chamber.
- The mobile phase travels slowly from top to bottom in the stationary phase. The sample goes with the solvent and its components are separated.
- The isolated sample mixture is observed on paper as spots after drying. Coloring reagents are sprayed on paper to visualize the spot or viewed under ultraviolet light.
Difference between ascending and descending paper chromatography:
In paper chromatography, there are two methods for separating compounds: ascending and descending paper chromatography. The principle of separation, however, is the same in both forms. However, the principle of separation is the same in both types.
The major difference between ascending and descending paper chromatography is that in ascending paper chromatography the mobile phase is rises by capillary action through the stationary phase which is against the gravity, while in descending paper chromatography the mobile phase flows over the stationary phase and it moves by gravity.
Commonly asked questions on paper chromatography are as follows.
What is ascending and descending chromatography?
Both ascending and descending chromatography is a technique used in paper chromatography. In ascending chromatography, for the separation of components, the solvent moves upward by capillary action on the stationary phase. In descending chromatography, the mobile phase travels along with the gravity by which the separation takes place.
What are the advantages of ascending paper chromatography?
The major advantages of ascending paper chromatography include, that organic as well as inorganic compounds can be identified, it requires less amount of sample, it is an economical method, in the short time it separates the compounds, etc.
What are the major disadvantages of ascending paper chromatography?
This method is not suitable for a compound with a higher Rf value, as it does not have a long stationary phase. Another is, it does not save data for long periods.
What are the 4 types of chromatography?
There are four main chromatographic forms. These are liquid chromatography, thin-layer chromatography, gas chromatography, and paper chromatography.
You may also like this
No comments:
Post a Comment