Paper Chromatography – Comprehensive Guide
This article describes information about paper chromatography, its working principle, Different types of paper chromatography, Procedure, Factors affecting, applications, Limitations and advantages. Enlisted suitable frequently asked questions with answers.
What is Paper Chromatography and Its working Principle ?
Paper chromatography is a Type of chromatography Technique that uses a special paper, known as chromatography paper, as the stationary phase, and a liquid solvent as the mobile phase. This technique separates the components of a mixture based on their different rates of migration through the chromatography paper. It is one such method of chromatography that has been widely used for over 70 years to separate and analyze different components of a mixture.
It works on the principle of differential partitioning, which is the separation of components based on their different partition coefficients between the stationary and mobile phases. The stationary phase in this technique is chromatography paper, while the mobile phase is the liquid solvent.
History of Paper Chromatography
Paper chromatography was first introduced by Martin and Synge in 1941, and were awarded Nobel Prize in Chemistry in 1952 for their work on the development of partition. Since then, this technique has been extensively used for the separation and identification of different compounds in a mixture.
Types of Paper Chromatography
It is is a chromatographic technique that uses paper as the stationary phase to separate and analyze different components in a mixture. There are three types of such as : ascending, descending, and horizontal paper chromatography.
- Ascending Paper Chromatography
- Descending Paper Chromatography
- Horizontal Paper Chromatography
Detailed Descriptions on Types of Paper Chromatography
- Ascending paper chromatography: In this type of chromatography, the paper is placed vertically in a container, and the sample is applied to a spot at the bottom of the paper. The solvent is added to the container, and as it rises up the paper, it carries the components of the sample with it. The components move at different rates depending on their affinity for the stationary phase and the mobile phase, resulting in separation.
- Descending paper chromatography: Descending paper chromatography is the opposite of ascending paper chromatography. The paper is hung vertically, and the sample is applied at the top. The solvent is added to the bottom of the paper, and as it flows down, it separates the components of the sample based on their affinity for the stationary phase and the mobile phase.
- Horizontal paper chromatography: In this type of chromatography, the paper is placed horizontally, and the sample is applied at the bottom of the paper. The solvent is added to one end of the paper, and it flows horizontally, carrying the components of the sample with it. The components move at different rates, depending on their affinity for the stationary phase and the mobile phase, leading to separation.
In short conclusion Each type of paper chromatography has its own advantages and disadvantages, and the choice of which type to use depends on the specific requirements of the experiment. Ascending chromatography is the most commonly used type and is suitable for separating small samples, while descending chromatography is preferred for larger samples. Horizontal chromatography is used for special applications, such as separating complex mixtures.
Procedure of Paper Chromatography
The procedure of this Chromatography Technique involves the following steps:
- Preparation of the sample to be separated.
- Application of the sample on the chromatography paper.
- Placement of the chromatography paper in the developing chamber containing the mobile phase.
- Development of the chromatogram by allowing the solvent to move up the paper.
- Visualization of the separated components using various techniques.
Detailed Description on procedure of paper chromatography :
- Preparation of the sample: The first step is to prepare the sample to be separated. The sample can be a liquid or a solid, but it needs to be dissolved or suspended in a suitable solvent to create a solution. The solution should be clear and free from any impurities.
- Application of the sample on the chromatography paper: A small spot of the sample solution is applied at the bottom of the chromatography paper using a capillary tube or a micropipette. The spot should be small and concentrated to ensure good separation.
- Placement of the chromatography paper in the developing chamber: The chromatography paper is then placed in a container, such as a beaker or a jar, containing the mobile phase. The mobile phase is a solvent that moves up the paper by capillary action, carrying the components of the sample with it.
- Development of the chromatogram: The solvent is allowed to move up the paper, separating the components of the sample based on their affinity for the stationary phase (the paper) and the mobile phase (the solvent). The development should be stopped before the solvent reaches the top of the paper.
- Visualization of the separated components: The separated components can be visualized using various techniques, such as spraying the paper with a reagent or exposing it to UV light. The spots can then be marked and their Rf values (the ratio of the distance traveled by the spot to the distance traveled by the solvent) can be calculated to identify the components.
Further more Visualization of separated components can be visualized using various techniques , Some of the techniques described below .
- UV light: Some components in the mixture absorb UV light and appear as dark spots under UV light. This technique is commonly used in analyzing nucleic acids, proteins, and other biomolecules.
- Ninhydrin Reagent : Ninhydrin is a chemical reagent that reacts with amino acids and other nitrogen-containing compounds, producing a blue or purple color. This technique is often used in analyzing amino acids and peptides.
- Iodine Vapor: Iodine vapor can be used to detect the presence of lipids, fatty acids, and other nonpolar compounds. The components appear as yellow or brown spots on the paper.
- Silver nitrate stain : Silver nitrate is a sensitive stain that reacts with halides, amino acids, and other organic compounds, producing a brown or black color. This technique is often used in analyzing amino acids and proteins.
- Dragen Dorff’s Reagent: Dragen Dorff’s reagent is a chemical stain that reacts with alkaloids, producing an orange or brown color. This technique is often used in analyzing plant extracts and other natural products.
- Bromocresol green dye : Bromocresol green is a pH-sensitive dye that changes color from yellow to blue depending on the acidity of the medium. This technique is often used in analyzing acids and bases.
Factors affecting Paper Chromatography
Various factors can affect the separation and resolution of the components in paper chromatography. These include nature of the sample, Type of solvent used, Temperature, pH, and the concentration of the mobile phase.
Applications of Paper Chromatography
It is used for the separation and identification of different components in complex mixtures.
Advantages of Paper Chromatography
It has several advantages over other chromatography techniques. It is simple, inexpensive, and requires only a small amount of sample. It can also separate compounds with a high degree of resolution.
Limitations of Paper Chromatography
This type of chromatography also has certain limitations, such as the inability to separate compounds that have similar properties, low sensitivity, and the possibility of sample contamination.
Frequently Asked Questions (FAQs)
What is the difference between paper chromatography and thin-layer chromatography?
In this Technique uses chromatography paper as the stationary phase, while thin-layer chromatography uses a thin layer of silica gel or alumina on a glass plate.
What is the mobile phase in paper chromatography?
The mobile phase in this chromatography is a liquid solvent.
What are the advantages of paper chromatography?
It is simple, inexpensive, and requires only a small amount of sample. It can also separate compounds with a high degree of resolution.
What are the limitations of paper chromatography?
It has low sensitivity, cannot separate compounds with similar properties, and has a risk of sample contamination.
What are the different types of paper chromatography?
The three types of this chromatography are ascending, descending, and horizontal paper chromatography.
