Function of chromatography
Function of chromatography
The term chromatography was originally applied by Micheal Tswett, a Russian botanist in 1906.It may be defined as the technique of separation of substances according to their partition coefficients (i.e., their relative solubilities) two immiscible phases. In this method the separation of the components of a mixture is a function of their different affinities for a fixed or stationary phase (such as a solid or liquid) and their differential solubility in a moving or mobile phase (such as a liquid or a gas). Chromatography is of several types. Physics Wallah Biology Doubts page consist of more questions for reference.
1) Adsorption or column chromatography
The stationary phase consists of a column of charcoal, silica, alumina, calcium carbonate or magnesium oxide. The solution is made to percolate through this column when different chemicals get adsorbed at various levels. This technique is useful for separation of tissue lipids.
2) Thin layer chromatography
The stationary phase consists of a thin plate of cellulose powder, alumina or silica. As a few drops of mixture are poured over it the different chemical spread to different distances. By this method amino acids, nucleotides and other low molecular weight products can be separated.
3) Paper chromatography
A paste of mixture is applied near one end of a chromatographic paper. The lower end below the paste is dipped in a solvent. As the solvent rises in chromatographic paper, the different chemicals of the mixture spread to different distances. The paper can be rotated to obtain two dimensional chromatogram. This method is especially useful for the detection and separation of amino acids nucleotides and low molecular weight products.
4) Ion exchange chromatography
Beads of cellulose and other materials, having negative and positive charges are placed in a column. The mixture is poured over the column. As the mixture passes through the column, its constituents become separate according to their charges. This technique is used in purification of insulin, plasma fractionation and separation of proteins.
5) Gel filteration chromatography
This is also called molecular sieve chromatography. Dextran gel (Sephadex) is available with various pore sizes. A mixture is poured over a column of sephadex. The various chemicals pass through the pores and come out of the column. The larger molecules come out first, followed by progressively smaller sized molecules. This technique is used in determining the molecular weight of proteins.
6) Affinity chromatography
In this technique stationary phase consists of a column of ligands. Ligands are molecules that bind to other specific molecules at particular sites. Mixture is allowed to pass through the column. The chemical linkages occur between ligands and their specific chemicals while others pass out of the column. Cellular enzymes, immunoglobulins, mRNAs etc. are separated by affinity chromatography.