Chromatography And Its Types Pdf

chromatography and its types pdf

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Chromatography is a process for separating components of a mixture. To get the process started, the mixture is dissolved in a substance called the mobile phase, which carries it through a second substance called the stationary phase.

What is Chromatography and How Does it Work?

Are you a chemistry student? Visit A-Level Chemistry to download comprehensive revision materials - for UK or international students! Chromatography is an analytical technique used to separate mixture of chemical substances into its individual compounds. Different types of chromatography are used in lab. Chromatography consists of two phases: one mobile phase and one contiguous stationery phase.

The stationery phase is liquid or solid and the mobile gas is gas or liquid. The compound mixture moves along with the mobile phase through stationery phase and separates depending on the different degree of adhesion to the silica of each component in the sample or the compound mixture. Similar procedure used in all chromatography. Lets explain the process of column chromatography to understand the general procedure.

This is also known as liquid column chromatography. In this type of chromatography silica or alumina is used as stationery phase and a suitable solvent is used as a mobile phase. The column is packed with the slurry of silica or alumina. Here the each compounds in the mixture or sample is separated depending on the polarity of the molecules.

Thin layer chromatography is the most widely used method in chemical or biochemical lab. Here as a stationery phase a thin layer of silica is used on a glass, metal or rigid plastic. The solvent rises up the chromatography taking each component of the sample with it. The components travel with the solvent depends on their polarity.

The non polar components travel faster than the polar component. Thus separated bands of components are observed under UV-light. The similar technique is used on paper chromatography. Difference is, instead of using a thin layer of silica on metal, it uses a special type of chromatography paper as stationery phase. This paper is made of cellulose.

Drops or line of reaction mixture is placed on to it and then placed into a sealed container of swallow solvent. The non polar compound travels fast in this chromatography. Polar compound make bond with cellulose and thus does not go far. Definition of chromatography Chromatography is an analytical technique used to separate mixture of chemical substances into its individual compounds.

Fig: Column chromatography Principles of chromatography Chromatography consists of two phases: one mobile phase and one contiguous stationery phase. Chromatography terms A chromatography is a physical method of separation while chromatograph is an equipment to separates mixture of compounds into its components. A mobile phase is the solvent that moves through the column. A stationery phase is the solid e.

Eluent is the fluid that enter into the column. Eluate is the fluid that leaves the column or collects into a flask. Process Similar procedure used in all chromatography. A column is firstly packed with silica gel, the stationery phase. Then a solvent mobile phase is passed through the column. The sample is poured slowly on the top of the silica. A suitable solvent is poured on the sample to flow through the silica under the gravity pressure.

It separates the components of the compound mixture depending on different degrees of adhesion to the silica. The separated compounds are then collected in different flask or test tube for identification. Types of chromatographpy Column chromatography This is also known as liquid column chromatography.

Thin layer chromatography TLC Thin layer chromatography is the most widely used method in chemical or biochemical lab. Fig: Thin layer chromatography Paper chromatography The similar technique is used on paper chromatography. Summary Chromatography is an analytical technique used to separate mixture of chemical substances into its individual compounds.

The mobile phase flows along with the each component through the stationery phase depending on the degree of adhesion of each component to the stationery phase. Thus they are separated.

Gas chromatography

The science of chromatography began early in the twentieth century, with the Russian botanist Mikhail Tswett, who used a column packed with calcium carbonate to separate plant pigments. The method was developed rapidly in the years after World War II, and began to be applied to environmental problems with the invention of the electron capture detector ECD in by James Lovelock. This detector, with its specificity and very high sensitivity toward halogenated organic compounds, was just what was needed to determine traces of pesticides in soils, food and water and halocarbon gases in the atmosphere. This happened at exactly the time when the effect of anthropogenic chemicals on many environmental systems was becoming an issue of public concern. Within a year, it was being applied to pesticide analysis. The pernicious effects of long lived, bioaccumulating pesticides, such as DDT, would have been very difficult to detect without the use of the ECD.

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Gas chromatography GC is a common type of chromatography used in analytical chemistry for separating and analyzing compounds that can be vaporized without decomposition. Typical uses of GC include testing the purity of a particular substance, or separating the different components of a mixture the relative amounts of such components can also be determined. In some situations, GC may help in identifying a compound. In preparative chromatography , GC can be used to prepare pure compounds from a mixture. In gas chromatography, the mobile phase or "moving phase" is a carrier gas , usually an inert gas such as helium or an unreactive gas such as nitrogen.


Chromatography is an important biophysical technique that enables the separation, identification, and purification of the components of a mixture for qualitative and quantitative analysis. Proteins can be purified based on characteristics such as size and shape, total charge, hydrophobic groups present on the surface, and binding capacity with the stationary phase. Four separation techniques based on molecular characteristics and interaction type use mechanisms of ion exchange, surface adsorption, partition, and size exclusion.

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Gas Chromatography

Gas chromatography is a term used to describe the group of analytical separation techniques used to analyze volatile substances in the gas phase. In gas chromatography, the components of a sample are dissolved in a solvent and vaporized in order to separate the analytes by distributing the sample between two phases: a stationary phase and a mobile phase. The mobile phase is a chemically inert gas that serves to carry the molecules of the analyte through the heated column. Gas chromatography is one of the sole forms of chromatography that does not utilize the mobile phase for interacting with the analyte. The stationary phase is either a solid adsorbant, termed gas-solid chromatography GSC , or a liquid on an inert support, termed gas-liquid chromatography GLC. In early s, Gas chromatography GC was discovered by Mikhail Semenovich Tsvett as a separation technique to separate compounds.

Liquid chromatography is a technique used to separate a sample into its individual parts. This separation occurs based on the interactions of the sample with the mobile and stationary phases. Liquid-solid column chromatography, the most popular chromatography technique and the one discussed here, features a liquid mobile phase which slowly filters down through the solid stationary phase, bringing the separated components with it.

This article throws light upon the twelve types of chromatographic techniques used in biochemistry. There are different kinds of chromatographic techniques and these are classified according to the shape of bed, physical state of mobile phase, separation mechanisms. Apart from these there are certain modified forms of these chromatographic techniques involving different mechanisms and are hence categorized as modified or specialized chromatographic techniques. It is the preparative application of chromatography. It is used to obtain pure chemical compounds from a mixture of compounds on a scale from micrograms up to kilograms using large industrial columns. The classical preparative chromatography column is a glass tube with a diameter from 5 to 50 mm and a height of 50 cm to 1 m with a tap at the bottom. Slurry is prepared of the eluent with the stationary phase powder and then carefully poured into the column.

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Ion-Exchange Chromatography and Its Applications

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Column Chromatography.