Chromatography Essay

Chromatography is the laboratory technique for separating mixtures into their components for analyzing, identifying, purifying or quantifying the mixtures or components. The mixture dissolved in the mobile phase (gas or liquid) is passed through the stationary phase (liquid or solid), which separates the analyte from the other molecules in the mixture. The differences in the migration rate of the compounds in these two phases effects the separation.

Chromatography is either preparative or analytical. Preparative chromatography is a form of purification because the components separated from the mixture are for further use. On the other hand, the purpose of Analytical chromatography is done normally is for measuring the relative proportions of analytes in a mixture. It is done using smaller amount of materials.

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Column Chromatography:

Column Chromatography is an adsorption type of chromatography. The separation depends on the adsorption to the stationary phase. Here, the stationary phase is a solid material and the mobile phase is the liquid. It is used for purifying liquids and solids. In this technique, the stationary bed is embedded within the tube. The mixture of mobile phase and the sample that has to be separated are entered through the top of the column. The components in the mixture move with different rates. The substances with lower adsorption towards the stationary phase travel quickly and eluted out first while the substances with greater adsorption travel slowly and eluted out at the end.

Rate of movement is determined by the R.F value:

Retention Factor:

It is the ratio of distance travelled by solute and the distance moved by solvent.

Planar Chromatography:

Planar chromatography is a technique using the st...

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...h the help of the second buffer which increases the ionic strength of the solution. The charge of the buffer can also be modified to alter the rate of interaction between proteins and the resin. If the proteins are negatively charged at our corresponding given P.H, we should use anion exchange chromatography and if it is positively charged, we should use cation exchange chromatography.

Size Exclusion Chromatography

Size Exclusion Chromatography is the separation technique which depends on the molecular size of the components present in the mixture. The molecules are separated due to their different rate of moving out through the pores of the packed material when they pass through the bed of porous particles. The principle of size exclusion chromatography is the non-adsorptive behavior of the sample, which increases the retention of the molecules under analysis.