Chromatography is a biochemical technique, in which the compounds of the Mixture are separated based on their differential migration and solubility. Thin layered chromatography is a modified paper chromatography. Thin layer chromatography is simply called TLC.
Procedure of Thin Layer chromatography
- In thin layer chromatography silica gel or alumina powder is used as a thin layer On a glass plate and hence the name thin layer chromatography.
- It is an adsorption chromatography.
- In thin layer chromatography the compounds of a mixture are separated between Solid and liquid phases.
- The glass plate functions as the supporting phase.
- The silica gel or the alumina powder functions as the solid stationary phase.
- A suitable solvent is used as the liquid mobile phase.
- The silica gel is allowed to spread as a thin layer on the glass plate. It is allowed to Dry At room temperature. It is activated by heating in an oven at 250°C.
- The mixture is spotted at one end of the thin layer.
- The glass plate is kept in a jar in a slanding position.
- A suitable solvent is added to the jar. The solvent level should be below the level Of the spot.
- The solvent moves up in the thin layer carrying the compounds.
- After 30 minutes the glass plate is removed and is dried.
- A suitable reagent is sprayed to locate the compounds.
- Rf (resolution front) is calculated. Rf=Distance travelled by amino acid / Distance travelled by solvent
Significance of TLC
- It is an easy technique than paper chromatography.
- It requires less time.
- It is a sensitive technique.
- It is more efficient.
- It is faster than paper chromatography.
- It is used to separate not only amino acids but a variety of chemicals.
