Thin Layer Chromatography (TLC)

-It is open bed technique, substances are separated by the differential migration that occurs when a solvent flow (by capillary action) along a thin layer of fine powder spread on a glass or plastic plate(TLC). -Basic principle behind the differential migration is adsorption. -A solute undergoes distribution between two phases: i)a stationary phase acting through adsorption. ii)a mobile phase in the form of a liquid. -Adsorbent: thin, uniform layer of dry finely powdered material. -Adsorbent is applied to a glass, plastic or metal sheet or plate. -Separation principles may be: a)Adsorption, b)Partition(including reverse phase partition) c)Ion exchange, d)Molecular exclusion(size-exclusion), -Combination mechanisms may involve. -Mechanism is evident from the nature of the thin layer & solvent system. Coating material: Dried silica or alumina, Mobile phase/solvent system: Organic solvents. Mechanism: Adsorption. -In adsorption: The more the affinity , slowly the component moves along the plate. Lesser the affinity faster the movement of the component

Evaluation of the chromatogram: -Spots are detected visually, UV irradiation or other chemical technique.

i)Qualitative ii)Quantitative i)Qualitative, -Rf value which is the ratio of distance travelled by the solute (Component) and solvent (Mobile phase movement) -It is unique for a pure component in that solvent system -Thus serves as the reference parameter in Qualitative analysis. ii)Quantitative: 1)Direct techniques: Visual assessment, Determiantion by measurements of spot area, Quantitative TLC incorporating densitometry, Direct spectrophotometry on thin-layer chromatograms. 2)Indirect techniques: Gravimetry, UV/Vis spectrophotometry, Fluorimetry, Polarography, Coulometry, Flamephotometry, Radiometry, Vapour-phase chromatography etc.