TLC: Thin Layer Chromatography is quick and inexpensive. Indicates how many components
there are in mixture.
Phase: Stationary phase is solid adsorbent, usually silica. Stationary phase lies on TLC place,
usually glass or plastic. Mobile phase is liquid solvent.
Process Adsorption: The process adsorption is where the silica holds the different components
in mixture to its surface. Separation achieved by relative adsorptions of components with
stationary phase, as components have different affinities for the absorbent. Adsorption is the
process that separates substances.
Experiment for TLC: Pencil base line near bottom of TLC plate. Using capillary tube, spot sample
onto base line. Prepare chromatography tank- put a solvent into beaker. Place TLC plate in
beaker and make sure solvent doesn’t cover spot. Cover beaker with watch glass.
Allow solvent to rise up plate until near the top. Mark solvent front- where solvent reaches.
Allow plate to dry.
One spot may contain two or more compounds, particularly if the spot is over a large regiondue to similar adsorption/ similar Rf value.
If an impure substance, the substance will break up into a couple of spots. However if small
impurity, will break up into a smaller spot than the other spots.
Invisible Spots: Sometimes invisible spots, so use UV lamp to see spots OR spray plate with a
locating agent e.g. iodine to see spots. Use locating agent ninhydrin for amino acids and related.
Retention Factor Rf: TLC chromatography analysed by calculating Rf value. Compare Rf value of
each component to data tables values- IDENTIFY using same solvent and absorbent.
Rf = distance moved by component/ distance moved by solvent front = measure x/ measure y.
Amino acids will be more soluble if more polar solvent used, will travel further up plate and Rf
values larger.
Remember to measure from base line and NOT solvent front
