2.2.10 Chromatography

1) Mobile Phase 2) Stationary Phase

• For most form soft chromatograph --> components of mixture are separated based on their different attraction/affinities for each of the two phases.

  • Intermolecular forces

1. Paper Chromatography

2. Liquid Column Chromatography

3. Gas-Liquid Chromatography

Research Skills

• Chromatography methods are divided into categories depending on format

1. Planar chromatography

   1. Paper --> separates based on partition

   2. Thin-layer --> based on absorption

2. Column chromatography

• or mechanism of separation

1. Partition

2. absorption

3. size exclusion

4. ion exchange chromatography

Paper Chromatography

• Can investigate mixtures of leaf pigments or amino acids

• To demonstrate this form, lets separate the pigment of ink

• Stationary: Rectangular piece of chromatography paper ( hydrated cellulose ) with any "suitable" solvent

• Mixture is placed on a dotted start line near the beginning of the paper

• This paper is placed at the bottom of a chamber like a beaker that contains a small amount of the solvent inside

• Placing the lid prevents solvent loss and saturates the atmosphere with solvent vapour

• Solvent is allowed to move to a certain point along the stationary phase and the components of the mixture will move different distances up the paper according to their relative affinities for both the SP and the MP

  • Components that dissolve in the solvent better are transported further up the paper

• The result of a chromatography experiment is called chromatogram

• Why hydrated celluose?

  • Contain many $-OH$ groups

    • Very polar and attract water molecules tightly

      • The surface of the cellulose is covered in water

  • Therefore, the components of the mixture are partitioned between the water layer and the less polar solvent

    • Less polar components travel with the solvent up the paper

    • Polar components is left behind by the solvent and stay in the water layer

Thin layer chromatography ( TLC )

• More expensive but greater sensitivity than latter

• Stationary: Rectangular plate made up glass or metal coated with silica ( silicon Dioxide, $SiO_2$) or alumina ( Aluminum oxide, $Al_2O_3$ )

  • Silica and alumina very polar & contains many hydroxyl $(-OH)$ group

• In a eluting chamber contains a non-polar solvent

  • Polar components stick to the polar coating surface and $\therefore$ travel slowly

  • Non-polar components dissolve in solvent and travel further up plate

Experiments

• Colourless substances --> Amino Acids

  • Sprayed w/ locating agent

    • Reacts with colourless components to form coloured products

  • Show up in UV light?

Retardation factor ( $R_F$ )