chromatography

• Chromatography

  An analytical technique that separates components in a mixture between a mobile phase and a stationary phase
• Mobile phase


  • May be a liquid or a gas
• Stationary phase

  • May be a solid (as in thin-layer chromatography, TLC) or either a liquid or solid on a solid support (as in gas chromatography, GC)
• Solid stationary phase

  Separates by adsorption
• Liquid stationary phase

  Separates by relative solubility
• If the stationary phase was polar and the moving phase was non-polar e.g. hexane

  Non-polar compounds would pass through the column more quickly than polar compounds as they would have a greater solubility in the non-polar moving phase
• Separation by column chromatography


  Depends on the balance between solubility in the moving phase and retention in the stationary phase
• Gas-liquid chromatography GC


  • Mobile phase is an inert gas such as nitrogen, helium, argon
  • Stationary phase is a liquid on an inert solid
• Types of chromatography


  • Thin-layer chromatography (TLC)
  • Column chromatography (CC)
  • Gas chromatography (GC)
• Retention times

  Used to identify different substances
• Rf values

  Used to identify different substances
• Thin-layer chromatography method

  1. Draw pencil line 1 cm above bottom of TLC plate

  2. Mark spots for each sample along line

  3. Use capillary tube to add tiny drop of each solution to a spot

  4. Allow plate to air dry

  5. Add solvent to chamber, place plate in chamber with solvent below pencil line

  6. When solvent reaches 1 cm from top, remove plate and mark solvent level

  7. Allow plate to dry in fume cupboard

  8. Place plate under UV lamp to see spots, draw around them in pencil

  9. Calculate Rf values
• Two-directional chromatography

  Separates a complex mixture by using two different solvents in sequence
• HPLC


  • Stationary phase is a solid silica
  • Mobile phase is a liquid
• Column chromatography (CC)

  1. Glass tube filled with stationary phase (usually silica or alumina)

  2. Filter or plug used to retain solid in tube

  3. Solvent added to cover powder

  4. Mixture to be analysed dissolved in minimum solvent and added to column

  5. Solvent or solvent mixture run through column

  6. Time for each component to reach end of column recorded (retention time)
• Gas-liquid chromatography


  • Mobile phase is an inert gas such as helium
  • Stationary phase is a high boiling point liquid absorbed onto a solid
• Retention time


  Time taken for a particular compound to travel from injection to detector
• Some compounds have similar retention times so will not be distinguished
• Basic gas-liquid chromatography


  • Tells how many components in mixture by number of peaks
  • Tells abundance of each substance by area under each peak
• Gas-liquid chromatography can be combined with mass spectrometry, IR or NMR to identify all components in a mixture
• GC-MS


  Used in analysis, forensics, environmental analysis, airport security and space probes
• If the temperature or the flow rate is higher


  Substance will move more quickly through the column to give shorter retention times
• It is important to use an inert carrier gas such as helium or nitrogen