29.1 - chromatography and functional group analysis

• Chromatography


  Chromatography is used to separate individual components from a mixture of substances. All forms of chromatography have a stationary phase and a mobile phase.
• Stationary phase in thin layer chromatography


  The phase that does not move.

  - Commonly a thin metal sheet coated in alumina or silica.
• What happens in the stationary phase of thin layer chromatography?


  - The solute molecules adsorb onto the surface.

  - The relative adsorptions of the separated components determines how far they will travel along the stationary phase.

  - The more they interact with the stationary phase, the more they will 'stick' to it.

  - The more adsorbent a substance, the less distance it will travel.
• Mobile phase in thin layer chromatography


  The phase that does move.

  - Flows over the stationary phase.

  - It is a polar or non-polar liquid (solvent) that carries components of the compound being investigated.

  - Polar solvents - water or alcohol.

  - Non-polar solvents - alkanes.
• What is chromatography used for?


  Used in analysis of drugs, plastics, flavourings, air samples, and has applications in forensic science.
• Advantages of thin layer chromatography (TLC)


  Quick and inexpensive.
• Thin layer chromatography (TLC)


  An analystic technique that indicates how many components are in a mixture.
• TLC plate


  A plastic sheet or glass coated with a thin layer of a solid adsorbent substence - usually silica.
• What is the stationary phase in TLC?


  The adsorbent (usually silica).
• How does TLC work?


  The different components in the mixture have different affinities for the absorbent and bind with differing strengths to its surface.
• Adsorption


  The process by which the solid silica holds the different substances in the mixture to its surface. Separation is achieved by the relative adsorptions of subtances with the stationary phase.
• Steps of carrying out TLC


  1. On a TLC plate, draw a horizontal line at the bottom edge (in pencil). This is called the baseline.

  
  

  2. Using a capillary tube, spot a small amount of a solution of the sample onto the baseline on the plate.

  
  

  3. Prepare a chromatography tank for the TLC plate by adding a small quantity of solvent to a beaker, with a watch glass placed on top.

  
  

  4. Place the TLC plate inside the beaker with solvent (making sure the solvent does not cover the spot) and place a watch glass to cover the beaker. The solvent will travel up the plate, dissolving the compound as it does.

  
  

  5. As the solvent reaches the top, remove the plate and draw another pencil line where the solvent has reached. This is the solvent front.

  
  

  6. If the sample components are coloured, their spots are easily identifiable on the chromatogram.

  - If the sample components are not coloured, we can locate them using UV light and draw around them in pencil.
• Whta value can a TLC plate be used to calculate for compounds?


  Rf values.
• How to calculate Rf values


  Distance travelled by component / distance travelled by solvent.
• Why must the baseline be drawn in pencil?


  Any other medium would interact with the sample component and solvents used in the analysis process.
• What is gas chromatography useful for?


  Separating and identifying volatile organic compounds present in a mixture.
• Stationary phase in gas chromatography


  A high boiling liquid adsorbed onto an inert solid support (capillary column).
• Mobile phase in gas chromatography


  An inert carrier gas, such as helium or neon, moves the sample molecules through the stationary phase.
• How does gas chromatography work?


  - The sample is injected into the gas chromatogram.

  - The mobile carrier gas carries the components through the capillary column.

  - The components slow down as they interact with the liquid stationary phase.

  - The more soluble the component is in the liquid stationary phase, the slower it moves.

  - Components are separated depending on their solubility in the liquid stationary phase.

  - Compounds in the mixture reach the detector at different times.

  - The compound retained in the column for the shortest time is detected first and has the lowest retention time.
• Retention time


  The time taken for each component to travel through the column.
• What two pieces of information can be obtained from a gas chromatogram?


  1. Retention times can be used to identify the components present in the sample by comparing these to retention times for known components.

  
  

  2. Peak integrations (the area under each peak) can be used to determine the concentrations of components in the sample.
• How are concentrations of components found from a gas chromatogram?


  By comparing components peak integration (area) with values obtained from standard solutions of the component.
• Steps to find the concentration of components in a gas chromatogram using standard solutions?


  1. Prepare standard solutions of known concentrations of the compounds being investigated,

  
  

  2. Obtain gas chromatograms for each standard solution.

  
  

  3. Plot a calibration curve of peak area against concentration. This is called external calibration and offers a method for converting peak area into concentration.

  
  

  4. Obtain a gas chromatogram of the compound being investigated under the same conditions.

  
  

  5. Use the calibration curve to measure the concentration of the compound.

  
• Qualitative analysis of organic functional groups


  Reactions can be carried out on a test-tube scale to identify the functional groups in organic compounds.
• Chemical test for alkenes and observations


  Add bromine water drop-wise. Bromine water decolourised from orange to colourless.
• Chemical test for haloalkane and observations


  Add silver nitrate and ethanol and warm to 50°C in a water bath. Chloroalkane = white precipitate, bromoalkane = cream precipitate, iodoalkane = yellow precipitate.
• Chemical test for carbonyls and observations


  Add 2,4-dinitrophenylhydrazine. Orange precipitate.
• Chemical test for aldehydes and observations


  Add Tollens' reagent and warm. Silver mirror.
• Chemical test for primary and secondary alcohol, and aldehyde and observations

  Add acidified potassium dichromate (VI) and warm in a water bath. Colour change from orange to green.
• Chemical test for carboxylic acid and observations


  Add aqueous sodium carbonate. Effervescence.