The range of over-the-counter analgesics (painkillers) is quite extensive, learners establish which brands are most commonly used and use these in the analysis

It is useful to introduce this activity by discussing the use of painkillers and their other effects: antipyretic, anti-inflammatory and antirheumatic. This will contextualise the learner’s experience of the chemistry involved in the experiment.


  • Thin-layer chromatography plate and a pencil (not a biro or felt tip pen)
  • Test tubes in a stand: method of labelling the test-tubes
  • Capillary tubes for use as micropipettes
  • Chromatography chamber. Either a screw top jar tall enough to take the tlc plate, a small beaker with a petri dish for a lid or a commercial tank
  • Access to a fume cupboard and short wavelength UV lamp


  • Dissolving solvent: ethanol and dichloromethane
  • Aspirin standard: 1 g aspirin in 20 cm3 dissolving solvent
  • Caffeine standard: 1 g of caffeine in 20 cm3 dissolving solvent
  • Reference standard: a 1:1 mixture of the aspirin and caffeine standards
  • Ethyl ethanoate as chromatography medium

Health, safety and technical notes

  • Read our standard health and safety guidance here.
  • Wear eye protection
  • Ethanol is flammable, see CLEAPSS Hazcard HC040a
  • Dichloromethane is harmful by inhalation. Avoid breathing vapour and avoid contact with skin and eyes, see CLEAPSS Hazcard HC028 
  • Ethyl ethanoate is volatile, highly flammable and the vapour may irritate the eyes and respiratory system. Avoid breathing the vapour and avoid contact with the eyes. Keep away from flames. See CLEAPSS Hazcard HC043a
  • Short wave UV may cause skin cancer and eye damage. Do not observe directly. The viewer should be screened from direct radiation.


You need two tlc plates and six capillary tubes as micropipettes. Make sure that you do not touch the surface of the tlc plates with your fingers during this activity. Handle the plate only by the edges, and use tweezers if possible.

Part A

  1. Take a tlc plate and using a pencil (not a biro or felt tip pen) lightly draw a line across the plate about 1 cm from the bottom. Mark off three equally spaced points.
  2. You are provided with reference solutions which contain, respectively, aspirin, caffeine and a known mixture of aspirin and caffeine. Use three of the micropipettes to spot samples of these reference solutions onto the tlc plate. Allow the spots to dry and then repeat three more times. The spots should be about 1–2 mm in diameter.
  3. When all the spots are dry, place the tlc plate in the developing tank making sure that the original pencil line is above the level of the developing solvent – ethyl ethanoate. Put a lid on the tank and allow to stand in a fume cupboard until the solvent front has risen to within a few millimetres of the top of the plate.
  4. Remove the plate from the tank and quickly mark the position of the solvent front. Allow the plate to dry.
  5. Observe the plate under a short wavelength UV lamp and lightly mark with a pencil any spots observed.
  6. Place the plate in a jar or beaker containing a few iodine crystals. Put a cover on the jar and warm gently on a steam bath until spots begin to appear. Do this in a fume cupboard.

Part B

  1. Prepare a tlc plate with four points on the base line.
  2. Place half a tablet of one of the analgesics to be analysed on a piece of paper and crush it with a spatula. Transfer it to a small labelled test-tube and add 5 cm3 of the solvent (a 1:1 mixture of ethanol and dichloromethane). Warm gently on a steam bath to dissolve as much of the tablet as possible. Any residue is likely to be a binding agent: allow it to settle for a few minutes. This may be starch. How could you confirm this? Repeat this procedure to make solutions of the other two analgesics.
  3. Using similar procedures to Part A:2, spot a sample of each of the clear solutions onto your prepared tlc plate. On the fourth spot, place the reference mixture used in Part A.
  4. Repeat 3–6 from Part A. Draw diagrams to show how your results appear under UV light and in iodine.


  • Analgesics contain aspirin and should produce a spot corresponding to the known aspirin standard.
  • Analgesics containing caffeine should produce a spot corresponding to the known caffeine reference.
  • Any spots not corresponding to aspirin or caffeine represent other medicines such as ibuprofen. This is less polar than aspirin and therefore moves further up the tlc plate.
  • Caffeine is a central nervous system stimulant and, in low doses, induces wakefulness and improves mental sharpness.
  • Advantages of tlc in medicine analysis:
    • Able to separate closely related compounds;
    • Only small samples required;
    • Quick and easy to carry out, and 
    • Cheap

Further investigations

  • Give the students an unknown sample to identify.
  • Do starch tests on tablets to verify the nature of the binder. Why is starch used?