Thin Layer Chromatography

Spots of the substances move upwards with different speeds by immersing the plate in a solvent

Scientific name: Substances are differently distributed between the absorbent layer and the eluent flowing through it, whereby the distances of shifting at the same time of these substances are different


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Wear eye protection goggles. Use gloves.

Always follow general safety recommendations. Please note that conducting chemistry experiments you must comply with the relevant legal procedures in your country.

Step-by-step instruction

  1. Take a small beaker.
  2. Cut the TLC-plate so that it is fully placed in the beaker.
  3. Draw a straight line at a distance of 5-10 mm from the lower edge of the plate.
  4. Apply a spots of the substances using a capillary. There are several options. Firstly, it is possible to apply a few colored substances on the line at a distance. Secondly, it is possible to apply several substances in one point. Third, the spot may be applied from a mixture of substances. You can use anything as an analytes. For example, you can use markers or you can prepare an extract of plant dyes from leaves .
  5. Prepare the eluent. Mix hexane and ethyl acetate in a ratio of 1: 1. Pour the eluent into the beaker so its level is below the deposition of substances on the plate.
  6. Gently lower the plate in the eluent.
  7. Wait for the moment when the eluent will pass more than 2/3 of the length of the plate, but not until the end!
  8. Observe the colored spots located at the different distances from the lower edge of the plate.
  9. If the substance are still at the start, replace the eluent, increasing the proportion of ethylacetate, and if the substances run up to the end, it is necessary to increase the proportion of hexane. Repeat the steps 1-8.
  10. Separation of colorless substances that glow in ultraviolet can be observed using an UV-lamp.

Scientific background

Thin-layer chromatography is a chromatographic method based on the use of a thin layer of adsorbent as the stationary phase. It is based on the fact that the separated substances are distributed differently between the sorbent and eluent flowing through it, whereby the distances of shifting at the same time of these substances are different. TLC provides great opportunities for analysis and separation of substances, as adsorbent and the eluent can vary within wide limits. Series of plates is commercially available with various sorbents, it makes it possible to quickly and routinely use the method. There are more reliable and reproducible kind of TLC, like high-performance thin-layer chromatography (HPLC), which uses special plates and sophisticated equipment.

TLC was established in 1889 and essentially developed from the middle of the XX century to the present time and is widely used in pharmaceutical, medical and food sectors, as well as in the academic and industrial science.


Published on 15 May 2015

  • Fire
  • Heating with fire
  • Explosion
  • Poisoned gas
  • Organic
  • Electricity
  • Solution
  • Oxidation reduction
  • Color change
  • Precipitate
  • Gassing
  • Catalyst