Thin Layer Chromatography

Thin Layer Chromatography

Basics of Thin Layer Chromatography

  • Thin Layer Chromatography (TLC) is a type of chromatography used to separate the components of a mixture.
  • It relies on a stationary phase on a thin layer, which is typically a silica gel, and a mobile phase which transports the components up the stationary phase.
  • The principle behind TLC is adsorption, not absorption. Components are attracted and adhered to the surface of the solid stationary phase.
  • Molecules with higher polarity stay closer to the stationary phase and move more slowly, while less polar components move faster and farther.

Running a Thin Layer Chromatography

  • A pencil is used to draw the baseline onto the TLC plate, so it does not interfere with the development stage.
  • The mixture is dotted onto this line, and the plate is then stood in a shallow pool of solvent (the mobile phase).
  • As the solvent travels up the plate by capillarity, it carries the different components of the mixture with it.

Understanding TLC Results

  • Different components will travel different distances, depending on their attraction to the mobile or stationary phase.
  • After the run, the plate is dried and can be visualised using ultraviolet light, or by staining.
  • Using a Rf value (ratio of distance moved by the compound to distance moved by solvent), different components can be identified.
  • This value should be constant for a particular substance in a given solvent and room temperature.

Applications in Medicine

  • TLC can be used to identify and quantify pharmaceutical substances in a drug product.
  • It serves as an effective qualitative analysis tool for the separation and identification of drugs or metabolites in body fluids.
  • Additionally, it is used to detect adulterants or contaminants in drug substances.
  • It’s also utilized as a part of purity tests and stability studies during the development and manufacture of pharmaceuticals.

Limitations of TLC

  • TLC is not as accurate as other chromatography techniques such as high performance liquid chromatography (HPLC), due to varying flow rates of solvent and inconsistencies in the spotting of samples.
  • TLC cannot provide active compound confirmation on its own; spectroscopic methods are required for confirmation.
  • It has the potential for sample contamination during handling and the solvent evaporation stage.