Thin-Layer Chromatography - TLC Introduction


What is TLC - Fast Separation of a Broad Range of Substances in a Single Run

Thin-Layer Chromatography PlatesDefinition: Thin-Layer chromatography (TLC) is an easy-to-use, fast and highly versatile separation technique for qualitative and quantitative analysis. Thin-Layer Chromatography is performed on a glass, plastic, or aluminum plate, which is coated with a thin layer of adsorbent material. The sample is applied on one end of the plate, and a suitable solvent is allowed to rise up the plate by capillary forces. Since different substances are moving up the Thin-Layer Chromatography plate at different rates, they can be separated, identified and analyzed.

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Thin-Layer Chromatography - Features and Benefits

  • Fast and highly economical separation method, requires no sophisticated instruments for simple qualitative results
  • Simple sample preparation with disposable plates
  • Multiple samples can be run simultaneously under identical conditions
  • Easy two-dimensional separation using two distinct mobile phases in different directions

Types of Thin-Layer Chromatography

The term Thin-Layer Chromatography is generally used to describe the method in all its forms, including manual or semiautomatic operation on conventional, high-performance or modified layers. TLC techniques include HPTLC, PLC and TLC-MS, as well as various detection methods.

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Thin-Layer Chromatography - Applications Fields

TLC can be used to analyze virtually any class of substances, including pesticides, steroids, alkaloids, lipids, nucleotides, glycosides, carbohydrates, fatty acids and many others.

Check each application area to learn more

Analyzing Chromatogram Results

A finished chromatogram provides information about the migration behavior of the separated substances. It also allows to calculate the retardation factor (Rf), which is the ratio of the distance traveled by the substance to the distance traveled by the solvent front. Under identical conditions, the Rf value is characteristic for each compound. Hence, it can be used to confirm the identity of a compound.

Rf = Zs / (Zf – Z0)

  • Rf = retardation factor
  • Zs = distance of the substance zone from the starting line [mm]
  • Zf = distance of the solvent front from the solvent liquid level [mm]
  • Z0 = distance between the solvent liquid level and the starting line [mm]
Calculation of the TLC Rf Value

Preconditions for Thin-Layer Chromatography

When can you use Thin-Layer Chromatography? It may be used if:

  • Substances to be analyzed are soluble in a solvent or mixture of solvents
  • The substances are non-volatile or of low volatility
  • The substances are strongly polar, of medium polarity, nonpolar, or ionic
  • A large number of samples must be analyzed simultaneously, cost-effective, and within a limited period of time
  • All sample components are individually detectable (remain at the start or migrate with the front) after separation, or can be subjected to various detection methods (e.g. in drug screening)
  • The substances cannot be detected by liquid chromatography (LC) or gas chromatography (GC), or only with great difficulty
  • Samples to be analyzed would damage LC or GC columns
  • The solvents used would attack the sorbents in LC columns

Our Literature Recommendations

  • Sabine Bladt and Veronika Rickl: Plant Drug Analysis: A Thin Layer Chromatography Atlas; May 2009
  • Bernard Fried: Practical Thin-Layer Chromatography: A Multidisciplinary Approach; May 1996
  • Raymond P. W. Scott: Thin Layer Chromatography (Chrom-Ed Series); February 2012

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