Thin Layer Chromatography

• Introduction

The thin layer chromatography (TLC) is a kind of chromatography where the stationary phase is a thin layer of material deposited on a support, this set is called a plate. The mobile phase is a solvent mixture, call eluent, that it moves for the stationary phase by means of capillarity. Generally, the plates are formed by a layer of silica gel or another material with adsorbent properties.

• Obtaining the chromatogram

The sample solves in few quantity of a volatile solvent. If it’s going to be in use patrons of some substance it’s prepared of the same way. Later it’s drawn a base line with a pencil and a few points of reference where the samples will be puncture with a capillary.



It’s necessary to prepare the container in which the chromatography is going to be realized. This container is a simple glass container where is added the eluent that is going to be in use, always in a level lower than the base line that it has drawn in the plate. In the container there gets a strip of filter paper in order that the eluent vapor is for all the container. In this moment the plate gets carefully in the container and it wraps up, hoping that the level of the eluent ascends for the plate.


When the level is near to top, the plate is extracted and a new line is drawn in this level. The volatile solvent is being evaporates in a little time leaving the plate dry. Now it’s necessary find a method to visualize the result of the chromatogram and see the separation of components punctured in the plate. In case that the substance is colored there would be no problem since the components would be seen. For other compounds, the commercial plates are in the habit of taking a fluorescent substance that if it’s radiated by UV light, the components can see as dark circular spots. These spots must be marked on the plate with a pencil. Another method can be make react the components with some substance that colour these compounds to give a visual sign.



• Factors that influence in the elution

The factor that more influences in the separation of the components in this kind of chromatography is the polarity of these components. The plate is formed by a layer of gel of silica, a very polar substance, and therefore, it will attract in major measurement to more polar components.
On having fulfilled the chromatogram, the most polar substances will adsorbed more with the support and its displacement speed will be smaller, this means that on having revealed the plate of chromatography, the most polar component will be in a place nearer to base line that we have drawn, and on the contrary, the least polar component will move more rapidly and will be more far from the base line.


The eluent polarity is a factor that get that the components move more rapid or slower in a chromatography. That is to say, for an eluent with greater polarity, this one will be more attracted by the support and the components will be freer to move better, for that, on having the plate revealed, the components will go out more far from the base line. For the same reason, with a less polar eluent, this one almost doesn’t adsorbed for the polar support, and the compounds will have a greater interaction with the support, for that its speed will be smaller and on having finished the chromatography, it will have moved not much from the base line. For these factors, there is advisable the election of a good eluent for the compounds separation. For every experience the kind of eluent will be different, depending on the components to separate, therefore, the best thing will be to realize several tests with different kind of eluent, this way consider the most ideal eluent for our chromatography.



• Applications
  • Identification of some substance known in a mixture for its comparison with a patron of pure substance.
  • Measures the purity degree of a sample, if several spots exist in the chromatogram, it’s a sign that exist several components in the sample.
  • Obtain a semiquantitative measurement of some component. Puncturing the same quantity in a plate, as much bigger is the spot obtained in the chromatogram, the concentration of this substance will be bigger.

More information about TLC
More images of TLC

Linux Code

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Johannes Van der Waals

He’s famous “for his work in the equation of state of gases and the liquids”, for which he obtain the Nobel prize of Physics in 1910.

Van der Waals was the first one in realizing the need to take in consideration the volume of the molecules and the intermolecular forces establishing the relation among pressure, volume and temperature of the gases and the liquids.

Vía Wikipedia.

Introduction to analytical instrumental methods

The instrumental methods are based on the measurement of some physicist-chemistry property. Its use has increased with the development of the electronics, for the facility of detecting changes in the physicist-chemistry properties and transform them to an understandable language for the humans. An analytical instrument is the support in which is developed this transformation.

This text show a small introduction for the most important instrumental methods.

• 1. Spectroscopic methods
  • 1.1. Atomic Spectroscopy
    • 1.1.1. Atomic Absorption Spectroscopy

    The sample atomize by means of high temperature medium that provides a high energy. The atomizer can be a flame or a graphite furnace.
    It’s radiated the atomic vapor and the atoms are able of absorbing to a characteristic length of wave. The measured absorbance will be a function of the concentration in the atomic vapor.
    It’s used a hollow cathode lamp in order that the radiation that affects in the sample is typical of the element that it’s wanted to quantify.
    Example of Atomic Absorption Spectrum.

    • 1.1.2. Atomic Emission Spectroscopy

    The most used atomizers are plasmas and the electric arch and spark. These atomizers obtain a temperature of up to 10000K.
    When the sample it’s atomized with this temperature, the atoms will be in an excited state and when they return to the fundamental state they will emit a typical radiation that will be a function of the element quantity in the sample.

    Example of atomic emission spectrum.

    • 1.1.3. Atomic Mass Spectrometry

    This method consists in atomization of the sample, conversion the atoms to ions, separation of these ions by means of the difference of the relation mass/charge (m/z) and count of the ions of every kind.
    For the separation and count of ions, a mass spectrometer is used.

    Example of atomic mass spectrum.

  In this kind of methods, the present substances in a sample, turn into atoms or elementary ions in gaseous state, for the later measurement of the analytical property.

  • 1.2. Molecular Spectroscopy
    • 1.2.1. Molecular Absorption Spectroscopy

    The substances can absorb to a certain wavelength. If it’s radiated certain one λ, the intensity that comes to the detector will be smaller that the intensity with which we radiate the sample. This absorbance is proportional to concentration of analite, according to Beer’s Law.
    Usually there is in use the region UV and Visibly of electromagnetic spectrum.

    Example of Molecular Absorption Spectrum.

    • 1.2.2. IR Absorption Spectroscopy

    On having applied a IR radiation to a molecule, the molecule can change from some states of vibrational energy and rotational to others.
    These changes are defined by a typical energy, or that is the same thing, to a certain wavelength, and it depend on the atoms that correspond to bond, to the kind of this bond and to the kind of vibration or rotation movements.

    Example of IR Spectrum.

    • 1.2.3. Nuclear Magnetic Resonance

    It’s based on the radiation absorption on the region of the radio frequency. In this process the atomic nucleus are involved.
    It’s a method for structural determination very well, since the signal of every atom, it will go out to bigger or smaller displacement, depending on the atoms that it’s around in the molecule.

    Example of NMR Spectrum.

    • 1.2.4. Molecular Mass Spectroscopy

    It’s based on the separation of different radical or molecular ions by the difference of the relation mass/charge among them.
    It’s a method of structural determination and can give information over complex mixtures, among other applications.

    Example of Mass Molecular Spectrum.

  Are spectroscopic methods where analite that wants determinate it’s in molecular form.

Spectroscopy is a science that treats the interaction of the electromagnetic radiation, or other particles, with the matter.

• 2. Electrochemical Methods
  • 2.1. Potentiometric Methods

  It’s based on the measurement of equilibrium potential on a galvanic cell, on which we have two electrodes, indicator and reference.
  We measure the potential difference between the electrodes, this potential difference between the electrodes is related to analite concentration that we want to measure.

  • 2.2. Voltammetric Methods

  Are methods in which the analite concentration is deduced from measurements of current intensity depending on the applied potential.
  It has big advantages like that the analite doesn’t lost on having realized the experiment.

It’s based on the electrical properties of an analite on dissolution. They possess specially low limits of detection.

• 3. Chromatographics Methods
  • Gas Chromatography, where the mobile phase is an inert gas and the stationary phase can be solid or liquid.
  • Liquid Chromatography, where the mobile phase is one o more liquid and the stationary phase is solid.

The chromatography is a separation method of substances in complex mixtures. It’s in use generally for separation, but some component connected to the instruments for the later determination by some analytical instrumental method.
The mixture to resolving it’s introduced in a system formed by a fluid (mobile phase), which circulates in contact with the stationary phase.
The mixture components that possess a major affinity for the stationary phase, its advance speed will become smaller and the separation will be achieved.
There are two important kinds of cromatografía:

Example of chromatogram.

There are more instrumental methods of the described ones and these present some major complications of exposed in this text. But for a small introduction the presented here is valid.
In a future I will write an amplification of this text with more methods and a major extent in the contents.