high performance liquid chromatography, broad peaks

Adsorption Chromatography - Principle and Procedure

What is Adsorption Chromatography?

  1. What is Adsorption Chromatography?
    1. Adsorption Chromatography Principle
    2. Types of Adsorption Chromatography
    3. AC Procedure
    4. Apparatus for TLC method

It is one of the oldest chromatography types and is most widely used in the organic laboratories in the form of column chromatography and thin-layer chromatography. TLC is used to analyze the compounds and to check reactions progress and column chromatography performs separation and purification of compounds. In adsorption chromatography, the stationary phase is a fine solid partitioned to maximize surface area. The analytes are interacting differently with the adsorbent since each has a different affinity. Hence, they separate different retention times.

Adsorption Chromatography Principle

Adsorption chromatography(AC) works on the principle that certain solid material, called adsorbent can hold compound onto their surface. The adsorbent beds will distinguish between the adsorbate based on their binding strength as eluent is continuously run over the stationary phase in the column; variations in the flow rate of the compound ultimately lead to separation of the analytes. The most widely used adsorbents in adsorption chromatography include silica, alumina, and charcoal. Based on their relative polarities the elution order of components from the adsorptive stationary phases can often be estimated. On polar adsorbents, compounds with the more polar functional groups are strongly retained and are therefore eluted earliest.

Types of Adsorption Chromatography

There are three main types of AC, such as column chromatography, thin-layer chromatography (TLC) and gas-solid chromatography.

Column chromatography is a commonly used technique of separating the compounds from the sample mixture. It used on small or large scans to separate and purify compounds. The mixture of samples moves through the stationary phase with the mobile phase and based on various degrees of adhesion it separates the compounds.

Thin-layer chromatography (TLC) is an analytical method of separation used for qualitative analysis and monitoring of the reaction as well as identifying unknown compounds.

Gas-solid chromatography (GC) is an analytical method for the separation of volatile compounds in which the mobile phase is gas and the stationary phase is suitable for solid support. The gas-solid chromatography, work on the principle of separation is adsorption and generally used for samples that had less solubility in the stationary phase.

AC Procedure

Before beginning an AC experiment, we must recognize the various components essential to perform the process.

Stationary phase: The stationary the phase of adsorption chromatography is adsorbent.

Mobile Phase: In AC, either a liquid or a gas is used as a mobile phase.

Apparatus for TLC method

TLC jar: It helps to keep the atmosphere proper during isolation.

TLC plate: It is used as the stationary phase coated with a thin layer of silica, alumina, on glass plastic or aluminum.

Iodine chamber or UV chamber: It helps visualize the spots on the thin layer chromatography plate.

Capillary tube: It is used to apply the sample mixture.

Thin Layer Chromatography Diagram

Adsorption Chromatography
Thin Layer Chromatography Diagram

Experimental Procedure of Adsorption Chromatography

The following steps are involved in the TLC process.

  1. Prepare the developing container: Pour the solvent to a depth just below 0.5 cm inside the chamber to further saturate with the mobile phase or solvent vapor.
  2. Prepare the TLC plate: TLC plates are usually commercially available; just you need to cut them to the required size.
  3. Spot the TLC plate: Dip the micro-capillary into the sample solution, and subsequently touch its end gently on the plate at the suitable place.
  4. Develop the plate: In the chamber, place the prepared plate, cover it with the lid and leave untouched. Via capillary action, the solvent will raise the TLC plate.
  5. Visualize the spots: Remove the plate from the chamber, and if the plate has some colored spots, mark them. Most of the analytes are not colored and should be visualized by placing them in an iodine chamber or UV cabinet. Finally, calculate the Rf values for all the separated compounds.

Adsorption Chromatography Applications

  • It is used for the separation of amino acids.
  • In this technique, the analytes are purified from a sample mixtures according to certain physicochemical properties.
  • It is used to determine the concentration of a molecule.
  • It is used for the separation and identification of isomers.
  • The adsorption technique is useful for the identification of carbohydrates.
  • It is used for the separation and identification of various mixtures of volatile compounds.

The Advantages of AC are as Follows.

  • Adsorption Chromatography is an important tool for separating many molecules that cannot be distinguished from other methods.
  • Adsorption chromatography uses a broad range of mobile phases.
  • Very few types of equipment are used as opposed to other separation methods.
  • The molecules in the complex mixture can be easily separated.

The Disadvantages of AC are as Follows.

  • Automation is making it more complicated and expensive.
  • The major disadvantage of adsorption chromatography is that some solutes have longer retention times.
  • It can cause catalytic variations in the sample.
  • Results obtained by some methods of adsorption chromatography is difficult to reproduce.

Commonly asked questions on Adsorption chromatography are as follows.

  • What is AC?

AC is a separation method, where the stationary phase is an adsorbent.

  • What is the basic principle of AC?

In chemistry, AC is a method of chromatography based on the adsorption principle.

  • What type of chromatography is AC?

Adsorption chromatography is a form of liquid chromatography in which components are retained at the support surface based on their adsorption and desorption.

  • What is the major advantage of AC?

The major advantage of AC is that it separates the mixture into individual compounds.

The major difference between adsorption and partition chromatography is that the sample materials are physically interacting with the stationary and mobile phases used.


A. Wikipedia: https://en.wikipedia.org/wiki/Chromatography

B. Knauer: https://www.knauer.net/en/search?q=chromatography

C. Kromasil: https://www.kromasil.com/support/faq.php

D. Shimadzu: https://www.shimadzu.com/an/service-support/technical-support/analysis-basics/basic/what_is_hplc.html

E. ChemistryView: https://www.chemistryviews.org/details/education/9464911/What_is_HPLC/


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