AAT Bioquest

What are the main types of chromatography techniques?

Posted April 18, 2023


The main types of chromatography techniques include: column chromatography, paper chromatography, gas chromatography, affinity chromatography, ion exchange chromatography.

Column chromatography is a technique utilized to differentiate a single chemical compound from a mixture dissolved in a fluid. This type of chromatography separates substances based on differential adsorption of compounds to the adsorbent as compounds diffuse through the column at distinct rates. This allows them to get differentiated in fractions. Column chromatography may be utilized in small to large scale experiments to purify substances. 

Paper chromatography is a technique used for the separation of compounds based on the differential solubility of the stationary and mobile phase (solvent). It is a cost-efficient technique that requires very small amounts of substance. In this technique, the solvent travels across the paper and carries amino acids with it, which are separated based on differential solubility. Retention factor is used to identify specific amino acids in the mixture during separation and compared with the standard Rf value chart. 

Gas chromatography uses gas flow through a glass or metal column which separates volatile mixtures based on their differences of boiling point and polarity. In this type of chromatography, the components of a sample are dissolved in a solvent and vaporized to separate the analytes. This is done by dispersing the sample between two phases: mobile phase and stationary phase. The mobile phase is a chemically inert gas that is used to carry molecules of the analyte through the heated column. The stationary phase is either a solid adsorbent (gas-solid chromatography) or a liquid on an inert support (gas-liquid chromatography). 

Affinity chromatography is a separation technique based on molecules specific interactions with immobilized ligands. Some examples of these interactions are enzyme/substrate, enzyme/inhibitor, and antibody/antigen interactions. The molecule of interest is typically dissolved in the mobile phase and a ligand is typically immobilized in the stationary phase.

Ion exchange chromatography is used to separate charged ionizable molecules on the basis of differences in charge properties. More specifically, it is based on the different degrees of electrostatic interactions between the stationary phase and solutes.

Additional resources

Separation techniques: Chromatography

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