5.1: Overview of Chromatography

Chromatography is a technique used to separate the components of a mixture. It can be used as an analytical technique to gain information about what is present in a mixture, or as a purification technique to separate and collect the components of a mixture. Chromatography in the organic chemistry laboratory can be classified into several broad categories. These techniques follow the same general principles in terms of how they are able to separate mixtures, and so will be discussed collectively in this section.

The general idea of chromatography can be demonstrated with food dyes in your kitchen. Commercial green food dye does not contain any green colored components at all, and chromatography can show that green food dye is actually a mixture of blue and yellow dyes. If a drop of green food dye is placed in the middle of a paper towel followed by a few drops of water, the water will creep outwards as it wets the paper. As the water expands, the dye will travel with it. If you let the dye expand long enough, you'll see that the edges will be tinted with blue. This is the beginning of the separation of the blue and yellow components in the green dye by the paper and water.

Time-lapse sequence of a drop of green food dye on a paper towel, followed by a drop of water.

A complete separation of the green food dye can be accomplished using paper chromatography. A dilute sample is deposited on the bottom edge of a piece of paper, the paper is rolled in a cylinder, stapled, and placed vertically in a closed container containing a small amount of solvent. The solvent is allowed to wick up the paper through capillary action (called "elution"), and through this method complete separation of the blue and yellow components can be achieved.

Paper chromatography of diluted green food dye: a) Before elution, b) After elution, c) Zoom in before elution, d) Zoom in after elution.

In all chromatographic methods, a sample is first applied onto a stationary material (stationary phase) that either absorbs or adsorbs the sample: adsorption is when molecules or ions in a sample adhere to a surface, while absorption is when the sample particles penetrate into the interior of another material. The sample is then exposed to a liquid or gas traveling in one direction (mobile phase). The sample may overcome its intermolecular forces with the stationary surface and transfer into the moving material. The sample will later readsorb to the stationary material, and transition between the two materials in a constant equilibrium.

Generic chromatography sequence for compound X: a) Adsorption, b) Exposure to a mobile phase, c) Sample X breaks its attachment to the stationary phase, d) Movement with the mobile phase, e) Reattachment to the stationary phase. Steps c-e are equilibrium steps and constantly repeated.

The main principle that allows chromatography to separate components of a mixture is that components will spend different amounts of time interacting with the stationary and mobile phases. A compound that spends a large amount of time in the mobile phase will move quickly away from its original location, and will separate from a compound that spends a larger amount of time stationary. The main principle that determines the amount of time spent in the phases is the strength of intermolecular forces experienced in each phase. If a compound has strong intermolecular forces with the stationary phase it will remain adsorbed for a longer amount of time than a compound that has weaker intermolecular forces. This causes compounds with different strengths of intermolecular forces to move at different rates.

Adapted from Chromatography Generalities by Lisa Nichols.