Chromatography is a means of separating molecules of varying size and mass. It is not a definitive means of measuring mass, mass spectrometers are necessary for these measurements. Nor does it analyse the actual molecules' structure, it simply splits a mixture into component molecules.
The technique goes back a long way to when Michail Tswett separated plant pigments using a column of chalk about 100 years ago.
Actual chromatograph output screen captured courtesy of Tas Alkaloids
The process has two main parts, a moving part or phase, which is either a gas or liquid, and a stationary phase which may be a solid or liquid supported on a gel.
A very primitive but still effective piece of chromatography is felt-tip pen ink dropped on filter paper with a drop of water on top. The ink will separate into colours with time as the solution moves out into the paper. The inks interact with the paper, and the different molecules pass through at differing rates leading to separation.
The mobile phase is the diluted ink, the stationary phase, the paper. Here the ink components are measured at deference positions at a fixed time.
In column chromatography, the liquids or gases are passed along a fixed length, a column, and separate out to arrive at the end at different times.
FACTORS INFLUENCING CHROMATOGRAPHY.
Interaction of mobile to stationary phases. This process is often adsorption. ( Adsorption is a great word - absorption is the soaking INTO a solid, adsorption is the "locking" of mobile molecules ONTO the surface of the solid.) If a molecule adsorbs to the stationary surface better than another, it will take longer to pass down the column.
Length of column. Longer columns separate better but take longer to produce results.
Pressure under which the liquid/gas is forced through the column. HPLC is "High Pressure Liquid Chromatography" ( sometimes HPerformanceLC ) as the use of pumps to push the liquid/gas makes for better performance. The flow rate is increased.
Sensitivity of the detector to the separated molecules.
The stationary phase can typically be Silica Gel, alumina, cellulose, various resins
Detection can be through
ultraviolet absorption - UV is shone through the emerging column and changes in UV intensity noted
refractive index - when different chemicals emerge, refractive index changes are noted
conductivity - ionic compounds and inorganic ions are conductive, thus monitoring the resistance in a small cell detects materials flowing through.
Increasingly, the outputs of chromatographs are linked to mass spectrometers so the separated molecules can be analysed with respect to molecular mass. This aids identification.
