Chromatography

Chromatography separates the parts of a mixture so they can be identified and measured one by one. Every kind has a mobile phase that moves (a liquid or gas) and a stationary phase that stays put (paper, a coated plate, the packing in a column). Each substance spends some of its time stuck to the stationary phase; the more it sticks, the slower it travels, so the mixture spreads out.

In paper or thin-layer chromatography (TLC) a spot of sample is put near the bottom of a strip, and solvent creeps up past it. When the solvent has nearly reached the top, you mark how far it got (the solvent front). Each substance's retention factor, Rf = distance it moved ÷ distance the solvent moved, is between 0 and 1 and is fairly constant for a given substance, plate and solvent.

Gas chromatography (GC) pushes a vaporized sample through a long, thin coated column with a carrier gas such as helium. Liquid chromatography, usually high-performance (HPLC), pumps a liquid through a column packed with tiny particles at high pressure. GC suits things that vaporize without breaking down (fuels, solvents); HPLC handles the rest (drugs, sugars, proteins).

A detector at the end draws a chromatogram: a flat baseline with a peak each time something comes out. The time a substance takes is its retention time; matched against a standard run the same way, it helps identify it. The area under the peak is proportional to the amount, so with a calibration you can measure concentration too.

Good separation needs peaks that don't overlap. You can change the column, the solvent mix, the flow rate or (in GC) the temperature program. Often a mass spectrometer is the detector, so each peak gets identified as it comes out (GC-MS, LC-MS).