Fractionation is a separation process in which a certain quantity of a mixture (of gasses, solids, liquids, enzymes, or isotopes, or a suspension) is divided during a phase transition, into a number of smaller quantities (fractions) in which the composition varies according to a gradient. Fractions are collected based on differences in a specific property of the individual components. A common trait in fractionations is the need to find an optimum between the amount of fractions collected and the desired purity in each fraction. Fractionation makes it possible to isolate more than two components in a mixture in a single run. This property sets it apart from other separation techniques.

Fractionation is widely employed in many branches of science and technology. Mixtures of liquids and gasses are separated by fractional distillation by difference in boiling point. Fractionation of components also takes place in column chromatography by a difference in affinity between stationary phase and the mobile phase. In fractional crystallization and fractional freezing, chemical substances are fractionated based on difference in solubility at a given temperature. In cell fractionation, cell components are separated by difference in mass.