Size-Exclusion Chromatography

Size-exclusion chromatography, or SEC, involves separating molecules based on size.

Here, the stationary phase can be silica particles or cross-linked polymer resin beads that contain a network of uniform pores.

Large molecules are excluded from entering the pores and eluted first, whereas very small molecules are retained the longest and eluted last.

Intermediate-sized molecules are separated in the retention window and undergo fractionation.

The retention time of the molecules is directly influenced by their size.

The experimental calibration curve obtained using standard substances is then used to estimate the unknown molecular mass.

Size exclusion chromatography using hydrophilic packing is known as gel filtration and is applied to water-soluble samples. Hydrophobic packing materials are used in gel permeation chromatography for substances soluble in less-polar organic solvents.

Both these methods are useful for the separation of polymers or macromolecules.