Size Exclusion Chromatography / Gel Permeation Chromatography: An Introduction in 30 Minutes

The bulk physical properties such as strength and toughness of materials such as synthetic and natural polymers, or the activity and efficacy of proteins and biopharmaceuticals are strongly dependent on their molecular properties. In a large number of industries, there is a clear and strong desire to control molecular properties such as molecular weight and structure in order to better manipulate and control the bulk properties of the material. It is therefore necessary to have a reliable technique for making such measurements. 

Size Exclusion Chromatography (SEC), also referred to as Gel Permeation Chromatography (GPC) or Gel Filtration (GF), is a separations technique and subset of High Performance Liquid Chromatography (HPLC), whereby polymer molecules are separated based on their hydrodynamic volume. Sample molecules, dissolved in a suitable solvent, pass through a gel packing matrix within a column and diffuse into and out of pores within the gel. Smaller molecules diffuse more frequently and deeper into the pores of the packing matrix so that their progress through the column is impaired, whereas larger molecules penetrate fewer of the pores. In this way, molecules in a mixed sample, such as a polydisperse polymer or a mixture of protein molecules, are organized according to their size. After the separation, one or more detectors is used to characterize the sample. This always involves a concentration detector and may or may not include advanced detectors such as light scattering and intrinsic viscosity. The primary goal of this characterization is to measure the molecular weight and molecular weight distribution but may also include the measurement of other properties such as size and structure. The measured values can then be used to understand the sample’s bulk properties.