Size exclusion chromatography

Size Exclusion Chromatography (SEC) consists on the separation technique based on the molecular size of the components, used for semi-preparative purifications and various analytical assays. It is a separation technique which takes the advantage of the difference in size of the molecules. separation is achieved by the differential exclusion from the pores of the packing material, of the sample molecules as they pass through a bed of porous particles. The principle feature of SEC is its gentle non-adsorptive interaction with the sample, enabling high retention of biomolecular activity.

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 Principe

Size exclusion chromatography (SEC) is the separation of a mixture based on the molecular size of the components in this mixture. The separation is achieved by the differential exclusion or inclusion of solutes as they pass through stationary phase consisting of pores of different sizes cross linked polymeric gels or beads. The process is based upon different permeation rates of each solute molecule into the interior of gel particles. Size exclusion chromatography involves gentle interaction with the sample, enabling high retention of biomolecular activity. The separation of organic polymers in non-aqueous systems is called gel permeation chromatography (GPC).

A column of  porous matrix is in equilibrium with a suitable mobile phase for the molecules to be separated. Large molecules are excluded from the pores will pass through the space  between the matrix and will come first in the effluent. Smaller molecules will get distributed in between the mobile phase of in and outside the molecular sieve and will then pass through the column at a slower rate,so appear later in effluent.

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