How does SEC work?
Posted: Wed 29 Jun 2016 12:22How does SEC work?
FAQ from TOSOH
Molecules in solution assume 1 of 3 possible configurations: linear, e.g. PEG; branched, e.g. dextran; and globular, e.g. protein. As such a given molecule will have a unique hydrodynamic radius in solution. It is the differences in molecular dimensions that enables separation of compounds in a mixture by means of size exclusion liquid chromatography. Whereas large molecules will have limited or no access to the pores in an SEC column, smaller species will fit in the pores and be retained. Thus, the molecules will elute from the column in reverse order of molecular weight. Unretained compounds will come out in the column’s void volume (the space between particles in the resin bed). A fully retained molecule will elute at the column’s included (or mobile phase) volume which is the sum of the void volume and pore volume.
FAQ from TOSOH
Molecules in solution assume 1 of 3 possible configurations: linear, e.g. PEG; branched, e.g. dextran; and globular, e.g. protein. As such a given molecule will have a unique hydrodynamic radius in solution. It is the differences in molecular dimensions that enables separation of compounds in a mixture by means of size exclusion liquid chromatography. Whereas large molecules will have limited or no access to the pores in an SEC column, smaller species will fit in the pores and be retained. Thus, the molecules will elute from the column in reverse order of molecular weight. Unretained compounds will come out in the column’s void volume (the space between particles in the resin bed). A fully retained molecule will elute at the column’s included (or mobile phase) volume which is the sum of the void volume and pore volume.