Ion exchange chromatography (IEC)

Ion Exchange Chromatography (IEC)

The N and S ends on the magnet attract each other while N and N or S and S ends will repeal each other. The similar phenomenon occurs for electronics: Positive charge and negative charge attract each other while positive and positive or negative and negative charges repeal each other. When compounds dissociate and become ions, they will have electronic charges. Positive ion is called cation and negative ion is called anion.

Ion exchange mode uses ionic attraction and repulsion forces. What is different from magnet is that N end of magnet is always N, but molecular ion like protein can be an anion or a cation depending on the surrounding conditions (mobile phase). By changing the electronic characteristics of the mobile phase, the electronic charge of each component in the sample also changes. They may change from cation to anion (or vice versa) and this creates the interaction between the component and the packed gel. One time it may cause attraction and other time it may cause repulsion. Since the components that experience repulsion will be eluted out (and attraction causes retention) from the column, each component is separated based on their ionic strengths.

It is common to change the mobile phase during an analysis for ion exchange mode. Such method (changing mobile phase) is called gradient method. Most frequent method used for gradient elution is to prepare two mobile phases and change the ratio of two solvents over a time. Contrast to gradient method, the method uses single solvent throughout the run is called isocratic method.

Packed gel for ion exchange column is modified with either anion or cation functional groups. Commonly used functional groups are followings.

Quaternary ammonium (QA) : Strong anion exchanger

Diethyl aminoethyl (DEAE) : Weak anion exchanger

Sulfopropyl (SP) : Strong cation exchanger

Carboxylmethyl (CM) : Weak cation exchanger

Gels generally used are porous gel (having pores on the surface). However for the ion-exchange gel, small-sized gel (2.7 um) without pores is sometimes used. Gels without pores are called non-porous gel and it is effective for fast analysis. Both polymer-based and silica-based columns are used for ion-exchange columns. Though unlike RP columns that mainly uses silica base, ion-exchange columns requires use of alkali conditions sometimes, so importance of polymer-based materials is higher with ion-exchange columns than RP columns. Shodex IEC columns are made of Polyhydroxymethacrylate or Polyvinyl alcohol. Ion exchange is frequently used in biochemistry area such as separation of protein, peptide, and nucleic acids.