chiral HPLC column

Chiral HPLC columns are made by immobilising single enantiomers onto the stationary phase. Resolution relies on the formation of transient diastereoisomers on the surface of the column packing. The compound which forms the most stable diastereoisomer will be most retained, whereas the opposite enantiomer will form a less stable diastereoisomer and will elute first.

The forces that lead to this interaction are very weak and require careful optimisation by adjustment of the mobile phase and temperature to maximise selectivity. Typically a free energy of interaction difference of only 0.03 kJ/mol between the enantiomers and the stationary phase will lead to resolution.

The effect of temperature is important in chiral HPLC. Lower temperature will increase chiral recognition, but as it alters the kinetics of mass transfer, it may actually make the chromatography worse by broadening peaks. There is often an optimum temperature for a separation and knowledge of this gives the analyst another factor to exploit in the method development process

The type of column used for separating a class of enantiomer is often very specific, this combined with the high cost of chiral columns, makes the choice of which column to use very critical.