Enantiomeric separation of amino alcohols on protein phases using statistical experimental design. A comparative study.

Two LC supports often used for giving enantioselective retention were tested and compared in the reversed-phase mode using statistical experimental design. The two supports contain two different proteins, alpha1-acid glycoprotein or cellulase immobilised to silica particles, as the chiral selectors. The two chromatographic columns are commercially available as Chiral-AGP and Chiral-CBH. Twelve closely structurally related amino alcohols were used as the testing solutes. For each column three important mobile phase descriptors, that improve the chiral recognition, were chosen as variables and retention and separation factors were used as responses. All the tested solutes were separated using the two protein based supports. However, the highest enantioselectivities, i.e., separation factors higher than 10 were obtained using the Chiral-CBH column. The solute structure, e.g., distance between the nitrogen atom and the chiral carbon atom, and position as well as type of substituent in the aromatic ring highly influence the enantioselectivity on both columns. For one of the solutes the choice of mobile phase composition could be used to control the retention order of the two enantiomers.