RELATIONSHIP BETWEEN THE ASSOCIATION CONSTANT AND ENANTIOSELECTIVITY ON THE FLAVOPROTEIN-CONJUGATED CHIRAL STATIONARY-PHASE FOR HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

The relationship between the enantioselectivity and association consta nts of enantiomers on high-performance liquid chromatography with a pr otein-conjugated chiral stationary phase (CSP) was investigated using a flavoprotein-conjugated CSP, with ketoprofen (KP) as a model drug. T he association constants of the enantiomers for native flavoprotein we re different; it has been found that they contribute to the chiral sep aration on a protein CSP. Equations including the association constant s were developed to calculate, the capacity factors and enantioselecti vities on protein CSPs; they were validated by means of experiments us ing a flavoprotein column in combination with variable numbers of non- specific columns in series. The capacity factors and enantioselectivit ies calculated with the developed equations showed reasonable agreemen t with those obtained by chromatography employing a conventional flavo protein column (150 mmX4.6 mm i.d.) with a mobile phase containing met hanol as an organic modifier. Our results imply that protein-conjugate d CSPs are complex-stationary phases consisting of both a specific pha se for chiral separation and a non-specific phase, and that the resolu tion of the enantiomers depends upon the balance of interactions of so lute molecules with the two phases.