RESOLUTION OF ENANTIOMERS OF ALCOHOLS AND AMINES BY HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY AFTER DERIVATIZATION WITH A NOVEL FLUORESCENT CHIRAL REAGENT

4-(2-Chloroformylpyrrolidin-1-yl)-7-nitro-2,1,3-benzoxadiazole [R(+)-NBD-Pro-COCl and S(-)-NBD-Pro-COCl], optically active tagging reagents, have been synthesized for resolution of enantiomers of amines and alcohols by high-performance liquid chromatography. The reagents react with amino and hydroxyl functional groups in the presence of pyridine to produce the corresponding diastereomers. The optimum excitation and emission wavelengths for the diastereomers in water-acetonitrile (1:1) were approximately 485 nm and 530 nm, respectively. The excitation and emission wavelengths were independent of the amine or alcohol portion of the diastereomer. The resulting diastereomers can usually be efficiently resolved by normal-phase chromatography with n-hexane-ethyl acetate as the eluent. When R(+)-NBD-Pro-COCl was used as the derivatization reagent, the diastereomers corresponding to the R-configurations of amines and alcohols were eluted faster than those from the S-configuration. The elution order was reversed when the diastereomers were prepared with S(-)-NBD-Pro-COCl. The R(s) values of the diastereomers derived from amines and alcohols by normal-phase chromatography are in the range of 3.23-4.32 and 2.99-4.10, respectively. After derivatization with NBD-Pro-COCls the alcohol enantiomers were also separated adequately by a reversed-phase column with a water-acetonitrile mixture.