Preparation and Application of a New Ion-Pairing Chiral Stationary Phase for the Liquid Chromatographic Resolution of N-(3,5-Dinitrobenzoyl)-α-amino Acids

Abstract

A liquid chromatographic chiral stationary phase (CSP) was prepared starting from (1S, 2S)-1,2-diaminocyclohexane. The CSP containing two simple functional groups, such as a primary amino group and an ureide tethering linkage on a cyclohexane ring, was quite successful with the use of 60% 2-propanol in hexane containing 0.5% acetic acid as a mobile phase for the resolution of various N-(3,5-dinitrobenzoyl)-α-amino acids, the separation (α) and the resolution factors (R _S ) being in the range of 1.12–1.23 and 1.33–2.99, respectively. In contrast, N-(3,5-dinitrobenzoyl)-α-amino acid ethyl esters and N-(3,5-dinitrobenzoyl)-α-amino acid amides were not resolved at all, or resolved very poorly on the CSP. From these results, it was concluded that the ionic or ion-paring interaction between the CSP and analytes plays a very important role for the chiral recognition. The chromatographic behaviors for the resolution of N-(3,5-dinitrobenzoyl)-α-amino acids on the CSP were found to be controlled with the variation of the type and the content of alcohol and/or acid in hexane.

Keywords:

• Chiral separation
• Ion pairing chiral stationary phase
• Liquid chromatography
• N-(3,5-Dinitrobenzoyl)-α-amino acids

ACKNOWLEDGMENTS

This work was supported by grant-in-aids for the National Core Research Center Program (No. R15-2007-022-03001-0) and for the National Research Laboratory (NRL) Program (No. M1-0318-00-0005) from MOST and KOSEF.

Notes

^a Mobile phase: 60% 2-propanol in hexane + 0.5% CH₃COOH. Flow rate: 0.5 mL/min. Detection: 254 nm UV. Temperature: 20°C. k ₁: Retention factor of the first eluted enantiomer (the absolute configuration of the first eluted enantiomer is presented in the parenthesis). k ₂: Retention factor of the second eluted enantiomer (the absolute configuration of the second eluted enantiomer is presented in the parenthesis). α: Separation factor. R _S : Resolution factor.

^a Mobile phase: 60% 2-propanol in hexane +0.5% CH₃COOH. Flow rate: 0.5 mL/min. Detection: 254 nm UV. Temperature: 20°C. k ₁: Retention factor of the first eluted enantiomer. k ₂: Retention factor of the second eluted enantiomer. α: Separation factor. R _S : Resolution factor.

^a Mobile phase: 60% 2-propanol in hexane +0.5% CH₃COOH. Flow rate: 0.5 mL/min. Detection: 254 nm UV. Temperature: 20°C. k ₁: Retention factor of the first eluted enantiomer. α: Separation factor. R _S : Resolution factor.

^a Flow rate: 0.5 mL/min. Detection: 254 nm UV. Temperature: 20°C. k ₁: Retention factor of the first eluted enantiomer. k ₂: Retention factor of the second eluted enantiomer. α: Separation factor. R _S : Resolution factor.