Chiral separation of selected proline derivatives using a polysaccharide type stationary phase by high-performance liquid chromatography

Abstract

Proline derivatives, such as Boc-proline, Boc-2-methylproline, Boc-2-methylproline benzyl ester and Boc-2-methyl-4-hydroxy-proline benzyl ester, have been widely used as a building block leading to a variety of pharmaceutical compounds. Therefore, there is a wide interest in the chiral separation of these compounds. High-performance liquid chromatography (HPLC) methods were developed using a Chiralpak AD-H column to separate enantiomers of these proline derivatives. The effect of mobile phase composition and column temperature was studied. For the proline derivatives studied in this work, good resolution was achieved using a mobile phase composition of hexane, ethanol and 0.1% TFA. For prolines containing carboxyl or hydroxy group, resolution was changed dramatically corresponding to changes as little as 1% of ethanol in the mobile phase, suggesting that the dominant chiral recognition is from hydrogen bonding interactions. On the other hand, for prolines containing a benzyl ester instead of hydroxy group next to the chiral center, resolution was not affected as significantly with the changes of ethanol content in the mobile phase, indicating a different leading chiral recognition mechanism, such as inclusion, steric effect, or possible π–π interaction. Linearity, precision and limit of detection were also measured for Boc-2-methylproline and Boc-2-methylproline benzyl ester.