Comparative study on separation of diastereomers by HPLC

Summary Reversed (RP-HPLC) and normal phase chromatographic (NP-HPLC) separations have been developed for diastereomers ofN-acyl-1-methyl-1,2,3,4-tetrahydo-β-carbolines which are acylated derivatives of simple natural β-carboline alkaloids. Separations of derivatives having different acyl moieties in theO,O-diacyl-tartaric acid ester subtituent differed remarkably. Little or no resolution in either NP-HPLC or RP-HPLC could be achieved with the diacetyl-tartrate derivative. Base-line separation by RP-HPLC but no separation by NP-HPLC was possible with the bulkier and more apolar dipivaloyl derivative. Retention order of the bis-benzoylated diastereomers was reversed and separation time increased dramatically by RP-HPLC. Good separation of the medium polarity, but rigid,N-camphanyl derivative by NP-HPLC has been achieved, whereas RP-HPLC could not be used for separation of these diastereomers. Separability of different diastereomers was highly dependent on polarity and rigidity of the derivatizingN-acyl moieties. Conformational analysis by molecular mechanics and comparison of the lowest energy conformational states of the diastereomers was applied to rationalise separation-retention behaviour of stereoisomers by RP-HPLC. Presented at Balaton Symposium '01 on High-Performance Separation Methods, Siófok, Hungary, September 2–4, 2001