Directly Coupled Chiral HPLC−NMR and HPLC−CD Spectroscopy as Complementary Methods for Structural and Enantiomeric Isomer Identification:  Application to Atracurium Besylate

Abstract

Directly coupled HPLC−NMR spectroscopy has now become a standard, commercially available technique for mixture characterization. Here the extension of the technique to chiral HPLC separation is reported and it is shown that HPLC−NMR together with HPLC−CD provide complementary approaches for the identification of structural isomers and enantiomers. The general approach has been exemplified using the neuromuscular blocking agent atracurium besylate, which comprises a mixture of 10 isomers in various proportions as four racemic pairs and two meso compounds. Diagnostic reporter resonances in the 1H NMR spectrum of atracurium besylate were assigned using a combination of one-dimensional and two-dimensional NMR experiments at 750 MHz. Stop-flow 750-MHz 1H NMR spectroscopy was used on-line after chiral column HPLC separation to identify the enantiomeric pairs, to distinguish the meso compounds, and to identify key configurational features of the isomers. The parallel HPLC−CD experiments served to assign the enantiomers based upon the known CD and absolute stereochemistry of (R)-laudanosine hydrochloride, an analogue with the same tetrahydroisoquinoline structural unit as atracurium. It is thereby demonstrated that high-field HPLC−NMR and HPLC−CD is a powerful combination of techniques which could be combined on-line for mixture characterization.

Development of an HPLC-NMR method for the determination of PAHs in soil samples – a comparison with conventional methods

“…b According to the IUPAC nomenclature [ As shown by us [14] and others [19,20,21], the coupling of HPLC and NMR leads to a significant enhancement of selectivity especially for aromatic compounds. As an example, Fig.…”

A Photometric Screening Method for Dimeric Naphthylisoquinoline Alkaloids and Complete On-Line Structural Elucidation of a Dimer in Crude Plant Extracts, by the LC−MS/LC−NMR/LC−CD Triad

“…In different combinations, these hyphenated techniques have already proven their value for the analysis of complex matrixes. [20][21][22][23][24] The on-line elucidation of relative or absolute configurations of chiral molecules right from crude extracts, however, without the use of chiral chromatographic phases, is still a demanding task, which has been accomplished only rarely. [25][26][27][28][29] In this paper, we present a screening procedure for this pharmacologically promising class of dimeric naphthylisoquinoline alkaloids.…”

High‐performance liquid chromatographic enantioseparation of naphthol‐substituted tetrahydroisoquinolines on polysaccharide‐based chiral stationary phases

“…The normal‐phase separation of ( S )‐ and ( R )‐1‐phenyl‐1,2,3,4‐Tiq was accomplished using an immobilized polysaccharide‐based Chiralpak IC column with different propan‐2‐ol or ethanol– n ‐hexane–ethanolamine mobile phases (Kazoka et al ., ). After chiral column separation 750 MHz H‐1 NMR spectroscopy was used on‐line to identify the enantiomeric pairs of atracurium besylate (Mistry et al ., ).…”