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Abstract
The use of vancomycin as a chiral selector was introduced by D. W. Armstrong. This macrocyclic antibiotic was used either free in solution for thin-layer chromatography (TLC) and capillary electrophoresis (CE) or covalently bonded to silica gel in the case of high performance liquid chromatography (HPLC). Its possible use in countercurrent chromatography (CCC) or centrifugal partition chromatography (CPC) is also of interest since these techniques have preparative capabilities which make them attractive for multi-gram scale production of enantiomers.
Two immiscible liquid phases coexist: an aqueous stationary phase containing vancomycin, and an organic mobile phase in which the racemic compound is more partitioned. This has a number of advantages over HPLC: lower stationary phase costs, ease of recovery of the antibiotic and exchange with a fresh batch, and, especially, improved resolution due to higher selectivity (α).
Surprisingly, our experiments showed that chiral recognition in CCC & CPC is strongly solvent-sensitive. Acetonitrile, used by D. W. Armstrong at 10% or higher concentrations, was found to inhibit enantio-selectivity at levels as low as 5%. Inhibition was observed with most solvents, with the notable exception of toluene and related aromatics bearing no other heteroatom than halogens in their molecule. Exceptionally high selectivity factors (α > 30) were observed for dansyl-norleucine in toluene-, ethylbenzene- and hexafluorobenzene- vancomycin 0.1 M (pH 4.7) systems. In a CPC run using toluene, injection of 50 mg dansyl-norleucine led to the isolation of 20.4 mg L isomer () and 24.3 mg D isomer (
). In HPLC using the commercial vancomycin Chiroptic V® column, both enantiomers were found to be free of contamination (ee # 100%).
ACKNOWLEDGMENTS
We are grateful to Sophie Boisrobert for skillful assistance during preliminary studies, and to Dr. R. D. G. Cooper, from E. Lilly Research Laboratories, for kindly providing us with the pKa values of pure vancomycin.