Immobilisation and evaluation of a vancomycin chiral stationary phase for capillary electrochromatography

The macrocyclic antibiotic, vancomycin, is covalently bonded to LiChrospher ® diol silica packed columns and evaluated in capillary electrochromatography (CEC) both in the reversed-phase and polar organic mode. Initially, capillaries were packed with 5 μm LiChrospher ® 100 Å diol silica and evaluated in CEC with a reversed-phase biphenyl-pyrene achiral test resulting in resolution and efficiency values of ca. 2.5 and 100 000 plates meter −1, respectively. Repeatability for this test (resolution and efficiency) was also examined and found to be acceptable for both run-to-run ( n=5, 0.74% and 1.5%) and column-to-column ( n=5, 3.4% and 9.0%), respectively. Similar results were obtained when the 10 μm LiChrospher ® 1000 Å diol silica was examined with the exception of efficiency, where a reduced plate height value of four times lower was obtained compared to the 100 Å material. A simple three step in-situ vancomycin immobilisation procedure was subsequently carried out on these packed diol columns. Selectivity was obtained for thalidomide enantiomers on this vancomycin chiral stationary phase in reversed-phase CEC with resolution and efficiency values of ca. 2.5 and 80 000 plates meter −1, with acceptable repeatability ( n=8) 0.9% and 3.0%, respectively. Selectivity was also obtained for thalidomide enantiomers on this phase in the polar organic mode with resolution and efficiency values of ca. 2.5 and 120 000 plates meter −1, with acceptable repeatability ( n=7) 0.9% and 2.0%, respectively. It was possible to deduce from a chemometric design carried out for evaluating the mobile phase component effects that organic modifier ratio, MeOH/MeCN, played a significant role in controlling both resolution and efficiency. It was also possible to separate a number of basic analytes including four β-adrenergic blocking agents in the polar organic mode albeit with lower resolution and efficiency values, ca. 1.5 and 45 000 plates meter −1, respectively.