Preparation And Evaluation Of Vancomycin-boned Stationary Phase

Chromatographic stationary phase is the core of the chromatographic separation. The stationary phases prepared by bonding functional groups on the surface of the substrate with different modification methods can provide different selective separation effects. Vancomycin as a macrocyclic antibiotic, has complex and unique molecular structure. As a surface modification agent, it can produce a variety of intermolecular interactions with the sample molecules. In this paper, according to the unique molecule structure of vancomycin, vancomycin chiral capillary monolithic column and vancomycin bonded silica stationary phase were prepared by monomer derived method and indirect method, respectively. The latter was appied to the reverse, hydrophilic and ion exchange modes.Monolithic column has the characteristics of simple preparation, high column capacity and easy modification on the surface of the stationary phase. Vancomycin chiral capillary monolithic column was obtained based on polyacrylamide integral substrate by monomer derivative method. The optimization of column preparation conditions were using formamide as solvent; under alkaline conditions; vancomycin: GMA=1:5(mol/mol); reaction temperature=50℃; reaction time=6h. Chromatographic column were used to separate amino acid enantiomers on the constant column pressure mode of micro column liquid chromatography. When buffer solution ionic strength was10mmol/L, pH=4.5and the column pressure was5.0Mpa, resolution of phenylalanine enantiomers was up to1.49; while buffer solution ionic strength was10mmol/L, pH=4.5and column pressure was4.7Mpa, resolution of tyrosine and phenylalanine enationmers were2.99and1.95, respectively.Vancomycin-boned chiral stationary phase was prepared, by using amino bonded silica particles of10μm as matrix and glutaraldehyde as spacer. The column showed good separation ability after repeating injection of300times. Application in the reversed, hydrophilic and anion exchange modes confirmed that the column not only can be used in different separation modes for achiral substances. Good separation performance was obtained for the separations of nepafenac, drugs, stevia sugar and organic acid samples in different separation modes. Hence, it further expands the application range of vancomycin-bonded stationary phase and provides guidance for designation of new type stationary phase and improve method development of chromatographic stationary phase modified by special compound with complex construction in corresponding separation mode.