AQbD-Oriented UHPLC/MS/MS Method Development for Glycopeptides Assessment in Pharmaceutical Forms.

Abstract

Vancomycin and teicoplanin are glycopeptide antibacterials that inhibit the bacteria cell wall synthesis showing activity against gram-positive bacteria. Development of the sensitive method is of great importance for quality control of these drugs that are fermentation products. Modification of the fermentation conditions could cause the differences in the relative amount of the total substance or component, as it is the case with teicoplanin. The main objective of this study was development of the sensitive and effective ultra high performance liquid chromatography - tandem mass sprectrometry (UHPLC-MS/MS) method for simultaneous quantification of vancomycin, all six subcomponents of teicoplanin, and its pharmacopoeial impurity A in pharmaceutical forms. The scientific-based Quality by Design approach was implemented in the MS and UHPLC method development. Detection and quantification of analytes were carried out in positive electrospray ion mode by multiple reaction monitoring. Capillary voltage, cone voltage and collision energy were optimized by implementing experimental design methodology and optimal values for each fragment ion were obtained by performing experiments according to 'Rechtschaffen' design matrix. An ACQUITY CSH Phenyl-hexyl (2.1 × 50mm, particle size 1.7μm) column was chosen for the separation under the gradient elution mode with the mobile phase consisted of 0.1% formic acid in water (mobile phase A) and acetonitrile (mobile phase B). Optimal gradient elution parameters were achieved by applying 'Rechtschaffen' design too. Method operable design regions were constructed for investigated MS and chromatographic parameters. The method was fully validated, and its applicability was confirmed throughout the ability to follow the behavior of vancomycin and teicoplanin under stress conditions.