Combination of capillary electrophoresis, Quality by Design, NMR and molecular modeling for impurity profiling: definition of the design space and investigation of intermolecular affinities, complexation and separation mechanisms

Quality by Design principles have been applied for the set-up of various capillary electrophoresis (CE) methods for the determination of drugs, of their related substances and their enantiomeric purity. The use of a risk-based approach in the development of a separation method for impurity assay has been recently reported with success, as it makes it possible an in-depth understanding of parameters affecting analytical method performances. The design space (DS) concept requires the definition of a multidimensional region where the quality of analytical data is assured with a selected probability and can be defined by means of mathematical models representing a step forward in the comprehension of the electrophoretic behaviour of the analytes. Pseudostationary phases involving micelles, microemulsions and cyclodextrins (CDs) were used for CE separation. Molecular Dynamics (MD) and NMR studies were employed to elucidate separation mechanisms, host-guest interactions and intermolecular affinities and confirmed the CE experimental results. Aspects such as the affinity pattern of analytes towards various CDs as well as the equilibrium constants and the structure of complexes were addressed. The effect of cosurfactant n-butanol and the role of ionic surfactant sodium dodecyl sulphate (SDS) on separation selectivity were investigated. Capacity factors and effective mobilities of the solutes were calculated and compared with the potential and the gain energy of inclusion complexes of analytes with surfactant and cosurfactant. MD simulations and NMR experiments underlined the ability of CDs of including the SDS monomer forming inclusion complexes and the pivotal role of this surfactant in modulating the different affinities of the analytes for chiral selectors.