Adaptation of high-performance liquid chromatographic methods to multiple spectral analyses

Abstract

Adapting well-characterized existing liquid chromatographic separations for compatibility with mass spectrometric detection often requires changes in chemical and/or chromatographic conditions. Non-volatile solvent modifiers, such as phosphates or alkylsulfonates, have been widely used in chromatography because of their buffer or ion-pairing capacities, coupled with advantageous low-UV-wavelength absorption characteristics. Non-volatile components in the eluent stream are generally avoided in mass spectrometry as they quickly decrease the detector response. In addition to issues such as buffer capacity and effective ion-pairing, consideration should be given to the impact of stationary phase and column geometry. The work presented here illustrates advantages of a new high-purity, high-pore-volume stationary phase used in Waters Symmetry columns when attempting to accommodate a wide range of conditions imposed by different detection schemes. The effect of column geometry under a variety of typical LC-MS conversion situations is also included. The studies include resolution of critical hydrophilic pairs and stationary-phase loading performance relative to converting existing separations for LC-MS analysis while achieving maximum sensitivity and maintaining chromatographic fidelity.