Rationale: Mixed-mode reversed-phase/anion exchange liquid chromatography is useful for separations of mixtures containing anions (e.g. ionized acids). However, when using this form of liquid chromatography with mass spectrometry detection, the bleed of amine-containing hydrolysis products from the columns may cause ion suppression or enhancement.
Methods: Using electrospray ionization tandem quadrupole mass spectrometry detection, we determined the ion suppression or enhancement caused by column bleed for three mixed-mode reversed-phase/weak anion-exchange columns containing stationary phases that differ in chemical structure. Two of the stationary phases are based on silica particles, while the third uses ethylene-bridged hybrid organic/inorganic particles, which have improved hydrolytic stability. Mixtures of acidic and basic analytes were combined with the chromatography flow postcolumn, both with and without a column, and their mass spectrometry ion signal responses (peak areas) were determined. The ratio of signal response with and without a column is the matrix factor. Positive ion electrospray measurements were carried out using 0.1% formic acid (pH ~ 2.7) as a mobile phase additive, and 10mM ammonium formate (pH ~ 6.4) was used for negative ion electrospray detection.
Results: The matrix factors under both positive and negative ionization modes were closest to 1 (0.74-1.16) for the hybrid particle-based columns, showing minimal ion suppression or enhancement. In contrast, the silica-based columns gave matrix factors ranging from 0.04 to 1.86, indicating high levels of ion suppression or enhancement. These results may be explained by the differences in the structures of the stationary phases, which affect the relative amounts of hydrolysis products that elute from the columns.
Conclusions: The low levels of mass spectrometry ion suppression or enhancement caused by column bleed from the hybrid particle-based columns should allow for accurate quantitative mass spectrometric detection combined with mixed-mode reversed-phase/weak anion-exchange chromatography.
© 2021 The Authors. Rapid Communications in Mass Spectrometry published by John Wiley & Sons Ltd.