Doctor of Philosophy
Maryanne M. Collinson
Choosing a stationary phase is the first step in developing a liquid chromatography (LC) separation method, where the selectivity is governed by the differential interactions of the analytes with the stationary and mobile phases. Introducing a gradient in stationary phase functionality allows for tuning of analyte retention, translating to a possible improvement in selectivity and an increase in resolution versus that offered by uniform stationary phases.
In this work, C18-silica, phenylbutyl-silica, and phenylbutyl-ammonium opposed continuous stationary phase gradients were fabricated using controlled rate infusion (CRI) on particle packed LC columns. Characterization of the stationary phase was carried out using spectroscopy and LC analysis to relate the ligand density gradient profile to the observed chromatographic parameters.
C18-silica gradients were created with a time-dependent acid hydrolysis infusion and demonstrated an increase in resolution when combined with a mobile phase gradient. Phenylbutyl-silica and phenylbutyl-ammonium gradients were produced using an in-situ silanization CRI method. Phenylbutyl-silica gradients were confirmed to be stable and reproducible; however, produced tailing peak shapes. Phenylbutyl-ammonium gradients were utilized to incorporate an ion exchange model into a simulator built by Jeong et al. The phenylbutyl-ammonium gradient was not reproducible but did exhibit an increase in resolution when combined with a mobile phase gradient. Also, the ion exchange model was successfully added within the simulator, with percent differences for retention prediction all under 5 %. This dissertation serves as a proof-of-concept for gradient stationary phases on particle packed LC columns.
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Available for download on Tuesday, May 07, 2024