Preparation Of HILIC Stationary Phases And Their Applications

Along with the rapid development of bioscience, medical science, food chemistry and environmental science, the separation of polar compounds is becoming a hot topic in chromatographic science. However, traditional chromatographic technology such as normal phase liquid chromatography (NPLC), reverse phase liquid chromatography (RPLC), ionic exchange chromatography (IPC) and Mixed-mode chromatography (MMC) face huge challenges in the separation of these polar compounds.Hydrophilic interaction chromatography (HILIC) provides a perfect solution for separation of polar compounds. It overcomes the drawbacks of the poor retention and selectivity in RPLC. For the mobile phase employed, HILIC can be conveniently coupled to electrospray ionization mass spectrometry (ESI-MS). Peak symmetry of basic compounds in the HILIC often improves in comparison to RPLC. Benefit from these advantages, HILIC has gradually found useful applications in the analysis of various polar compounds. However, with the increased diversity and complexity of biological and environmental samples, it's very necessary to develop new stationary phases to widen the available selectivity range in HILIC.In this dissertation, the author prepared some new HILIC stationary phases with different functionalities via several synthetic methods. A series of standard polar compounds were employed to investigate the retention mechanisms of these phases. Good separation performances were observed when these phases were further applied to separate some standard polar compounds, demonstrating excellent application potential in HILIC. The major contents of the dissertation include the following aspects.1. A novel phosphate ester-bonded silica (PES) stationary phase was prepared based on "thiol-ene" click chemistry. Due to its hydrophilic phosphate-ester groups and short hydrophobic alkyl chains, the PES stationary phase exhibited dual retention mechanism via complexation and hydrophobic interactions with phenols. Benefiting from this special interaction mechanism, the newly synthesized PES stationary phase showed good selectivity in the separation of phenols. Furthermore, the separations of 10 nucleosides and nucleobases on the PES stationary phase were achieved in both RPLC mode and HILIC mode.2. A novel zwitterionic stationary phase (APS) was synthesized by further reaction on di-amino-modified porous silicon surface. A series of standard polar compounds were employed to investigate the retention mechanisms. Effective separation of several small polar compounds including water-soluble vitamins, nucleosides and nucleobases, and organic acids was achieved in the HILIC mode.3. A new type of zwitterionic stationary phases with controllable ratio of positively charged tertiary amine groups and negatively charged carboxyl groups were synthesized in a facile manner. The elemental analysis of different batches of ACS phases demonstrated good reproducibility of the preparation strategy. Various kinds of mixtures were employed to evaluate the separation selectivity of the three ACS stationary phases. The three ACS columns exhibited different selectivity and retention behavior for different polar analytes, demonstrating their excellent application potential in the analysis of polar compounds in HILIC.4. A new type of N-acryloytris(hydroxymethyl) aminomethane-bonded silica (NAS) stationary phase was synthesized based on "thiol-ene" click chemistry. A series of standard polar compounds were employed to investigate the retention mechanisms. Due to its hydrophilic triolacrylamide groups and short hydrophobic alkyl chains, the NAS phase was successfully applied in both RPLC mode and HILIC mode. Good chromatographic performance for the separation of alkylbenzenes and nucleosides was obtained under RPLC and HILIC mode respectively.