Epoxy-based monoliths for capillary liquid chromatography of small and large molecules

A versatile epoxy-based monolith was synthesised by polycondensation polymerisation of glycidyl ether 100 with ethylenediamine using a porogenic system consisting of polyethylene glycol, M(w) = 1000, and 1-decanol. Polymerisation was performed at 80 °C for 22 h. A simple acid hydrolysis of residual epoxides resulted in a mixed diol-amino chemistry. The modified column was used successfully for hydrophilic interaction liquid chromatography (HILIC) of small molecule probes such as nucleic acid bases and nucleosides, benzoic acid derivatives, as well as for peptides released from a tryptic digest of cytochrome c. The mixed-mode chemistry allowed both hydrophilic partitioning and ion-exchange (IEX) interactions to contribute to the separation, providing flexibility in selectivity control. Residual epoxide groups were also exploited for incorporating a mixed IEX chemistry. Alternatively, the surface chemistry of the monolith pore surface rendered hydrophobic via grafting of a co-polymerised hydrophobic hydrogel. The inherent hydrophilicity of the monolith scaffold also enabled high performance separation of proteins under IEX and hydrophobic interaction modes and in the absence of non-specific interactions.