Streamlined Sample Cleanup: Small Molecule Fractionation and Extraction Via Low-Volume Polymer Monolithic Columns for In-Line Analysis

Polymer monoliths are stationary-phase materials for liquid chromatography and solid-phase extraction. Their porous structure, tuneability and simple synthesis enable tailoring to specific analysis requirements in analytical chemistry. Typically, polymer monoliths are used to separate larger biomolecules. Due to their lower binding capacity, the applications of polymer monoliths for the chromatographic separation of small molecules remain limited. However, recent literature has shown that polymer monoliths have the potential for the extraction of small molecules. In this research, butyl methacrylate-co-ethylene glycol dimethacrylate polymer monoliths were synthesized using localized UV polymerization in capillaries. The performance of reversed-phase polymer monoliths in automated in-line solid-phase extraction-mass spectrometry was demonstrated by the analysis of endocannabinoids from neat standard mixes and spiked cell culture media without prior sample preparation. The synthesized monoliths exhibited a binding capacity of 1896 pmol. Furthermore, we showed the repeatability of the monolith synthesis, with a variance in permeability of 19%. The system's stability is demonstrated through the analysis of multiple batches, comparing different monoliths and reusing the same monolith repeatedly, resulting in relative standard deviations (RSDs) below 20% for all extracted compounds. This automated method with hyphenated mass spectrometry improves throughput over previous manual monolithic extractions for small molecules.