Capillary methacrylate-based monoliths by grafting from/to gamma-ray polymerization on a tentacle-type reactive surface for the liquid chromatographic separations of small molecules and intact proteins

[object Object]tCapillary methacrylate-based monoliths were prepared for the high performance liquid chromatography(HPLC) separation of both small molecules and large biomolecules. An efficient grafting from/to syntheticapproach was adopted introducing a network of activated sites in the inner wall surface using the newsilanization agent (N-trimethoxysilylpropyl)-polyethylenimine. Copolymerization of lauryl methacrylatemonomer and 1,6-hexanediol dimethacrylate cross-linker in the presence of porogenic solvents wasobtained under continuous -ray exposure with high conversion yield.The morphology and porous structure of the resulting monoliths have been investigated by ScanningElectron Microscopy (SEM) and1H NMR cryoporosimetry. By chromatographic investigation, the newcapillary columns attested high kinetic performance (with efficiency larger than 100,000 theoreticalplate/m for small molecules at optimum mobile phase linear velocity of about 0.5 mm/s) and also excellentmechanical stability and repeatability.The new methacrylate-based monolithic capillary columns have been successfully employed for effi-cient reversed-phase separation of intact proteins and peptides.© 2016 Elsevier B.V. All rights reserved.1. IntroductionMonolithic materials are versatile adsorbents widely employedin separation science, sample preparation and as supports for flow-through applications (e.g. heterogeneous catalysis, ion-exchange,solid-phase extraction, etc.) [1-4]. Interest around their prepara-tion and applications has been rapidly growing in recent years.Some reviews about the use of monoliths as separation media foranalytical chromatography have been published [5-10]. Two of themost important groups of monoliths are based on inorganic silicachemistry, and those derived from organic monomers. Silica-basedmonoliths consist of a bi-continuous mesoporous skeleton as resultof the sol-gel preparation method designed by Tanaka in the 90 s[11]. On the other hand, polymeric organic ones have a normally*Corresponding author.E-mail address: francesco.gasparrini@uniroma1.it (F. Gasparrini).globule-like backbone. They are most commonly obtained by asingle-step polymerization process starting from a bulk mixtureof monomers, cross-linkers (difunctional monomers) and poro-gens. In both cases, monoliths are characterized by a single-bodymesoporous structure with interconnected channels (flow-throughpores).Thanks to the possibility of modulating the skeleton thicknesswith respect to the width of the flow-through pores, monolithscombining high efficiency and high permeability can be prepared.They have been proven to be particularly suitable for high effi-ciency separations of large biomolecules, which are excluded bythe mesoporous network and do not experience the usually slowmass transfer therein [12-14]. A further considerable advantagealso includes the simplicity of in-situ preparation and, conse-quently, monolithic chromatographic columns of virtually anygeometry and shape can be easily prepared. This flexibility allows toovercome the constraints related to both packing and miniaturiza-tion of the conventional particle-based chromatographic column