Shape-anchored porous polymer monoliths for integrated online solid-phase extraction-microchip electrophoresis-electrospray ionization mass spectrometry.

We report a simple protocol for fabrication of shape-anchored porous polymer monoliths (PPMs) for on-chip SPE prior to online microchip electrophoresis (ME) separation and on-chip (ESI/MS). The chip design comprises a standard ME separation channel with simple cross injector and a fully integrated ESI emitter featuring coaxial sheath liquid channel. The monolith zone was prepared in situ at the injection cross by laser-initiated photopolymerization through the microchip cover layer. The use of high-power laser allowed not only maskless patterning of a precisely defined monolith zone, but also faster exposure time (here, 7 min) compared with flood exposure UV lamps. The size of the monolith pattern was defined by the diameter of the laser output (∅500 µm) and the porosity was geared toward high through-flow to allow electrokinetic actuation and thus avoid coupling to external pumps. Placing the monolith at the injection cross enabled firm anchoring based on its cross-shape so that no surface premodification with anchoring linkers was needed. In addition, sample loading and subsequent injection (elution) to the separation channel could be performed similar to standard ME setup. As a result, 15- to 23-fold enrichment factors were obtained already at loading (preconcentration) times as short as 25 s without sacrificing the throughput of ME analysis. The performance of the SPE-ME-ESI/MS chip was repeatable within 3.1% and 11.5% RSD (n = 3) in terms of migration time and peak height, respectively, and linear correlation was observed between the loading time and peak area.

Interfacing microchip isoelectric focusing with on-chip electrospray ionization mass spectrometry.

In this work, we demonstrate the interfacing of microchip capillary isoelectric focusing (cIEF) with online mass spectrometric (MS) detection via a fully integrated, on-chip sheath flow electrospray ionization (ESI) emitter. Thanks to the pH-dependent surface charge of the SU-8 polymer cIEF can be successfully run in native SU-8 microchannels without need for surface pretreatment prior to analysis. On the other hand, the inherent electroosmotic flow (EOF) taking place in SU-8 microchannels at high pH can be exploited to electrokinetic mobilization of the focused pH gradient toward the MS and no external pumps are required. In addition to direct coupling of a cIEF separation channel to an ESI emitter, we developed a two-dimensional separation chip for two-step, multiplex cIEF-transient-isotachophoretic (tITP) separation. In this case, cIEF is performed in the first dimension (effective L=20mm) and tITP in the second dimension (L=35mm) followed by ESI/MS. As a result, the migration order is affected by both the pI values (cIEF) and the intrinsic electrophoretic mobilities (tITP) of the sample components. The selectivity of the separation system was shown to be different from pure cIEF or pure ITP, which allows at best for baseline separation of two compounds with nearly identical pI values. The repeatabilities of the migration times of the two-step cIEF-tITP separation were 3.1-6.8% RSD (n=3). Thanks to the short separation channel, relatively short focusing times of 60-270s (depending on the applied focusing potential) were sufficient for establishment of the pH gradient and cIEF separation of the sample components, yielding total analysis times (including loading, focusing, and mobilization) well below 10min.