(601b) Monolithic Columns in Thin Layer Format for TLC-MS Separations

Lamprou, A., Lawrence Berkeley National Laboratory
Lin, Z., Lawrence Berkeley National Laboratory
Lu, Y., Lawrence Berkeley National Laboratory
Svec, F., University of California at Berkeley

Porous polymer
monoliths are applicable as stationary phases for a variety of chromatographic
modes. Among them, thin layer chromatography (TLC) is a facile and inexpensive
separation method, where the mobile phase is propagating through a porous layer
by capillary actuation. Owing to its flat format, TLC may be coupled with
advanced mass spectrometric (MS) detection methods, like matrix-assisted laser
desorption/ionization (MALDI) and laser ablation electrospray ionization
(LAESI). It also offers the advantage of increased throughput by permitting to
run several samples in parallel lines, while the developed plates can be stored
for archiving. Motivated by the increasing attention that TLC is receiving
during the last decade, very thin polymeric monolith layers have been prepared on
suitable supports and applied for the separation of biomolecules, i.e. peptides,
proteins and nucleosides.

The performance of such monolithic thin layers is controlled by both their porous structure and
surface chemistry. Thus polystyrene-based, as well as
polymethacrylate-based layers have been prepared using
UV- or thermally-initiated polymerizations, in the presence of different
porogenic mixtures. Their pore surface was
subsequently modified via photografting or attachment of functional moieties
via chemical reactions.

the styrene-based thin layers were hypercrosslinked
by Friedel-Crafts alkylation, in order to create a
multiplicity of additional mesopores (see Figure 1). Hence, excellent
separations of small molecules were achieved (see Figure 2). The methacrylate-based layers
were photografted with different functional monomers,
whereby the exposure to the initiating UV light was spatially controlled in a
fashion that enabled the formation of a diagonal hydrophobicity gradient.
Through this adjustment, during two sequential developments in reversed phase
and ion-exchange mode, the solutes were able to encounter the gradient twice. Thus
truly 2-dimensional separations, with superior resolution and selectivity were

In another
implementation, noble metal nanoparticles were attached on the
methacrylate-based thin layers, followed by a final modification with polar
branched polymer chains, enabling the separation of peptides and nucleosides in
hydrophilic interaction (HILIC) mode. Moreover, a functional monomer bearing
weakly acidic groups, together with conjugated double bonds may be directly
copolymerized with a crosslinker and appropriate porogens. The resulting
monolithic thin layers can be used


Figure 1. Hydrophobic hypercrosslinked polystyrene-based thin
monolithic layer (Scanning Electron Microscopy image).

Figure 2. Separation of three peptides on a polystyrene-based thin
monolithic layer (left) and subsequent analyte detection by MALDI (right). Mobile phase was 65% acetonitrile in water, with 0.1% trifluoroacetic
acid, Development time was 15 min.

as weak-cation exchangers, as well
as assist the energy transfer from a laser beam to the adsorbed analytes, thus
being ideal substrates for MS detection immediately after the separation. The desorption/ionization of the separated
compounds from such functional plates, as well as from those
bearing noble metal nanoparticles was actually facilitated
without the addition of any matrix medium, which is typically applied. Coupling
TLC separation with direct MS detection provides the advantages of simplified
sample preparation, as well as improved analytical sensitivity and

Han, Y.; Levkin, P.; Abarientos, I.; Liu, H.; Svec,
F.; Frechet, J. M. J., Analytical Chemistry 2010, 82 (6), 2520-2528.

2. Urbanova, I.; Svec, F., J.
Sep. Sci. 2011, 34, 2345-2351.