Page 277 - Multidimensional Chromatography

272 Multidimensional Chromatography

selectivity which may be needed when analytes have to be determined at the trace
level and or characterization and confirmatory information is required. Cai and
Henion (111) used an immunoaffinity-SPE–SPE–LC–MS–MS system for the
analysis of LSD and its analogues and metabolites in human urine. The on-line chro-
matographic system involved an immunoaffinity pre-column, a packed capillary
trapping column and a packed capillary analytical column. With the trapping column
as intermediate the immuno pre-column could be operated at high flow rates, while
the analytical column was maintained at 3.5 l/min, thereby facilitating the coupling
with MS by electrospray. The urine of LSD users was analysed and concentrations of
LSD and analogues at the low-ppt level could be measured. A similar system was
used for the analysis of five -agonists in bovine urine yielding LOQs in the 10–50
ppt range, and applied to the determination of the bovine renal elimination of clen-
buterol over a period of 15 days after administration (112). For the selective extrac-
tion of propanolol from urine, a protein G pre-column was primed with drug-specific
antibodies and coupled on-line to an SPE–LC–MS system which comprised a CN
analytical column for separation and an ion-spray source for LC–MS interfacing.
By using the single-ion-monitoring mode, propanolol could be specifically detected
down to 1 ng/ml in 20 ml urine. Creaser et al. (108) used on-line immunoaffinity-
SPE–LC–ion-trap MS with a particle beam interface for the determination of corti-
costeroids in equine urine.

11.3.4 METAL-LOADED SORBENTS AND
MOLECULARLY IMPRINTED POLYMERS IN SPE–LC

An interesting–but not often used–option to increase SPE selectivity is the applica-
tion of metal-loaded sorbents that are able to bind certain organic compounds which
form specific complexes with the immobilized metal ions such as Cu(II), Ag(I),
Hg(II) and Pd(II). Nielen et al. (113) showed that metal-loaded phases can be used
for selective on-line sample handling and trace enrichment in LC. Some bioanalyti-
cal applications were reported by Irth and co-workers 114–116). A pre-column with
a silver-loaded thiol phase was used in an on-line fashion for the extraction of 5-fluo-
rouracil (a chemotheurapetic agent) from plasma (114). SPE sorption of the analyte
by complexation with Ag(I) occured at high pH, while desorption to the ODS analyt-
ical column was based on analyte protonation at low pH. For plasma samples, either
a second pre-column packed with PLRP-S was placed before the Ag(I)-thiol pre-
column in order to remove macromolecular and apolar components, or the samples
were deproteinated before analysis. By using UV detection, a detection limit of 3 nM
was obtained for 5-fluorouracil for direct injection of 1 ml samples. The same
SPE–LC–UV system with a silver-loaded pre-column was used for the trace-level
determination of 3´-azido-3´-deoxythymidine (AZT), a potential drug for the treat-
ment of AIDS, in plasma (115), and also for the analysis of four barbiturates in
plasma (116).
Recently, molecularly imprinted polymers (MIPs) have gained attention as
new, selective sorbents for chromatography and SPE. The cavities in the polymer