Page 272 - Multidimensional Chromatography

Biomedical and Pharmaceutical Applications 267

on-line applications will be treated in which special formats and/or selective SPE
sorbents are used in combination with LC.

11.3.1 AUTOMATED SPE–LC

In order to achieve more cost-effective analyses, biomedical laboratories aim for a
higher sample throughput and, thus, also for faster sample pretreatment procedures.
This aspiration has led to an increasing interest in the automation of sample prepara-
tion by using robotic devices. The whole off-line SPE sequence can be automated
with instruments such as the ASPEC (Gilson), Microlab (Hamilton) and Rapid Trace
(Zymark). Some of these devices can be coupled to LC in an at-line fashion, as they
provide the possibility for automated injection of the final extract. A good illustration
of the high degree of automation that can be accomplished today is the determination
of the glucocorticosteroid budesonide in plasma samples, as described by Kronkvist
et al. (88). Their automated method comprised a Tecan robot for initial sample
manipulation (e.g. pipetting and mixing), an ASPEC XL for SPE of the samples, an
on-line trace enrichment system for further purification and concentration of the
sample extracts, and a gradient LC–MS–MS system for separation and selective
detection of the analyte and the internal standard. The total system can analyse up to
800 samples a week with satisfactory accuracy, yielding an LOQ of 15 pM for 1 ml
plasma samples.
In 1996, the 96-wells SPE technology was introduced for high-throughput analy-
sis; this allows the simultaneous processing of 96 samples in a standard microtiter
plate format (89). Such technology, in combination with a pipetting robot, was used
to design a high-throughput SPE system followed by RPLC with UV detection for
the determination of cimetidine (a histamine H2-receptor antagonist) in plasma (90).
Validation results were found to be adequate and a good correlation between the
results obtained with the automated method and with a manual method was demon-
strated. The average sample preparation time decreased from 4 min to 0.6 min per
sample and a sample throughput of 160 samples a day was achieved. Recently, Jemal
et al. (91) made a comparison of plasma sample pretreatment by manual liquid–
liquid extraction, automated 96-well liquid–liquid extraction and automated 96-well
SPE for the analysis by LC–MS–MS. The 96-well methods were three times faster
than the manual method. The time required for the 96-well SPE could be further
reduced by 50 % when the extracts were injected directly on to the LC (i.e. not incor-
porating drying and reconstitution steps in the SPE procedure).
In principle, on-line SPE–LC can be automated quite easily as well, for instance,
by using such programmable on-line SPE instrumentation as the Prospekt (Spark
Holland) or the OSP-2 (Merck) which have the capability to switch to a fresh dispos-
able pre-column for every sample. Several relevant applications in the biomedical
field have been described in which these devices have been used. For example, a
fully automated system comprising an autosampler, a Prospekt and an LC with a UV