Page 79 - Advances in Forensic Applications of Mass Spectrometry - Jehuda Yinon

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ion suppression from late-eluting components, a dual-column switching sys-
tem was applied and the analysis time was shortened to 2.5 min. Column
switching after homogenization and protein precipitation was applied for
determination of various drugs (benzodiazepines, felodipine, nitrendipine,
nicardipine) in animal tissues to study drug distribution. The drugs were
separated on short (2.1 ¥ 10 mm) columns and analyzed with ESI or
ACPI/MS/MS. 17
From all above-mentioned studies, two general conclusions may be
drawn: the extracts should be of high quality, and the chromatographic
separation should not be neglected or sacrificed. LC/MS is not the panacea
that will replace the optimal sample pretreatment and separation.

2.2.1.2 Composition of the Mobile Phase and Type of Column
The composition of the mobile phase may greatly affect the sensitivity of
18
LC/MS analysis. Temesi and Law studied the influence of mobile phase
on electrospray response in ESI/MS, using 35 various acidic, neutral, and
basic drugs as test substances. Methanol and ACN were used as organic
modifiers, and ammonium acetate, ammonium formate, and TFA were used
as electrolytes. Generally, in the positive ionization mode, methanol gave
stronger signals than ACN. Electrospray response decreased with increasing
concentration of ammonium acetate/formate. Very large quantitative indi-
vidual differences between drugs were observed. The authors concluded
that a thorough optimization of all eluent parameters is essential for single
analyte analysis.
According to Naidong et al., reversed-phase chromatography, although
19
widely used, is less compatible with MS/MS detection than normal-phase
chromatography. This is particularly true for highly polar compounds, which
are hardly retained on reversed-phase columns, even on the mobile phase
containing mostly water. A low percentage of organic phase modifier affects
the sensitivity due to nonoptimal spraying conditions. Conversely, normal-
phase columns are used with a mobile phase containing high percentage of
organic solvent, assuring better dispersing and evaporation of electrospray
droplets. That was demonstrated in series of comparative experiments with
nicotine, cotinine, and albuterol, analyzed with LC/ESI/MS/MS after separa-
tion on C18 and silica columns. The mobile phase for a reversed-phase
column contained 10% acetonitrile, and for a normal-phase column, 70%
acetonitrile. The sensitivity observed after normal-phase separation was dis-
tinctly higher due to the longer retention times of polar analytes and the
separation from matrix compounds causing ion suppression. A similar
approach was used by the same group of authors for the analysis of morphine
22
and its glucuronides in serum, 20,21 for fentanyl, and for hydrocodone/hydro-
morphone. The combination of straight-phase short (50 ¥ 3 mm) column
23

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