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Evaluation of an LC-MS/MS Research Method
for the Analysis of 33 Benzodiazepines and
their Metabolites
Valérie Thibert
1
, Norbert Dirsch
2
, Johannes Engl
2
, Martin Knirsch
2
1
Thermo Fisher Scientific, Courtaboeuf, France;
2
RECIPE Chemicals + Instruments GmbH, Munich, Germany
Application Note 593
Key Words
TSQ Vantage, Transcend TLX-1, benzodiazepines
Goal
The goal of this work was to evaluate the RECIPE
®
ClinMass
®
LC-MS/MS
Complete Kit for research purposes using an online method analysis of
33 benzodiazepines and benzodiazepine metabolites in serum.
Introduction
Benzodiazepines are prescribed for the management of
anxiety, sleeping disorders, muscle spasms, and seizures.
Benzodiazepines are widely viewed as safe drugs that have
relatively few side effects. However, high dosages over
prolonged periods can lead to tolerance, which leads to a
loss of efficacy and/or physical and psychological
dependence, resulting in severe withdrawal symptoms.
Benzodiazepines can also be abused in cases of crime,
suicide, and drug-facilitated sexual assault. While
benzodiazepine intoxication alone is rarely fatal,
concurrent use with alcohol or other drugs can be
life-threatening. For all of these reasons, additional
research into dosage and effects of benzodiazepines
is necessary.
Benzodiazepines are active at very low concentrations and
have short half lives; therefore, it is of great interest to
clinical researchers and forensic toxicologists to
simultaneously analyze benzodiazepines and their
metabolites in biological samples. However,
the analysis is not always easy because of very low blood
concentrations and the complexity of detecting multiple
drugs at the same time, especially in a biological matrix.
Typically, benzodiazepines are quantified in serum.
In this work, the RECIPE kit for benzodiazepines analysis
in serum was evaluated for research purposes using a
Thermo Scientific
™
Transcend
™
TLX-1 system for
performing both the online solid-phase extraction (SPE)
and the chromatography. The Transcend system was
coupled to a Thermo Scientific
™
TSQ Vantage
™
triple-stage
quadrupole mass spectrometer for quantitative analysis.
The RECIPE kit allows the analysis of 33 benzodiazepines
and their metabolites in 12–15 minutes for research
applications, such as those discussed in this application
note. It includes the SPE column, analytical column,
mobile phases, optimization mixtures, calibrators, quality
controls, and an internal standards solution that integrates
20 deuterated internal standards. One kit is sufficient for
up to 200 samples.