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High-Throughput Quantitative
LC-MS/MS Analysis of 6 Opiates
and 14 Benzodiazepines in Urine
Bill Yu, Kristine Van Natta, Marta Kozak, Thermo Fisher Scientific, San Jose, CA
Application Note 588
Key Words
Opiates, benzodiazepines, Prelude SPLC
Goal
Develop a high-throughput, low solvent consumption, easy-to-run
method for quantitative forensic analysis of six opiates and fourteen
benzodiazepines in urine.
Introduction
Analyses of opiate and benzodiazepine panels are some of
the highest-volume applications in forensic toxicology
labs. In order to meet the need for high throughput, a fast,
simple, and cost-effective method was developed,
consisting of hydrolysis, simple urine dilution, separation
by liquid chromatography (LC), and analysis by mass
spectrometry (MS). The method incorporated the
Thermo Scientific
™
Prelude SPLC
™
system (Figure 1),
which features two independent channels of sample
preparation and liquid chromatography (SPLC). With
the Prelude SPLC system, LC methods can be executed
in parallel with a different method on each channel
(Figure 2) or the same method on both channels
(Figure 3) and multiplexed into a mass spectrometer
for serial detection. Serial MS detection of multiplexed
methods improves mass spectrometer utilization time,
increases throughput of forensic toxicology laboratories,
and reduces analysis cost. The syringe pumps and
high-pressure, low-volume gradient mixing used in
Prelude SPLC system provide enhanced HPLC
performance: improved peak shape and resolution as well
as stable retention times.
Figure 2. Parallel analysis of 6 opiates (10 ng/mL) and 14 benzodiazepines (25 ng/mL) in multiplexed mode
Figure 1. Prelude SPLC system
Conventional LC-MS/MS
0.0 1.0 2.0 3.0 4.0
5.0 6.0 7.0 8.0 9.0
10.0 11.0 12.0 13.0
0.0 1.0 2.0 3.0 4.0
5.0 6.0 7.0 8.0 9.0
10.0 11.0 12.0 13.0
0.0 1.0 2.0 3.0 4.0
5.0 6.0 7.0 8.0 9.0
10.0 11.0 12.0 13.0
Injection 1
Injection 2
14 Benzodiazepines
(25 ng/mL each)
14 Benzodiazepines
Injection 3
Prelude SPLC multiplexing
Injection 1
Injection 2
14 Benzodiazepines
(25 ng/mL each)
14 Benzodiazepines
Channel 1
Injection 3
Injection 1
Injection 2
6 Opiates
Channel 2
6 Opiates
(10 ng/mL each)
Acquisition of relevent MS data