CLINICAL/FORENSICS ARTICLE

Assure LC/MS/MS System Performance for Drug Analyses

Using a System Suitability Test Mix

By Kristi Sellers, Clinical/Forensic Innovations Chemist and Houssain El Aribi, Ph.D., LC/MS Product Specialist, MDS Sciex

Increase sample throughput and data quality with easy, reliable verification of LC/MS/MS performance.
Extensively documented standard preparation assures accurate, consistent solutions.
Method included in Cliquid® Drug Screen & Quant Software—automatically generates test reports.

Sample throughput is a critical issue in drug toxicology, and it can be adversely affected by inferior system performance. Poor system performance can produce unreliable data, increase downtime, and necessitate sample reanalysis, which ultimately decreases sample throughput. To ensure that your LC/MS/MS system is running properly, a system suitability mix should be analyzed on a regular basis before case samples are analyzed.

Restek and Applied Biosystems have developed a system suitability mix specifically for drug testing that contains compounds covering a wide range of molecular weights, polarities, and retention times (Table I). This standards mix is designed to verify system performance and identify system problems. Figure 1 shows a representative chromatogram (+MRM transitions) of this suitability mix analyzed on an Applied Biosystems API 3200™ LC/MS/MS system. This simple test evaluates the entire analytical system, including the autosampler, column, HPLC pumps, and mass spectrometer. The data is automatically compared to expected results by Applied Biosystem’s Cliquid® Drug Screen & Quant Software to identify system problems.

Use this system suitability mix for drug analyses to assure system performance and simplify trouble-shooting.

The Cliquid® Drug Screen & Quant Software automates this test and generates a verification report which highlights failures. Peak area, peak shape, retention time reproducibility, fragmentation, and library search function all are evaluated through the software by comparing the test mix data to expected results. For example, full scan linear ion trap MS/MS data for diazepam and caffeine are compared to the library to assess fragmentation. A mass spectral match of 80% or more must be achieved to pass this portion of the system suitability test. Otherwise, the failure will be highlighted on the automated report.

Analyzing this system suitability mix for drug analysis on a regular basis assures system performance, improves data quality, increases sample throughput, and simplifies troubleshooting. Moreover, the Cliquid® Drug Screen & Quant Software for Routine Forensic Toxicology enables nonexpert LC/MS/MS users to employ this system suitability test with little effort.

Acknowledgement

Method and data supplied by Applied Biosystems.

Table I: Mix components vary in chemical properties, providing a rigorous system performance test.
Analyte	MW	RT (min.)	Q1	Q3
Amiodarone	645	12.30	646.0	58.0
Amphetamine	135	4.21	136.1	91.1
Caffeine	194	1.72	195.1	122.9
Codeine	299	3.47	300.2	165.2
Diazepam	284	5.25	285.1	193.1
Doxepin	279	8.72	280.2	107.1
Haloperidol	375	9.08	376.1	123.0
Morphine	285	2.24	286.1	165.1

Figure 1: Increase sample throughput by verifying system readiness with a drug standard system suitability mix. (MRM transitions)

Sample:

system suitability mix

Inj.:

30µL

Conc.:

amiodarone	10µg/mL
amphetamine	10
caffeine	10
codeine	10
diazepam	10
doxepine	10
haloperidol	1
morphine	10

Sample Diluent:

methanol

Column:

Allure® PFP Propyl (cat.# 9169552)

Dimensions:

50mm x 2.1mm

Particle size:

5µm

Pore size:

60Å

Conditions:

Instrument:

Shimadzu Prominence HPLC

Mobile Phase:

A. 0.2% formic acid in water and 2mM ammonium formate in water

B. 0.2% formic acid in water and 2mM ammonium formate in acetonitrile

Time (min.)	Flow (mL/min.)	%B
0.0	0.5	10
10.0	1.0	90
15.0	1.0	90
15.50	0.5	10
17.50	0.5	10

Flow:

see gradient table

Temp.:

40°C

Det.:

Applied Biosystems API 3200™ LC/MS/MS system

Ion Source:

electrospray, positive

Ion Spray Voltage:

4000

Gas 1:

40psi

Gas 2:

70psi

Source Temp:

500°C

Mass Spectrometer Conditions:

Analyte	MW	%RT (min)	Q1	Q3
Amiodarone	645	12.30	646.0	58.0
Amphetamine	135	4.21	136.1	91.1
Caffeine	194	1.72	195.1	122.9
Codeine	299	3.47	300.2	165.2
Diazepam	284	5.25	285.1	193.2
Doxepin	279	8.72	280.2	107.1
Haloperidol	375	9.08	376.1	123.0
Morphine	285	2.24	286.1	165.1

LC_PH0468

References

H. El Arbi, T. Sasaki, A. Schreiber, K. Sellers, K. Herwehe. Development of an LC/MS/MS System Suitability Test for Forensic Toxicology Applications. Applied Biosystems/MDS Sciex, 2007.