MSACL 2024 Abstract

Development and Analytical Performance of 43 Drug of Abuse Panel in Urine using the SCIEX Triple Quad™ Citrine LC-MS/MS

Michael Breckenridge(1), Mahesheema Ali (1,2)
(1) Department of Pathology, The MetroHealth System, Cleveland, OH, USA. (2) Department of Pathology, Case Western Reserve University School of Medicine, Cleveland, OH, USA

Mahesheema Ali, PhD, NRCC (Presenter) The MetroHealth Medical Center

Presenter Bio: Dr. Mahesheema Ali is the Medical Director of the Core laboratory, Toxicology& POCT at MetroHealth Systems and an Associate Professor of Laboratory Medicine at Case Western Reserve University School of Medicine. She is board-certified by the American Board of Clinical Chemistry and is a fellow of the ADLM Academy. Dr. Ali's academic interests focus on implementing innovative laboratory tests for in vitro diagnostic testing in areas such as automated chemistry, toxicology, mass spectrometry, and point-of-care testing (POCT). She is also committed to promoting excellence and enhancing the quality of laboratory testing.

Relevant Financial Disclosures (within past 24 months, reported on Mar 25, 2025) No relevant financial relationship(s) to disclose. Conflict mitigation NOT REQUIRED.

Background: This study reports the analytical performance of the SCIEX Citrine Triple Quad MS/MS system to analyze 43 drugs of abuse in human urine matrix monitoring over 125 MRM transitions (including internal standards).

Method: The evaluation of performance involved verifying identity, selectivity, linearity, the limit of detection (LOD), lowest limits of quantification (LLOQ), accuracy, precision, extraction recovery, matrix effect, and stability. The method uses the SCIEX Exion HPLC system and the SCIEX Triple Quad™ Citrine LC-MS/MS system to ensure fast, sensitive, and reproducible results were utilized, and data was processed using Multiquant MD software 3.0.3 version. To evaluate the drug compounds, commercially available urine spiked with Cerilliant was used and processed under certain conditions with an optimized scheduled MRM.

Sample Prep Conditions
To analyze the 50 μL urine sample, it needs to be hydrolyzed. Use 15 µL of IMCS Rapid Hydrolysis Buffer and 5 µL of IMCSzyme for this purpose, and let it sit for 15 minutes at room temperature. After hydrolysis, add 10 µL of internal standard with 135 µL of water. Next, add 300 µL of sample diluent, which should be a mix of 3 parts mobile phase A and 1 part mobile phase B. Centrifuge the mixture at 15,000 rpm for 15 minutes and the sample is now ready for analysis.

Liquid Chromatography Conditions
Column: Phenomenex Kinetex Phenyl-Hexyl and Security Guard: ULTRA Phenyl cartridge are used in this method. The mobile phase A is made up of water and 0.1% formic acid, whereas mobile phase B contains methanol and 0.1% formic acid. The flow rate is set at 0.8 mL/min, and the injection volume is 2 uL. A linear gradient is used from 2% to 98% B over 6.4 minutes, with a retention time of 1 to 6.5 minutes.

Mass Spectrometry Conditions
Duration of the Method: 7.5 minutes. Polarity can be Positive or Negative. Electrospray Scheduled MRM detection window is 50 seconds. Transitions are compound dependent. Source Conditions are flow rate optimized.

Results: The regression coefficients (r2) for the calibration curves in the study were ≥0.99. Intra and intraassay imprecision were found to be <10%. Comparison with a reference laboratory revealed <20% bias for all 42 analytes, deeming correlation coefficients >0.990. Linearity ranges were established from the lowest to upper limit calibrator concentrations with 10- to 100-fold maximum dilution.

Conclusion: An analytical method was developed to quantify over 43 drug components in human urine. This method provides an effective strategy for improving TAT in drug testing and confirmation of screen results.