Toxicology Drug Screening in Blood by CE-MSn - a Feasibility Study

Significance of the topic

The reliable detection and identification of drugs in whole blood are essential in forensic toxicology for legal, clinical, and workplace safety applications. Traditional workflows often require separate extraction, derivatization, and multiple instrument platforms for screening and confirmation, extending analysis time and resource requirements. Integrating capillary electrophoresis with tandem and multistage mass spectrometry (CE-MSn) promises a streamlined approach that combines separation, sensitive detection, and structural identification in a single run.

Objectives and overview

This feasibility study evaluates whether CE-MSn using an Agilent LC/MSD Trap system can:

• Detect a panel of 17 model drugs spiked into whole blood at low nanogram-per-milliliter levels.
• Generate MS, MS2, and MS3 spectra sufficiently robust for unambiguous compound identification.
• Analyze authentic forensic case extracts with minimal method optimization.

Methodology

Drug residues were obtained from porcine blood spiked at 20 ng/mL and from three authentic forensic blood samples previously characterized by ELISA and GC/MS. Extraction followed a published liquid-liquid procedure involving phosphate buffer, 1-chlorobutane partitioning, evaporation, and reconstitution. CE separations employed bare fused-silica capillaries (50 µm ID, 21.5 cm to UV, 84 cm to MS), 100 mmol/L phosphate buffer at pH 2.38, electrokinetic injection (12 kV, 16 s), and a gradient to 20 kV over 0.15 s at 25 °C in 25 min runs.

Used Instrumentation

• CE system: Agilent G1600A 3D CE with diode-array detection (200 nm / 4 nm bandwidth; reference 375 nm / 75 nm).
• Mass spectrometer: Agilent LC/MSD Trap XCT with ESI source in positive mode, UltraScan range 50–500 m/z, drying gas 7 L/min at 130 °C, nebulizer 8–12 psi, trap drive 27 V, Auto MSn with three stages.
• Sheath liquid: 0.5% formic acid in 50/50 methanol/water at 7.5 µL/min.

Main results and discussion

1. Sensitivity and matrix tolerance: CE-MSn detected all 17 drugs in porcine blood at 20 ng/mL with clear extracted ion electropherograms and negligible matrix interference. Limits of detection for select analytes are expected to be below 20 ng/mL.
2. Case studies:

• Case 1: Amphetamine, methamphetamine, and cocaine were detected and correctly identified by MS2 and MS3 despite partial co-migration of the amphetamines.
• Case 2: MDA and MDMA showed co-elution yet were distinguished unambiguously through MS2 and MS3 library matching after removal of common background fragments.
• Case 3: Authentic toxicology sample contained quinine, cocaine, cocaethylene, bupropion metabolites, and others. Auto MSn unambiguously identified major drugs; minor or overlapping metabolites required manual library searches due to limited database entries.

These findings demonstrate that multistage mass spectra overcome co-elution and provide definitive structural confirmation within a single analytical workflow.

Advantages and practical applications

• Unified platform: Screening and confirmatory identification are achieved in one injection, reducing sample handling and time.
• High selectivity: MSn data resolve co-moving analytes and suppress matrix interferences more effectively than UV detection alone.
• Potential for automation: Auto MSn capabilities enable targeted detection lists and reduced manual intervention, boosting throughput.

Future trends and applications

CE-MSn could be extended to larger drug panels, including novel psychoactive substances, by enriching spectral libraries. Further method validation will refine limits of detection, robustness, and quantitation. Combining CE-MSn with automated sample preparation and data processing pipelines may transform routine forensic toxicology, clinical drug monitoring, and pharmaceutical impurity screening.

Conclusion

This feasibility study confirms that CE coupled with multistage mass spectrometry on a single instrument provides sensitive, selective screening and reliable identification of drugs in whole blood. With further validation and library expansion, CE-MSn is poised to streamline forensic and clinical toxicological workflows by merging separation, detection, and confirmation into an integrated platform.

Reference

• Hudson JC, Golin M, Malcolm M, Whiting CF. Capillary electrophoresis with diode-array detection for forensic drug screening. Can Soc Forens Sci J. 1998;31:1–29.
• Phan DT, Harrsch PB. Application of CE-MS to illicit drug analysis. Agilent Technol Appl Note. 2005. Publication 5968-9221E.
• Knisley D, Hetherington M, McKay G. Analysis of drugs by CE-MSn. Agilent Technol Appl Note. 2005. Publication 5989-2910EN.
• Compendium of Pharmaceuticals and Specialties. 35th ed. Canadian Pharmacists Association; 2000.