Optimized SPE for UPLC-MS/MS and GC-MS/MS Determination of THC and Metabolites in Urine and Blood

Significance of the topic

Accurate quantification of THC and its metabolites in biological fluids is critical for forensic toxicology, clinical diagnostics, workplace testing and compliance with legal limits. Rapid and reliable analysis ensures effective monitoring of recent cannabis use and aids in impairment assessment.

Objectives and Study Overview

The study aimed to develop a single solid-phase extraction (SPE) protocol compatible with both UPLC-MS/MS and GC-MS/MS workflows for simultaneous determination of Δ9-tetrahydrocannabinol (THC), hydroxy-THC (OH-THC) and carboxy-THC (COOH-THC) in blood and urine. It further sought to optimize both tandem mass spectrometry methods to deliver high sensitivity, reproducibility and throughput.

Methodology

• Sample Pre-treatment: Blood samples were protein precipitated with acetonitrile, centrifuged and diluted with 1% ammonia; urine samples were alkaline hydrolyzed at 60°C, neutralized and mixed with acetonitrile.
• SPE Cleanup: Oasis MAX cartridges were conditioned, loaded, washed with water/methanol and eluted with hexane-based solvents tailored to retain analytes and remove polar interferences.
• UPLC-MS/MS Analysis: Acquity UPLC with BEH C18 column, 400 μL/min gradient from 60% to 90% acetonitrile in 4 min; electrospray ionization in positive and negative modes, monitoring specific MRM transitions.
• GC-MS/MS Analysis: Derivatization with BSTFA/TMCS at 70°C, analysis on an A6890 GC with RTX-5MS column; temperature ramp to 200°C then 240°C, electron impact MRM detection.

Instrumentation

• Acquity UPLC system coupled to a Quattro Premier XE tandem mass spectrometer.
• Agilent A6890 gas chromatograph with RTX-5MS column and tandem mass spectrometer.

Main Results and Discussion

The method achieved recoveries between 85% and 93% for all analytes, with ion suppression below 10%. Calibration curves were linear from 0 to 100 ng/mL (r2 >0.993). Limits of quantification were below 5 ng/mL. Intraday and interday reproducibility of internal standard areas showed relative standard deviations below 10%. UPLC-MS/MS analysis was completed in five minutes, while GC-MS/MS required 25 minutes per run.

Benefits and Practical Applications

• Unified SPE protocol reduces sample handling and simplifies workflows by serving both LC and GC platforms.
• High sensitivity and robustness make it suitable for forensic, clinical and workplace drug testing.
• Rapid LC-MS method supports high-throughput screening, while optional GC-MS offers confirmatory analysis.

Future Trends and Applications

Advances may include automation of SPE steps, miniaturized and high-throughput configurations, integration with high-resolution mass spectrometry for untargeted metabolite profiling, and portable MS systems for on-site testing.

Conclusion

The optimized Oasis MAX SPE protocol demonstrates performance equal or superior to conventional silica-based cartridges and supports both UPLC-MS/MS and GC-MS/MS analysis of THC and its metabolites. The method delivers rapid, sensitive and reproducible results for blood and urine screening.