Analysis of THC Metabolites in Urine by GC/MS-Scan

Importance of the Topic

Cannabis use and its regulation are evolving globally, creating a need for reliable confirmation methods to detect its psychoactive component, Δ9-tetrahydrocannabinol (Δ9-THC), and its metabolites in biological samples. Confirmatory analysis in urine is critical for forensic, clinical, and workplace drug testing, as it provides definitive evidence of cannabis exposure following initial screenings.

Objectives and Overview of the Study

This study demonstrates a gas chromatography/mass spectrometry scan (GC/MS-scan) method for quantifying Δ9-THC, Δ8-THC, their hydroxy and carboxy metabolites in urine. The goals were to evaluate sensitivity at sub-ppb levels, verify mass spectrum-based confirmation capability, and assess quantitative accuracy and repeatability across relevant concentration ranges.

Sample Pretreatment

A liquid-liquid extraction protocol was employed for urine samples. Target analytes included Δ9-THC, Δ8-THC, their primary hydroxy-metabolites (Δ9-OH-THC and Δ8-OH-THC), and carboxy-metabolites (Δ9-THC-COOH and Δ8-THC-COOH). Deuterated analogs (Δ8-THC-d3, Δ9-THC-d3, Δ9-OH-THC-d3, Δ9-THC-COOH-d3) served as internal standards to correct for extraction and analytical variability.

Instrumentation

• Gas Chromatograph/Mass Spectrometer: Shimadzu GCMS-TQ 8040 NX (alternatively GCMS-QP2020 NX or GCMS-QP2050 NX)
• Auto-injector: AOC-30i
• Column: SH-I-5Sil MS, 30 m × 0.25 mm I.D. × 0.25 µm film thickness

Analytical Conditions

Injection temperature 280 °C in splitless mode, column oven program from 150 °C (1 min) at 15 °C/min to 320 °C (3 min). Carrier gas linear velocity set at 45.6 cm/s. MS operated in scan mode (m/z 45–600) with ion source at 230 °C and interface at 280 °C. Data acquisition event time 0.3 s.

Main Results and Discussion

• Chromatographic separation yielded clear total ion current peaks for each analyte at 100 ng/mL spiking levels, with distinct mass chromatograms despite close retention times.
• Mass spectra recorded at 15 ng/mL showed signal-to-noise ratios ≥ 68 for all targets, enabling spectrum-based confirmation without full subtraction.
• Calibration curves over 1–100 µg/mL exhibited linearity with correlation coefficients (R) ≥ 0.995 for all compounds.
• Intra-day precision (n=5) at 15 ng/mL and 250 ng/mL demonstrated accuracy between 86 % and 113 % and repeatability (%RSD) below 5 %, confirming quantitative reliability.

Benefits and Practical Application

• Provides full mass spectral data for definitive confirmation of cannabis metabolites.
• Achieves ppb-level quantification sufficient for forensic and clinical confirmation testing.
• Offers robust repeatability and accuracy to support routine QA/QC workflows.

Future Trends and Possibilities

Advancements may include integration of automated spectral library matching software to streamline confirmatory workflows, coupling GC/MS-scan with high-sensitivity detectors or tandem MS to enhance detection limits, and extending methods to other classes of drugs and metabolites in biological matrices.

Conclusion

The GC/MS-scan method described delivers sensitive, repeatable, and reliable quantification and confirmation of THC metabolites in urine. While slightly less sensitive than tandem MS approaches, it compensates by providing comprehensive mass spectra for high-confidence verification of initial screening outcomes.

Reference

• II-5 Cannabis Test Methods, Ed. by The Pharmaceutical Society of Japan: Toxicological Test Methods and Annotations 2017
• Analysis of THC Metabolites in Urine by GC-MS/MS (01-00860)