Determining Carboxy-THC in Hair Using Agilent Captiva EMR— Lipid Cleanup with LC/MS/MS

Significance of the Topic

The analysis of 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (carboxy-THC) in hair is critical for forensic and clinical applications due to the extended detection window, resistance to adulteration, and regulatory requirements such as the Society of Hair Testing cutoff of 0.0002 ng/mg. Sensitive, selective, and robust methods are essential to reliably confirm cannabis use at trace levels within the complex hair matrix.

Study Objectives and Overview

This work aimed to develop and validate a sample preparation and LC/MS/MS protocol for quantifying carboxy-THC in human hair, compare the performance of Agilent 6470A and 6495C triple quadrupole mass spectrometers, and assess the impact of Agilent Captiva EMR–Lipid cleanup on method sensitivity and robustness. The target quantification limit was set at 0.2 ng/g hair (0.0002 ng/mg) in line with SoHT guidelines.

Methodology

Hair samples (25 or 40 mg) were washed, dried, and cut into small pieces. Samples were spiked with isotopically labeled internal standard (carboxy-THC-d3), digested in 1 M NaOH at 100 °C for 30 minutes, and subjected to liquid–liquid extraction (LLE) with n-hexane/ethyl acetate (9:1, v/v). Extracts were dried under nitrogen, reconstituted in 80:20 MeOH/water, and cleaned using Agilent Captiva EMR–Lipid solid-phase extraction cartridges. Final extracts were concentrated and injected for LC/MS/MS analysis using gradient elution.

Used Instrumentation

• Agilent 1290 Infinity II LC system with Poroshell 120 Phenyl-Hexyl column (3.0 × 150 mm, 1.9 µm) and inline filter.
• Agilent 6495C and 6470A triple quadrupole mass spectrometers with Jet Stream ESI source in negative MRM mode.

Key Results and Discussion

The calibration curve was linear (R2 > 0.99) over 0.15–3.2 ng/g with limits meeting the 0.2 ng/g target. Recoveries averaged 101–110% with RSDs below 9% at key QC levels. The 6495C system showed approximately threefold higher response than the 6470A at the 0.2 ng/g cutoff. Efficient lipid removal by Captiva EMR–Lipid reduced matrix suppression, improved signal stability, and enhanced method robustness relative to classical LLE and ion-exchange SPE approaches.

Benefits and Practical Applications

• No derivatization is required, enabling direct screening and confirmation workflows.
• High recovery and low RSD support reliable quantification at trace levels.
• Robust cleanup minimizes instrument contamination and downtime.
• Meets forensic and workplace testing criteria for cannabis use verification.

Future Trends and Applications

Advances may include automation of sample preparation, reduced sample sizes, integration with high-resolution mass spectrometry for enhanced specificity, and expansion to other low-abundance biomarkers in hair or alternate keratinous matrices. Emerging EMR sorbents and microextraction techniques will further streamline workflows and increase throughput.

Conclusion

A validated LC/MS/MS method employing Agilent Captiva EMR–Lipid cleanup on hair samples delivers sensitive, precise, and robust quantification of carboxy-THC at forensic cutoff levels. The superior performance of the 6495C platform underscores its suitability for routine hair analysis in clinical, forensic, and workplace testing laboratories.

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