Rapid β-Glucuronidase Hydrolysis of 11-nor-9-carboxy-Δ⁹-THC: Optimizing m/z 345.2 → 299.1 Transitions using “Click” Chemistry (T ≈ 37°C)

Cannabis use is usually confirmed by looking for a breakdown product that the body locks away behind a sugar molecule. Freeing it again is the slow step in the test. This work replaces the enzyme step with a faster chemical reaction and shows the result holds up against the standard method, cutting the time a laboratory needs from hours to minutes.

Introduction: The quantification of 11-nor-9-carboxy-Δ⁹-tetrahydrocannabinol (THC-COOH) in biological matrices is limited by slow enzymatic hydrolysis. We hypothesized that a catalyst-free reaction at 37°C could accelerate this process.

Methods: Urine samples (n = 50) were fortified with THC-COOH-glucuronide. Hydrolysis was compared using E. coli β-glucuronidase vs. our novel “Click” reagent. Detection was performed on a Triple Quadrupole MS monitoring the 345.2 → 299.1 and 345.2 → 193.1 transitions.

Results: The novel method achieved >95% hydrolysis in 15 min (vs. 2 h for enzymatic). Linearity was maintained from 1–500 ng/mL (R² = 0.998).

Conclusion: This protocol significantly reduces preparation time while maintaining sensitivity (µg/L levels) required for forensic workflows.