Determination of ketamine and norketamine in hair samples by µ-QuEChERS and gas chromatography coupled to tandem mass spectrometry

• A µ-QuEChERS method was developed for the detection of ketamine and norketamine in hair samples using GC–MS/MS. • High sensitivity with detection limits of 0.01 ng/mg for ketamine and 0.8 ng/mg for norketamine. • µ-QuEChERS is a cost-saving, eco-friendly clean-up method, and complies with sustainable green chemistry principles. • Reliable method, allowing detecting both chronic and occasional ketamine users. • First study using µ-QuEChERS in hair testing, with potential for future use in forensic scenarios. Ketamine has gained popularity in Europe as a new psychoactive substance due to its dissociative effects, which are sought after by users at raves, for unprescribed treatment of psychological issues, and in drug-facilitated crimes. Forensic experts often face challenges in proving drug use, and hair offers distinct advantages as a sample, revealing both long-term consumption and recent abuse. However, due to its complex nature, hair requires pretreatment to remove interferences and enhance analytical sensitivity. This study employs a miniaturised version of the “Quick, Easy, Cheap, Rugged, and Safe” (QuEChERS) extraction method, commonly used in food and environmental analyses. The method uses 1 mL of 5 % formic acid in acetonitrile and 250 mg ammonium formate as solvent and salt partitioning agents, along with 50 mg magnesium sulphate and 10 mg primary-secondary amine for dispersive solid-phase extraction. After this clean-up process, samples were analysed using gas chromatography coupled with tandem mass spectrometry. Following optimisation, the method was validated for detecting ketamine and its primary metabolite, norketamine. A limit of detection of 0.01 ng/mg was achieved for ketamine, and 0.8 ng/mg for norketamine, with limits of quantification of 0.05 ng/mg and 0.8 ng/mg, respectively. This established a working range of 0.05–5 ng/mg for ketamine and 0.8–5 ng/mg for norketamine. Recovery rates ranged from 47–76 % for ketamine and 14–27 % for norketamine. The method was then applied to real samples from drug users, proving the effectiveness of the µ-QuEChERS technique as a sustainable clean-up method. Despite its miniaturisation, it retained the performance of the original QuEChERS protocol and allowed obtaining high sensitivity, enabling analyte detection at a concentration below the Society of Hair Testing’s cut-off value of 0.2 ng/mg.