O22: Quantification of 22 synthetic cannabinoids and 10 metabolites in human hair

Abstract

Introduction Synthetic cannabinoids are the most common drugs among an expanding array of compounds that mimic the effects of traditional illicit psychoactive substances. A few methods have been developed for the determination of the parent synthetic cannabinoids in the keratin matrix, opening the scientific debate about how to interpret the quantitative results. In particular, the discrimination between (i) passive or active exposure, and (ii) chronic consumption and occasional use, appears to be essential within the forensic context. Just as occurred for the traditional drugs of abuse, the detection of drug metabolites is likely to represent the key factor to support the decision procedure. Therefore, we updated our existing method [ A. Salomone et al., Drug Testing and Analysis, 2014, 6, 126–134 ] in order to complement the determination of 22 synthetic cannabinoids in human hair with 10 of their metabolites. Afterwards, we applied the new method to 15 real hair samples which previously tested positive for at least one synthetic cannabinoid [ A. Salomone et al., cited ], in order to verify the presence of metabolites. Methods The analytical method was validated in accordance with the criteria and recommendations of national and international guidelines. The following parameters were investigated: selectivity, specificity, linearity range, detection and quantification limits (LOD and LOQ), intra-assay and inter-assay precision and accuracy. Carry-over effect, recovery and matrix effects were also investigated. Two groups of subjects, namely driving re-licensing and drug abuse/withdrawal control subjects, provided hair samples that had previously resulted positive to at least one common drug of abuse and one synthetic cannabinoid. In this procedure, the samples were added with 1 mL NaOH 1N and subsequently incubated at 95 °C for 10 min. Immediately afterwards, the samples were extracted with 5 mL of n-hexane/ethylacetate 90:10 (v/v). The aqueous phase was newly added with glacial CH 3 COOH to reach a pH of 3.5–4.0 and again extracted with n-hexane/ethylacetate 90:10. The two organic phases were unified, dried under a nitrogen flow at 70 °C and reconstituted with 50 μL of methanol. An aliquot of 1 μL methanol solution was directly injected into the UHPLC-MS/MS system. Results The optimized UHPLC – MS/MS method allowed the simultaneous determination of 22 synthetic cannabinoids, 10 metabolites and 6 internal standards. The whole chromatographic run, comprehensive of the time required for column re-equilibration before the following injection, was completed in 8.0 min. Retention times ranged between 2.2 min (WIN 48,098) and 5.5 min (JWH- 020). Two samples out of 15 resulted positive to one metabolite, namely the N-(5-hydroxypentyl) metabolite of JWH-122. The calculated concentrations were 2.5 pg/mg and 0.72 pg/mg. In the same samples, the drug parent concentrations were, respectively, 2800 pg/mg and 760 pg/mg. Conclusion The present preliminary study describes for the first time an extensive search of metabolites of synthetic cannabinoids in human hair. Undoubtedly, the detection of metabolites in hair may represent the decisive factor to support the interpretation of hair analyses, particularly with respect to potential sources of external contamination. More samples will be analyzed in order to (i) verify to what extent the presence of metabolites can prove the active consumption, (ii) evaluate the concentration ratios between parent drugs and metabolites, and (iii) elaborate tentative cut-offs for parent drug and metabolites.