Comprehensive Profiling of Phytocannabinoids and Semisynthetic Cannabinoids in Seized Materials from Northern Italy

In recent years, semisynthetic cannabinoids (SSCs) have emerged in commercial and illicit markets as alternatives designed to circumvent regulations targeting Δ⁹-tetrahydrocannabinol (Δ⁹-THC). However, their pharmacological properties, potency, and toxicological profiles remain poorly characterized, raising concerns about potential risks to consumers. In the present study, a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the identification and quantification of 31 phytocannabinoids and SSCs, including acetylated forms, inCannabis -derived products. The initial identification of Δ⁸- and Δ⁹-tetrahydrocannabinol acetate (Δ⁸-THCOAc, Δ⁹-THCOAc), 9R- and 9S-hexahydrocannabinol acetate (9R,9S-HHCOAc), and cannabinol acetate (CBNOAc) was achieved using in-house synthesized reference materials. The LC separation successfully resolved multiple epimeric and isomeric cannabinoid structures, and the validated method was applied to 151Cannabis-derived products seized in Northern Italy between 2024 and 2025. Over the study period, a total of 93% of the seizedCannabis -derived products contained SSCs (>LOQ), including 49% of flowers and 97% of resins. Δ⁹-THCOAc was the most frequently identified SSC (85%), followed by Δ⁹-tetrahydrocannabiphorol (Δ⁹-THCP) (61%), Δ⁸-tetrahydrocannabinol (Δ⁸-THC) (18%), and Δ⁸-THCOAc (6%). In addition, molecular docking simulations were performed on cannabinoid receptors 1 and 2 (CB1 and CB2) to provide a structure-based framework for interpreting how acetylation and side-chain modification influence ligand-receptor interactions, indicating increased binding propensity for acetylated and side-chain-extended derivatives. SSCs represent an emerging class of cannabinoids with a rapidly increasing popularity and distinct receptor binding affinities. Despite their widespread availability, systematic investigations are urgently needed to assess their safety profiles and their potential public health implications.