Characterisation of the designer cathinone N-isopropylbutylone by a multi-technique approach and its metabolic profiling and stability assessment using dried matrix spots

Synthetic cathinones are one of the most prevalent new psychoactive substances on the drug market over the last 20 years. Their structural diversity, rapid turnover and variable stability in biological matrices pose significant challenges for laboratories. This study applied a comprehensive, multi-analytical workflow to characterise, profile the metabolism, and assess the stability of the synthetic cathinone N-isopropylbutylone, with a particular focus on dried matrix spot approaches. Structural elucidation was achieved through the combined use of GC–MS, LC–HRMS and NMR spectroscopy. This provided complementary information, enabling the compound to be identified with confidence. Investigating the metabolic profile in an animal model led to the identification of multiple phase I and phase II metabolites, including products of N-dealkylation, demethylation followed by O-methylation and glucuronidation. Differences in metabolite detection windows were noted between blood and urine. The applicability of dried blood spots (DBS) and dried urine spots (DUS) for metabolite detection and analyte stability was evaluated over a 90-day period. Dried matrix spots proved to be a valuable technique for analysing cathinones and their metabolites, and their enhanced stability in this medium stored at room temperature was confirmed, with most of the analytes being detectable throughout the entire study period. This study highlights the potential of dried matrix spot approaches as practical and effective tools for analysing synthetic cathinones and their metabolites. It also provides new analytical and metabolic data on N-isopropylbutylone. The proposed workflow could be used in future applications in forensic and clinical toxicology.