A Sensitive and Comprehensive LC-MS/MS Method for the Analysis of Hallucinogens, Synthetic Cathinones, and Synthetic Cannabinoids in Urine Samples

Abstract

The laboratory analysis of new psychoactive substances and related drugs is crucial for accurate clinical and forensic diagnosis of poisonings. Given this, a new LC-MS/MS method for analyzing hallucinogens, synthetic cathinones, and synthetic cannabinoids in urine was developed. Urine samples were extracted using a liquid–liquid extraction protocol optimized via a multivariate experimental design. An aliquot of 400 μL of urine was enzymatically hydrolyzed with β-glucuronidase and extracted with 700 μL of ethyl acetate. The resulting extracts were analyzed using LC-MS/MS, with a total chromatographic run time of 8 min. The method was validated according to the ANSI/ASB Standard 036 guideline and was applied to 24 samples from suspected poisoning cases. The lower limits of quantification ranged from 0.1 to 1 ng/mL. Within-run and between-run precision (CV) were < 16%, and bias ranged from −12.8% to 19.8%. Nine of the 20 analytes investigated showed significant ionization suppression or enhancement (> 25%). Only two analytes (2C-E and 2-oxo-3-OH-LSD) had a recovery rate lower than 70%. Among the 24 analyzed urine samples, one tested positive for 25B-NBOH, one for LSD, and two for 2-oxo-3-OH-LSD. The developed method enables the simultaneous quantification of 20 illicit drugs, serving as an efficient diagnostic tool for clinical and forensic laboratories.