Synthetic cathinones continue to proliferate in clandestine drug markets worldwide. N-ethylnorpentylone (also known as N-ethylpentylone or ephylone) is a popular emergent cathinone, yet little information is available about its toxicology and pharmacology. Here we characterize the analytical quantification, clinical presentation, and pharmacological mechanism of action for N-ethylnorpentylone. Liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) was used to quantify N-ethylnorpentylone in blood obtained from human cases. Clinical features exhibited by the intoxicated individuals are described. The activity of N-ethylnorpentylone at plasma membrane transporters for dopamine (DAT), norepinephrine (NET) and 5-HT (SERT) was assessed using in vitro assays measuring uptake inhibition and evoked release of [ H] neurotransmitters in rat brain synaptosomes. Our LC-MS/MS method assayed N-ethylnorpentylone concentrations with limits of detection and quantification of 1 and 5 ng/mL, respectively. Quantitation was linear from 5 to 500 ng/mL, and the method displayed specificity and reproducibility. Circulating concentrations of N-ethylnorpentylone ranged from 7 to 170 ng/mL in clinical cases, and the associated symptoms included palpitations, tachycardia, agitation, hallucinations, coma and death. N-Ethylnorpentylone was a potent inhibitor at DAT (IC = 37 nM), NET (IC = 105 nM) and SERT (IC = 383 nM) but displayed no transporter releasing activity. We present a validated method for quantifying N-ethylnorpentylone in human case work. The drug is a psychomotor stimulant capable of inducing serious cardiovascular and neurological side-effects which can be fatal. In vitro findings indicate that N-ethylnorpentylone exerts its effects by potent blockade of DAT and NET, thereby elevating extracellular levels of dopamine and norepinephrine in the brain and periphery.

New psychoactive substances (NPS) are a major public health problem, primarily due to the increased number of acute poisoning cases. Detection of these substances is a challenge. The aim of this research was to develop and validate a sensitive screening method for 104 drugs of abuse, including synthetic cannabinoids, synthetic cathinones, fentanyl analogues, phenethylamines, and other abused psychoactive compounds (i.e., THC, MDMA, LSD and their metabolites) in oral fluid by liquid chromatography–tandem mass spectrometry (LC–MS-MS). The Quantisal™ oral fluid device was used to collect oral fluid samples. The oral fluid elution buffer mixture (500 μL sample) was extracted with t-butyl methyl ether (MTBE), and chromatographic separation was performed on a Raptor™ biphenyl column (100 mm x 2.1 mm ID, 2.7 μm), with a total run time of 13.5 min. Limits of detection were established at three concentrations (0.05, 0.1 or 1 ng/mL) for most analytes, except for acetyl norfentanyl and mescaline (5 ng/mL). Matrix effects were generally less than 20% and overall extraction recoveries greater than 60%. The highest matrix effect was observed within the synthetic cannabinoids group (PB-22, −55.5%). Lower recoveries were observed for 2C-T (47.2%) and JWH-175 (58.7%). Recoveries from the Quantisal™ device were also evaluated for all analytes (56.7 to 127%), with lower recoveries noted for 25I-NBOMe, valerylfentanyl and mCPP (56.7, 63.0 and 69.9%, respectively). Drug stability in oral fluid was evaluated at 15, 60 and 90 days and at 25 °C, 4 °C and − 20 °C. As expected, greater stability was observed when samples were stored at −20 °C, but even when frozen, some NPS (e.g., synthetic cannabinoids) showed more than 20% degradation. The method was successfully applied to the analysis of 7 authentic oral fluid samples positive for 16 different analytes. The method achieved good sensitivity and simultaneous detection of a wide range of NPS.

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Purpose We developed and validated a method for quantitative analysis of 50 psychoactive substances and metabolites (antidepressants, benzodiazepines and opioids) in oral fluid samples using simple liquid–liquid extraction procedure followed by liquid chromatography–tandem mass spectrometry (LC–MS/MS). Method Oral fluid samples were collected using Quantisal™ device and extracted by liquid–liquid extraction with 1.0 mL of methyl tert-butyl ether and then analyzed using LC–MS/MS. Results The method attended method validation criteria, with limits of quantification as low as 0.5 and 1.0 ng/mL, and linearity between 0.5–50.0 ng/mL for antidepressants, 0.5–25.0 ng/mL for benzodiazepines and 1.0–50.0 ng/mL to opioids. During method validation, bias and imprecision values were not greater than 16 and 20%, respectively. Ionization suppression/enhancement bias results were not greater than 25%. No evidence of carryover was observed. Sample stability studies showed that almost all analytes were stable at 25 °C for 3 days and at 4 °C for 7 days. Freeze–thaw cycles stability showed that most antidepressants and opioids were stable under these conditions. Autosampler stability study showed that all analytes were stable for 24 h, except for nitrazepam and 7-aminoclonazepam. Thirty-eight authentic oral fluid samples were analyzed; 36.8% of the samples were positive for 2 drugs. Citalopram was the most common drug found, followed by venlafaxine. Conclusions The method was validated according to international recommendations for the 50 analytes, showing low limits of quantification, good imprecision and bias values, using simple liquid–liquid extraction, and was successfully applied to authentic oral fluid samples analysis.