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Introduction: NBOMes and NBOHs are psychoactive drugs derived from phenethylamines and have hallucinogenic effects due to their strong agonism to serotonin 5-HT2A receptors. Although cases of toxicity associated with the recreational use of substituted phenethylamines are frequently reported, there is a lack of information on the possible neurotoxic effects of NBOMe and NBOH in the brain hippocampus, a major neurogenesis region. Objectives: This study aimed at assessing the phenotypic and molecular effects of prolonged exposure of the hippocampus to the drugs 25H-NBOMe and 25H-NBOH. Methods: The ex vivo organotypic culture model of hippocampal slices (OHC) was used to investigate, by immunofluorescence and confocal microscopy, and transcriptome analyses, the mechanisms associated with the neurotoxicity of 25H-NBOMe and 25H-NBOH. Results: Reduction in the density of mature neurons in the OHCs occurred after two and seven days of exposure to 25H-NBOMe and 25H-NBOH, respectively. After the withdrawal of 25H-NBOMe, the density of mature neurons in the OHCs stabilized. In contrast, up to seven days after 25H-NBOH removal from the culture medium, progressive neuron loss was still observed in the OHCs. Interestingly, the exposure to 25H-NBOH induced progenitor cell differentiation, increasing the density of post-mitotic neurons in the OHCs. Corroborating these findings, the functional enrichment analysis of differentially expressed genes in the OHCs exposed to 25H-NBOH revealed the activation of WNT/Beta-catenin pathway components associated with neurogenesis. During and after the exposure to 25H-NBOMe or 25H-NBOH, gene expression patterns related to the activation of synaptic transmission and excitability of neurons were identified. Furthermore, activation of signaling pathways and biological processes related to addiction and oxidative stress and inhibition of the inflammatory response were observed after the period of drug exposure. Conclusion: 25H-NBOMe and 25H-NBOH disrupt the balance between neurogenesis and neuronal death in the hippocampus and, although chemically similar, have distinct neurotoxicity mechanisms.
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Drugs of abuse are psychoactive substances illicitly distributed and used worldwide. In Rio de Janeiro, Brazil, they represent a public health issue and are directly related to several social problems. The recent increase in appearances of new psychoactive substances (NPS), derived from structural modifications of existing psychoactive substances, poses a threat to public health and forensic laboratories worldwide, as little is known about these substances. This study aimed to chemically and geographically map drugs of abuse from blotter papers seized by the Civil Police of Rio de Janeiro State (PCERJ) between 2006 and 2019. High-performance analytical techniques, such as gas chromatography-mass spectrometry (GC-MS) and Orbitrap mass spectrometry (Orbitrap-MS), combined with statistical analyses, were employed to characterize the seized samples. The most common chemical compounds in NPS found in this study were synthetic phenethylamines, i.e., molecules from the 25I-NBOH (2-(((4-iodo-2,5-dimethoxyphenethyl)amino)methyl)phenol) and 25I-NBOMe (2-(4-iodo-2,5-dimethoxyphenyl)-N-(2-methoxybenzyl)ethanamine) families. Prior to 2014, the majority of seized blotter papers contained lysergic acid diethylamide (LSD) and were concentrated in the Metropolitan region. An upsurge in blotter paper seizures was observed from 2014 to 2017; the most common substances during this time were from the NBOMe family. NBOH compounds emerged in 2016 in coastal regions with high tourism, reaching over 1300 items only in 2017. Only one synthetic cannabinoid was found among the blotter papers seized in Rio de Janeiro between 2006 and 2019. The assembled chemical data and statistical analyses allowed the mapping and monitoring of the chemical profiles of the seized blotter papers, providing a strong foundation for the understanding of the origins and movement of these drugs around the RJ State.
Assuntos
Tráfico de Drogas/estatística & dados numéricos , Controle de Medicamentos e Entorpecentes/estatística & dados numéricos , Papel , Psicotrópicos/química , Brasil , Canabinoides/química , Formas de Dosagem , Fentanila/análogos & derivados , Fentanila/química , Cromatografia Gasosa-Espectrometria de Massas , Humanos , Estrutura MolecularRESUMO
The 25R-NBOH family is a group of thermally labile compounds that are relevant for forensic sciences and traditionally analyzed by GC-MS after derivatization - a step that is time consuming in a routine work. In this paper, the use of short analytical columns (4 and 10 m) showed to decrease compound degradation in the GC oven during chromatographic separation and to allow the analysis of non-derivatized 25R-NBOH compounds by GC-MS. A shorter column demanded a higher gas flow rate, and both factors decreased residence time of the analytes in the column and their degradation. The inlet temperature (250° C or 280°C) did not impact the response of 25R-NBOH. A 25R-NBOH fragmentation pathway by electron ionization was also presented for the first time. The GC-MS method with a 4 m column was successfully applied to other compounds of forensic interest, and it can be tested in the analysis of biological samples in toxicological investigations.
Assuntos
Etanolaminas/análise , Patologia Legal/métodos , Cromatografia Gasosa-Espectrometria de Massas , Toxicologia/métodos , TemperaturaRESUMO
NBOMe and NBOH are new psychoactive substances with potent activity on serotonin 5-HT2a receptors causing serious toxic effects, including serotonin toxidrome and death. The aim of this work was to develop a comprehensive MS/MS protocol, using triple quadrupole mass spectrometers coupled to LC and GC, for rapid screening and quantitation of NBOMes and NBOHs in seized blotter papers. Different scan methods (neutral loss, precursor ion or multiple reaction monitoring) were used to obtain structural information of phenylethylamine class. The developed protocol was validated for qualitative and quantitative analysis, showing a satisfactory limit of detection (1 ng/mL), with excellent selectivity, imprecision (intra and interday imprecision lower than 1.2 % RSD) and accuracy (between -7.1 and +5.6 %, n = 15), as well as bias values. The analysis of real samples shown that NBOH compounds were the most frequently detected, with concentrations ranging from 0.1 to 1,929 µg per blotter sample. Triple quadrupole mass spectrometers can be a useful tool for identification of new psychoactive substances. A comprehensive protocol using both LC-MS/MS and GC-MS/MS, with different scanning modes, have been developed and showed to be useful to screening NBOMe and NBOH in blotter papers.
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A new potent serotonin 5-HT2A receptor agonist was identified in blotter papers by several state level forensic laboratories in Brazil. The 25I-NBOH is a labile molecule, which fragments into 2C-I when analyzed by routine seized material screening gas chromatography (GC) methods. GC-mass spectrometry (MS), liquid chromatography-quadrupole time-of-flight-MS, and Fourier transform infrared and nuclear magnetic resonance analyses were performed to complete molecular characterization. Individual doses range from 300 to 1000 µg. Despite its being a potent 5-HT2A receptor agonist, 25I-NBOH is neither registered in the United Nations Office on Drugs and Crime (UNODC) nor classified as a scheduled substance in most countries. Sweden and Brazil seem to be the only countries to control 25I-NBOH. To our knowledge, this is the first scientific report dealing with identification of 25I-NBOH in actual seizures.
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The increasing number of new psychoactive substances (NPS) and their quick worldwide spreading, often only slightly modified in the form of new derivatives and analogues, have brought the need for fast, wide-ranging, and unequivocal identification methods in clinical and forensic investigations. Because it usually provides secure results, gas chromatography coupled to mass spectrometry (GC-MS) has been routinely employed as the standard technique for the detection of NPS in blotter papers. For 25I-NBOH (N-(2-hydroxybenzyl)-2-(4-iodo-2,5-dimethoxyphenyl)ethan-1-aminium), however, GC-MS analysis of an blotter paper extract leads to incorrect results. In this work, we investigated whether easy ambient sonic-spray mass spectrometry imaging (EASI-IMS), and ambient ionization MS method can be applied directly to the surface of the sample requiring therefore no extraction or sample preparations, would serve as an efficient, sensitive, and secure alternative for 25I-NBOH screening.