| 1. |
- Castaneto, Marisol S., et al.
(författare)
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Identification of AB-FUBINACA metabolites in human hepatocytes and urine using high-resolution mass spectrometry
- 2015
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Ingår i: Forensic Toxicology. - : Springer Verlag (Germany). - 1860-8965 .- 1860-8973. ; 33:2, s. 295-310
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Tidskriftsartikel (refereegranskat)abstract
- AB-FUBINACA, N-(1-amino-3-methyl-1-oxobutan-2-yl)-1-(4-fluorobenzyl)-1H-indazole-3-carboxamide, is an indazole synthetic cannabinoid identified in drug seizures around the world. Few metabolism data are available, despite the need for human urinary markers to detect AB-FUBINACA intake. Our main objective was to identify suitable analytical targets by analyzing human hepatocyte incubation samples with high-resolution mass spectrometry (HRMS) and to confirm the results in authentic urine specimens. We also determined AB-FUBINACAs metabolic stability in human liver microsomes (HLMs) and compared hepatocyte and urine results with in silico predictions. The metabolic stability of AB-FUBINACA was determined in pooled HLMs (1 A mu mol/l, up to 1 h). The metabolite profile of human hepatocytes (10 A mu mol/l, 1 and 3 h) and urine samples from two subjects were determined by HRMS using information-dependent tandem-mass spectrometry (MS-MS) acquisition. Data were analyzed with MetabolitePilot (TM) software utilizing different processing algorithms, including generic peak finding, mass defect filtering, neutral loss, and product ion filtering. In silico metabolite prediction was performed with MetaSite (TM) software. AB-FUBINACAs half-life in HLMs was 62.6 +/- A 4.0 min. AB-FUBINACA produced 11 metabolites (2 glucuronides) in human hepatocytes and 10 were identified in authentic human urine. Major metabolic pathways were terminal amide hydrolysis, acyl glucuronidation and hydroxylation at the aminooxobutane moiety. Epoxidation followed by hydrolysis, hydroxylation at the indazole moiety and dehydrogenation were minor pathways. Defluorination did not occur. Seventeen first-generation metabolites were predicted in silico, of which seven were observed in vitro and eight in vivo. We recommend AB-FUBINACA carboxylic acid, hydroxy AB-FUBINACA carboxylic acid, dihydrodiol AB-FUBINACA and dihydrodiol AB-FUBINACA carboxylic acid as suitable urinary markers.
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| 2. |
- Diao, Xingxing, et al.
(författare)
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In vitro and in vivo human metabolism of a new synthetic cannabinoid NM-2201 (CBL-2201)
- 2017
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Ingår i: Forensic Toxicology. - : SPRINGER. - 1860-8965 .- 1860-8973. ; 35:1, s. 20-32
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Tidskriftsartikel (refereegranskat)abstract
- In 2014, NM-2201 (CBL-2201), a novel synthetic cannabinoid (SC), was detected by scientists at Russian and US laboratories. It has been already added to the list of scheduled drugs in Japan, Sweden and Germany. Unfortunately, no human metabolism data are currently available, which makes it challenging to confirm its intake, especially given that all SCs investigated thus far have been found to be extensively metabolized. The present study aims to recommend appropriate marker metabolites by investigating NM-2201 metabolism in human hepatocytes, and to confirm the results in authentic human urine specimens. For the metabolic stability assay, 1 A mu M NM-2201 was incubated in human liver microsomes (HLMs) for up to 1 h; for metabolite profiling, 10 A mu M of NM-2201 was incubated in human hepatocytes for 3 h. Two authentic urine specimens from NM-2201-positive cases were subjected to beta-glucuronidase hydrolysis prior to analysis. The identification of metabolites in hepatocyte samples and urine specimens was achieved with high-resolution mass spectrometry via information-dependent acquisition. NM-2201 was quickly metabolized in HLMs, with an 8.0-min half-life. In human hepatocyte incubation samples, a total of 13 NM-2201 metabolites were identified, generated mainly from ester hydrolysis and further hydroxylation, oxidative defluorination and subsequent glucuronidation. M13 (5-fluoro PB-22 3-carboxyindole) was found to be the major metabolite. In the urine specimens, the parent drug NM-2201 was not detected; M13 was the predominant metabolite after beta-glucuronidase hydrolysis. Therefore, based on the results of our study, we recommend M13 as a suitable urinary marker metabolite for confirming NM-2201 and/or 5F-PB-22 intake.
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| 3. |
- Diao, Xingxing, et al.
(författare)
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Strategies to distinguish new synthetic cannabinoid FUBIMINA (BIM-2201) intake from its isomer THJ-2201: metabolism of FUBIMINA in human hepatocytes
- 2016
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Ingår i: Forensic Toxicology. - : SPRINGER. - 1860-8965 .- 1860-8973. ; 34:2, s. 256-267
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Tidskriftsartikel (refereegranskat)abstract
- Since 2013, a new drugs-of-abuse trend attempts to bypass drug legislation by marketing isomers of scheduled synthetic cannabinoids (SCs), e.g., FUBIMINA (BIM-2201) and THJ-2201. It is much more challenging to confirm a specific isomers intake and distinguish it from its structural analog because the isomers and their major metabolites usually have identical molecular weights and display the same product ions. Here, we investigated isomers FUBIMINA and THJ-2201 and propose strategies to distinguish their consumption. THJ-2201 was scheduled in the US, Japan, and Europe; however, FUBIMINA is easily available on the Internet. We previously investigated THJ-2201 metabolism in human hepatocytes, but human FUBIMINA metabolism is unknown. We aim to characterize FUBIMINA metabolism in human hepatocytes, recommend optimal metabolites to confirm its consumption, and propose strategies to distinguish between intakes of FUBIMINA and THJ-2201. FUBIMINA (10 mu M) was incubated in human hepatocytes for 3 h, and metabolites were characterized with high-resolution mass spectrometry (HR-MS). We identified 35 metabolites generated by oxidative defluorination, further carboxylation, hydroxylation, dihydrodiol formation, glucuronidation, and their combinations. We recommend 5-OH-BIM-018 (M34), BIM-018 pentanoic acid (M33), and BIM-018 pentanoic acid dihydrodiol (M7) as FUBIMINA specific metabolites. THJ-2201 produced specific metabolite markers 5-OH-THJ-018 (F26), THJ-018 pentanoic acid (F25), and hydroxylated THJ-2201 (F13). Optimized chromatographic conditions to achieve different retention times and careful selection of specific product ion spectra enabled differentiation of isomeric metabolites, in this case FUBIMINA from THJ-2201. Our HR-MS approach should be applicable for differentiating future isomeric SCs, which is especially important when different isomers have different legal status.
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| 4. |
- Jones, A Wayne, et al.
(författare)
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Heroin poisoning deaths with 6-acetylmorphine in blood: demographics of the victims, previous drug-related offences, polydrug use, and free morphine concentrations in femoral blood
- 2012
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Ingår i: FORENSIC TOXICOLOGY. - : Springer Verlag (Germany). - 1860-8965 .- 1860-8973. ; 30:1, s. 19-24
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Tidskriftsartikel (refereegranskat)abstract
- This article discusses cases of drug-poisoning death in which 6-acetylmorphine (6-AM) was identified in blood as evidence for recent use of heroin. We report the demographics of the victims, previous drug-related offences, polydrug use, and the concentrations of free morphine in peripheral blood. After solid-phase extraction, morphine, codeine, and 6-AM were determined in blood samples by isotope-dilution gas chromatography-mass spectrometry (GC-MS) using limits of quantitation of 0.005 mg/l for each opiate. The victims of heroin poisoning were mainly men (88%), with a mean age of 35.4 +/- A 8.4 years (+/- SD) and no significant gender difference in age (men 35 +/- A 8.4 years; women 35 +/- A 8.6 years). The median concentration of free morphine in blood (n = 671) was 0.25 mg/l (66% andgt; 0.20 mg/l) and women had a higher concentration (0.30 mg/l) than men (0.24 mg/l) (P andlt; 0.05). No significant difference (P andgt; 0.05) was found for the concentration of free morphine in blood when heroin was the only drug taken (median 0.26 mg/l, n = 53) compared with multidrug deaths (median 0.24 mg/l, n = 618) (P andgt; 0.05). The coingested drugs most commonly identified in heroin-related deaths were ethanol (44%), diazepam (27%), cannabis (20%), and flunitrazepam (19%). We found that 61% of victims had previous drug-related offences ranging from 1 to 48 times. The close agreement between the concentrations of free morphine in blood when heroin was the only drug taken and multidrug deaths suggests that differences in tolerance to opiates is more important in causing death than adverse drug-drug interactions.
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| 5. |
- Leong, Huey Sze, et al.
(författare)
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Monitoring metabolism of synthetic cannabinoid 4F-MDMB-BINACA via high-resolution mass spectrometry assessed in cultured hepatoma cell line, fungus, liver microsomes and confirmed using urine samples
- 2021
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Ingår i: Forensic Toxicology. - : SPRINGER. - 1860-8965 .- 1860-8973. ; 39:1, s. 198-212
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Tidskriftsartikel (refereegranskat)abstract
- Purpose A tert-leucinate derivative synthetic cannabinoid, methyl (2S)-2-([1-(4-fluorobutyl)-1H-indazole-3-carbonyl]amino)-3,3-dimethylbutanoate (4F-MDMB-BINACA, 4F-MDMB-BUTINACA or 4F-ADB) is known to adversely impact health. This study aimed to evaluate the suitability of three different modes of monitoring metabolism: HepG2 liver cells, fungus Cunninghamella elegans (C. elegans) and pooled human liver microsomes (HLM) for comparison with human in-vivo metabolism in identifying suitable urinary marker(s) for 4F-MDMB-BINACA intake. Methods Tentative structure elucidation of in-vitro metabolites was performed on HepG2, C. elegans and HLM using liquid chromatography-tandem mass spectrometry and high-resolution mass spectrometry analysis. In-vivo metabolites obtained from twenty authentic human urine samples were analysed using liquid chromatography-Orbitrap mass spectrometry. Results Incubation with HepG2, C. elegans and HLM yielded nine, twenty-three and seventeen metabolites of 4F-MDMB-BINACA, respectively, formed via ester hydrolysis, hydroxylation, carboxylation, dehydrogenation, oxidative defluorination, carbonylation or reaction combinations. Phase II metabolites of glucosidation and sulfation were also exclusively identified using C. elegans model. Eight in-vivo metabolites tentatively identified were mainly products of ester hydrolysis with or without additional dehydrogenation, N-dealkylation, monohydroxylation and oxidative defluorination with further oxidation to butanoic acid. Metabolites with intact terminal methyl ester moiety, i.e., oxidative defluorination with further oxidation to butanoic acid, were also tentatively identified. Conclusions The in-vitro models presented proved useful in the exhaustive metabolism studies. Despite limitations, HepG2 identified the major 4F-MDMB-BINACA ester hydrolysis metabolite, and C. elegans demonstrated the capacity to produce a wide variety of metabolites. Both C. elegans and HLM produced all the in-vivo metabolites. Ester hydrolysis and ester hydrolysis plus dehydrogenation 4F-MDMB-BINACA metabolites were recommended as urinary markers for 4F-MDMB-BINACA intake.
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| 6. |
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| 7. |
- Ujvary, Istvan, et al.
(författare)
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Acryloylfentanyl, a recently emerged new psychoactive substance: a comprehensive review
- 2017
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Ingår i: Forensic Toxicology. - : SPRINGER. - 1860-8965 .- 1860-8973. ; 35:2, s. 232-243
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Forskningsöversikt (refereegranskat)abstract
- N-(1-Phenethylpiperidin-4-yl)-N-phenylacrylamide, or acryloylfentanyl (acrylfentanyl), is a synthetic opioid and a close structural analogue of fentanyl, which is widely used in medicine as an adjunct to general anaesthesia during surgery and for pain management. Until recently, acryloylfentanyl was known only from the scientific literature, but in 2016 this non-controlled substance became available on the illicit drug market as a powder and nasal spray in Europe and the USA. By the end of 2016, detection of acryloylfentanyl in six European countries, including 47 deaths associated with the drug, had been reported to the European Monitoring Centre for Drugs and Drug Addiction (EMCDDA) through the European Union Early Warning System, which is a part of the system designed to identify and respond to the appearance of new psychoactive substances that may pose potential public health risks similar to drugs controlled under the United Nations drug control conventions. Herein we review what is known about this potent narcotic opioid. In addition to describing its chemical properties and the synthetic routes, analytical methodologies for the identification of the substance, as well as the limited information on the biological properties, including in vitro and in vivo pharmacological studies with the substance, are summarised. Analytically confirmed acute intoxications show that the signs and symptoms of acryloylfentanyl poisoning correspond to the opioid overdose triad of decreased consciousness, miosis and respiratory depression. Importantly, naloxone works as an antidote in life-threatening poisoning. The major human urinary metabolites identified in fatal overdose cases were nor-acryloylfentanyl, as well as mono- and dihydroxylated derivatives and their conjugates.
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| 8. |
- Watanabe, Shimpei, et al.
(författare)
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Metabolism of the benzodiazepines norflurazepam, flurazepam, fludiazepam and cinolazepam by human hepatocytes using high-resolution mass spectrometry and distinguishing their intake in authentic urine samples
- 2020
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Ingår i: Forensic Toxicology. - : SPRINGER. - 1860-8965 .- 1860-8973. ; 38:1, s. 79-94
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Tidskriftsartikel (refereegranskat)abstract
- Purpose Norflurazepam, also a metabolite of other prescription benzodiazepines, appeared on the new psychoactive substances (NPS) drug market recently, complicating the interpretation of NPS findings. The aims of the study were to tentatively identify potential metabolites of norflurazepam and structural analogues (flurazepam, fludiazepam and cinolazepam) produced by hepatocytes and in authentic human samples and to discuss the possibility to differentiate drug consumption. Methods Each drug (5 mu mol/L) was incubated with pooled human hepatocytes, and metabolites were identified using liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Similarly, urine with/without hydrolysis and blood/serum from three flurazepam and seven presumptive norflurazepam cases were analysed by LC-HRMS. Results No metabolites were detected for norflurazepam in hepatocytes, but six metabolites for flurazepam, two for fludiazepam and three for cinolazepam were found. In human specimens collected after flurazepam ingestion, a total of eight metabolites, in good agreement with hepatocyte metabolites, were detected. In specimens of presumptive norflurazepam intake, norflurazepam and its metabolites (four hydroxy metabolites and one glucuronide of a hydroxy metabolite) were found. Conclusions Based on the results, hydroxy metabolites for norflurazepam, N-(hydroxyethyl), desethyl and didesethyl for flurazepam, hydroxy for fludiazepam and glucuronides and N-(hydroxyethyl) for cinolazepam are recommended for monitoring. While flurazepam, fludiazepam and cinolazepam were metabolised by hepatocytes at side chain, norflurazepam was not, which seems to indicate that hepatocytes have difficulty in modifying the benzene/diazepine rings of some 1,4-benzodiazepines. As for confirming the intake of norflurazepam, the urine ratio of 3-hydroxy-norflurazepam/norflurazepam might be the key; a high ratio might be correlated to norflurazepam intake, thereby enabling the differentiation.
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