Postmortem analysis of three methoxyacetylfentanyl-related deaths in Denmark and in vitro metabolite profiling in pooled human hepatocytes

Mardal, Marie; Johansen, Sys Stybe; Davidsen, Anders Bork; Telving, Rasmus; Jornil, Jakob; Dalsgaard, Petur Weihe; Hasselstrøm, Jørgen B.; Øiestad, Åse Marit Leere; Línnet, Kristían; Andreasen, Mette Findal

doi:10.1016/j.forsciint.2018.07.020

Cited by 28 publications

(33 citation statements)

References 17 publications

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“…Consequently, 4-ANPP was detected in all cases with fentanyl consumption, except for case #25 (urine). 4-ANPP is a major metabolite of methoxyacetylfentanyl (Mardal et al, 2018) and furanylfentanyl (Watanabe et al, 2017) and a minor metabolite of acetylfentanyl (Watanabe et al, 2017), and was detected in every blood and urine samples when one of the three analogs was taken. 4-ANPP blood, urine, and hair concentrations were lower than those of fentanyl, methoxyacetylfentanyl, acetylfentanyl, and their nor-metabolites when one of the three analogs was taken alone, making it an inefficient and non-specific marker of consumption.…”

Section: Discussionmentioning

confidence: 99%

Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry Assay for Quantifying Fentanyl and 22 Analogs and Metabolites in Whole Blood, Urine, and Hair

et al. 2019

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Recently, synthetic opioid-related overdose fatalities, led by illicitly manufactured fentanyl and analogs, increased at an alarming rate, posing a global public health threat. New synthetic fentanyl analogs have been constantly emerging onto the drug marked for the last few years, to circumvent the laws and avoid analytical detection. Analytical methods need to be regularly updated to keep up with the new trends. In this study, we aimed to develop a new method for detecting the newest fentanyl analogs with a high sensitivity, in whole blood, urine, and hair. The method is intended to provide to clinical and forensic toxicologists a tool for documenting consumption. We developed a comprehensive ultra-high-performance liquid chromatography-tandem mass spectrometry method for quantifying fentanyl and 22 analogs and metabolites. Urine samples were simply diluted before injection; a liquid-liquid extraction was performed for blood testing; and a solid phase extraction was performed in hair. The chromatographic separation was short (8 min). The method was validated with a high sensitivity; limits of quantifications ranged from 2 to 6 ng/L in blood and urine, and from 11 to 21 pg/g in hair. The suitability of the method was tested with 42 postmortem blood, urine, or hair specimens from 27 fatalities in which fentanyl analogs were involved. Average blood concentrations (±SD) were 7.84 ± 7.21 and 30.0 ± 18.0 μg/L for cyclopropylfentanyl and cyclopropyl norfentanyl, respectively ( n = 8), 4.08 ± 2.30 μg/L for methoxyacetylfentanyl, ( n = 4), 40.2 ± 38.6 and 44.5 ± 21.1 μg/L for acetylfentanyl and acetyl norfentanyl, respectively ( n = 3), 33.7 and 7.17 μg/L for fentanyl and norfentanyl, respectively ( n = 1), 3.60 and 0.90 μg/L for furanylfentanyl and furanyl norfentanyl, respectively ( n = 1), 0.67 μg/L for sufentanil ( n = 1), and 3.13 ± 2.37 μg/L for 4-ANPP ( n = 9). Average urine concentrations were 47.7 ± 39.3 and 417 ± 296 μg/L for cyclopropylfentanyl and cyclopropyl norfentanyl, respectively ( n = 11), 995 ± 908 μg/L for methoxyacetylfentanyl, ( n = 3), 1,874 ± 1,710 and 6,582 ± 3,252 μg/L for acetylfentanyl and acetyl norfentanyl, respectively ( n = 5), 146 ± 318 and 300 ± 710 μg/L for fentanyl ( n = 5) and norfentanyl ( n = 6), respectively, 84.0 and 23.0 μg/L for furanylfentanyl and furanyl norfentanyl, respectively ( n = 1), and 50.5 ± 50.9 μg/L for 4-ANPP ( n = 10). Average hair concentrations were 2,670 ± 184 and 82.1 ± 94.7 ng/g for fentanyl and norfentanyl, respectively ( n = 2), and 10.8 ± 0.57 ng/g for 4-ANPP ( n = 2).

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Section: Discussionmentioning

confidence: 99%

Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry Assay for Quantifying Fentanyl and 22 Analogs and Metabolites in Whole Blood, Urine, and Hair

et al. 2019

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“…This compound is among the latest fentanyl analogues emerged onto the recreational drug scene, potentially being sold to unsuspecting users as a contaminant or substitute for heroin. As a new active substance, it has been linked to several intoxication cases, mostly lethal [31,32]. To our knowledge, we present hereby the first tentative identification of this analyte in hair, based on literature data which should be confirmed as soon as a commercial reference standard will be available.…”

Section: Retrospective Untargeted Analysis Of Real Samplesmentioning

confidence: 75%

Targeted and untargeted detection of fentanyl analogues and their metabolites in hair by means of UHPLC-QTOF-HRMS

Salomone

Corcia²,

Negri³

et al. 2020

Anal Bioanal Chem

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Detection of new psychoactive substances and synthetic opioids is generally performed by means of targeted methods in mass spectrometry, as they generally provide adequate sensitivity and specificity. Unfortunately, new and unexpected compounds are continuously introduced in the illegal market of abused drugs, preventing timely updating of the analytical procedures. Moreover, the investigation of biological matrices is influenced by metabolism and excretion, in turn affecting the chance of past intake detectability. In this scenario, new opportunities are offered by both the non-targeted approaches allowed by modern UHPLC-HRMS instrumentation and the investigation of hair as the matrix of choice to detect long-term exposure to toxicologically relevant substances. In this study, we present a comprehensive and validated workflow that combines the use of UHPLC-QTOF-HRMS instrumentation with a simple hair sample extraction procedure for the detection of a variety of fentanyl analogues and metabolites. A simultaneous targeted and untargeted analysis was applied to 100 real samples taken from opiates users. MS and MS/MS data were collected for each sample. Data acquisition included a TOF-MS high-resolution scan combined with TOF-MS/MS acquisition demonstrating considerable capability to detect expected and unexpected substances even at low concentration levels. The predominant diffusion of fentanyl was confirmed by its detection in 68 hair samples. Other prevalent analogues were furanylfentanyl (28 positive samples) and acetylfentanyl (14 positive samples). Carfentanil, methylfentanyl, and ocfentanil were not found in any of the analyzed samples. Furthermore, the retrospective data analysis based on untargeted acquisition allowed the identification of two fentanyl analogues, namely β-hydroxyfentanyl and methoxyacetylfentanyl, which were not originally included in the panel of targeted analytes.

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“…4‐ANPP (or despropionylfentanyl), both a precursor and metabolite of fentanyl 12 and other analogs such as furanylfentanyl, 31 methoxyacetylfentanyl 32 and acetylfentanyl 33 was proposed as an alternative marker of fentanyl intake rather than nor‐metabolites, even if the few available data showed controversial findings. In the two cases reported by Busardò et al, 4‐ANPP was quantified at 10.4 and 11.2 pg/mg, respectively, making it an inefficient and non‐specific marker of consumption according to the authors.…”

Section: Discussionmentioning

confidence: 99%

Development and validation of liquid chromatography‐tandem mass spectrometry targeted screening of 16 fentanyl analogs and U‐47700 in hair: Application to 137 authentic samples

Larabi

Martin

Etting

et al. 2020

Drug Testing and Analysis

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This study was to validate a LC–MS/MS method for the determination of 17 new synthetic opioids (NSOs) in hair including 3‐fluorofentanyl, 3‐methylfentanyl, acetylfentanyl, acetylnorfentanyl, alfentanyl, butyrylfentanyl, butyrylnorfentanyl, carfentanil, fentanyl, furanylfentanyl, furanylnorfentanyl, methoxyacetylfentanyl, norcarfentanil, norfentanyl, ocfentanil, sufentanil, and U‐47700, and to apply it to 137 authentic samples. Twenty milligrams of hair was decontaminated in dichloromethane and underwent liquid extraction. 10 μL of the reconstituted residue were injected onto the system. The separation was performed in 12 minutes in a gradient mode at a flow rate of 300 μL/min using a Hypersyl Gold PFP column (100 × 2.1 mm i.d., 1.9 μm) maintained at 30°C. Compounds were detected in positive ionization and MRM modes using a TSQ Endura mass spectrometer (ThermoFisher). The method was validated according to EMA guidelines. The LLOQ was in the range 1–50 pg/mg, and the calibration ranged from the LLOQ‐1000 pg/mg. Intra‐ and inter‐day accuracy (bias) and precision were < 15%. Extraction recoveries of parent drugs and metabolites were 74–120% and 7–62%, respectively. The matrix effect was in the range 59–126% (CVs ≤ 12.9%). Fentanyl was found in six cases at concentrations of < 1–1650 pg/mg (n = 14 segments). Five fentanyl analogs were quantified in two cases: 3‐fluorofentanyl (25–150 pg/mg, n = 5), furanylfentanyl (15–500 pg/mg, n = 5), methoxyacetylfentanyl (500–600 pg/mg, n = 2), acetylfentanyl (1 pg/mg, n = 2), carfentanyl (2.5–3 pg/mg, n = 2). This fully validated method allowed us to test for the first time 3‐fluorofentanyl and norcarfentanil in hair among 15 other NSOs, and brings new data regarding 3‐fluorofentanyl and methoxyacetylfentanyl hair concentrations.

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Postmortem analysis of three methoxyacetylfentanyl-related deaths in Denmark and in vitro metabolite profiling in pooled human hepatocytes

Cited by 28 publications

References 17 publications

Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry Assay for Quantifying Fentanyl and 22 Analogs and Metabolites in Whole Blood, Urine, and Hair

Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry Assay for Quantifying Fentanyl and 22 Analogs and Metabolites in Whole Blood, Urine, and Hair

Targeted and untargeted detection of fentanyl analogues and their metabolites in hair by means of UHPLC-QTOF-HRMS

Development and validation of liquid chromatography‐tandem mass spectrometry targeted screening of 16 fentanyl analogs and U‐47700 in hair: Application to 137 authentic samples

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