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Metabolism of Carfentanil, an Ultra-Potent Opioid, in Human Liver Microsomes and Human Hepatocytes by High-Resolution Mass Spectrometry

ABSTRACT

Carfentanil is an ultra-potent synthetic opioid. No human carfentanil metabolism data are available. Reportedly, Russian police forces used carfentanil and remifentanil to resolve a hostage situation in Moscow in 2002. This alleged use prompted interest in the pharmacology and toxicology of carfentanil in humans. Our study was conducted to identify human carfentanil metabolites and to assess carfentanil’s metabolic clearance, which could contribute to its acute toxicity in humans. We used Simulations Plus’s ADMET Predictor™ and Molecular Discovery’s MetaSite™ to predict possible metabolite formation. Both programs gave similar results that were generally good but did not capture all metabolites seen in vitro. We incubated carfentanil with human hepatocytes for up to 1 h and analyzed samples on a Sciex 3200 QTRAP mass spectrometer to measure parent compound depletion and extrapolated that to represent intrinsic clearance. Pooled primary human hepatocytes were then incubated with carfentanil up to 6 h and analyzed for metabolite identification on a Sciex 5600+ TripleTOF (QTOF) high-resolution mass spectrometer. MS and MS/MS analyses elucidated the structures of the most abundant metabolites. Twelve metabolites were identified in total. N-Dealkylation and monohydroxylation of the piperidine ring were the dominant metabolic pathways. Two N-oxide metabolites and one glucuronide metabolite were observed. Surprisingly, ester hydrolysis was not a major metabolic pathway for carfentanil. While the human liver microsomal system demonstrated rapid clearance by CYP enzymes, the hepatocyte incubations showed much slower clearance, possibly providing some insight into the long duration of carfentanil’s effects.

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Acknowledgments

The authors would like to acknowledge Drs. Alison Director-Myska, Eric Moore, Neil Jensen, and Joseph Corriveau for their unwavering and continued support of this research effort. We would also like to acknowledge Simulations Plus and Molecular Discovery for the use of their software for the in silico metabolite predictions. Finally, we would like to acknowledge Tim Moeller of BioreclamationIVT for his help with the primary hepatocyte culturing procedures and technical support.

This work was funded by the Defense Threat Reduction Agency (DTRA) under project number CB3281.

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Correspondence to Michael G. Feasel.

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Michael G. Feasel and Ariane Wohlfarth contributed equally to this work.

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Feasel, M.G., Wohlfarth, A., Nilles, J.M. et al. Metabolism of Carfentanil, an Ultra-Potent Opioid, in Human Liver Microsomes and Human Hepatocytes by High-Resolution Mass Spectrometry. AAPS J 18, 1489–1499 (2016). https://doi.org/10.1208/s12248-016-9963-5

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KEY WORDS

  • carfentanil
  • metabolism
  • norcarfentanil
  • opioid