Evidence of enzyme-mediated transesterification of synthetic cannabinoids with ethanol: potential toxicological impact

Orapan Apirakkan, Ivana Gavrilovic, Giuseppe Floresta, Cheyenne Pierre, Annelies Cannaert, Christophe Stove, P I Dargan, David A. Cowan, Lewis Couchman, Vincenzo Abbate

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Synthetic cannabinoids (SCs) represent a large proportion of novel psychoactive substances on the black market and have caused a number of deaths. Polydrug use including combination of SCs and ethanol could further complicate the toxicological impact. To the best of our knowledge, there have been no reports presenting evidence of transesterification between SCs and ethanol in vitro.

The in vitro metabolism of the four carboxylate SCs PB-22, NPB-22, 5-fluoro-PB-22 (5F-PB-22), and 5-fluoro-NPB-22 (5F-NPB-22) in the presence of ethanol using human liver microsomes with and without appropriate enzyme inhibitors was studied. Newly identified SC ethyl esters were chemically synthesised and fully characterised. The activity of these SCs and their ethanol transesterification products were assessed using cannabinoid receptor (CB1 and CB2) activation assays.

SCs/ethanol transesterification products were detected and studied using liquid chromatography–high-resolution mass spectrometry. We have shown that the SC ethyl ester formation is mediated by human carboxyl esterase enzymes. The ethyl esters exhibited a reduced activity for the CB receptors compared with their parent compounds.

These novel ethyl esters may be useful additional markers of cannabinoid administration, and especially so if they prove to have longer half-lives than their parent compounds.
Original languageEnglish
Pages (from-to)95-107
Number of pages13
JournalForensic Toxicology
Issue number1
Early online date13 Jul 2019
Publication statusPublished - 1 Jan 2020


  • 5F-NPB-22
  • Biomarkers
  • Carboxylate synthetic cannabinoids
  • Ethanol
  • In vitro drug metabolism
  • Transesterification


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