Analysis of 17 fentanyls in plasma and blood by UPLC-MS/MS with interpretation of findings in surgical and postmortem casework

Abstract

The opioid crisis is linked to an increased misuse of fentanyl as well as fentanyl analogs that originate from the illicit drug market. Much of our current understanding of fentanyl and fentanyl analog use in our communities comes from postmortem toxicology findings. In the clinical settings of addiction medicine and pain management, where the opioid abuse potential is high, the use of fentanyl, as well as specific fentanyl analogs, may be underestimated due to limited plasma testing and limited availability of assays with suitable analytical sensitivity and selectivity to detect misuse of fentanyls. We report plasma and blood assays for 17 fentanyls (these include fentanyl, fentanyl analogs, fentanyl metabolites and synthetic precursors) in clinical, and medical examiner, casework. A mixed-mode solid phase extraction of diluted plasma or precipitated blood was optimized for maximum recovery of the fentanyls with minimized matrix effects. Analysis was performed using a Waters ACQUITY UPLC I-Class interfaced with a Waters Xevo TQ-S micro tandem quadrupole mass spectrometer. Method parameters were optimized and validated for precision, accuracy, carryover, linearity and matrix effects. Application studies were performed in postmortem blood obtained in 44 fentanyl-related fatalities and in serial plasma samples from 18 surgical patients receiving intravenous fentanyl therapy while undergoing parathyroidectomy. Fentanyls found in postmortem cases included fentanyl, norfentanyl, despropionyl-fentanyl (4-ANPP), beta-hydroxy fentanyl (β-OH fentanyl), acetyl fentanyl, acetyl norfentanyl, methoxyacetyl fentanyl, furanyl fentanyl, cyclopropyl fentanyl, and para-fluorobutyryl fentanyl, with fentanyl, norfentanyl, 4-ANPP and β-OH fentanyl predominating in frequency. Fentanyl concentrations ranged from 0.2 to 56 ng/mL and fentanyl was nearly always found with 4-ANPP, norfentanyl and β-OH fentanyl. Concentrations of other fentalogs ranged from <1 to 84 ng/mL (extrapolated). In the surgical cases, fentanyl was detected and quantified along with norfentanyl and β-OH fentanyl, but without detection of 4-ANPP in any of the samples. The association and relative concentrations of β-OH fentanyl, fentanyl and norfentanyl in the postmortem and clinical studies indicated a metabolic, rather than an illicit, source of β-OH fentanyl.