The increased availability of these analgesics subsequently caused widespread misuse of both prescription and non-prescription opioids. In 2018, more than 47,000 deaths in the USA were caused by opioid overdose, resulting in the US Department of Health and Human Services to declare a public health emergency the following year.
The increasing demand for opioids has resulted in an alarming level of novel psychoactive substances (NPS) in the illicit market in recent years. In particular, the level of fentanyl and its analogues is on the rise, which is thought to be due to its lower cost in comparison to heroin and its wide distribution and availability online.
Fentanyl is a μ-receptor agonist and binds 80–100 times more strongly than morphine, and its halogenated derivatives have been found to have increased psychotropic effects due to their greater affinity for specific receptors. These factors increase the potential of fatal intoxications when these opioids are abused. Therefore, it is crucial that known and emerging NPS can be rapidly identified once they have been seized by authorities to gain a greater understanding of their effects.
Scientists at Sapienza University of Rome, Italy, have been able to use a combination of analytical techniques, including nuclear magnetic resonance (NMR), to identify two fentanyl derivatives from a seized NPS sample.
Preliminary analysis of the unknown powder was performed by infrared (IR) spectroscopy. By comparing the data with IR compound libraries, the group were able to obtain a correspondence of 55% with fentanyl, suggesting a fentanyl analogue. The sample also showed a large presence of mannitol, a substance commonly used as a diluent in powder formulations.
Raman spectroscopy was then performed and detected the presence of two different substances in the unknown sample. As IR spectroscopy gave a partial match with fentanyl, these two substances were compared with known fentanyl analogues. One compound had some bands indicative of butyrylfentanyl, while the other had some bands characteristic of furanylfentanyl. These data verified the IR spectroscopy analysis of the sample consisting of fentanyl derivatives.
Gas chromatography-mass spectrometry (GC-MS) was used to elucidate further information on these two compounds. The chromatographic peak at retention time of 23.0 minutes was the most intense peak and gave a positive match to butyrylfentanyl upon comparison with MS compound libraries. Further analysis by liquid chromatography high-resolution MS/MS (LC–HRMS/MS) produced a mass spectrum with an observed mass peak at m/z 351.2433 (C23H31N2O+), which corresponds to the exact mass of butyrylfentanyl.
GC-MS also identified a second, less intense, chromatographic peak at retention time 25.8 minutes and the mass spectrum was found to correspond to fluoro-furanylfentanyl. Further analysis with LC–HRMS/MS gave a mass spectrum with a mass peak m/z 393.1973 (C24H26FN2O2+), the mass of fluoro-furanylfentanyl.
These analytical techniques were able to elucidate useful information about the components of the seized sample. However, the exact molecular structures of these compounds, in particular the position of the fluorine of the fluoro-furanylfentanyl, was not achieved. Therefore, NMR was performed to determine definite structures.
Using a Bruker BioSpin AVANCE III spectrometer, the team performed one-dimensional 1H NMR as well as two-dimensional homonuclear total correlation spectroscopy (1H-1H TOCSY) and heteronuclear single-quantum correlation spectroscopy (1H-13C HSQC). The proton spectrum was able to determine that the butyrylfentanyl was isobutyrylfentanyl, due to the presence of a doublet at 0.91 ppm. This multiplicity is not possible for a CH3 belonging to a linear chain, but only to an iso-butyl one. The information obtained from the three NMR techniques were able to identify that the fluorine is in position 4 of the fluorophenyl moiety of the fluoro-furanylfentanyl.
This case study demonstrated the importance of analytical techniques in the identification of illicit opioids. While all the techniques used where able to suggest the seized sample contained fentanyl derivatives, only NMR analysis was able to provide a definite structure of isobutyrylfentanyl and 4-fluoro-furanylfentanyl.
Isobutyrylfentanyl has previously been reported in seized samples, but 4-fluoro-furanylfentanyl was a newly identified compound. As a result of these data, formal notification of these substances to the national early warning system (NEWS) and the European Monitoring Center for Drugs and Drug Addiction (EMCDDA) was sent to continue to further the identification and understanding of illicit NPS and ultimately fight the opioid epidemic.
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