Improved separation of fentanyl isomers using metal cation adducts and high‐resolution ion mobility‐mass spectrometry

Abstract

Fentanyl is a potent synthetic opioid that has attracted significant attention due to its illegal production and distribution, resulting in misuse, overdose, and fatalities. Because numerous fentanyl analogs, including structural isomers, with different potency have been discovered in the field, there is a critical need to continue developing analytical methodologies capable of accurate identification in forensic and clinical laboratories. This study aimed to develop a rapid method for detecting and separating fentanyl isomers based on ion mobility–mass spectrometry (IM‐MS), where IM separates gas‐phase ions based on differences in their size, shape, and charge. Several strategies for improved differentiation were implemented, including using unconventional cation adducts (e.g., alkali and transition metals) and data post‐processing by high‐resolution demultiplexing. A collection of collision cross section (CCS) values for the various metal ion adducts was gathered, which can be used to improve confidence of identification in future samples. Notable examples, such as [M + Cu]+ and [M + Ag]+ adducts, contributed to significant improvement of resolution between isomers. Furthermore, the addition of high‐resolution post‐processing provided resolving power of >150, which constitutes a significant increase in comparison with the normal 50–60 obtained with low‐resolution drift tube instruments. Collectively, these improved separation strategies allowed for confident detection and subsequent quantitative analysis. The optimized IM‐MS method resulted in quantification of fentanyl in human urine with limits of detection and quantification of 13 pg/mL and 40 pg/mL, respectively.

Identification of Unique Fragmentation Patterns of Fentanyl Analog Protomers Using Structures for Lossless Ion Manipulations Ion Mobility-Orbitrap Mass Spectrometry

“…These data were acquired over >50 scans to ensure data averaging was not the contributor to the difference. To further validate that this finding was real, we performed the same experiments using two lower collision energies (20,25).…”

“…The mass isolation window of the QMS was centered on the exact mass of each ion, and the narrowest possible quadrupole mass isolation window (±0.2 amu) was used. Ions were then sent to the HCD cell, and higher-energy fragmentation was performed using three different non-normalized collision energies (20,25,30). After fragmentation, ions were sent to the Orbitrap mass analyzer for high-resolution mass analysis (mass resolution setting = 140k).…”

“…Many positional isomers of fentanyls often cannot be distinguished by fragmentation alone, although multistage mass spectrometry (MS n ) experiments have shown potential in this area . Therefore, researchers have begun analyzing mixtures of fentanyl and fentanyl analogs with ion mobility (IM) spectrometry, which separates gas-phase ions based on their size (rotationally averaged collision cross section (CCS)) and electrical charge. − Zaknoun et al used a portable commercial drift tube (DT) IM spectrometer and atmospheric pressure chemical ionization (APCI) ionization to detect six novel fentanyl analogs in samples seized by law enforcement . Interestingly, each of the six fentanyl analogs exhibited two IM separable distributions with different intensities.…”

“…The same claims were later echoed by Aderorho and Chouinard while studying the effects of using metal cation adducts to achieve better IM separation between structurally similar fentanyl analogs. 20 Unfortunately, metal cationization does not prove or disprove the existence of protomers simply because there are no protonated species to evaluate, and many metal cations collapse the bimodal distribution observed with protonated species into a single peak, which distorts any conclusions that can be drawn about protomers. While these previous hypotheses were based on experimental observations, Lau et al addressed this question by using density functional theory (DFT) to theoretically explore the fragmentation pathways of three possible fentanyl protomers ((1) amide oxygen, (2) amide nitrogen, (3) piperidine nitrogen; see Supporting Information Figure S1 for chemical structure of fentanyl with labeled protonation sites).…”

Separation and Characterization of Synthetic Cannabinoid Metabolite Isomers Using SLIM High-Resolution Ion Mobility-Tandem Mass Spectrometry (HRIM-MS/MS)

“…All measurements were made in triplicate to report repeatability (in terms of the standard deviation). Further details of this system have been published elsewhere …”