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

Abstract

Synthetic cannabinoids, a subclass of new psychoactive substances (NPS), are laboratory-made substances that are chemically similar to those found naturally in the cannabis plant. Many of these substances are illicitly manufactured and have been associated with severe health problems, prompting a need to develop analytical methods capable of characterizing both known and previously undetected compounds. This work focuses on a novel Structures for Lossless Ion Manipulations (SLIM) IM-MS approach to the differentiation and structural characterization of synthetic cannabinoid metabolites, specifically MDA-19/BUTI-NACA, JWH-018, and JWH-250 isomer groups. These different compound classes are structurally very similar, differing only in the position of one or a few functional groups; this yielded similarity in measured collision cross section (CCS) values. However, the high resolution of SLIM IM provided adequate separation of many of these isomers, such as sodiated JWH-250 metabolites N-4-OH, N-5-OH, and 5-OH, which displayed CCS of 187.5, 182.5, and 202.3 Å 2 , respectively. In challenging cases where baseline separation was precluded due to nearly identical CCS, such as for JWH-018 isomers, simple derivatization by dansyl chloride selectively reacted with the 6-OH compound to provide differentiation of all isomers using a combination of CCS and m/z. Finally, the opportunity to use this method for structural elucidation of unknowns was demonstrated by using SLIM IM mobility-aligned MS/MS fragmentation. Different MDA-19/BUTINACA isomers were first mobility separated and could then be individually activated, yielding unique fragments for both targeted identification and structural determination. Overall, the described SLIM IM-MS/MS workflow provides significant potential as a rapid screening tool for the characterization of emerging NPS such as synthetic cannabinoids and their metabolites.

SLIM-Based High-Resolution Ion Mobility Reveals New Structural Insights into Isomeric Vitamin D Metabolites and their Isotopologues

“…SLIM traveling wave (TW) conditions were optimized (frequency and amplitude) for resolving power and signal abundance, by systematically ramping frequency or amplitude while keeping the other parameter constant; this yielded optimized TW conditions of 45 V and 35 000 Hz (Figure S1). SLIM CCS measurements required calibration using the Agilent Tune Mix, as previously, using these specifically optimized TW conditions. , Unlike in those previous studies, where SLIM demonstrated minor, yet easily correctable, SLIM CCS N2 bias (roughly ∼2–3% positive bias for lipids and ∼1% negative bias for synthetic cannabinoid metabolites), these values differed from their matching DT CCS N2 (Table S4) by an average of <0.1% (Table ); all values agreed with their corresponding DT CCS N2 values to within ±0.5%, with the only exception being a previously unresolved peak that will be discussed later. Additionally, the standard deviation of replicate measurements was ±0.2 Å 2 for nearly all compounds.…”

“…This recently developed IM technique allows for measurement of mobility as ions are propelled by a traveling wave through serpentine paths patterned on printed circuit boards. − The serpentine nature allows for long-path separations (≥13 m) that provide resolving power of R p > 200. Numerous small molecule applications have been demonstrated including separation of isomeric glycans, amino acids and peptides, , bile acids, lipids, , and most recently synthetic cannabinoid metabolites . An important consideration for the proposed coupling of high-resolution IM with quantitative clinical analysis is the anticipated arrival time shift for isotopically labeled internal standards relative to their unlabeled equivalents (i.e., isotopologues), based on differences in their reduced mass. − Such separations have been observed previously with SLIM for tetraalkylammonium ions, amino acids, and peptide isotopologues, , as well as with other low-field − and high-field IM platforms. − Williamson et al used a cyclic IMS (cIMS) device to measure structurally specific mass distribution-based isotopic shifts for 25-hydroxyvitamin D3, among other compounds, and interestingly they determined that the previously identified “closed” and “open” conformations had different relative shifts, with the “closed” showing only a 0.17% relative increase in arrival time in comparison with the 0.28% shift for the “open” conformer .…”

Rapid differentiation of xylazine metabolites using SLIM IM-MS

“…Our group has reported on a recently commercialized high-resolution IM platform, Structures for Lossless Ion Manipulations (SLIM), 17–19 for the rapid differentiation of synthetic cannabinoid isomers. 20…”

“…1B). To overcome this challenge, these compounds were derivatized using the common dansyl chloride reaction, 20,21 which selectively dansylated amines and phenolic alcohols. The success of this reaction (which was performed by combining the individual metabolites with dansyl chloride in a pH 9.0 buffer solution maintained at 60 °C for 5 minutes) was monitored by the singly dansylated product at m / z 470.157 (Fig.…”