The Screening and Confirmation of NBOMe Designer Drugs on Blotter Paper by Ambient Ionization Mass Spectrometry and FTIR Chemical Imaging Spectroscopy Amanda Moore1 , Sabra Botch-Jones1, David Barajas1, Tom Byron2, Ryan Smith2, Jamie Foss2,3, Jason Weisenseel2, Kevin Tyvoll2, Frank Kero2

1. Boston University School of Medicine, Biomedical Forensic Sciences, 72 E. Concord St., Boston, MA 02118 2. PerkinElmer Environmental Health , 2651 Warrenville Road, Ste. 100 , Downers Grove, IL 60515

3. Maine Health and Environmental Testing Laboratory, 221 State St., Augusta, ME 04333

Novel AspectsNo prep analysis of fortified blotter paper samples for illicit designer drugs of abuse using FTIR chemical imaging for orthogonal analysis versus ambient ionization mass spectrometry.

MAFS 2016 *For complete method details, contact: frank.kero@perkinelmer.com

LSD/ paper components mixture

LSD reference spectrum

NBOMe reference

Figure 12: Spatial determination of illicit substances by FTIR library matching

NBOMes

Figure 2. Structures of relevant designer drugs in this study

• NBOMe drugs (Figure 2) are also known as new psychoactive substances (NPS).

• N-(methoxybenzyl)-phenethylamine compounds are typically consumed on blotter paper.

• The user experience has been described as a more potent, but similar high when compared to LSD.

• These substances are emerging as an issue of public health as an increase in cause of death case studies have found a link to NBOMes.

• Reference spectra is provided in Figure 10

LSD

Introduction

Previous work from this group: N-(methoxybenzyl)-phenethylamine compounds (also known as “NBOMe’s”) are an emerging threat to public health with fatality and severe injury related investigations increasing throughout the country. The NBOMe compounds are obtained as “legal” highs through unregulated internet purchases and are typically administered via blotter paper. The user experience has been described as a more potent, but a similar high when compared to lysergic acid diethylamide (LSD). Users often believe they are ingesting LSD when in fact it was one or a combination of the NBOMe compounds. Previous reports from this group have demonstrated the utility of non-targeted high resolution mass spectrometry platforms for this application paired with both ambient ionization and electrospray ionization sources since authentic case work samples are often obtained as mixtures of true unknowns (reported at NEAFS 2015). This previous work employed a minimal sample preparation via a 5 min liquid extraction of the target compound(s) from the blotter paper. This work was completed in collaboration with the State of Maine Department of Health and Human Services (Augusta, Maine), the University of Central Florida (Orlando, Florida ) and Boston University School of Medicine Biomedical Forensic Sciences Program (Boston, Massachusetts). The next step: In an extension of this work, chemical imaging using Fourier Transform Infrared Spectroscopy (FTIR) was evaluated to provide an orthogonal workflow solution for confirmation and to leverage the advantage of Direct Sample Analysis-Time-of-Flight Mass Spectrometry (DSA-TOFMS, PerkinElmer, Waltham, MA, USA) for screening (e.g.to reduce the speed of analysis, reduced cost in consumables and waste etc.). FTIR chemical imaging method development variables considered included variation in sampling depth and scan modes to generate a spatial chemical map of the sample. Method: Certified reference material of 25C-NBOME (Cerilliant, Round Rock, TX, USA) was analyzed as “cast” films by allowing a liquid solution in methanol to evaporate before analysis. In addition, 25C-NBOMe was spiked onto blotter paper. Further, certified reference material of LSD (Cerilliant) was also analyzed for comparison. A macro Geranium (Ge) Attenuated Total Reflectance (ATR) sampling accessory was used. The scan range was 4,000-748 cm-1, spectral resolution was 8 cm-1, scans per pixel was 4, field of view was 500 x 500 um, special resolution 1.56 µm, measures spectra of 102,4000, and the total number of scans were 64. By utilizing the Ge ATR, the diffraction limits were 2.5 um at 1,000 cm-1.

Utilizing the spectral library and chemical imaging, LSD was identified and able to be separated from the blotter paper components which were identified as polyethylene/carbonate, cellulose, hydrocarbon/fluorocarbon/carbonate mixtures. The correlation for the NBOMe was determined to be ~0.78 and the same components were identified from the blotter paper. There are indications that this technique may be used to identify the presences of LSD and NBOMe on a paper matrix. High spatial resolution is needed to isolate the chemical entity from the paper constituents, however it results in a small field of view or area measured limiting the ability to examine the whole matrix. If drug is not well dispersed within paper matrix, technique may produce a negative result do to limitation of area measured. It is anticipated that this workflow would be of broad-based interest to the forensic community on the topic of emerging drugs of abuse, designer and synthetic drugs. More data on case samples is needed to confirm conclusions on the utility of FTIR imaging for this matrix.

Figure 1: Chemical Structure of LSD

• Lysergic acid diethylamide = LSD (Figure 1)

• Well reported hallucinogenic substance

• Culture of abuse emerged in the 1950’s

• Clear or white odorless material

• Sublingual dosage on blotter paper

• Reference spectra is provided in Figure 10.

An overview of the DSA source and instrument housing is provided in Figures 5 and 5. Method details

• Probe Temp: 300°C• Nebulizer N2 Gas flow: 80psi• Aux Gas: 4 L/min• Dry Gas Heater 25°C• Corona current: 6 µAmp• Scan rate: 10 scans / second• Cap entrance: -800 V• Endplate: -200 V• Capillary Exit Voltage: 120 V• Note: DSA uses APCI tune mix for lock mass ions for daily

calibration of the TOFMS

Figure 4. DSA source schematic

Figure 5. DSA-TOFMS (left) DSA (top right) DSA ion source with mesh target screen (bottom right)

Blotter paper samples

Figure 4. Samples as received

• Blotter paper samples were fortified at Boston University (Figure 3) to support method development.

• This concentration was selected to be comparable with current dosages relevant for consumption.2

• Solution standards obtained from Cayman Chemical (Ann Arbor, Michigan) were included in the shipment for method development (Figure 4)

Figure 3. a) blotter paper b) fortified at Boston University

Untreated paper NBOMe treaded paper LSD treaded paper

Figure 11 Chemical imaging of fortified blotter paper with hallucinogenic drugs of abuse

Analysis of an Authentic Forensic Case Sample: Suspected LSD

[C18H22ClNO3 +H]+

[C8H9O]+

[C7H7]+

m/z 121

m/z 91

Figure 8. Fragmentation pathway to m/z 121 and m/z 91 ions

Figure 7. (Top) Singe dosage unit analyzed under initial instrument conditions with a capillary exit setting of 100V (10 μL sample volume). Mass Accuracy = 1.2 ppm. (Bottom) Single dosage unit analyzed under adjusted instrument conditions with a capillary exit setting of 120V showing CID fragmentation (10 μL sample volume). Mass accuracy = 0.60 ppm.

Figure 6. Suspected LSD blotter paper seized in Maine.

Under the hood• Does not require N2 purge• MIR/NIR• 1.56µm spatial resolution• Mode switching• Hot and cold stage options• Software automated atmospheric compensation

Figure 9: FTIR Chemical Imaging instrumentation

PerkinElmer Spotlight 400

A)

B)

Real case sample: A perforated blotter paper was seized in Bangor, ME and submitted to the Maine Health and Environmental Testing Laboratory as suspected LSD for analysis by DSA-TOFMS.

• Sample prep – 5 min liquid extraction with DCM• Sample volume for analysis - 10 µL • The analysis time per replicate was 15 sec

DSA-TOFMS identified the sample as 25C-NBOMe. Results were confirmed by GC-MS….. This method has a relatively long analysis time by comparison.

Q: Can no prep FTIR imaging do better? A: Possibly (more data needed to confirm)

Figure 10: FTIR reference spectra for the Selected analytes

Please note: This DSA-TOFMS case study has recently been published in Forensic Science International 267 (2016) 89–95

Stand alone FTIR reference spectra

LSD

25C-NBOMe