5F-cumyl-PINACA in ‘e-liquids’ for electronic cigarettes – A new type of synthetic cannabinoid in a trendy product Institute of Forensic Medicine Verena Angerer, Bjoern Moosmann, Florian Franz and Volker Auwärter Forensic Toxicology Insitute of Forensic Medicine, Forensic Toxicology, Medical Center – University of Freiburg, Germany Introduction: In recent years, e-liquids used in electronic cigarettes have become an increasingly attractive alternative to smoking tobacco. Especially among young people e-cigarettes are becoming more and more popular [1]. A new trend is the use of e-liquids containing synthetic cannabinoids instead of nicotine as active ingredients. In the frame of the EU-Projects ‘SPICE’, ‘SPICE II Plus’ and ‘SPICE Profiling’, which comprise a systematic monitoring of the online market of ‘legal highs’, e-liquids were bought from online retailers who also sell herbal blends. Product monitoring Metabolism Shop X Method Method in vivo (human): Buying cumyl-PINACA serum & urine (0.6 mg oral intake) Extraction with n-Hexane Library match in vitro: GC-MS (in-house library) 5F-cumyl-PINACA incubation 1 h, 37 °C (n = 16) cumyl-PINACA pooled human liver n = 21 microsomes LC-MS/MS analysis (pHLM) Unknown Unknown Comparison to Match (3 E-liquids) (n = 5) EDND-Database (n = 2) Results HRMS, MSn After ingestion of 0.6 mg cumyl-PINACA serum concentration 0.1 O orally, the volunteer did not experience • Elemental composition 0.08 NH any drug-related symptom. Cumyl- • Structural information 0.06 N 0.04 N PINACA itself could be detected in serum c [ng/ml] e.g. 0.02 Isolation with semi- over a period of about 17 h. The Fig 1: 0 preparative HPLC or maximum concentration observed was 0 10203040 Flash Chromatographie time [h] 5F-cumyl-PINACA 0.1 ng/ml (6 h after ingestion, Fig. 2). Pure substance F The main metabolites of cumyl-PINACA Fig.2: Serum concentration of (reference material) showed mono- and dihydroxylation at the cumyl-PINACA after 0.6 mg (oral) pentyl moiety (in-vivo and in-vitro) and Results could be detected for at least 31 h (Fig.3) 47 % of the e-liquids contained only nicotine as active ingredient. The Main metabolites of cumyl-PINACA other liquids contained one or more synthetic cannabinoids, e.g. O O O 5F-APINACA, AB-PINACA or 5F-PB-22. Three of the liquids sold as NH NH NH ‘c-liquids’ (all from one retailer) contained 5F-cumyl-PINACA (Fig. 1). N N Hydroxylation N Hydroxylation N N N in vitro + in vivo in vitro + in vivo To assess the relative potency of this new substance class – carrying M01 M07 (probably a cumyl-moiety - a set of synthetic cannabinoids (AB-CHMINACA, HO M02 HO 5-hydroxy-Pentyl) M08 M10 AB-FUBINACA, AB-PINACA, AM-2201, cumyl-PINACA, EG-018, HO JWH-018, MDMB-CHMICA, THJ-2201) were characterised using the cumyl-PINACA Oxidation Oxidation cAMP Biosensor Assay with CB1 as target (DiscoveRx, Fremont, 1.40E+07 in vitro + in vivo in vitro USA). The results of the Biosensor Assay are listed in Table 1. 1.20E+07 M07 M08 1.00E+07 O M10 NH O Assay 8.00E+06 M09 NH Compound Name EC50 (nM) Curve Top Max Response M01 Format 6.00E+06 M02 N 4.00E+06 N AB-CHMINACA Agonist 0.28 94.8 94.827 Area/Crea100 N Hydroxylation N 2.00E+06 AB-FUBINACA Agonist 0.89 97.4 97.82 0.00E+00 in vitro AB-PINACA Agonist 1.74 92.3 95.254 0.0 10.0 20.0 30.0 40.0 Time [h] AM-2201 Agonist 0.45 103 101.41 O HO Fig.3: main metabolites of cumyl- M09 Cumyl-PINACA Agonist 0.06 93.7 92.86 PINACA in urine after 0.6 mg (oral) O EG-018 Agonist 40.7 74.3 71.826 JWH-018 Agonist 1.13 97.6 97.392 Main metabolites of 5F-cumyl-PINACA (in vitro only) MDMB-CHMICA Agonist 0.14 94.8 94.57 O THJ-2201 Agonist 1.68 95.6 91.92 NH O Table.1: results of the cAMP Biosensor Assay NH N N O N NH Conclusion N N HO The increasing popularity of e-liquids particularly among young people N and the extreme potency of the added synthetic cannabinoids pose a F serious threat to public health. There is a high risk of unintended poisoning, and in the long term prevalence of these drugs could rise in F the younger population due to introduction of trendy products. 5F-cumyl-PINACA HO Acknowledgement: References: Contact: The authors would like to thank Sonja Klemenc from National Forensic Laboratory Ljubljana, Slovenia for providing [1] Camenga DR, Cavallo DA, Kong G, et al. Verena Angerer cumyl-PINACA. Adolescents' and young adults' perceptions of Institute of Forensic Medicine, Forensic Toxicology, electronic cigarettes for smoking cessation: A focus This publication has been produced with the financial support of Medical Center – University of Freiburg group study. Nicotine & tobacco research: official the European Commission (JUST/2011/ Albertstr. 9, 79104 Freiburg, Germany DPIP/AG/3597 and JUST/2013/ISEC/DRUGS/ journal of the Society for Research on Nicotine and AG/6421), the German Federal Ministry of Tobacco 2015. [email protected] Health and the City of Frankfurt/Main..