Biotransformation and detectability of the new psychoactive substances N,N-diallyltryptamine (DALT) derivatives 5-fluoro-DALT, 7-methyl-DALT, and 5,6-methylenedioxy-DALT in urine using GC-MS, LC-MSn, and LC-HR-MS/MS

Abstract

Derivatives of N,N-diallyltryptamine (DALT) can be classified as new psychoactive substances. Biotransformation and detectability of 5-fluoro-DALT (5-F-DALT), 7-methyl-DALT (7-Me-DALT), and 5,6-methylenedioxy-DALT (5,6-MD-DALT) are described here. Their metabolites detected in rat urine and pooled human liver microsomes were identified by liquid chromatography (LC)-high resolution (HR)-tandem mass spectrometry (MS/MS). In addition, the human cytochrome-P450 (CYP) isoenzymes involved in the main metabolic steps were identified and detectability tested in urine by the authors' urine screening approaches using GC-MS, LC-MS n ,or LC-HR-MS/MS. Aromatic and aliphatic hydroxylations, N-dealkylation, N-oxidation, and combinations could be proposed for all compounds as main pathways. Carboxylation after initial hydroxylation of the methyl group could also be detected for 7-Me-DALT and O-demethylenation was observed for 5,6-MD-DALT. All phase I metabolites were extensively glucuronidated or sulfated. Initial phase I reactions were catalyzed by CYP1A2, CYP2B6, CYP2C9, CYP2C19, CYP2D6, CYP3A4, and CYP3A5. Rat urine samples were analyzed following two different low dose administrations. GC-MS was not able to monitor consumption reliably, but all three drugs are predicted to be detectable in cases of overdose. The LC-MS n and LC-HR-MS/MS approaches were suitable for detect an intake of all three compounds mainly via their metabolites. However, after the lowest dose, a reliable monitoring could only be achieved for 5-F-DALT via LC-MS n and LC-HR-MS/MS and for 7-Me-DALT via LC-HR-MS/MS. The most abundant targets in both LC-MS screenings were one of two hydroxy-aryl metabolites and both corresponding glucuronides for 5-F-DALT, one N-deallyl hydroxy-aryl, the carboxy, and one dihydroxy-aryl metabolite for 7-Me-DALT, and the demethylenyl metabolite, its oxo metabolite, and glucuronide for 5,6-MD-DALT.

Toxicology and Analysis of Psychoactive Tryptamines

“…Table 2 summarizes the analytical methods reported for the determination of tryptamines in biological matrices, including information on sample size, sample preparation, instrumentation and validation parameters. Different biological matrices were employed for forensic purposes including conventional matrices urine [102,112,114,118,[121][122][123]125,126], blood [110,[113][114][115][117][118][119]121,123], plasma [109,111,112,118] serum [112] and unconventional matrices oral fluid [120] and hair [116,124]. Protein precipitation is a simple sample preparation method, providing sufficient extraction efficiency while simplifying extraction and preparation time.…”

“…Different protein precipitation methods [115,117,119,121] include different solvents for blood extraction but methanol is the most common solvent. Different sample preparation techniques include protein precipitation [113,116,117,119] for urine, serum and blood samples, acid hydrolysis [102,121,122] for urine, centrifugation [102,111,112,[114][115][116][117]119,[121][122][123][124]126] for blood, plasma and urine samples, washing [109,114,116,125] for plasma, blood and hair samples, enzymatic hydrolysis for urine samples [112], solid phase extraction (SPE) [109][110][111]114], liquid-liquid extraction (LLE) [114,120], dispersive liquid-liquid microextraction (DLLME) [115] and a solvent extraction for dried blood spots (DBS) [113]. Tryptamine is a relatively labile analyte but it remains stable in an acidic environment.…”

“…Most analytical methods included chromatographic separations and mass spectrometry detection. LC-MS/MS [109,[111][112][113][114][115][116][117]119,123,124], LC-HRMS/MS [102,122], GC-MS [102,118,122,123], LC-MS [118,123] and ultra-performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS/MS) [121] analytical methods are most common for forensic toxicology purposes. Different approaches utilized triple quadrupole full-scan mode [102,126], triple quadrupole multiple reaction monitoring (MRM) [109][110][111]113,116,119,124] with data dependent acquisition (DDA) [102] and photodiode array acquisition PDA [118], single quadrupole time of flight (TOF) [121,125].…”

“…Low limits of detection (LOD) are key for identifying tryptamines; two analytical methods achieved LODs of 0.10-0.15 ng/mL for 5-MeO-DiPT, with the same sample volume and LC-MS/MS technology but different sample preparation techniques [117,119]. Michely et al had LODs of 10-50 ng/mL for DALT and 5-MeO-DALT by GC-MS, with a 2.5 mL urine sample [102] but later achieved LODs of 1-5 ng/mL using LC-HRMS/MS and fifty times lower sample volume [122]. Also, Martin et al determined psilocin in 500 µL plasma with a 0.1 ng/mL LOD [111], whereas in their more recent method they present an LOD of 0.05 ng/mL using 1 mL of the same matrix with a different sample preparation [112].…”

Receptor binding profiles and behavioral pharmacology of ring-substituted N,N-diallyltryptamine analogs

“…The following drugs were tested: N,N -diallyltryptamine hydrochloride (DALT), 5-methoxy- N,N -diallyltryptamine hydrochloride (5-MeO-DALT), 5-fluoro- N,N -diallyltryptamine hydrochloride (5-F-DALT), 5-bromo- N,N -diallyltryptamine hydrochloride (5-Br-DALT), 4-hydroxy- N,N -diallyltryptamine fumarate (4-HO-DALT), 4-acetoxy- N,N -diallyltryptamine fumarate (4-AcO-DALT), 2-phenyl- N,N -diallyltryptamine hydrochloride (2-Ph-DALT), 5-methoxy-2-methyl- N,N -diallyltryptamine hydrochloride (5-MeO-2-Me-DALT), 5-fluoro-2-methyl- N,N -diallyltryptamine hydrochloride (5-F-2-Me-DALT), and 7-ethyl- N,N -diallyltryptamine hydrochloride (7-Et-DALT). 4-AcO-DALT fumarate and 4-HO-DALT hemifumarate were obtained from Scientific Supplies (London, UK); the other tryptamines were synthesized, fully characterized, and available from previous studies (Meyer et al 2014; Michely et al 2015; Dinger et al 2016; Brandt et al 2017a; Caspar et al 2017; Michely et al 2017).…”