Qualitative and Quantitative Analysis of Tryptamines in the Poison of Incilius alvarius (Amphibia: Bufonidae).

Rising numbers of psychoactive tryptamine derivatives have become available on the drug market over the last decade, making these naturally occurring or synthetically manufactured compounds highly relevant for forensic analyses. One of these compounds is 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT), a constituent of the dried poison of Incilius alvarius (Colorado River toad), which has a history of ritual and/or recreational use. Still, comprehensive and validated qualitative as well as quantitative analytical data on the psychoactive components of this poison are scarce. In this study, samples of the poison of Incilius alvarius were collected from live toads in the Sonoran Desert, Arizona (USA), and analyzed with a set of complementary methods. Acetone/water (70/30, v/v) proved to be the solvent of choice for the extraction of tryptamine derivatives. Trace compounds were enriched, and overload with 5-MeO-DMT was prevented by chromatographic separation of 5-MeO-DMT prior to qualitative analyses. The method for quantification was validated. Attenuated total reflection-Fourier transform infrared spectroscopy was suitable to identify 5-MeO-DMT as the main tryptamine in samples of the poison. The combined evaluation of analytical data gained from gas chromatography-mass spectrometry (GC-MS), high-performance liquid chromatography-quadrupole time-of-flight high-resolution MS (HPLC-qToF-HRMS) and HPLC-MS-MS confirmed the presence of 5-MeO-DMT, 5-MeO-N-methyltryptamine, 5-MeO-tryptamine, 5-MeO-tryptophol, 2-(5-methoxy-1H-indol-3-yl)-acetic-acid (5-MIAA), 5-HO-N-methyltryptamine, bufotenin, DMT and tryptophan. For the first time, 5-MeO-tryptamine and two positional isomers of hydroxylated MeO-DMT were detected in the poison of Incilius alvarius. The tryptamine present in the highest concentrations was 5-MeO-DMT (mean ± SD: 410,000 ± 30,000 μg/g). Mean concentrations of 5-MeO-tryptamine (490 ± 260 μg/g), 5-HO-N-methyltryptamine (270 ± 120 μg/g), bufotenin (2,800 ± 1,900 μg/g) and DMT (250 ± 80 μg/g) showed a relatively high variability between individual samples. The comprehensive analytical reference data of Incilius alvarius poison presented here might prove useful for forensic chemists.