The 4-AcO-DMT of new psychoactive substances is constantly evolving, with arylalkylamines being one of the largest groups among them.

4-AcO-DMT binds to the dopamine transporter in rat brain cells with a lower affinity than cocaine. In silico data suggests that the primary action on dopamine is through reversal of the transporter to release dopamine.. 4-AcO is available from online vendors selling “research chemicals” and was first seize by German authorities in 2020. No scientific data about its pharmacological effect and its fate in the human body are available so far. Therefore we investigated the in vivo and in vitro metabolism of 4-AcO-DMTafter detection of this compound in a fatal intoxication.

Method 4-AcO-DMT online

For tentative identification of phase-I metabolites 4-AcO-DMT was incubate. with poole human liver microsomes (pHLM). Additionall, an authentic urine sample was worke up use protein precipitation .with and without prior glucuronide cleavage. Analyse for metabolite identification were carri out use high resolution LC-ESI(+)-QTOF-MS.

Chromatographic separation was achieve on a Kinetex Biphenyl column (100 × 2.1 mm, 2.6 μm). For the post-mortem investigation of the fatal intoxication. a standar addition approach use automate SPE for sample workup .was performe to quantify 4-AcO-DMT in femoral blood with LC-MS/MS.

Results

In the 4-AcO-DMT assay three phase-I metabolites were identifie, of which only two where confirme in the available authentic urine sample.

In addition six further phase-I and two phase-II metabolites were detecte in the analyze urine sample. Predominant metabolic reactions were hydroxylation, oxidation, N-dealkylation, glucuronidation and combination thereof. In urine an oxo-hydroxy metabolite, that was not generate in the 4-AcO-DMT assay. was detecte with the highest abundance. The concentration of  in the femoral blood sample was 7.100 ± 600 ng/mL.

Conclusion

The in vitro 4-AcO-DMT assay was not suitable to predict the main metabolites. DMT as confirmed in the available real case sample. The majority of the metabolic reaction and in particular the oxidation of 4-AcO-DMT appear to be catalyze by enzyme.

that are not present in an active state in the 4-AcO-DMT assay. Analyse of an authentic urine sample suggest DMT and N-demethyl as suitable biomarker for  consumption. To confirm the exaxt chemical strucuture of these markers isolation and structure elucidation, e.g. by NMR, or the synthesi of reference compound would be neede. In the fatal intoxication case, the elevate blood concentration of  4-AcO was considere likely to have contribute .