Chemicals and reagents used in analyses were acetonitrile, methanol, and water (Chem Solve LC–MS; Witko, Łódź, Poland); ammonium formate (Fluka, Seelze, Germany); formic acid, 4-methylmethcathinone (4-MMC)-d3 and dimethyl sulfoxide (DMSO)-d6 (Sigma-Aldrich, Darmstadt, Germany).
Analyzed compound has been seized by the police during the search of parcels at the post office in May 2020. The secured evidence in the form of grey crystalline powder (Fig. 2) was packed in sachets of two different sizes: one of them contained 0.9 g, and the other 25.0 g. Performed analyses showed that each of the evidence contained only one psychoactive substance—etazene.
X-ray quality crystals were obtained by recrystallization of the crude sample (sachet content) from methanol. Suitable crystal was mounted on a thin quartz fiber attached to a copper pin and immediately transferred to the diffractometer. Crystallographic data were collected at 100 (2) K on a KUMA X-ray diffractometer equipped with an Oxford Cryosystems open-flow nitrogen cryostat, using ω-scan and graphite-monochromated Mo Kα (λ = 0.71073 Å) radiation (Agilent, Yarnton, Oxfordshire, UK). Cell refinement, data reduction and analysis were carried out with CRYSALISPro software (Agilent).
Structure was solved with direct methods using with SHELXS  and refined with full-matrix least-squares techniques on F2 with SHELXL . The H atoms were included in idealized geometry riding on their parent atoms with C–H = 0.95–0.99 Å, N–H = 0.88–1.00 Å, and with Uiso(H) = 1.2Ueq(CH, CH2, NH) or 1.5Ueq(CH3) except for water H atoms, which were located in the Fourier maps, refined with O–H distances restrained to 0.860(1) Å and then constrained parent atoms (AFIX 3 instruction). The molecular structure plots were prepared using Diamond . Full details can be found in the crystallographic information file (CIF) file CCDC 2016899.
Powder X-ray diffraction of the crude sample (sachet content) was performed using a Bruker D8 Advance X-ray diffractometer operating with Cu Kα radiation (λ = 1.5418 Å), equipped with a VÅNTEC-1 PSD (Bruker AXS GmbH, Karlsruhe, Germany). Data were recorded on a silicon low background sample holder in the range 5–60° 2θ with an angular step size of 0.016° and a counting time of 1 s per step.
NMR spectra were recorded in DMSO-d6 at room temperature on a Bruker 500 MHz Avance II spectrometer (Bruker, Billerica, MA, USA) (1H frequency 500.13 MHz, 13C 125.7 MHz). Spectra were referenced to the residual solvent signals (2.50 and 39.5 ppm). 2D experiments (correlation spectroscopy, heteronuclear single-quantum correlation spectroscopy, and heteronuclear multiple-bound correlation spectroscopy) were performed by means of standard Bruker software, recorded with 2048 data points in the t2 domain and up to 1024 points in the t1 domain, with a 0.5–1 s recovery delay. Analyzed sample was dissolved in DMSO-d6 with addition of gas HCl.
A total of 100 mg of the sample was dissolved in 5 mL of methanol and diluted 1000-fold with methanol. A total of 10 μL of diluted solution was transferred to a 150 μL vial with a cap containing 80 μL of methanol and 10 μL of internal standard (IS) (4-MMC-d3, 1 μg/mL), mixed on a vortex and analyzed by LC–MS/MS as below.
Analysis was performed using an ultra-high-performance liquid chromatograph (Nexera X2; Shimadzu, Kyoto, Japan). The separation was done using a Kinetex XB-C18 (150 × 2.1 mm i.d., 2.6 μm particle size column; Phenomenex, Torrance, CA, USA) with the thermostat set at 40 °C. The mobile phase consisted of a mixture of 10 mM ammonium formate and 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B). The gradient elution was carried out at a constant flow of 0.4 mL/min. The gradient applied was as follows: 0 min, 5% B; 12 min, 98% B; 14 min, 98% B; and 15 min, 5% B. The return to the initial gradient compositions (95% A/5% B) was held for 5 min.
Detection of the investigated compound was achieved using a triple-quadrupole mass spectrometer (LCMS-8050; Shimadzu). The spectrometer was equipped with an electrospray ionization source; determination of the investigated substance was carried out in the scan mode with positive ionization, and mass range of m/z 50–1000. The following MS parameters were fixed: nebulizing gas flow, 3 L/min; heating gas flow, 10 L/min; interface temperature, 250 °C; desolvation line temperature, 200 °C; heat block temperature, 350 °C; and drying gas flow, 10 L/min.
A 100 mg sample was dissolved in 5 mL of methanol and analyzed by GC–MS/MS. The GC–MS/MS analysis was performed using a gas chromatograph coupled with a triple quadrupole mass spectrometer (GC–MS-TQ8040; Shimadzu). The injector was maintained at 250 °C. Sample injection (2 µL) was in the splitless mode. Separation was conducted using the SH-5MSi column (30 m length, 0.25 mm inner diameter and 0.25 µm film thickness; Shimadzu). Helium was used as a carrier gas at the flow rate of 0.94 mL/min. The mass detector was set to positive electron ionization (EI) mode and the electron beam energy was 70 eV. The mass detector was operating in a full scan mode in the 40–500 amu range.
UV-VIS and FTIR spectroscopies
The UV-VIS absorption spectrum was recorded in the methanol solution using the UV-1900i UV/VIS spectrophotometer (Shimadzu), and the spectrum was collected in the range of wavelength 190–1100 nm at room temperature. The UV-VIS absorption spectrum was recorded in 1 cm quartz cuvette. The FTIR spectrum was recorded in the KBr tablet using the VERTEX 70 FTIR spectrometer (Bruker). Spectrum was collected in range of 4000–400 cm−1. Sample analysis was conducted at room temperature.