Polyethylene microcentrifuge tubes

Sample preparation followed a published protocol 29. Briefly, ground Verbenae herba plant material (VO‐1 to VO‐3) was frozen in liquid nitrogen and homogenized with an analytical ball mill (Mikro‐Dismembrator, Sartorius, Göttingen, Germany). 100.0 ± 0.1 mg plant material was weighed into 1.5 mL polyethylene microcentrifuge tubes (Eppendorf, Hamburg, Germany), mixed with 1.0 mL ethanol/water (1:1, v/v) on a Vortex mixer (VWR, Vienna, Austria) and extracted by sonication for 10 min. After centrifugation (10621 x g for 5 min) supernatants were placed in a 5 mL volumetric flask. This procedure was repeated four more times, and the flask filled up to the final volume with the extraction solvent. All sample solutions were prepared in triplicated and filter prior analysis.

Gentian root batches
(R-1–R-13) were finely pulverized to homogeneity with a coffee
mill (3 × 1 min grinding time). The material (200.0 ± 0.1
mg) was weighed into 5.0 mL polyethylene microcentrifuge tubes (Eppendorf,
Hamburg, Germany), mixed with 2.0 mL methanol on a Vortex mixer (VWR,
Vienna, Austria), and sealed. Extraction was consequently conducted
by sonication for 10 min at ambient temperature (Bandelin Sonorex,
Berlin, Germany). To avoid temperature elevation, the water in the
ultrasonic bath was changed after each extraction step. After the
extraction, the samples were centrifuged at 10 600g for 5 min and the supernatant was transferred into a 10 mL volumetric
flask. To ensure complete extraction (absolute recovery), the whole
procedure was repeated four more times, the supernatants were combined,
and the flask was filled up to volume with methanol.
Regarding
the liquid preparations, 1.5 mL of each liqueur and clear spirit was
evaporated to dryness and redissolved in 0.75 mL of MeOH. Recovery
of this step was evaluated by the analogous workup of reference material
samples dissolved in 40% EtOH/water (v/v). Due to the high concentration
of 5, samples L-3 and L-7 had to be diluted 1:1 (v/v)
with methanol before quantitation. All sample solutions were prepared
in triplicate, filtered (0.45 μm cellulose acetate membrane,
VWR, Vienna, Austria), and stored at 4 °C until analysis.