Ammonia solution 32

Poly(propylene imine) dendrimer second-generation, phenol, 9-chloroacridine, ammonia solution (32%) and anhydrous magnesium sulfate were obtained from Sigma-Aldrich and used as received. All organic solvents: tetrahydrofurane, ethylacetat, chloroform, dichloromethane, acetonitrile, methanol, ethanol, used in this study were used as obtained from Sigma-Aldrich without any additional purifications. Dimethyl sulfoxide (DMSO) for molecular biology was used for antibacterial screening (Sigma–Aldrich). The antibacterial activity of the compounds was tested in vitro with Gram (+) bacterial strain Bacillus cereus (B. cereus) ATCC 11778 and Gram (−) bacterial strain Pseudomonas aeruginosa (P. aeruginosa) 1390, and the antifungal activity was screened on fungi strain Candida lipolytica (C. lipolytica) 7618. Microorganisms were obtained from the collection of the Institute of Microbiology, Bulgarian Academy of Sciences, Sofia, Bulgaria. Before experimental use, cultures from solid medium were sub cultivated in a liquid medium (meat-peptone broth, MPB), incubated (orbital shaker incubator) and used as a source of inoculums for each experiment.

All analytical standard solutions were purchased from Cerilliant (Texas, USA) at a concentration of 1.0 mg/mL (native chemicals) or 100 µg/mL (mass-labelled chemicals) in either methanol or acetonitrile. The native chemicals included amphetamine, benzoylecgonine, cocaine, ketamine, mephedrone, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), 3,4-methylenedioxyethylamphetamine (MDEA), methylenedioxymethamphetamine (MDMA), 6-monoacetylmorphine (6-MAM) and norketamine. The corresponding deuterated chemicals included 6-MAM-D₆, amphetamine-D₁₁, benzoylecgonine-D₈, cocaine-D₃, methamphetamine-D₁₁ and MDMA-D₅. The working solution was prepared in methanol at 10 µg/mL for a native-mixture standard and at 1 µg/mL for a mass-labelled-mixture standard (internal standard, IS) stored at −20 °C in amber vials. Ultrapure Milli-Q water was generated from the Milli-Q water purification system (Milli-Q IQ 7000, Merck, Darmstadt, Germany). Concentrated hydrochloric acid (HCl), LC-grade methanol, ammonia solution (32%) and Whatman glass microfiber filters (Grade GF/D) were purchased from Sigma Aldrich. Optima LC–MS grade formic acid was acquired from Fisher Scientific (New Hampshire, USA). Oasis MCX cartridges (150 mg, 6 cc) for solid-phase extraction (SPE) and the SPE manifold with 20 ports were acquired from Waters (Massachusetts, USA).