Studies on the metabolism and detectability of the emerging drug of abuse diphenyl‐2‐pyrrolidinemethanol (D2PM) in rat urine using GC‐MS and LC‐HR‐MS/MS

In the last years, the number of new psychoactive substances, so‐called ‘legal highs’, has enormously increased. They are sold via online shops often with inaccurate and false information about the content. The aim of this work was to study the metabolism and the detectability of the drug of abuse diphenyl‐2‐pyrrolidinemethanol (D2PM) in rat urine using gas chromatography‐mass spectrometry and liquid chromatography‐high resolution‐tandem mass spectrometry. Five phase I and two phase II metabolites were identified suggesting hydroxylation at the pyrrolidine and diphenyl part as the main metabolic steps. Assuming similar kinetics, an intake of D2PM should be detectable in human urine mainly via its metabolites. Copyright © 2013 John Wiley & Sons, Ltd.     

Multiple‐injection high‐throughput gas chromatography analysis

Multiple‐injection techniques have been shown to be a simple way to perform high‐throughput analysis where the entire experiment resides in a single chromatogram, simplifying the data analysis and interpretation. In this study, multiple‐injection techniques are applied to gas chromatography with flame ionization detection and mass detection to significantly increase sample throughput. The unique issues of implementing a traditional “Fast” injection mode of multiple‐injection techniques with gas chromatography and mass spectrometry are discussed. Stacked injections are also discussed as means to increase the throughput of longer methods where mass detection is unable to distinguish between analytes of the same mass and longer retentions are required to resolve components of interest. Multiple‐injection techniques are shown to increase instrument throughput by up to 70% and to simplify data analysis, allowing hits in multiple parallel experiments to be identified easily.     

Efficient preparative separation of 6‐(4‐aminophenyl)‐5‐methyl‐4, 5‐dihydro‐3(2H)‐pyridazinone enantiomers on polysaccharide‐based stationary phases in polar organic solvent chromatography and supercritical fluid chromatography

6‐(4‐Aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone is a key synthetic intermediate for cardiotonic agent levosimendan. Very few studies address the use of chiral stationary phases in chromatography for the enantioseparation of this intermediate. This study presents two efficient preparative methods for the isolation of (R)(?)‐6‐(4‐aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone in polar organic solvent chromatography and supercritical fluid chromatography using polysaccharide‐based chiral stationary phases and volatile organic mobile phases without additives in isocratic mode. Under optimum conditions, Chiralcel OJ column showed the best performance (α = 1.71, Rs = 5.47) in polar organic solvent chromatography, while Chiralpak AS column exhibited remarkable separations (α = 1.81 and Rs = 6.51) in supercritical fluid chromatography with an opposite enantiomer elution order. Considering the sample solubility, runtime and solvent cost, the preparations were carried out on Chiralcel OJ column and Chiralpak AS column (250 × 20 mm i.d.; 10 µm) in polar organic mode and supercritical fluid chromatography mode with methanol and CO₂/methanol as mobile phases, respectively. By utilizing the advantages of chromatographic techniques and polysaccharide‐based chiral stationary phases, this work provides two methods for the fast and economic preparation of (R)(?)‐6‐(4‐aminophenyl)‐5‐methyl‐4,5‐dihydro‐3(2H)‐pyridazinone, which are suitable for the pharmaceutical industry.     

Combination of ultrasound‐assisted ethyl chloroformate derivatization and switchable solvent liquid‐phase microextraction for the sensitive determination of l‐methionine in human plasma by GC–MS

A green and fast analytical method for the determination of l ‐methionine in human plasma is presented in this study. Preconcentration of the analyte was carried out by switchable solvent liquid phase microextraction after ethyl chloroformate derivatization reaction. Instrumental detection of the analyte was performed by means of gas chromatography–mass spectrometry. N,N‐Dimethyl benzylamine was used in the synthesis of switchable solvent. Protonated N,N‐dimethyl benzylamine volume, volume/concentration of sodium hydroxide, and vortex period were meticulously fixed to their optimum values. Besides, ethyl chloroformate, pyridine, and ethanol volumes were optimized in order to get high derivatization yield. After the optimization studies, limit of detection and quantitation values were attained as 3.30 and 11.0 ng/g, respectively, by the developed switchable solvent liquid phase microextraction gas chromatography–mass spectrometry method that corresponding to 76.7‐folds enhancement in detection power of the gas chromatography–mass spectrometry system. Applicability and accuracy of the switchable solvent liquid phase microextraction–gas chromatography–mass spectrometry method were also checked by spiking experiments. Percent recovery results were ranged from 97.8 to 100.5% showing that human plasma samples could be analyzed for its l ‐methionine level by the proposed method.     

Mass spectrometric properties of N‐(2‐methoxybenzyl)‐2‐(2,4,6‐trimethoxyphenyl)ethanamine (2,4,6‐TMPEA‐NBOMe), a new representative of designer drugs of NBOMe series and derivatives thereof

Emergence of new psychoactive substances, hallucinogenic phenethylamines in particular, in illicit market is a serious threat to human health in global scale. We have detected and identified N‐(2‐methoxybenzyl)‐2‐(2,4,6‐trimethoxyphenyl)ethanamine (2,4,6‐TMPEA‐NBOMe), a new compound in NBOMe series. Identification was achieved by means of gas chromatography/mass spectrometry (GC/MS), including high‐resolution mass spectrometry with tandem experiments (GC/HRMS and GC/HRMS²), ultra‐high performance liquid chromatography/high‐resolution mass spectrometry with tandem experiments (UHPLC/HRMS and UHPLC/HRMS²), and ¹H and ¹³C nuclear magnetic resonance spectroscopy. The peculiarities of fragmentation of the compound under electron ionization (EI) and collision‐induced dissociation were studied. Despite of the empirical rule denying migration of the hydrogen atom in McLafferty rearrangement to the benzene ring with substituents in the both ortho‐positions, it easily occurs for 2,4,6‐TMPEA‐NBOMe in EI conditions. We have noticed that electron‐donating substituents, e.g. methoxy groups in the both ortho‐positions and para‐positions favor the rearrangement. For specially synthesized N‐methyl and N‐acyl derivatives McLafferty rearrangement is not observed. N‐Acyl derivatives demonstrate McLafferty rearrangement, but the charge retains at the alternative fragment involving N‐acyl carbonyl group. We have also showed that the hydrogen atoms in 2,4,6‐trimethoxybenzene ring may be easily substituted for deuterium or for strong electrophiles like trifluoroacetyl. Analytical characteristics of 2,4,6‐TMPEA‐NBOMe and of some derivatives thereof which enable their determination in various criminal seizures are given. Copyright © 2016 John Wiley & Sons, Ltd.     

Synthesis of Deuterium Labeled (±)‐2,5‐Dimethoxyamphetamine (DMA), (±)‐4‐Bromo‐2,5‐Dimethoxyamphetamine (DOB), and (±)‐4‐Methoxyamphetamine (PMA) as Internal Standards for Quantitative Analyses by GC‐MS

DMA, DOB and PMA are increasingly abused central nervous system stimulants with neurotoxic properties. In recent years, many controlled substance analogs (designer drugs) with a large variety of structures have reached the illegal market, making their identification difficult. Therefore, this work studies the synthesis of DOB‐d₆, DMA‐d₆ and PMA‐d₃ as internal standards for use in gas chromatography‐mass spectrometry (GC‐MS) to identify controlled substances.     

Evaluating novel silica hydride‐based stationary phases for the analysis of phytocannabinoids and other psychoactive drugs

Three silica hydride based novel chromatographic phases chemically‐bonded with allyloxy‐DL‐alpha‐tocopherol, allylpentafluorophenyl, and 1‐eicosene moieties were evaluated as separation media for selected phytocannabinoids and other substances of abuse. In order to assess column selectivity, a series of reference standards was analyzed and detected by using liquid chromatography with mass spectrometry. Further, quantitative detections of cannabidiol and tetrahydrocannabinol were attempted for the extracts of cannabis plants and cannabidiol gummy formulation. For potential bioanalytical applications, the columns were evaluated for substance screening in a human urine matrix. In summary, the newly developed columns are functional and effective for the analysis of phytocannabinoids and various psychoactive drugs with or without the presence of biological matrices.     

Enantiodifferentiation of α-ketols in sherry by one- and two-dimensional HRGC techniques

The enantiomeric distrubution of solerone (5-oxo-4-hexanolide) 1, ethyl 4-hydroxy- 5-oxohexanoate 2, ethyl 5-hydroxy-4-oxohexanoate 3, 4-oxo-5-hexanolide 4, and solerol (5-hydroxy-4-hexanolide) 5 in sherry wines was determined by several HRGC techniques. While gas chromatography-mass spectrometry on chiral cyclodextrin phases (chiral GC-MS) prevented any racemization of α-ketols 2 and 3 caused by keto-enol tautomerization during analysis, multidimensional gas chromatography MDGC (coupled either by “live-T” switching or by “moving column stream switching” MCSS) led to rearranged constitutional-and stereoisomers. The stereochemical results are discussed regarding the biogenesis of sherry constituents 1–5.     

Self‐Assembly of Chiral Propeller‐like Supermolecules with Unusual “Sergeants‐and‐Soldiers” and “Majority‐Rules” Effects

Chiral amplification is an interesting phenomenon in supramolecular chemistry mainly observed in complicated systems in which cooperative effect dominate. Herein, chiral, supramolecular, propeller‐like architectures have been constructed through coassembly of an achiral disk‐shaped molecule and chiral amino acid derivatives driven by intermolecular hydrogen bonding. Both the “sergeants‐and‐soldiers” principle and “majority‐rules” effect are applicable in these discrete four‐component supermolecules, which are the simplest supramolecular system ever reported that exhibit chiral amplification.     

Chemometrics comparison of gas chromatography with mass spectrometry and comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry Daphnia magna metabolic profiles exposed to salinity

The performances of gas chromatography with mass spectrometry and of comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry are examined through the comparison of Daphnia magna metabolic profiles. Gas chromatography with mass spectrometry and comprehensive two‐dimensional gas chromatography with mass spectrometry were used to compare the concentration changes of metabolites under saline conditions. In this regard, a chemometric strategy based on wavelet compression and multivariate curve resolution–alternating least squares is used to compare the performances of gas chromatography with mass spectrometry and comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry for the untargeted metabolic profiling of Daphnia magna in control and salinity‐exposed samples. Examination of the results confirmed the outperformance of comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry over gas chromatography with mass spectrometry for the detection of metabolites in D. magna samples. The peak areas of multivariate curve resolution–alternating least squares resolved elution profiles in every sample analyzed by comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry were arranged in a new data matrix that was then modeled by partial least squares discriminant analysis. The control and salt‐exposed daphnids samples were discriminated and the most relevant metabolites were estimated using variable importance in projection and selectivity ratio values. Salinity de‐regulated 18 metabolites from metabolic pathways involved in protein translation, transmembrane cell transport, carbon metabolism, secondary metabolism, glycolysis, and osmoregulation.     

Determination of three kinds of banned drugs in milk powder by hybrid solid‐phase extraction purification combined with gas chromatography and mass spectrometry

A gradient clean‐up method for the quantification of five kinds of banned drugs (two hormones, two sedatives, and one chloramphenicol) in milk powder was developed. We used the combination of solid‐phase extraction purification with gas chromatography and mass spectrometry. Milk powder was initially hydrolyzed by β‐glucuronidase/arylsulfatase, and then the hydrolyzed solution was concentrated and purified using a C₈ and cation resin solid‐phase extraction column. To isolate hormones and chloramphenicol drugs, products from the previous step were diluted with methanol and further purified using a silica and diatomite solid‐phase extraction column. After derivatization, the drugs were analyzed by gas chromatography with mass spectrometry, and the hydrolyzed solution was diluted with 5% ammoniated methanol to purify sedatives before gas chromatography with mass spectrometry analysis. Results showed that after adding the banned drugs at concentrations of 0.3–10.0 μg/kg, the average recovery range was 78.2–97.3% with relative standard deviations of 5.3–12.5%. The limit of quantification of the banned drugs (S/N ≥ 10) was 0.3–5.0 μg/kg, whereas the limit of detection (S/N ≥ 3) was 0.1–2.0 μg/kg. The solid‐phase extraction gradient purification system was simple, rapid, and accurate, and could satisfy the detection requirements of hormone, sedatives, and chloramphenicol drugs when used together with gas chromatography and mass spectrometry.     

Studies on the metabolism of the Δ9‐tetrahydrocannabinol precursor Δ9‐tetrahydrocannabinolic acid A (Δ9‐THCA‐A) in rat using LC‐MS/MS,LC‐QTOF MS and GC‐MS techniques

In Cannabis sativa, Δ9‐Tetrahydrocannabinolic acid‐A (Δ9‐THCA‐A) is the non‐psychoactive precursor of Δ9‐tetrahydrocannabinol (Δ9‐THC). In fresh plant material, about 90% of the total Δ9‐THC is available as Δ9‐THCA‐A. When heated (smoked or baked), Δ9‐THCA‐A is only partially converted to Δ9‐THC and therefore, Δ9‐THCA‐A can be detected in serum and urine of cannabis consumers. The aim of the presented study was to identify the metabolites of Δ9‐THCA‐A and to examine particularly whether oral intake of Δ9‐THCA‐A leads to in vivo formation of Δ9‐THC in a rat model. After oral application of pure Δ9‐THCA‐A to rats (15 mg/kg body mass), urine samples were collected and metabolites were isolated and identified by liquid chromatography‐mass spectrometry (LC‐MS), liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) and high resolution LC‐MS using time of flight‐mass spectrometry (TOF‐MS) for accurate mass measurement. For detection of Δ9‐THC and its metabolites, urine extracts were analyzed by gas chromatography‐mass spectrometry (GC‐MS). The identified metabolites show that Δ9‐THCA‐A undergoes a hydroxylation in position 11 to 11‐hydroxy‐Δ9‐tetrahydrocannabinolic acid‐A (11‐OH‐Δ9‐THCA‐A), which is further oxidized via the intermediate aldehyde 11‐oxo‐Δ9‐THCA‐A to 11‐nor‐9‐carboxy‐Δ9‐tetrahydrocannabinolic acid‐A (Δ9‐THCA‐A‐COOH). Glucuronides of the parent compound and both main metabolites were identified in the rat urine as well. Furthermore, Δ9‐THCA‐A undergoes hydroxylation in position 8 to 8‐alpha‐ and 8‐beta‐hydroxy‐Δ9‐tetrahydrocannabinolic acid‐A, respectively, (8α‐Hydroxy‐Δ9‐THCA‐A and 8β‐Hydroxy‐Δ9‐THCA‐A, respectively) followed by dehydration. Both monohydroxylated metabolites were further oxidized to their bishydroxylated forms. Several glucuronidation conjugates of these metabolites were identified. In vivo conversion of Δ9‐THCA‐A to Δ9‐THC was not observed. Copyright © 2009 John Wiley & Sons, Ltd.     

Enantioselective micellar electrokinetic chromatography of dl‐amino acids using (+)‐1‐(9‐fluorenyl)‐ethyl chloroformate derivatization and UV‐induced fluorescence detection

Chiral analysis of dl ‐amino acids was achieved by micellar electrokinetic chromatography coupled with UV‐excited fluorescence detection. The fluorescent reagent (+)‐1‐(9‐fluorenyl)ethyl chloroformate was employed as chiral amino acid derivatizing agent and sodium dodecyl sulfate served as pseudo‐stationary phase for separating the formed amino acid diastereomers. Sensitive analysis of (+)‐1‐(9‐fluorenyl)ethyl chloroformate‐amino acids was achieved applying a xenon‐mercury lamp for ultraviolet excitation, and a spectrograph and charge‐coupled device for wavelength‐resolved emission detection. Applying signal integration over a 30 nm emission wavelength interval, signal‐to‐noise ratios for derivatized amino acids were up to 23 times higher as obtained using a standard photomultiplier for detection. The background electrolyte composition (electrolyte, pH, sodium dodecyl sulfate concentration, and organic solvent) was studied in order to attain optimal chemo‐ and enantioseparation. Enantioseparation of 12 proteinogenic dl ‐amino acids was achieved with chiral resolutions between 1.2 and 7.9, and detection limits for most derivatized amino acids in the 13–60 nM range (injected concentration). Linearity (coefficients of determination > 0.985) and peak‐area and migration‐time repeatabilities (relative standard deviations lower than 2.6 and 1.9%, respectively) were satisfactory. The employed fluorescence detection system provided up to 100‐times better signal‐to‐noise ratios for (+)‐1‐(9‐fluorenyl)ethyl chloroformate‐amino acids than ultraviolet absorbance detection, showing good potential for d ‐amino acid analysis.     

Enantioselective determination of (R)‐ and (S)‐lansoprazole in human plasma by chiral liquid chromatography with mass spectrometry and its application to a stereoselective pharmacokinetic study

A simple and enantioselective method was developed and validated for the simultaneous determination of (R)‐ and (S)‐lansoprazole in human plasma by chiral liquid chromatography with tandem mass spectrometry. Lansoprazole enantiomers and internal standard (esomeprazole) were extracted from plasma using acetonitrile as protein precipitating agent. Baseline chiral separation was achieved within 9.0 min on a Chiralpak IC column (150 mm × 4.6 mm, 5 μm) with the column temperature of 30°C. The mobile phase consisted of 10 mM ammonium acetate solution containing 0.05% acetic acid/acetonitrile (50:50, v/v). The mass spectrometric analysis was performed using a QTrap 5500 mass spectrometer coupled with an electrospray ionization source in positive ion mode. The multiple reactions monitoring transitions of m/z 370.1→252.1 and 346.1→198.1 were used to quantify lansoprazole enantiomers and esomeprazole, respectively. For each enantiomer, no apparent matrix effect was found, the calibration curve was linear over 5.00–3000 ng/mL, the intra‐ and inter‐day precisions were below 10.0%, and the accuracy was –3.8 to 3.3%. Analytes were stable during the study. No chiral inversion was observed during sample storage, preparation procedure and analysis. The method was applied to the stereoselective pharmacokinetic studies in human after intravenous administration of dexlansoprazole or racemic lansoprazole.     

Recent applications of gas chromatography with high‐resolution mass spectrometry

Gas chromatography coupled to high‐resolution mass spectrometry is a powerful analytical method that combines excellent separation power of gas chromatography with improved identification based on an accurate mass measurement. These features designate gas chromatography with high‐resolution mass spectrometry as the first choice for identification and structure elucidation of unknown volatile and semi‐volatile organic compounds. Gas chromatography with high‐resolution mass spectrometry quantitative analyses was previously focused on the determination of dioxins and related compounds using magnetic sector type analyzers, a standing requirement of many international standards. The introduction of a quadrupole high‐resolution time‐of‐flight mass analyzer broadened interest in this method and novel applications were developed, especially for multi‐target screening purposes. This review is focused on the development and the most interesting applications of gas chromatography coupled to high‐resolution mass spectrometry towards analysis of environmental matrices, biological fluids, and food safety since 2010. The main attention is paid to various approaches and applications of gas chromatography coupled to high‐resolution mass spectrometry for non‐target screening to identify contaminants and to characterize the chemical composition of environmental, food, and biological samples. The most interesting quantitative applications, where a significant contribution of gas chromatography with high‐resolution mass spectrometry over the currently used methods is expected, will be discussed as well.     

A target‐group‐change couple with mass defect filtering strategy to identify the metabolites of “Dogel ebs” in rats plasma,urine and bile

“Dogel ebs” was known as Sophora flavescens Ait., a classical traditional Chinese Mongolian herbal medicine, which had the effects on damp‐heat dysentery, scrofula, and syndrome of accumulated dampness toxicity. Although the chemical constituents have been clarified by our previous studies, the metabolic transformation of “Dogel ebs” in vivo was still unclear. To explore the mechanism of “Dogel ebs,” the metabolites in plasma, bile, and urine samples were investigated. A fast positive and negative ion switching technology was used for the simultaneous determination of flavonoids and alkaloids in “Dogel ebs” in a single run. And a target‐group‐change coupled with mass defect filtering strategy was utilized to analyze the collected data. 89 parent compounds and 82 metabolites were characterized by high‐performance liquid chromatography with quadrupole exactive Orbitrap mass spectrometry. Both phase I and phase II metabolites were observed and the metabolic pathways involved in oxidation, demethylation, acetylation, and glucuronidation. 69 metabolites of “Dogel ebs,” including three hydroxyls bonding xanthohumol, formononetin‐7‐O‐glucuronide, 2′‐hydroxyl‐isoxanthohumol decarboxylation metabolite, oxysophocarpine dehydrogen, 9α‐hydroxysophoramine‐O‐glucuronide, etc. were reported for the first time.     

A Convenient Method to Synthesize 1,3‐Selenazolidin‐2‐ones from 2‐Chloroethyl Amine Salts by Se/CO System under Atmospheric Pressure and Temperature

1,3‐Selenazolidin‐2‐ones were successfully synthesized by the reaction of 2‐chloroethyl amine salts with the Se/CO system via selenation and carbonylation in the presence of NaOH under atmospheric pressure and temperature, and were characterized by gas chromatography–mass spectrometry and nuclear magnetic resonance.     

Advanced mono‐ and multi‐dimensional gas chromatography–mass spectrometry techniques for oxygen‐containing compound characterization in biomass and biofuel samples

A wide variety of biomass, from triglycerides to lignocellulosic‐based feedstock, are among promising candidates to possibly fulfill requirements as a substitute for crude oils as primary sources of chemical energy feedstock. During the feedstock processing carried out to increase the H:C ratio of the products, heteroatom‐containing compounds can promote corrosion, thus limiting and/or deactivating catalytic processes needed to transform the biomass into fuel. The use of advanced gas chromatography techniques, in particular multi‐dimensional gas chromatography, both heart‐cutting and comprehensive coupled to mass spectrometry, has been widely exploited in the field of petroleomics over the past 30 years and has also been successfully applied to the characterization of volatile and semi‐volatile compounds during the processing of biomass feedstock. This review intends to describe advanced gas chromatography–mass spectrometry‐based techniques, mainly focusing in the period 2011–early 2020. Particular emphasis has been devoted to the multi‐dimensional gas chromatography–mass spectrometry techniques, for the isolation and characterization of the oxygen‐containing compounds in biomass feedstock. Within this context, the most recent advances to sample preparation, derivatization, as well as gas chromatography instrumentation, mass spectrometry ionization, identification, and data handling in the biomass industry, are described.     

Thio‐Bromo “Click” Reaction Derived Polymer–Peptide Conjugates for Their Self‐Assembled Fibrillar Nanostructures

The synthesis and self‐assembly of peptide–polymer conjugates into fibrillar nanostructures are reported, based on the amyloidogenic peptide KLVFF. A strategy for rational synthesis of polymer–peptide conjugates is documented via tethering of the amyloidogenic peptide segment LVFF (Aβ_17‐20) and its modified derivative FFFF to the hydrophilic poly(ethylene glycol) monomethyl ether (mPEG) polymer via thio‐bromo based “click” chemistry. The resultant conjugates mPEG‐LVFF‐OMe and mPEG‐FFFF‐OMe are purified via preparative gel permeation chromatography technique (with a yield of 61% and 64%, respectively), and are successfully characterized via combination of spectroscopic and chromatographic methods, including electrospray ionization time‐of‐flight mass spectrometry. The peptide‐guided self‐assembling behavior of the as‐constructed amphiphilic supramolecular materials is further investigated via transmission electron microscopic and circular dichroism spectroscopic analysis, exhibiting fibrillar nanostructure formation in binary aqueous solution mixture.     

Separation of enantiomers of new psychoactive substances by high‐performance liquid chromatography

New psychoactive substances are defined as compounds with consciousness‐changing effects and have been developed simultaneously with classical drugs. They arise through structural modifications of illegal substances and are mainly produced to circumvent laws. Availability is simple, since new psychoactive substances can be purchased from the Internet. Among them many chemical drug compound classes are chiral and thus the two resulting enantiomers can differ in their effects. The aim of this study is to develop a suitable chiral high‐performance liquid chromatography separation method for a broad spectrum of new psychoactive substances using cellulose tris(3,5‐dichlorophenylcarbamate) as a chiral selector. Experiments were performed by high‐performance liquid chromatography in normal‐phase mode under isocratic conditions using ultraviolet detection. Direct separation was carried out on a high‐performance liquid chromatography column (Lux® i‐Cellulose‐5, 3.5 μm, Phenomenex®), available since 2016. Excellent separation results were obtained for cathinones. After further optimization, even 47 instead of 39 out of 52 cathinones showed baseline separation. For amphetamine derivatives, satisfactory results were not achieved. Further, new psychoactive substances from other compound classes such as benzofuranes, thiophenes, phenidines, phenidates, morpholines, and ketamines were partially resolved, depending on the polarity and degree of substitution. All analytes, which were mainly purchased from the Internet, were proven to be traded as racemates.