Benefits of Ion Mobility Separation in GC-APCI-HRMS Screening: From the Construction of a CCS Library to the Application to Real-World Samples.

The performance of gas chromatography (GC) combined with the improved identification properties of ion mobility separation coupled to high-resolution mass spectrometry (IMS-HRMS) is presented as a promising approach for the monitoring of (semi)volatile compounds in complex matrices. The soft ionization promoted by an atmospheric pressure chemical ionization (APCI) source designed for GC preserves the molecular and/or quasi-molecular ion information enabling a rapid, sensitive, and efficient wide-scope screening. Additionally, ion mobility separation (IMS) separates species of interest from coeluting matrix interferences and/or resolves isomers based on their charge, shape, and size, making IMS-derived collision cross section (CCS) a robust and matrix-independent parameter comparable between instruments. In this way, GC-APCI-IMS-HRMS becomes a powerful approach for both target and suspect screening due to the improvements in (tentative) identifications. In this work, mobility data for 264 relevant multiclass organic pollutants in environmental and food-safety fields were collected by coupling GC-APCI with IMS-HRMS, generating CCS information for molecular ion and/or protonated molecules and some in-source fragments. The identification power of GC-APCI-IMS-HRMS for the studied compounds was assessed in complex-matrix samples, including fish feed extracts, surface waters, and different fruit and vegetable samples.

Improving Target and Suspect Screening High-Resolution Mass Spectrometry Workflows in Environmental Analysis by Ion Mobility Separation.

Currently, the most powerful approach to monitor organic micropollutants (OMPs) in environmental samples is the combination of target, suspect, and nontarget screening strategies using high-resolution mass spectrometry (HRMS). However, the high complexity of sample matrices and the huge number of OMPs potentially present in samples at low concentrations pose an analytical challenge. Ion mobility separation (IMS) combined with HRMS instruments (IMS-HRMS) introduces an additional analytical dimension, providing extra information, which facilitates the identification of OMPs. The collision cross-section (CCS) value provided by IMS is unaffected by the matrix or chromatographic separation. Consequently, the creation of CCS databases and the inclusion of ion mobility within identification criteria are of high interest for an enhanced and robust screening strategy. In this work, a CCS library for IMS-HRMS, which is online and freely available, was developed for 556 OMPs in both positive and negative ionization modes using electrospray ionization. The inclusion of ion mobility data in widely adopted confidence levels for identification in environmental reporting is discussed. Illustrative examples of OMPs found in environmental samples are presented to highlight the potential of IMS-HRMS and to demonstrate the additional value of CCS data in various screening strategies.

Understanding the pharmacokinetics of synthetic cathinones: Evaluation of the blood-brain barrier permeability of 13 related compounds in rats.

Synthetic cathinones are the second most commonly seized new psychoactive substance family in Europe. These compounds have been related to several intoxication cases, including fatalities. Although the pharmacological effects, metabolism, and pharmacokinetics of cathinones have been studied, there is little information about the permeability of these compounds through the blood-brain barrier (BBB). This is an important parameter to understand the behavior and potency of cathinones. In this work, 13 selected cathinones have been analyzed in telencephalon tissue from Sprague-Dawley rats intraperitoneally dosed at 3 mg/kg. Our results revealed a direct relationship between compound polarity and BBB permeability, with higher permeability for the more polar cathinones. The chemical moieties present in the cathinone had an important impact on the BBB permeability, with lengthening of the α-alkyl chain or functionalization of the aromatic ring with alkyl moieties resulting in lower concentration in telencephalon tissue. Our data suggest that transport of cathinones is a carrier-mediated process, similar to cocaine transport across the BBB.

Mass spectrometric strategies for the investigation of biomarkers of illicit drug use in wastewater.

The analysis of illicit drugs in urban wastewater is the basis of wastewater-based epidemiology (WBE), and has received much scientific attention because the concentrations measured can be used as a new non-intrusive tool to provide evidence-based and real-time estimates of community-wide drug consumption. Moreover, WBE allows monitoring patterns and spatial and temporal trends of drug use. Although information and expertise from other disciplines is required to refine and effectively apply WBE, analytical chemistry is the fundamental driver in this field. The use of advanced analytical techniques, commonly based on combined chromatography-mass spectrometry, is mandatory because the very low analyte concentration and the complexity of samples (raw wastewater) make quantification and identification/confirmation of illicit drug biomarkers (IDBs) troublesome. We review the most-recent literature available (mostly from the last 5 years) on the determination of IDBs in wastewater with particular emphasis on the different analytical strategies applied. The predominance of liquid chromatography coupled to tandem mass spectrometry to quantify target IDBs and the essence to produce reliable and comparable results is illustrated. Accordingly, the importance to perform inter-laboratory exercises and the need to analyze appropriate quality controls in each sample sequence is highlighted. Other crucial steps in WBE, such as sample collection and sample pre-treatment, are briefly and carefully discussed. The article further focuses on the potential of high-resolution mass spectrometry. Different approaches for target and non-target analysis are discussed, and the interest to perform experiments under laboratory-controlled conditions, as a complementary tool to investigate related compounds (e.g., minor metabolites and/or transformation products in wastewater) is treated. The article ends up with the trends and future perspectives in this field from the authors' point of view. © 2016 Wiley Periodicals, Inc. Mass Spec Rev 37:258-280, 2018.

Prediction of Collision Cross-Section Values for Small Molecules: Application to Pesticide Residue Analysis.

The use of collision cross-section (CCS) values obtained by ion mobility high-resolution mass spectrometry has added a third dimension (alongside retention time and exact mass) to aid in the identification of compounds. However, its utility is limited by the number of experimental CCS values currently available. This work demonstrates the potential of artificial neural networks (ANNs) for the prediction of CCS values of pesticides. The predictor, based on eight software-chosen molecular descriptors, was optimized using CCS values of 205 small molecules and validated using a set of 131 pesticides. The relative error was within 6% for 95% of all CCS values for protonated molecules, resulting in a median relative error less than 2%. In order to demonstrate the potential of CCS prediction, the strategy was applied to spinach samples. It notably improved the confidence in the tentative identification of suspect and nontarget pesticides.

Monitoring a large number of pesticides and transformation products in water samples from Spain and Italy.

Assessing the presence of pesticides in environmental waters is particularly challenging because of the huge number of substances used which may end up in the environment. Furthermore, the occurrence of pesticide transformation products (TPs) and/or metabolites makes this task even harder. Most studies dealing with the determination of pesticides in water include only a small number of analytes and in many cases no TPs. The present study applied a screening method for the determination of a large number of pesticides and TPs in wastewater (WW) and surface water (SW) from Spain and Italy. Liquid chromatography coupled to high-resolution mass spectrometry (HRMS) was used to screen a database of 450 pesticides and TPs. Detection and identification were based on specific criteria, i.e. mass accuracy, fragmentation, and comparison of retention times when reference standards were available, or a retention time prediction model when standards were not available. Seventeen pesticides and TPs from different classes (fungicides, herbicides and insecticides) were found in WW in Italy and Spain, and twelve in SW. Generally, in both countries more compounds were detected in effluent WW than in influent WW, and in SW than WW. This might be due to the analytical sensitivity in the different matrices, but also to the presence of multiple sources of pollution. HRMS proved a good screening tool to determine a large number of substances in water and identify some priority compounds for further quantitative analysis.

Analytical strategy to investigate 3,4-methylenedioxypyrovalerone (MDPV) metabolites in consumers' urine by high-resolution mass spectrometry.

The potential of high-resolution mass spectrometry (HRMS) for the investigation of human in vivo metabolism of 3,4-methylenedioxypyrovalerone (MDPV) using urine collected from a consumer (this is, in non-controlled experiments) has been investigated. As a control sample was not available, the common approach based on the comparison of a control/blank sample and samples collected after drug intake could not be used. Alternatively, an investigation based on common fragmentation pathways was applied, assuming that most metabolites share some fragments with the parent drug. An extension of this approach was also applied based on the fragmentation pathway of those metabolites identified in urine samples in the first step. The use of MS(E) experiments (sequential acquisition of mass spectra at low and high collision energy) has been crucial to this aim as it allowed promoting fragmentation in the collision cell without any previous precursor ion selection. MDPV belongs to the group of new psychoactive substances (NPS), being known as the "cannibal drug". This substance is being abused more and more and is associated with dangerous side effects. The human metabolites (both phase I and phase II) were detected and tentatively identified by accurate mass full-spectrum measurements using ultra-high performance liquid chromatography coupled to hybrid quadrupole time-of-flight mass spectrometry (UHPLC-QTOF MS). Following this strategy, up to 10 phase I metabolites, together with some glucuronides and sulphates, were detected and tentative structures were proposed. Several compounds identified in this work have not been previously reported in the literature.

Mass spectrometric evaluation of mephedrone in vivo human metabolism: identification of phase I and phase II metabolites, including a novel succinyl conjugate.

In recent years, many new designer drugs have emerged, including the group of cathinone derivatives. One frequently occurring drug is mephedrone; although mephedrone was originally considered as a "legal high" product, it is currently banned in most Western countries. Despite the banning, abuse of the drug and seizures are continuously reported. Although the metabolism of mephedrone has been studied in rats or in vitro using human liver microsomes, to the best of our knowledge, no dedicated study with human volunteers has been performed for studying the in vivo metabolism of mephedrone in humans. Therefore, the aim of this study was to establish the actual human metabolism of mephedrone and to compare it with other models. For this purpose, urine samples of two healthy volunteers, who ingested 200 mg mephedrone orally, were taken before administration and 4 hours after substance intake. The discovery and identification of the phase I and phase II metabolites of mephedrone were based on ultra-high-performance liquid chromatography coupled to hybrid quadrupole time-of-flight mass spectrometry, operating in the so-called MS(E) mode. Six phase I metabolites and four phase II metabolites were identified, four of them not previously reported in the literature. The structure of four of the detected metabolites was confirmed by synthesis of the suggested compounds. Remarkably, a mephedrone metabolite conjugated with succinic acid has been identified and confirmed by synthesis. According to the reviewed literature, this is the first time that this type of conjugate is reported for human metabolism.

Screening of pharmaceuticals and illicit drugs in wastewater and surface waters of Spain and Italy by high resolution mass spectrometry using UHPLC-QTOF MS and LC-LTQ-Orbitrap MS.

The existence of pharmaceuticals and illicit drugs (PIDs) in environmental waters has led many analytical chemists to develop screening methods for monitoring purposes. Water samples can contain a huge number of possible contaminants, commonly at low concentrations, which makes their detection and identification problematic. Liquid chromatography coupled with high resolution mass spectrometry (LC-HRMS) has proven itself effective in the screening of environmental contaminants. The present work investigates the use of the most popular HRMS instruments, quadrupole time-of-flight and linear trap quadrupole-Orbitrap, from two different laboratories. A suspect screening for PIDs was carried out on wastewater (influent and effluent) and surface water samples from Castellón, Eastern Spain, and Cremona, Northern Italy, incorporating a database of 107 PIDs (including 220 fragment ions). A comparison between the findings of both instruments and of the samples was made which highlights the advantages and drawbacks of the strategies applied in each case. In total, 28 compounds were detected and/or identified by either/both instruments with irbesartan, valsartan, benzoylecgonine and caffeine being the most commonly found compounds across all samples.

Improvements in analytical methodology for the determination of frequently consumed illicit drugs in urban wastewater.

Rapid and sensitive analytical methodology based on ultra high-performance liquid chromatography-tandem mass spectrometry has been developed for the determination of widely consumed drugs of abuse (amphetamines, MDMA, cocaine, opioids, cannabis and ketamine) and their major metabolites in urban wastewaters. Sample clean-up and pre-concentration was performed by a generic off-line SPE procedure using Oasis HLB. Special effort was made to incorporate amphetamine, which was found highly problematic in the wastewater samples tested, including an additional clean-up with Oasis MCX SPE and dispersive primary secondary amine. Correction for possible SPE losses or degradation during storage was made by the use of isotope-labelled internal standards (ILIS), available for all compounds, which were added to the samples as surrogates. Although ILIS were also efficient for matrix effects correction, the strong ionization suppression observed was not eliminated; therefore, a four-fold dilution prior to SPE was applied to influent wastewaters and a low injection volume was selected (3 µL), in order to reach a compromise between matrix effects, chromatographic performance and sensitivity. The method was validated at 25 and 200 ng L(-1) (effluent), and 100 and 800 ng L(-1) (influent), obtaining limits of quantification (i.e. the lowest level that the compound can be quantified and also confirmed with at least two MS/MS transitions) between 0.4-25 ng L(-1) (effluent) and 2-100 ng L(-1) (influent). The applicability of the method was demonstrated by analysis of 14 influent and 14 effluent wastewater samples collected over 2 weeks in Castellón (Spain) within a European collaborative study.

Discovery of Food Intake Biomarkers Using Metabolomics.

Due to the high impact of diet exposure on health, it is crucial the generation of robust data of regular dietary intake, hence improving the accuracy of dietary assessment. The metabolites derived from individual food or group of food have great potential to become biomarkers of food intake (BFIs) and provide more objective food consumption measurements.Herein, it is presented an untargeted metabolomic workflow for the discovery BFIs in blood and urine samples, from the study design to the biomarker identification. Samples are analyzed by liquid chromatography coupled to high-resolution mass spectrometry (LC-HRMS). A wide variety of compounds are covered by separate analyses of medium to nonpolar molecules and polar metabolites based on two LC separations as well as both positive and negative electrospray ionization. The main steps of data treatment of the comprehensive data sets and statistical analysis are described, as well as the principal considerations for the BFI identification.

A comprehensive study on diesel oil bioremediation under microcosm conditions using a combined microbiological, enzymatic, mass spectrometry, and metabarcoding approach.

This study aims at the application of a marine fungal consortium (Aspergillus sclerotiorum CRM 348 and Cryptococcus laurentii CRM 707) for the bioremediation of diesel oil-contaminated soil under microcosm conditions. The impact of biostimulation (BS) and/or bioaugmentation (BA) treatments on diesel-oil biodegradation, soil quality, and the structure of the microbial community were studied. The use of the fungal consortium together with nutrients (BA/BS) resulted in a TPH (Total Petroleum Hydrocarbon) degradation 42% higher than that obtained by natural attenuation (NA) within 120 days. For the same period, a 72 to 92% removal of short-chain alkanes (C12 to C19) was obtained by BA/BS, while only 3 to 65% removal was achieved by NA. BA/BS also showed high degradation efficiency of long-chain alkanes (C20 to C24) at 120 days, reaching 90 and 92% of degradation of icosane and heneicosane, respectively. In contrast, an increase in the levels of cyclosiloxanes (characterized as bacterial bioemulsifiers and biosurfactants) was observed in the soil treated by the consortium. Conversely, the NA presented a maximum of 37% of degradation of these alkane fractions. The 5-ringed PAH benzo(a)pyrene, was removed significantly better with the BA/BS treatment than with the NA (48 vs. 38 % of biodegradation, respectively). Metabarcoding analysis revealed that BA/BS caused a decrease in the soil microbial diversity with a concomitant increase in the abundance of specific microbial groups, including hydrocarbon-degrading (bacteria and fungi) and also an enhancement in soil microbial activity. Our results highlight the great potential of this consortium for soil treatment after diesel spills, as well as the relevance of the massive sequencing, enzymatic, microbiological and GC-HRMS analyses for a better understanding of diesel bioremediation.

Enhancing spectral quality in complex environmental matrices: Supporting suspect and non-target screening in zebra mussels with ion mobility.

Identification of bioaccumulating contaminants of emerging concern (CECs) via suspect and non-target screening remains a challenging task. In this study, ion mobility separation with high-resolution mass spectrometry (IM-HRMS) was used to investigate the effects of drift time (DT) alignment on spectrum quality and peak annotation for screening of CECs in complex sample matrices using data independent acquisition (DIA). Data treatment approaches (Binary Sample Comparison) and prioritisation strategies (Halogen Match, co-occurrence of features in biota and the water phase) were explored in a case study on zebra mussel (Dreissena polymorpha) in Lake Mälaren, Sweden's largest drinking water reservoir. DT alignment evidently improved the fragment spectrum quality by increasing the similarity score to reference spectra from on average (±standard deviation) 0.33 ± 0.31 to 0.64 ± 0.30 points, thus positively influencing structure elucidation efforts. Thirty-two features were tentatively identified at confidence level 3 or higher using MetFrag coupled with the new PubChemLite database, which included predicted collision cross-section values from CCSbase. The implementation of predicted mobility data was found to support compound annotation. This study illustrates a quantitative assessment of the benefits of IM-HRMS on spectral quality, which will enhance the performance of future screening studies of CECs in complex environmental matrices.

Are preserved coastal water bodies in Spanish Mediterranean basin impacted by human activity? Water quality evaluation using chemical and biological analyses.

The Spanish Mediterranean basin is particularly susceptible to climate change and human activities, making it vulnerable to the influence of anthropogenic contaminants. Therefore, conducting comprehensive and exhaustive water quality assessment in relevant water bodies of this basin is pivotal. In this work, surface water samples from coastal lagoons or estuaries were collected across the Spanish Mediterranean coastline and subjected to target and suspect screening of 1,585 organic micropollutants by liquid chromatography coupled to ion mobility separation and high resolution mass spectrometry. In total, 91 organic micropollutants could be confirmed and 5 were tentatively identified, with pharmaceuticals and pesticides being the most prevalent groups of chemicals. Chemical analysis data was compared with data on bioanalysis of those samples (recurrent aryl hydrocarbon receptor (AhR) activation, and estrogenic receptor (ER) inhibition in wetland samples affected by wastewater streams). The number of identified organic contaminants containing aromatic rings could explain the AhR activation observed. For the ER antagonistic effects, predictions on estrogenic inhibition potency for the detected compounds were used to explain the activities observed. The integration of chemical analysis with bioanalytical observations allowed a comprehensive overview of the quality of the water bodies under study.

In-depth comparison of the metabolic and pharmacokinetic behaviour of the structurally related synthetic cannabinoids AMB-FUBINACA and AMB-CHMICA in rats.

Synthetic cannabinoids receptor agonists (SCRAs) are often almost completely metabolised, and hence their pharmacokinetics should be carefully evaluated for determining the most adequate biomarker in toxicological analysis. Two structurally related SCRAs, AMB-FUBINACA and AMB-CHMICA, were selected to evaluate their in vivo metabolism and pharmacokinetics using male Sprague-Dawley rats. Brain, liver, kidney, blood (serum) and urine samples were collected at different times to assess the differences in metabolism, metabolic reactions, tissue distribution and excretion. Both compounds experimented O-demethyl reaction, which occurred more rapidly for AMB-FUBINACA. The parent compounds and O-demethyl metabolites were highly bioaccumulated in liver, and were still detected in this tissue 48 h after injection. The different indazole/indole N-functionalisation produced diverse metabolic reactions in this moiety and thus, different urinary metabolites were formed. Out of the two compounds, AMB-FUBINACA seemed to easily cross the blood-brain barrier, presenting higher brain/serum concentrations ratio than AMB-CHMICA.

Use of quadrupole time-of-flight mass spectrometry to determine proposed structures of transformation products of the herbicide bromacil after water chlorination.

The herbicide bromacil has been extensively used in the Spanish Mediterranean region, and although plant protection products containing bromacil have been withdrawn by the European Union, this compound is still frequently detected in surface and ground water of this area. However, the fast and complete disappearance of this compound has been observed in water intended for human consumption, after it has been subjected to chlorination. There is a concern about the possible degradation products formed, since they might be present in drinking water and might be hazardous. In this work, the sensitive full-spectrum acquisition, high resolution and exact mass capabilities of hybrid quadrupole time-of-flight (QTOF) mass spectrometry have allowed the discovery and proposal of structures of transformation products (TPs) of bromacil in water subjected to chlorination. Different ground water samples spiked at 0.5 µg/mL were subjected to the conventional chlorination procedure applied to drinking waters, sampling 2-mL aliquots at different time intervals (1, 10 and 30 min). The corresponding non-spiked water was used as control sample in each experiment. Afterwards, 50 µL of the water was directly injected into an ultra-high-pressure liquid chromatography (UHPLC)/electrospray ionization (ESI)-(Q)TOF system. The QTOF instrument enabled the simultaneous recording of two acquisition functions at different collision energies (MS(E) approach): the low-energy (LE) function, fixed at 4 eV, and the high-energy (HE) function, with a collision energy ramp from 15 to 40 eV. This approach enables the simultaneous acquisition of both parent (deprotonated and protonated molecules) and fragment ions in a single injection. The low mass errors observed for the deprotonated and protonated molecules (detected in LE function) allowed the assignment of a highly probable molecular formula. Fragment ions and neutral losses were investigated in both LE and HE spectra to elucidate the structures of the TPs found. For those compounds that displayed poor fragmentation, product ion scan (MS/MS) experiments were also performed. On processing the data with specialized software (MetaboLynx), four bromacil TPs were detected and their structures were elucidated. To our knowledge, two of them had not previously been reported.

Building an empirical mass spectra library for screening of organic pollutants by ultra-high-pressure liquid chromatography/hybrid quadrupole time-of-flight mass spectrometry.

Hybrid quadrupole time-of-flight mass spectrometry (QTOF MS) has gained wide acceptance in many fields of chemistry, for example, proteomics, metabolomics and small molecule analysis. This has been due to the numerous technological advances made to this mass analyser in recent years. In the environmental field, the instrument has proven to be one of the most powerful approaches for the screening of organic pollutants in different matrices due to its high sensitivity in full acquisition mode and mass accuracy measurements. In the work presented here, the optimum experimental conditions for the creation of an empirical TOF MS spectra library have been evaluated. For this model we have used a QTOF Premier mass spectrometer and investigated its functionalities to obtain the best MS data, mainly in terms of mass accuracy, dynamic range and sensitivity. Different parameters that can affect mass accuracy, such as lock mass, ion abundance, spectral resolution, instrument calibration or matrix effect, have also been carefully evaluated using test compounds (mainly pesticides and antibiotics). The role of ultra-high-pressure liquid chromatography (UHPLC), especially when dealing with complex matrices, has also been tested. In addition to the mass accuracy measurements, this analyser allows the simultaneous acquisition of low and high collision energy spectra. This acquisition mode greatly enhances the reliable identification of detected compounds due to the useful (de)protonated molecule and fragment ion accurate mass information obtained when working in this mode. An in-house empirical spectral library was built for approximately 230 organic pollutants making use of QTOF MS in MS(E) mode. All the information reported in this paper is made available to the readers to facilitate screening and identification of relevant organic pollutants by QTOF MS.

Retrospective LC-QTOF-MS analysis searching for pharmaceutical metabolites in urban wastewater.

The presence of pharmaceuticals in the environment is a matter of major concern because of their wide consumption and their potential negative effect on the water quality and living organisms. After human and/or veterinary consumption, pharmaceuticals can be excreted in unchanged form as the parent compound and/or as free or conjugated metabolites. These compounds seem not to be completely removed during wastewater treatments and might finally arrive to surface and ground waters. Consequently, both parent pharmaceuticals and metabolites are target analytes to be considered in analytical methodologies. The satisfactory sensitivity in full-acquisition mode, high-resolution, exact mass measurements and MS/MS capabilities of hybrid quadrupole time-of-flight (QTOF) mass spectrometry make of this technique a powerful analytical tool for the identification of organic contaminants. In this study, the use of QTOF-MS with the aid of specialised processing-data application managers has allowed the retrospective analysis of pharmaceuticals metabolites in urban wastewater without the need for additional injection of sample extracts. Around 160 metabolites have been investigated in wastewater samples previously analysed only for parent compounds using LC-QTOF under MS(E) mode (simultaneous recording of two acquisition functions, at low and high collision energy). The retrospective analysis was applied to search for pharmaceutical metabolites in parent-positive effluent wastewaters from the Spanish Mediterranean region. Five metabolites, such as clopidogrel carboxylic acid or N-desmethyl clarithromycin, were identified in the samples.

The key role of mass spectrometry in comprehensive research on new psychoactive substances.

New psychoactive substances (NPS) are a wide group of compounds that try to mimic the effects produced by the 'classical' illicit drugs, including cannabis (synthetic cannabinoids), cocaine and amphetamines (synthetic cathinones) or heroin (synthetic opioids), and which health effects are still unknown for most of them. Nowadays, more than 700 compounds are being monitored by official organisms, some of which have been recently identified in seizures and/or intoxication cases. Toxicological analysis plays a pivotal role in NPS research. A comprehensive investigation on NPS, from the first identification of a novel substance until its detection in drug users to help in diagnostics and medical treatment, requires the use of a wide variety of instruments and analytical strategies. This paper illustrates the key role of mass spectrometry (MS) along a comprehensive investigation on NPS. The synthetic cannabinoid XLR-11 and the synthetic cathinone 5-PPDi have been chosen as representative substances of the most consumed NPS families. Moreover, both compounds have been investigated at our laboratory in different stages of the three-step strategy considered in this article. The initial identification and characterisation of the compound in consumption products, the first reported metabolic pathway and the development of analytical methodologies for its determination (and/or their metabolites) in different toxicological samples are described. The analytical strategies and MS instruments are briefly discussed to show the reader the possibilities that MS instrumentation offer to analytical scientists. This publication aims to be a starting point for those interested on the NPS research field from an analytical chemistry point of view.

Use of ion mobility-high resolution mass spectrometry in metabolomics studies to provide near MS/MS quality data in a single injection.

The use of ion mobility separations (IMSs) in metabolomics approaches has started to be deeply explored in the last years. In this work, the use of liquid chromatography (LC) coupled to IMS-quadrupole time-of-flight mass spectrometry (QTOF MS) has been evaluated in a metabolomics experiments using single injection of the samples. IMS has allowed obtaining cleaner fragmentation spectra, of nearly tandem MS quality, in data-independent acquisition mode. This is much useful in this research area as a second injection, generally applied in LC-QTOF MS workflows to obtain tandem mass spectra, is not necessary, saving time and evading possible compound degradation. As a case study, the smoke produced after combustion of herbal blends used to spray synthetic cannabinoids has been selected as study matrix. The smoke components were trapped in carbon cartridges, desorbed and analyzed by LC-IMS-QTOF MS using different separation mechanisms (reversed phase and HILIC) and acquiring in both positive and negative mode to widen the chemical domain. Partial Least Squares-Discriminant Analysis highlighted several compounds, and ratio between N-Isopropyl-3-(isoquinolinyl)-2-propen-1-amine and quinoline allowed differentiating between tobacco and herbal products. These two compounds were tentatively identified using the cleaner fragmentation spectra from a single injection in the IMS-QTOF MS, with additional confidence obtained by retention time (Rt) and collisional cross section (CCS) prediction using artificial neural networks. Data from this work show that LC-IMS-QTOF is an efficient technique in untargeted metabolomics, avoiding re-injection of the samples for elucidation purposes. In addition, the prediction models for Rt and CCS resulted of help in the elucidation process of potential biomarkers.