The role of analytical chemistry in exposure science: Focus on the aquatic environment.

Exposure science, in its broadest sense, studies the interactions between stressors (chemical, biological, and physical agents) and receptors (e.g. humans and other living organisms, and non-living items like buildings), together with the associated pathways and processes potentially leading to negative effects on human health and the environment. The aquatic environment may contain thousands of compounds, many of them still unknown, that can pose a risk to ecosystems and human health. Due to the unquestionable importance of the aquatic environment, one of the main challenges in the field of exposure science is the comprehensive characterization and evaluation of complex environmental mixtures beyond the classical/priority contaminants to new emerging contaminants. The role of advanced analytical chemistry to identify and quantify potential chemical risks, that might cause adverse effects to the aquatic environment, is essential. In this paper, we present the strategies and tools that analytical chemistry has nowadays, focused on chromatography hyphenated to (high-resolution) mass spectrometry because of its relevance in this field. Key issues, such as the application of effect direct analysis to reduce the complexity of the sample, the investigation of the huge number of transformation/degradation products that may be present in the aquatic environment, the analysis of urban wastewater as a source of valuable information on our lifestyle and substances we consumed and/or are exposed to, or the monitoring of drinking water, are discussed in this article. The trends and perspectives for the next few years are also highlighted, when it is expected that new developments and tools will allow a better knowledge of chemical composition in the aquatic environment. This will help regulatory authorities to protect water bodies and to advance towards improved regulations that enable practical and efficient abatements for environmental and public health protection.

The classification of almonds (Prunus dulcis) by country and variety using UHPLC-HRMS-based untargeted metabolomics.

The aim of this study was to use an untargeted UHPLC-HRMS-based metabolomics approach allowing discrimination between almonds based on their origin and variety. Samples were homogenised, extracted with ACN:H2O (80:20) containing 0.1% HCOOH and injected in a UHPLC-QTOF instrument in both positive and negative ionisation modes. Principal component analysis (PCA) was performed to ensure the absence of outliers. Partial least squares - discriminant analysis (PLS-DA) was employed to create and validate the models for country (with five different compounds) and variety (with 20 features), showing more than 95% accuracy. Additional samples were injected and the model was evaluated with blind samples, with more than 95% of samples being correctly classified using both models. MS/MS experiments were carried out to tentatively elucidate the highlighted marker compounds (pyranosides, peptides or amino acids, among others). This study has shown the potential of high-resolution mass spectrometry to perform and validate classification models, also providing information concerning the identification of the unexpected biomarkers which showed the highest discriminant power.

Determination of selected endogenous anabolic androgenic steroids and ratios in urine by ultra high performance liquid chromatography tandem mass spectrometry and isotope pattern deconvolution.

An isotope dilution mass spectrometry (IDMS) method for the determination of selected endogenous anabolic androgenic steroids (EAAS) in urine by UHPLC-MS/MS has been developed using the isotope pattern deconvolution (IPD) mathematical tool. The method has been successfully validated for testosterone, epitestosterone, androsterone and etiocholanolone, employing their respective deuterated analogs using two certified reference materials (CRM). Accuracy was evaluated as recovery of the certified values and ranged from 75% to 108%. Precision was assessed in intraday (n=5) and interday (n=4) experiments, with RSDs below 5% and 10% respectively. The method was also found suitable for real urine samples, with limits of detection (LOD) and quantification (LOQ) below the normal urinary levels. The developed method meets the requirements established by the World Anti-Doping Agency for the selected steroids for Athlete Biological Passport (ABP) measurements, except in the case of androsterone, which is currently under study.

Updating the list of known opioids through identification and characterization of the new opioid derivative 3,4-dichloro-N-(2-(diethylamino)cyclohexyl)-N-methylbenzamide (U-49900).

New psychoactive substances have been rapidly growing in popularity in the drug market as non-illegal drugs. In the last few years, an increment has been reported on the use of synthetic alternatives to heroin, the synthetic opioids. Based on the information provided by the European Monitoring Centre for Drug and Drug Addiction, these synthetic opioids have been related to overdoses and deaths in Europe and North America. One of these opioids is the U-47700. A few months ago, U-47700 was scheduled in the U.S. and other countries, and other opioid derivatives have been appearing in order to replace it. One of these compounds is U-49900, an analog of U-47700. A white powder sample was obtained from an anonymous user in Spain. After an accurate characterization by gas chromatography-mass spectrometry, ultra-high performance liquid chromatography-high resolution mass spectrometry, nuclear magnetic resonance and single-crystal X-ray diffraction; and complemented by Fourier-transformed infrared spectroscopy, ultraviolet and circular dichroism spectrophotometry, the drug sample was unequivocally identified as U-49900. The information provided will be useful for the Early Warning System and forensic laboratories for future identifications of the U-49900, as well as in tentative identifications of other related opioids.

Evaluation of uncertainty sources in the determination of testosterone in urine by calibration-based and isotope dilution quantification using ultra high performance liquid chromatography tandem mass spectrometry.

Three quantification methodologies, namely calibration with internal standard (Cal-IS, non-weighted), weighted calibration with internal standard (wCal-IS) and isotope pattern deconvolution (IPD) have been used for the determination of testosterone in urine by LC-MS/MS. Uncertainty has been calculated and compared for the three methodologies through intra- and inter-laboratory reproducibility assays. IPD showed the best performance for the intra-laboratory reproducibility, with RSD and combined uncertainty values below 4% and 9% respectively. wCal-IS showed similar performance, while Cal-IS where not constant and clearly worse at the lowest concentration assayed (2ng/mL) reaching RSD values up to 16%. The inter-laboratory assay indicated similar results although wCal-IS RSD (20%) was higher than IPD (10%) and Cal-IS get worse with RSD higher than 40% for the lowest concentration level. Uncertainty budgets calculated for the three procedures revealed that intercept and slope were the most important factors contributing to uncertainty for Cal-IS. The main factors for wCal-IS and IPD were the volumes of sample and/or standard measured.

Proposal of 5-methoxy-N-methyl-N-isopropyltryptamine consumption biomarkers through identification of in vivo metabolites from mice.

New psychoactive substances (NPS) are a new breed of synthetically produced substances designed to mimic the effects of traditional illegal drugs. Synthetic cannabinoids and synthetic cathinones are the two most common groups, which try to mimic the effects of the natural compounds 9Δ-tetrahydrocannabinol and cathinone, respectively. Similarly, synthetic tryptamines are designer compounds which are based on the compounds psilocin, N,N-dimethyltryptamine and 5-methoxy-N,N-dimethyltryptamine found in some mushrooms. One of the most important tryptamine compounds found in seizures is 5-methoxy-N,N-diisopropyltryptamine, which has been placed as controlled substance in USA and some European countries. The control of this compound has promoted the rising of another tryptamine, the 5-methoxy-N-methyl-N-isopropyltryptamine, which at the time of writing this article has not been banned yet. So, it is undeniable that this new substance should be monitored. 5-methoxy-N-methyl-N-isopropyltryptamine has been reported by the Spanish Early Warning System and detected in our laboratory in two pill samples purchased in a local smart shop. This has promoted the need of stablishing consumption markers for this compound in consumers' urine. In the present work, the metabolism and pharmacokinetic of 5-methoxy-N-methyl-N-isopropyltryptamine has been studied by an in vivo approach, using adult male mice of the inbred strain C57BLJ/6. The use of ultra-high performance liquid chromatography coupled to high resolution mass spectrometry allowed the identification of four metabolites. After the pharmacokinetic study in serum and urine, the O-demethylated metabolite and the non-metabolised parent compound are proposed as consumption markers in hydrolysed urine. Data reported in this work will help hospitals and forensic laboratories to monitor the consumption and potential intoxication cases related to this tryptamine.

Behaviour of emerging contaminants in sewage sludge after anaerobic digestion.

Nowadays, there is an increasing concern over the presence of contaminants in the aquatic environment, where they can be introduced from wastewater after their incomplete removal in the treatment plants. In this work, degradation of selected emerging pollutants in the aqueous and solid phases of sewage sludge has been investigated after anaerobic digestion using two different digesters: mesophilic and thermophilic. Initially, sludge samples were screened by ultra-high-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC-QTOF MS) for identification of emerging contaminants in the samples. In a second step, a target quantitative method based on LC coupled to tandem MS was applied for selected pollutants identified in the previous screening. The behaviour of the compounds under anaerobic conditions was studied estimating the degradation efficiency and distribution of compounds between both sludge phases. Irbesartan and benzoylecgonine seemed to be notably degraded in both phases of the sludge. Venlafaxine showed a significant concentration decrease in the aqueous phase in parallel to an increase in the solid phase. The majority of the compounds showed an increase of their concentrations in both phases after the digestion. Concentrations in the solid phase were commonly higher than in the aqueous for most contaminants, indicating that they were preferentially adsorbed onto the solid particles.

Analytical methodologies based on LC-MS/MS for monitoring selected emerging compounds in liquid and solid phases of the sewage sludge.

In this work, two analytical methodologies based on liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) were developed for quantification of emerging pollutants identified in sewage sludge after a previous wide-scope screening. The target list included 13 emerging contaminants (EC): thiabendazole, acesulfame, fenofibric acid, valsartan, irbesartan, salicylic acid, diclofenac, carbamazepine, 4-aminoantipyrine (4-AA), 4-acetyl aminoantipyrine (4-AAA), 4-formyl aminoantipyrine (4-FAA), venlafaxine and benzoylecgonine. The aqueous and solid phases of the sewage sludge were analyzed making use of Solid-Phase Extraction (SPE) and UltraSonic Extraction (USE) for sample treatment, respectively. The methods were validated at three concentration levels: 0.2, 2 and 20 µg L(-1) for the aqueous phase, and 50, 500 and 2000 µg kg(-1) for the solid phase of the sludge. In general, the method was satisfactorily validated, showing good recoveries (70-120%) and precision (RSD < 20%). Regarding the limit of quantification (LOQ), it was below 0.1 µg L(-1) in the aqueous phase and below 50 µg kg(-1) in the solid phase for the majority of the analytes. The method applicability was tested by analysis of samples from a wider study on degradation of emerging pollutants in sewage sludge under anaerobic digestion. The key benefits of these methodologies are: • SPE and USE are appropriate sample procedures to extract selected emerging contaminants from the aqueous phase of the sewage sludge and the solid residue. • LC-MS/MS is highly suitable for determining emerging contaminants in both sludge phases. • Up to our knowledge, the main metabolites of dipyrone had not been studied before in sewage sludge.

Potential of atmospheric pressure chemical ionization source in gas chromatography tandem mass spectrometry for the screening of urinary exogenous androgenic anabolic steroids.

The atmospheric pressure chemical ionization (APCI) source for gas chromatography-mass spectrometry analysis has been evaluated for the screening of 16 exogenous androgenic anabolic steroids (AAS) in urine. The sample treatment is based on the strategy currently applied in doping control laboratories i.e. enzymatic hydrolysis, liquid-liquid extraction (LLE) and derivatization to form the trimethylsilyl ether-trimethylsilyl enol ether (TMS) derivatives. These TMS derivatives are then analyzed by gas chromatography tandem mass spectrometry using a triple quadrupole instrument (GC-QqQ MS/MS) under selected reaction monitoring (SRM) mode. The APCI promotes soft ionization with very little fragmentation resulting, in most cases, in abundant [M + H](+) or [M + H-2TMSOH](+) ions, which can be chosen as precursor ions for the SRM transitions, improving in this way the selectivity and sensitivity of the method. Specificity of the transitions is also of great relevance, as the presence of endogenous compounds can affect the measurements when using the most abundant ions. The method has been qualitatively validated by spiking six different urine samples at two concentration levels each. Precision was generally satisfactory with RSD values below 25 and 15% at the low and high concentration level, respectively. Most the limits of detection (LOD) were below 0.5 ng mL(-1). Validation results were compared with the commonly used method based on the electron ionization (EI) source. EI analysis was found to be slightly more repeatable whereas lower LODs were found for APCI. In addition, the applicability of the developed method has been tested in samples collected after the administration of 4-chloromethandienone. The highest sensitivity of the APCI method for this compound, allowed to increase the period in which its administration can be detected.

Validation of a qualitative screening method for pesticides in fruits and vegetables by gas chromatography quadrupole-time of flight mass spectrometry with atmospheric pressure chemical ionization.

A wide-scope screening method was developed for the detection of pesticides in fruit and vegetables. The method was based on gas chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer with an atmospheric pressure chemical ionization source (GC-(APCI)QTOF MS). A non-target acquisition was performed through two alternating scan events: one at low collision energy and another at a higher collision energy ramp (MS(E)). In this way, both protonated molecule and/or molecular ion together with fragment ions were obtained in a single run. Validation was performed according to SANCO/12571/2013 by analysing 20 samples (10 different commodities in duplicate), fortified with a test set of 132 pesticides at 0.01, 0.05 and 0.20mg kg(-1). For screening, the detection was based on one diagnostic ion (in most cases the protonated molecule). Overall, at the 0.01mg kg(-1) level, 89% of the 2620 fortifications made were detected. The screening detection limit for individual pesticides was 0.01mg kg(-1) for 77% of the pesticides investigated. The possibilities for identification according to the SANCO criteria, requiring two ions with a mass accuracy ≤±5ppm and an ion-ratio deviation ≤±30%, were investigated. At the 0.01mg kg(-1) level, identification was possible for 70% of the pesticides detected during screening. This increased to 87% and 93% at the 0.05 and 0.20mg kg(-1) level, respectively. Insufficient sensitivity for the second ion was the main reason for the inability to identify detected pesticides, followed by deviations in mass accuracy and ion ratios.

Mass spectrometric behavior of anabolic androgenic steroids using gas chromatography coupled to atmospheric pressure chemical ionization source. Part I: ionization.

The detection of anabolic androgenic steroids (AAS) is one of the most important topics in doping control analysis. Gas chromatography coupled to (tandem) mass spectrometry (GC-MS(/MS)) with electron ionization and liquid chromatography coupled to tandem mass spectrometry have been traditionally applied for this purpose. However, both approaches still have important limitations, and, therefore, detection of all AAS is currently afforded by the combination of these strategies. Alternative ionization techniques can minimize these drawbacks and help in the implementation of a single method for the detection of AAS. In the present work, a new atmospheric pressure chemical ionization (APCI) source commercialized for gas chromatography coupled to a quadrupole time-of-flight analyzer has been tested to evaluate the ionization of 60 model AAS. Underivatized and trimethylsylil (TMS)-derivatized compounds have been investigated. The use of GC-APCI-MS allowed for the ionization of all AAS assayed irrespective of their structure. The presence of water in the source as modifier promoted the formation of protonated molecules ([M+H](+)), becoming the base peak of the spectrum for the majority of studied compounds. Under these conditions, [M+H](+), [M+H-H2O](+) and [M+H-2·H2O](+) for underivatized AAS and [M+H](+), [M+H-TMSOH](+) and [M+H-2·TMSOH](+) for TMS-derivatized AAS were observed as main ions in the spectra. The formed ions preserve the intact steroid skeleton, and, therefore, they might be used as specific precursors in MS/MS-based methods. Additionally, a relationship between the relative abundance of these ions and the AAS structure has been established. This relationship might be useful in the structural elucidation of unknown metabolites.

Identification of new omeprazole metabolites in wastewaters and surface waters.

Omeprazole is one of the world-wide most consumed pharmaceuticals for treatment of gastric diseases. As opposed to other frequently used pharmaceuticals, omeprazole is scarcely detected in urban wastewaters and environmental waters. This was corroborated in a previous research, where parent omeprazole was not detected while four transformation products (TPs), mainly resulting from hydrolysis, were found in effluent wastewaters and surface waters. However, the low abundance of omeprazole TPs in the water samples together with the fact that omeprazole suffers an extensive metabolism, with a wide range of excretion rates (between 0.01 and 30%), suggests that human urinary metabolites should be investigated in the water environment. In this work, the results obtained in excretion tests after administration of a 40 mg omeprazole dose in three healthy volunteers are reported. Analysis by liquid chromatography coupled to hybrid quadrupole time-of-flight mass spectrometry (LC-QTOF MS) reported low concentrations of omeprazole in urine. Up to twenty-four omeprazole metabolites (OMs) were detected and tentatively elucidated. The most relevant OM was an omeprazole isomer, which obviously presented the same exact mass (m/z 346.1225), but also shared a major common fragment at m/z 198.0589. Subsequent analyses of surface water and effluent wastewater samples by both LC-QTOF MS and LC-MS/MS with triple quadrupole revealed that this metabolite (named as OM10) was the compound most frequently detected in water samples, followed by OM14a and OM14b. Up to our knowledge, OM10 had not been used before as urinary biomarker of omeprazole in waters. On the contrary, parent omeprazole was never detected in any of the water samples. After this research, it seems clear that monitoring the presence of omeprazole in the aquatic environment should be focused on the OMs suggested in this article instead of the parent compound.

Investigating the presence of omeprazole in waters by liquid chromatography coupled to low and high resolution mass spectrometry: degradation experiments.

Omeprazole is one of the most consumed pharmaceuticals around the world. However, this compound is scarcely detected in urban wastewater and surface water. The absence of this pharmaceutical in the aquatic ecosystem might be due to its degradation in wastewater treatment plants, as well as in receiving water. In this work, different laboratory-controlled degradation experiments have been carried out on surface water in order to elucidate generated omeprazole transformation products (TPs). Surface water spiked with omeprazole was subjected to hydrolysis, photo-degradation under both sunlight and ultraviolet radiation and chlorination. Analyses by liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QTOF MS) permitted identification of up to 17 omeprazole TPs. In a subsequent step, the TPs identified were sought in surface water and urban wastewater by LC-QTOF MS and by LC coupled to tandem mass spectrometry with triple quadrupole. The parent omeprazole was not detected in any of the samples, but four TPs were found in several water samples. The most frequently detected compound was OTP 5 (omeprazole sulfide), which might be a reasonable candidate to be included in monitoring programs rather than the parent omeprazole.

Direct liquid chromatography-tandem mass spectrometry determination of underivatized glyphosate in rice, maize and soybean.

The residue determination of the widely used herbicide glyphosate (GLY) is highly problematic due to its amphoteric character, low mass and lack of chemical groups that might facilitate its detection. Most methods developed up to now have employed pre-column or post-column derivatization to form fluorescent derivatives and/or to reduce the polar character of the analyte facilitating its chromatographic retention. The aim of this work is to evaluate the feasibility of performing the direct LC-MS/MS determination of GLY residues in vegetables. After testing several Hydrophilic Interaction Liquid Chromatography (HILIC) columns, Obelisc N was selected due to its better chromatographic retention. LC-MS/MS determination has been performed in negative ionization mode, monitoring up to four transitions to give high reliability to the identification/confirmation process. This approach has been evaluated for the determination of GLY residues in rice, maize and soybean samples, and the method validated at different concentrations in compliance with the maximum residue limits established in the current legislation. After sample extraction with water, a combination of extract dilution, partition with dichloromethane, and solid phase extraction (SPE) using Oasis HLB cartridges (depending on the sample matrix under analysis) was applied. Quantification was made by using isotope-labeled GLY as internal standard and calibration in solvent. The methodology developed allows the rapid determination of GLY residues avoiding the derivatization step typically applied for this herbicide. The most critical issue is the robustness of the Obelisc N column, which was found to suffer rapid degradation with time. Extreme care and continuous testing of retention times and peak shapes is required for a reliable determination.

Improvements in the analytical methodology for the residue determination of the herbicide glyphosate in soils by liquid chromatography coupled to mass spectrometry.

The determination of glyphosate (GLY) in soils is of great interest due to the widespread use of this herbicide and the need of assessing its impact on the soil/water environment. However, its residue determination is very problematic especially in soils with high organic matter content, where strong interferences are normally observed, and because of the particular physico-chemical characteristics of this polar/ionic herbicide. In the present work, we have improved previous LC-MS/MS analytical methodology reported for GLY and its main metabolite AMPA in order to be applied to "difficult" soils, like those commonly found in South-America, where this herbicide is extensively used in large areas devoted to soya or maize, among other crops. The method is based on derivatization with FMOC followed by LC-MS/MS analysis, using triple quadrupole. After extraction with potassium hydroxide, a combination of extract dilution, adjustment to appropriate pH, and solid phase extraction (SPE) clean-up was applied to minimize the strong interferences observed. Despite the clean-up performed, the use of isotope labelled glyphosate as internal standard (ILIS) was necessary for the correction of matrix effects and to compensate for any error occurring during sample processing. The analytical methodology was satisfactorily validated in four soils from Colombia and Argentina fortified at 0.5 and 5mg/kg. In contrast to most LC-MS/MS methods, where the acquisition of two transitions is recommended, monitoring all available transitions was required for confirmation of positive samples, as some of them were interfered by unknown soil components. This was observed not only for GLY and AMPA but also for the ILIS. Analysis by QTOF MS was useful to confirm the presence of interferent compounds that shared the same nominal mass of analytes as well as some of their main product ions. Therefore, the selection of specific transitions was crucial to avoid interferences. The methodology developed was applied to the analysis of 26 soils from different areas of Colombia and Argentina, and the method robustness was demonstrated by analysis of quality control samples along 4 months.

Qualitative validation of a liquid chromatography-quadrupole-time of flight mass spectrometry screening method for organic pollutants in waters.

A multiclass wide-scope screening of organic contaminants in natural and waste water has been developed and validated for qualitative purposes, i.e. detection and reliable identification of compounds detected in samples at a certain level of concentration. The screening is based on the use of liquid chromatography coupled to quadrupole-time of flight mass spectrometry (LC-QTOF MS) and has been applied to water samples of different origin and matrix composition (surface water, ground water and effluent urban wastewater). Water samples were spiked with a standard mixture of around 150 organic contaminants from different chemical families (including a number of relevant metabolites/transformation products (TPs), at 0.1 and 1 µg/L concentration levels. After solid-phase extraction with Oasis HLB cartridges, sample extracts were analyzed by LC-QTOF MS and the accurate-mass full-spectrum data were processed for qualitative analysis. The presence of at least two ions (typically the (de)protonated molecule and one fragment ion) accurate-mass measured was used for the reliable identification. The screening detection limit (SDL) and the limit of identification (LOI) were established as the main parameters of the screening method. Nearly all compounds could be detected at the lowest concentration tested, but identification was problematic for some compounds at 0.1 µg/L level, especially in wastewater samples. The screening procedure was finally applied to different water samples using a home-made database of around 1100 organic contaminants. It allowed the detection and identification of several antibiotics, anti-inflammatory/analgesics drugs and lipid regulators. Cocaine and its metabolite benzoylecgonine were also frequently detected. In addition, triazine herbicides and their TPs, and fungicides like thiabendazol, carbendazim or imazalil, were also identified in some of the samples.

Combined use of liquid chromatography triple quadrupole mass spectrometry and liquid chromatography quadrupole time-of-flight mass spectrometry in systematic screening of pesticides and other contaminants in water samples.

As a suitable way for routine screening of pesticides and control of other organic contaminants in water, the combination of liquid chromatography triple quadrupole tandem mass spectrometry (LC-QqQ-MS/MS) and liquid chromatography-hybrid quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) has been applied to the analysis of 63 surface and waste water samples after conventional solid-phase extraction (SPE). The extracts were screened for 43 pesticides or degradation products by LC-QqQ-MS/MS achieving limits of detection (LOD) ranged from 0.04 to 2 ng L(-1). Of the 43 selected pesticides, 33 were detected in water samples. The ESI-QTOF MS instrument was run using two simultaneous acquisition functions with low and high collision energy (MS(E) approach) and acquiring the full mass spectra. A home-made database containing more than 1100 organic pollutants was used for substance identification. Around 250 of these compounds were available at the laboratory as reference standards. Five pesticides and 3 of their degradation products, different to those selected in the QqQ method, were detected by QqTOF-MS. Thirteen pharmaceuticals and two drugs of abuse were also identified in the samples. In practice, the sample preparation proved to be suitable for both techniques and for a wide variety of substances with different polarity. Mutual confirmation and evidence of co-occurrence of several other organic contaminants were the main advantages of the combination of both techniques.

Advantages of atmospheric pressure chemical ionization in gas chromatography tandem mass spectrometry: pyrethroid insecticides as a case study.

Gas chromatography coupled to mass spectrometry (GC/MS) has been extensively applied for determination of volatile, nonpolar, compounds in many applied fields like food safety, environment, or toxicology. The wide majority of methods reported use electron ionization (EI), which may result in extensive fragmentation of analytes compromising selectivity and sensitivity. This might also complicate the application of tandem MS due to lack of specific/abundant precursor ions. Pyrethroids are examples of compounds with this behavior. In this work, the potential of atmospheric pressure chemical ionization (APCI), a softer form of ionization, combined with GC and a triple quadrupole mass analyzer was investigated, taking pyrethroids as a case study and their determination in fruit and vegetables as example application. Ionization and fragmentation behavior of eight pyrethroids (bifenthrin, cyfluthrin, cypermethrin, permethrin, λ-cyhalothrin, fluvalinate, fenvalerate, and deltamethrin) by APCI were studied. The formation of a highly abundant (quasi) molecular ion was the main goal because of the enhanced selectivity when used as precursor ion in tandem MS. The addition of water as a modifier was tested to promote the generation of protonated molecules, resulting in notable improvement of sensitivity and selectivity for most compounds. The excellent detectability (low detection limits (LODs) <20 fg achieved) when using APCI combined with state-of-the-art tandem MS was demonstrated for real samples. Additionally, matrix effects were evaluated in terms of signal enhancement/suppression. Depending on the matrix, different degrees of suppression were observed, on average reducing the signal in matrix to 55% of that in solvent. The results presented in this paper demonstrate the potential of APCI as new source for GC/MS that could be applied to other analytical problems apart from those illustrated in this work.

Importance of MS selectivity and chromatographic separation in LC-MS/MS-based methods when investigating pharmaceutical metabolites in water. Dipyrone as a case of study.

Pharmaceuticals are emerging contaminants of increasing concern because of their presence in the aquatic environment and potential to reach drinking-water sources. After human and/or veterinary consumption, pharmaceuticals can be excreted in unchanged form, as the parent compound, and/or as free or conjugated metabolites. Determination of most pharmaceuticals and metabolites in the environment is commonly made by liquid chromatography (LC) coupled to mass spectrometry (MS). LC coupled to tandem MS is the technique of choice nowadays in this field. The acquisition of two selected reaction monitoring (SRM) transitions together with the retention time is the most widely accepted criterion for a safe quantification and confirmation assay. However, scarce attention is normally paid to the selectivity of the selected transitions as well as to the chromatographic separation. In this work, the importance of full spectrum acquisition high-resolution MS data using a hybrid quadrupole time-of-flight analyser and/or a suitable chromatographic separation (to reduce the possibility of co-eluting interferences) is highlighted when investigating pharmaceutical metabolites that share common fragment ions. For this purpose, the analytical challenge associated to the determination of metabolites of the widely used analgesic dipyrone (also known as metamizol) in urban wastewater is discussed. Examples are given on the possibilities of reporting false positives of dypirone metabolites by LC-MS/MS under SRM mode due to a wrong assignment of identity of the compounds detected.