Investigation of aflatoxin and ochratoxin A contamination of seized cannabis and cannabis resin samples.

Recreational cannabis is being legalized in more and more countries, and methods for the determination of contaminants, thereunder mycotoxins, start to emerge in scientific literature. On the other hand, cannabis continues being available on the illegal market without any quality control at all. Today, no information about mycotoxin contamination of illegal cannabis is available in literature. Therefore, in order to increase knowledge about mycotoxin contamination of cannabis, aflatoxins (AF) and ochratoxin A (OTA) were analyzed in 142 samples of illegal cannabis seized on the local market using a method based on HPLC-FLD detection, after clean up with immuno-affinity cartridges. AF were derivatized prior to detection with a Kobra cell. No AF contamination (LOD = 0.04 µg/kg) was detected in any of the samples analyzed. OTA however was detected in about one-third of the samples with an average concentration of 4.30 µg/kg (range from 1.02 to 16.21 µg/kg). No significant difference was observed between resin and herbal samples. Overall, the concentrations remain low and do not suggest an issue to human health if the cannabis consumption remains moderate.

Epimerization of ergot alkaloids in feed.

Chronic intake of cereals contaminated with ergot alkaloids can cause ergotism and result in the loss of toes and fingers or even death. Today, due to common risk management practices, ergotism is rare as a human disease but remains a problem in livestock husbandry. Each alkaloid coexists under two forms (R and S), though only the R-form presents toxic effects. The epimerization occurs spontaneously but the mechanisms remain globally unknown. Therefore, different processing methods were evaluated for their respective influences on the epimerization. The results suggest that ergotamine and ergosine are very stable ergot alkaloids, and neither their concentrations, nor their respective R/S ratios, are significantly influenced by heating, protic solvents or UV light. In contrast, for ergocristine, ergokryptine, ergocornine and ergometrine, heating can decrease the concentrations of these alkaloids and heat, protic solvents and UV light influence the R/S ratio towards the S-form, though the respective influence on the epimerization of these compounds is variable. In addition, the total concentration of all ergot alkaloids is reduced through heating. However, all these effects are not strong enough to change the composition of ergot alkaloids in feed substantially and to transform toxic feed into non-toxic feed.

Development of a UHPLC-FLD method for the analysis of ergot alkaloids and application to different types of cereals from Luxembourg.

Ergot alkaloids are toxins produced by some species of fungi in the genus Claviceps, that may infect rye and triticale and, in a minor degree, other types of cereals. In this study, a new UHPLC-FLD method for the quantification of the six major ergot alkaloids as well as their corresponding epimers was developed. The sample preparation was done by a solid-liquid extraction with acetonitrile and clean-up via freeze-out. The method was fully validated and then applied to 39 samples (wheat, rye, triticale, and barley) harvested in Luxembourg in 2016. Samples were sieved (1.9 × 20 mm) prior to analysis in order to remove sclerotia, hosting the alkaloids. However, 23 samples still contained at least one ergot alkaloid > LOQ and concentrations of the sum of the 6 ergot alkaloids ranged from 0.3 to 2530.1 µg/kg. Interestingly, the highest concentrations were measured in wheat and not in rye or triticale, suggesting that all kinds of cereals should be included in monitoring programs. The outcome of this study allowed giving a first overview of ergot alkaloid concentrations in cereals harvested in Luxembourg, and the measured concentrations were in similar ranges than in other parts of the world (e.g., Canada, France, Germany).

Utilization of long duration high-volume sampling coupled to SPME-GC-MS/MS for the assessment of airborne pesticides variability in an urban area (Strasbourg, France) during agricultural application.

Atmospheric samples have been collected between 14 March and 12 September 2012 on a 2-week basis (15 days of sampling and exchange of traps each 7 days) in Strasbourg (east of France) for the analysis of 43 pesticides. Samples (particle and gas phases) were separately extracted using Accelerated Solvent Extraction (ASE) and pre-concentrated by Solid Phase Micro-Extraction (SPME) before analysis by gas chromatography coupled to tandem mass spectrometry (GC-MS/MS). Four SPME consecutive injections at distinct temperatures were made in order to increase the sensitivity of detection for the all monitored pesticides. Currently used detected pesticides can be grouped in four classes; those used in maize crops (acetochlor, benoxacor, dicamba, s-metolachlor, pendimethalin, and bromoxynil), in cereal crops (benoxacor, chlorothalonil, fenpropimorph, and propiconazole), in vineyards (tebuconazole), and as herbicides for orchards, meadows of green spaces (2,4-MCPA, trichlopyr). This is in accordance with the diversity of crops found in the Alsace region and trends observed are in accordance with the period of application of these pesticides. Variations observed permit also to demonstrate that the long time sampling duration used in this study is efficient to visualize temporal variations of airborne pesticides concentrations. Then, long time high-volume sampling could be a simple method permitting atmospheric survey of atmospheric contamination without any long analysis time and consequently low cost.

Monitoring of polycyclic aromatic hydrocarbons (PAHs) in the atmosphere of southern Luxembourg using XAD-2 resin-based passive samplers.

XAD-2 resin-based passive samplers (PAS) with dimensions adapted to 100 mL accelerated solvent extraction cells were used to study the temporal and spatial variations of 17 PAHs on five sites in the atmosphere of southern Luxembourg. This new design allowed extracting the PAS without emptying the resin from the shelter. PAH analyses were done with gas chromatography-tandem mass spectrometry. PAS were deployed for 1 year with varying sampling periodicities, and 16 PAHs were detected with concentrations ranging from 1 ng/PAS for chrysene to 9,727 ng/PAS for naphthalene. The PAS were found adapted to the monitoring of temporal and spatial variations for lightweight PAHs (up to four aromatic rings) though not for heavy PAHs with five aromatic rings or more, as these compounds are preferably in the particle phase of the atmosphere and the amount of these PAHs trapped on the PAS will be too low.

Determination of polycyclic aromatic hydrocarbons in smoked and non-smoked black teas and tea infusions.

This study describes the occurrence of polycyclic aromatic hydrocarbons (PAHs) in smoked tea and tea infusions, via the monitoring of benzo(a)anthracene, chrysene, benzo(b)fluoranthene and benzo(a)pyrene (PAH4) that have been chosen as indicators for the occurrence of PAHs in food by the European Food Safety Agency. The concentrations ranged from 1.2 µg/kg for benzo(b)fluoranthene to 125.0 µg/kg for benzo(a)anthracene in smoked tea leaves, and from 0.6 µg/L for benzo(a)anthracene to 1.2 µg/L for benzo(b)fluoranthene in smoked tea infusions. Benzo(a)pyrene was never detected in infusions. The concentrations in non-smoked tea leaves ranged from 0.6 µg/kg for benzo(a)anthracene to 10.8 µg/kg for benzo(b)fluoranthene. It was shown that the concentrations of benzo(a)anthracene and chrysene were higher in smoked tea than in non-smoked tea while no difference was observed for benzo(b)fluoranthene and benzo(a)pyrene. The concentrations of PAHs in tea infusions are low compared to other foodstuffs, but the migration rates from leaves into water are high (82-123%).

Low-level detections of Sudan I, II, III and IV in spices and Chili-containing foodstuffs using UPLC-ESI-MS/MS.

Sudan dyes are red, synthetic azo dyes that are not allowed in foodstuffs in the European Union (Council Directive 94/36/EC). However, subppm levels of Sudan dye in spices are regularly reported, and it is assumed that these appearances are due to cross-contamination. In this paper, we present a newly developed fast and sensitive method for the quantification of Sudan I, II, III, and IV, using liquid-liquid extraction and UPLC-MS/MS analysis, and giving quantification limits ranging from 2.5 to 200 µg/kg. The method was applied to 21 samples, and 17 of them contained Sudan dye at low concentrations (3.3-8 709 µg/kg). Interestingly, it was observed that the distribution of Sudan dye in the sample is not homogeneous, which may lead to false negatives or to overestimations of the concentration, and that the pretreatment (blending or not) of the sample seriously influences the final result of the analysis.

Application of XAD-2 resin-based passive samplers and SPME-GC-MS/MS analysis for the monitoring of spatial and temporal variations of atmospheric pesticides in Luxembourg.

Passive air sampling has been shown to be a very interesting alternative to high-volume sampling by overcoming its disadvantages (size, weight, expensiveness). However, to date, only limited data is available about passive air sampling of current-use pesticides. In order to test if passive samplers allow monitoring of spatial and temporal variations of atmospheric pesticide concentrations, five XAD-2-resin based passive air samplers were deployed at five locations in Luxembourg. Samplers were analyzed using accelerated solvent extraction coupled to solid-phase microextraction and gas chromatography with tandem mass spectrometry. Collected data was used to study the spatial and temporal variations of the concentrations of the compounds. Twenty two pesticides were detected between March and October, while no pesticides were detected from November to February. Highest concentrations were measured on the rural sites, suggesting that the used XAD-2 resin-based passive samplers allow the simultaneous monitoring of multiple current-use pesticides and identifying spatial and temporal variations.

Multi-class pesticide analysis in human hair by gas chromatography tandem (triple quadrupole) mass spectrometry with solid phase microextraction and liquid injection.

A method for the simultaneous detection and quantification of 22 pesticides from different chemical classes was developed using solid-phase microextraction (SPME) and gas chromatography tandem (triple quadrupole) mass spectrometry. Pesticides were extracted from 50mg of pulverized hair with acetonitrile. The extract was submitted to two successive steps of direct immersion-SPME at 30°C and 90°C or to a liquid injection without SPME in order to obtain optimized conditions for each of the 22 analytes investigated. Validation parameters were significantly influenced by both the injection mode (SPME vs liquid injection) and the temperature of SPME. Limits of quantification ranged from 0.05 pg mg(-1) for trifluralin to 10 pg mg(-1) for pentachlorophenol. The application of the validated method to the analysis of samples collected from non-occupationally exposed volunteers demonstrated the presence of pesticides in all the samples tested. Altogether, 13 different analytes were detected at concentration above the limit of quantification.

Simultaneous determination of nicotine and PAH metabolites in human hair specimen: a potential methodology to assess tobacco smoke contribution in PAH exposure.

The present methodology was developed to simultaneously assess chronic exposure to PAHs and to tobacco from the analysis of one hair specimen per examined individual. The method is a two step extraction of twelve mono-hydroxy-PAHs and of nicotine, and their separate analysis by optimized methods using gas chromatography-negative chemical ionization-mass spectrometry. After method validation and assessment of the hair decontamination procedure, 105 hair specimens from smokers and non-smokers were analyzed. All the hair samples tested positive for nicotine. Median concentration was 10.7ng/mg for smokers and 0.5ng/mg for non-smokers. 70% of the samples tested positive for OH-PAHs. The most common one was 2-naphthol (61%) and its concentration was significantly higher in smokers than in non-smokers (median: 111 vs 70pmol/g, p=0.006). 2-OH-benzo(c)phenanthrene and 6-OH-chrysene were only detected once in a non-smoker's hair. The concentration of the sum of all PAH-metabolites ranged from 24 to 67190pmol/g (median: 118pmol/g). Only six samples tested positive for more than two different metabolites. The simultaneous detection of nicotine and OH-PAHs in hair is possible and provides reliable results. This represents a useful tool for the accurate biomonitoring of chronic exposure to PAH and correct identification of the sources of exposure.

Determination of farm workers' exposure to pesticides by hair analysis.

In the present work, a highly sensitive method based on solid phase microextraction and gas chromatography tandem (triple quadrupole) mass spectrometry was used to test hair samples for 50 pesticides including 39 molecules from different chemical families currently used in agriculture and 11 organochlorines. The population investigated was composed of 18 farm workers who provided hair samples repeatedly collected during the entire treatment period (from March to November 2009). Among the 62 hair samples that were collected, 33 different target molecules were detected. The most frequently detected agricultural pesticides were Diflufenican and Pyrimethanil, two herbicides which were detected in 13 subjects. The concentration in volunteers' hair matched with agricultural activity and the highest concentration was observed for Cyprodinil (1161pg/mg), an anilinopyrimidine used as a fungicide. For organochlorines, p,p'-DDE and γ-HCH were the most frequently detected molecules as they were present in at least one of the hair samples provided by each of the 18 volunteers. The highest concentrations detected for these chemicals reached 21.0pg/mg for p,p'-DDE and 23.5pg/mg for γ-HCH, but the highest concentration of organochlorine was observed for ß-endosulfan (105pg/mg). The results suggest that farm workers have a weak, though constant exposure to organochlorine pesticides, especially to p,p'-DDE and γ-HCH, while exposure to currently used pesticides is strongly associated with occupation. Observations also suggest that spraying work would not necessarily be the only source of exposure to agricultural pesticides and that worker not directly involved in spraying can also be submitted to significant level of exposure.

Temporal variations of concentrations of currently used pesticides in the atmosphere of Strasbourg, France.

Atmospheric samples have been collected in Strasbourg between April 18 and May 29, 2007 and were analyzed for 71 current-use pesticides, of which 38 were detected. Average concentrations ranged from 0.09 ng m(-3) for Fenarimol to 110.42 ng m(-3) for Dimethachlor, which was slightly higher than the concentrations reported from other, comparable agricultural regions. Significant temporal variations were observed for 30 pesticides, and for most of them it could be shown that these were linked to time, temperature or atmospheric pressure. In several cases this helped to identify pesticide application just before or at the beginning of the sampling period, or ongoing treatment. Humidity, in contrast to previous reports, could not be linked to these variations. For the other 8 pesticides, only very little temporal variations were observed. Generally, these concentrations were low (less than 1 ng m(-3)), and it was assumed that they are not in use in Alsace at present.

Analysis of phenols and nitrophenols in rainwater collected simultaneously on an urban and rural site in east of France.

The behaviour of phenols and nitrophenols in the atmosphere is of concern for environmental research as these are known to be toxic to humans and wildlife. To increase the knowledge about these compounds, the concentrations of 20 phenols and nitrophenols in rainwater were studied in an urban (Strasbourg) and a rural (Erstein) site in Eastern France. The wet rain samples were collected on a weekly basis between January 2002 and July 2003, and analyzed for phenols and nitrophenols using SPME and gas chromatography coupled to mass spectrometry. The compounds were derivatized with MDBSTFA directly in the injection port of the gas chromatograph. Total concentrations ranged from 0 to 1383 microg L(-1) in Strasbourg and from 0 to 1215 microg L(-1) in Erstein. These concentrations are in the same range than those measured in other studies, but considerably higher than the concentrations measured for pesticides (about 10 times) and polycyclic aromatic hydrocarbons (about 1000 times) on the same sites and at the same period. Phenols, except for o-cresol, were generally less concentrated than nitrophenols. o-Cresol was one of the major compounds, probably due to its role as raw material in pesticide production. The concentrations of phenols and nitrophenols were found to vary with time. No seasonal trend was observed, but significant correlations with rainfall amount were identified.

Determination of hydroxylated metabolites of polycyclic aromatic hydrocarbons in human hair by gas chromatography-negative chemical ionization mass spectrometry.

The study describes the determination of mono-hydroxylated polycyclic aromatic hydrocarbons (OH-PAHs), metabolites of PAHs, in human hair. Twelve selected OH-PAHs from two to four rings, generally determined in urine analysis, were investigated as markers of human exposure to PAHs. Following hydrolysis of hair specimens of 50-300 mg with 1M NaOH, OH-PAHs were extracted using dichloromethane and submitted to an optimized derivatization with (2S,4R)-N-heptafluorobutyryl-4-heptafluorobutoyloxy-prolyl chloride. Compounds were then analyzed using gas chromatography-negative chemical ionization mass spectrometry (GC-NCIMS). The average inter-day and intra-day variability was 12% and 17%, respectively. The average recovery was 52% and the limits of detection and quantification ranged from 20 and 66 pmol/g for 1-OH-phenanthrene (i.e., 3.9 and 12.8 pg/mg) to 311 and 1030 pmol/g for 2-OH-benzo(c)phenanthrene (i.e., 75.9 and 251 pg/mg). The influence of hair washing with water as decontamination step, and enzymatic treatment (beta-glucuronidase) to hydrolyze conjugated derivatives were also tested. The application of the developed method to the analysis of 30 hair specimens (17 from non-smoker and 13 from smoker volunteers) demonstrated inter-individual qualitative and quantitative variations. According to the easiness of hair sampling and based on the extended detection windows provided by hair analysis, this method is proposed as a new promising tool for the assessment of human chronic exposure to PAHs.

Comparison of MTBSTFA and BSTFA in derivatization reactions of polar compounds prior to GC/MS analysis.

In this study, MTBSTFA and BSTFA, which are among the preferred derivatization reagents for silylation were both tested on derivatization of six different groups of polar chemicals to get information about usefulness in terms of sensitivity and specificity of both reagents. Tested compound groups were nitrophenols and methoxyphenols, sterols and sugars, dicarboxylic acids and hydroxylated polycyclic aromatic hydrocarbons. It was found that MTBSTFA-derivates produce characteristic fragmentation patterns presenting mainly the fragments [M](+), [M-57](+) and [M-131](+), of which [M-57](+) is generally dominant on the mass spectrogram. BSTFA-derivates mainly show the fragments [M](+), [M-15](+) and [M-89](+) whereof the molecular ion [M](+) is generally dominant. It was also found that steric hindrance and molecular mass play a very important role in the choice of the best suited derivatization reagent: compounds with sterically hindered sites derivatized with MTBSTFA produce very small analytical responses or no signal at all, and compounds with high molecular mass produce no characteristic fragmentation pattern when derivatization is performed with BSTFA. It was also found that MTBSTFA-derivatization facilitates separation of isomer analytes, suggesting its choice in combination to semi-polar columns, whilst BSTFA seems better for sterically hindered compounds. Findings were confirmed with applications of both reagents to biological and environmental matrices (urine and atmospheric aerosols).

Quantitative determination of ethyl glucuronide in sweat.

In this study, ethyl glucuronide (EtG), a specific metabolite of ethanol, was for the first time detected in sweat after alcohol consumption by human volunteers. Sweat was collected using a sweat patch (PharmChek). After collection, chemicals accumulated on the patch were extracted with water and extracts were purified by solid phase extraction. EtG was determined by gas chromatography with mass spectrometric detection in negative chemical ionization mode. In parallel, the amount of sodium deposited on the patch was determined by capillary electrophoresis and used as a correction factor to calculate the volume of sweat accumulated on the patch and, hence, the concentration of EtG in sweat. The EtG sweat concentration observed ranged from 1.7 to 103.0 microg/L for alcohol consumption from 38.0 to 154.6 g equivalent pure ethanol. No EtG was detected in subjects who did not consume alcohol. Our results demonstrate that after ethanol consumption, EtG is detectable in sweat collected using a sweat patch. The simultaneous determination of sodium allows the estimation of the volume of sweat accumulated on the patch and to calculate the concentration of EtG in sweat. This represents the first quantitative determination of a xenobiotic in sweat collected using a sweat patch. This study suggests that EtG determination in sweat could represent an interesting alternative to urine or serum analysis for the control of abstinence of patients included in treatment programs.

Determination of fentanyl in sweat and hair of a patient using transdermal patches.

Fentanyl is a synthetic opioid used to treat intense chronic pain. In this study, the authors report detection and quantification of fentanyl in sweat and hair of a patient receiving fentanyl (25 microg/h) via a transdermal therapeutic system (TTS) for 22 days. Sweat was collected using sweat patches every night on days 13-21 of the therapy, and hair was collected 12 weeks after the end of the treatment. Detection and quantification was performed with liquid chromatography-tandem mass spectrometry using electron spray ionization in selected reaction monitoring mode. Alfentanyl was used as internal standard for quantification in hair and in sweat. Sodium ions have been used as endogenous internal reference for determination of volume of sweat excreted on each patch. Results show presence of fentanyl in both matrices. Fentanyl concentrations in sweat varied from 0.17 to 1.02 ng/microL, and time-resolved segmented hair analysis showed a maximum fentanyl concentration of 0.48 ng/mg of hair during the period of the therapy.

Determination of the volume of sweat accumulated in a sweat-patch using sodium and potassium as internal reference.

In the present work, we assessed the suitability of sodium and potassium physiologically present in sweat, as internal reference allowing to re-calculate the corresponding volume of sweat collected on a PharmChek Patch. A method using capillary electrophoresis with indirect ultra-violet detection was developed for the determination of sodium and potassium in sweat. The concentrations determined in specimens collected from 12 females and 10 males, using a home-made system composed of polypropylene copolymer bag, were 1039+/-89 mg/L and 711+/-45 mg/L for sodium, and 489+/-293 mg/L and 474+/-196 mg/L for potassium, respectively. In parallel, for seven females and eight males, the comparison of the volume of sweat collected in the same way to the re-calculated volume of sweat accumulated in a patch using sodium as internal standard, gave an average agreement of 98.4+/-15.0%. Results demonstrated the usefulness of sodium as internal standard to determine the volume of sweat accumulated in a patch, and confirm the suitability of PharmChek patch for the collection and determination of cations in sweat.

Solid-phase microextraction and gas chromatography-mass spectrometry for analysis of phenols and nitrophenols in rainwater, as their t-butyldimethylsilyl derivatives.

Solid-phase microextraction (SPME) coupled with gas chromatography-mass spectrometry has been used for analysis of four phenols and sixteen nitrophenols in rainwater samples. Analytes were extracted from the water in the immersion mode and derivatised for 5 min during direct desorption in the GC injector. Before desorption, 2 microL N-(t-butyldimethylsilyl)-N-methyltrifluoroacetamide (MDBSTFA) was introduced into the injector, which was maintained at 280 degrees C. Different conditions affecting extraction efficiency were studied, including temperature, type of microextraction fibre, and effect of pH and ionic strength. Five different fibre coatings were tested: 85-mum polyacrylate (PA), 100-microm polydimethylsiloxane (PDMS), 65-mum Carbowax-divinylbenzene (CW-DVB), 75-microm Carboxen-polydimethylsiloxane (CAR-PDMS), and 65-microm polydimethylsiloxane-divinylbenzene (PDMS-DVB). The best conditions were use of PA fibres for 40 min at ambient temperature (75 g NaCl per 100 mL, pH 3.0). MDBSTFA was used as derivatising agent because it enables analysis of phenols derivatives with high confidence in identification, because in electron-impact mode TBDMS-phenol derivatives produce the specific M-57 ion. Quantification was achieved by using 4-nitrophenol-d4, at 1 mg L(-1), as internal standard. Linearity was good, with correlation coefficients in the range 0.9888 (o-cresol) to 0.9987 (dinitro-o-cresol, DNOC). Detection limits varied between 0.208 and 99.3 microg L(-1) and quantification limits between 0.693 and 331 microg L(-1). Uncertainties varied between 8.7% (phenol) and 17.9% (4-methyl-2-nitrophenol). The method was successfully applied to the analysis of rainwater collected at urban and rural sites in Alsace (East of France). Because of derivatisation in the injector and the associated high temperature, the lifetime of the fibre is severely reduced.