Development of a Methodology for Determination of Dioxins and Dioxin-like PCBs in Meconium by Gas Chromatography Coupled to High-Resolution Mass Spectrometry (GC-HRMS).

An analytical strategy was applied to investigate polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and dioxin-like polychlorinated biphenyls (dl-PCBs) in newborn meconium samples. The methodology includes extraction by selective pressurized liquid extraction (SPLE), followed by a clean-up multicolumn step. The samples were injected by gas chromatography coupled to a high-resolution mass spectrometer (GC-HRMS). The surrogate recoveries ranged from 68% to 95%, and the average of the limit of quantification (LOQ) ranged from 0.03 to 0.08 pg g-1 wet weight (ww) for PCDD/Fs and 0.2 to 0.88 pg g-1 ww for dl-PCBs. The strategy was applied to 10 samples collected in Valencia (Spain) in 2022. In total, 18 out of 29 analysed congeners were detected in at least one sample, whereas 6 of them were detected in all the samples (OCDD, PCB-123, PCB-118, PCB-105, PCB-167, and PCB-156). The levels for the sum of the 17 congeners of PCDD/Fs and 12 congeners of dl-PCBs in the upper-bound (UB), expressed as picograms of toxic equivalency quantity (TEQ) per gram of ww, ranged from 0.19 to 0.31 pg TEQ g-1 ww.

Exposure assessment of Spanish lactating mothers to acrylamide via human biomonitoring.

Acrylamide (AA) is an organic compound classified as "Probably carcinogenic to humans" (Group 2 A) that can be found principally in processed carbohydrate-rich foods and tobacco smoke. In humans, after exposure, AA is rapidly metabolized and excreted in urine, predominantly as N-acetyl-S-(2-carbamoylethyl)-l-cysteine (AAMA), N-acetyl-S-(2-carbamoyl-2-hydroxyethyl)-l-cysteine (GAMA3) and N-Acetyl-3-[(3-amino-3-oxopropyl)sulfinyl]-L-alanine (AAMA-Sul), which can be used as short-term biomarkers of exposure to AA. In this study, the presence of AA metabolites in urine samples of lactating mothers living in Spain (n = 114) was analyzed by "dilute and shoot" and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). All urinary metabolites were detected in 100% of the analyzed samples, with geometric means of 70, 33 and 15 ng ml-1, for AAMA, AAMA-Sul and GAMA3, respectively. The consumption of coffee, bread and precooked food products were found to be significant predictors of internal exposure to AA. An estimated daily intake (EDI) of AA based on its urinary metabolites was calculated, obtaining mean values between 1.2 and 1.9 µg AA·kg bw-1·day-1 in the target population. The risk assessment was evaluated using both reverse and forward dosimetry, showing an average margin of exposure (MOE) of 349 and a hazard quotient (HQ) of 5.5. Therefore, AA exposure should be considered a medium priority for risk assessment follow-up in the Spanish population, since a health concern with respect to non-neoplastic toxicity could not be discarded.

Exposure to non-persistent pesticides in the Spanish population using biomonitoring: A review.

Although Spain does not have a regular national human biomonitoring program yet, different research groups are active in evaluating the exposure of children and adults to chemicals. In the last seven years, several studies in Spain have evaluated the internal exposure of the population to currently used pesticides. The present review analyzes the scope of these studies, the employed analytical methods and the main results of the exposure and risk, mainly for children and mothers. The frequency of exposure to biomarkers of exposure to organophosphate pesticides is high. Some non-specific dialkyl phosphate metabolites (DAPs), such as the diethyl phosphate (DEP), present Detection Frequencies (DFs) in the range of 65-92% in various studies. Also, the specific biomarker of the chlorpyrifos (3,5,6-trichloro-2-pyridinol, TCPy), achieves Detection Frequencies between 74% and 100% in many studies. For pyrethroids, the metabolite 3-phenoxybenzoic acid (PBA) is present, in general, in more than the 65% of the studied samples. Highly polar herbicides were only assessed in one study and both glyphosate and its metabolite aminomethylphosphonic acid showed Detection Frequencies around 60%. However, putting the biomonitoring data in a risk assessment context, the mean Hazard Quotient (HQ), used as a metric for the individual risk, ranges from 0.0006 (glyphosate) to 0.93 in farm workers (parathion), which means that is unlike that the exposure poses a health concern (HQ < 1).

Liquid chromatography-Orbitrap Tribrid high-resolution mass spectrometry using data dependent-tandem mass spectrometry with triple stage fragmentation as a screening tool to perform identification and risk assessment of unknown substances in food contact epoxy resin.

A new, fast, and automatic approach has been applied for the tentative identification of unknown substances released by food contact epoxy resin after performing a migration test with food simulant. This approach combines intelligent data acquisition with AcquireX linked to liquid chromatography-Orbitrap Tribrid high-resolution mass spectrometry using data dependent-tandem mass spectrometry with triple stage fragmentation coupled to Compound Discoverer™ software for automated data processing and compound identification. The identification of the observed features was performed using a set of identification criteria, including exact mass, isotope pattern, tandem mass spectrometry spectra match, and retention time. With these criteria, 263 substances were tentatively identified. Most of the identified compounds were additives, such as plasticisers, stabilizers, and antioxidants, used in different plastic applications. However, metabolites, biological constituents with pharmacological activity, and other substances with industrial applications were also detected. In order to perform a risk assessment of the food contact epoxy resin, threshold of toxicological concern approach was applied for the identified compounds. There was not risk associated with the migration of the identified substances.

Determination of 60 Migrant Substances in Plastic Food Contact Materials by Vortex-Assisted Liquid-Liquid Extraction and GC-Q-Orbitrap HRMS.

A GC-HRMS analytical method for the determination of 60 migrant substances, including aldehydes, ketones, phthalates and other plasticizers, phenol derivatives, acrylates, and methacrylates, in plastic food contact materials (FCM) has been developed and validated. The proposed method includes migration tests, according to Commission Regulation (EU) 10/2011, using four food simulants (A, B, C, and D1), followed by vortex-assisted liquid-liquid extraction (VA-LLE) and GC-Q-Orbitrap HRMS analysis in selected ion monitoring (SIM) mode, with a resolving power of 30,000 FWHM and a mass accuracy ≤5 ppm. The method was validated, showing satisfactory linearity (R2 ≥ 0.98 from 40 to 400 µg L-1), limits of quantification (40 µg L-1), precision (RSD, 0.6-12.6%), and relative recovery (81-120%). The proposed method was applied to the analysis of field samples, including an epoxy-coated tin food can, a drinking bottle made of Tritan copolyester, a disposable glass made of polycarbonate, and a baby feeding bottle made of polypropylene, showing that they were in compliance with the current European regulation regarding the studied substances.

Biomonitoring of bisphenols A, F, S and parabens in urine of breastfeeding mothers: Exposure and risk assessment.

In the present study we used human biomonitoring to assess the internal exposure and the risk to four parabens and three bisphenols in 103 Spanish breastfeeding mothers participating in the BETTERMIILK project. Urinary methylparaben (MP), ethylparaben (EP), propylparaben (PP) and butylparaben (BP) presented detection frequencies ranging from 12% (BP) to 92% (MP), while bisphenol A (BPA), bisphenol F (BPF) and bisphenol S (BPS) were detected in 76% (BPA) and 20% (BPF, BPS) of the mothers. Average paraben concentrations (geometric mean) ranged from 0.021 ng mL-1 (BP) to 17.7 ng mL-1 (MP), whereas bisphenols had geometric means concentrations from 0.042 ng mL-1 (BPF) to 0.927 ng mL-1 (BPA). Except for BPA, the estimated daily intakes (EDI) were calculated in order to interpret urinary levels in a risk assessment context. The obtained EDIs ranged from 0.00042 mg/kg/day for PP to 0.0434 mg/kg/day for MP and EP. A hazard quotient (HQ) was calculated for BPA (0.0049) and parabens (0.001-0.004), showing no risk in the studied population. Sociodemographic characteristics, food consumption, and usage patterns of personal care products (PCPs) were investigated as possible determinants of exposure. Use of makeup and skincare products were associated with higher concentrations of MP and PP, respectively. Regarding dietary habits, MP was also associated with the consumption of packaged and bakery products.

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.

Determination of nitrofuran metabolites and nifurpirinol in animal tissues and eggs by ultra-high performance liquid chromatography-tandem mass spectrometry validated according to Regulation (EU) 2021/808.

In this work, an analytical method for the determination of eight non-allowed nitrofurans, including nifurpirinol and the metabolites of furazolidone, furaltadone, nitrofurantoin, nitrofurazone, nifursol, nitrovin, and nifuroxazide in animal tissues, including muscle (poultry, bovine, ovine, porcine, rabbit, and fish), kidney (bovine, ovine, porcine), and eggs, has been developed and validated according to Regulation (EU) 2021/808. The method was based on derivatization with 2-nitrobenzaldehyde in acid medium, followed by vortex-assisted liquid-liquid extraction and solid phase extraction for sample purification prior to ultra-high performance liquid chromatography-tandem mass spectrometry. Under selected conditions, the method was validated showing satisfactory relative matrix effects (CV ≤ 20 %), linearity (R2 ≥ 0.98), trueness (≤20 %, expressed as bias), accuracy (83-120 %), repeatability (1.7-19.9 %), reproducibility (1.9-25.7 %), specificity (blank signal ≤30 % at the LCL), and ruggedness. The decision limit for confirmation (CCα) for the target analytes ranged from 0.27 to 0.35 µg kg-1, all below the current reference point for action (RPA) of 0.5 µg kg-1 for the studied compounds. This validated method is currently accredited according to UNE-EN ISO/IEC 17025 by the Spanish National Accreditation Body (ENAC) to be implemented for official control analyses in the Public Health Laboratory of Valencia (Spain).

Risk assessment and predictors of the exposure to polycyclic aromatic hydrocarbons in Spanish adults by urinary human biomonitoring.

Polycyclic aromatic hydrocarbons (PAHs) are produced primarily during incomplete combustion of organic matter and in various industrial processes. They are widespread environmental pollutants that are of significant interest due to their potential toxicity. Humans can be exposed to PAHs through ingestion, inhalation and dermal contact. In the present study, ten urinary PAH biomarkers were determined in first-morning urine samples (n = 504) from the adult population (aged 18-65 years) residing in the Valencian Region of Spain. These samples were analysed using liquid-liquid extraction followed by high-performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS). All PAH biomarkers were quantified in more than 65 % of the urine samples. Naphthalene biomarkers, 1-hydroxynaphthalene (1OHNAP) and 2-hydroxynaphthalene (2OHNAP), exhibited the highest levels with geometric means (GMs) of 0.7 and 11.9 µg L-1, respectively. The 95th percentile of all PAH biomarkers ranged from 0.22 to 64.8 µg L-1. Estimated daily intakes (EDIs) for the analysed PAH families in the studied population ranged from 17 (pyrene) to 18581 (naphthalene) ng·kg-bw-1·day-1 (GM). Significant associations were observed between the quantified urinary metabolites of PAHs and smoking status, home location, annual household incomes, BMI, and the consumption of grilled food in the last 24 h. Hazard quotients (HQs) of naphthalene and consequently, hazard indexes (HIs) were above 1, especially for smokers. Therefore, potential health risks associated with PAH exposure in the target population could not be discarded.

Analysis of microplastics in commercial vegetable edible oils from Italy and Spain.

In this work, assessment of microplastics (MPs) in commercial vegetable edible oils from Italy and Spain, including extra-virgin olive oil, olive oil, sunflower oil, and mixed seed oil, has been conducted for the first time. The method was based on sample dilution with ethanol:n-hexane (1:3, v/v), homogenization, vacuum filtration on macroporous silicon filters with 5 µm pore diameter to collect MPs, and automatic µ-FTIR spectroscopy for MPs detection and characterization. In the analysis of oil samples, a mean MPs abundance of 1140 ± 350 MPs/L was found. Observed MPs were characterized, being most of them fragments (81.2 %), with particle sizes < 100 µm (77.5 %), and mainly composed of polyethylene (50.3 %) and polypropylene (28.7 %), among others. Statistical analysis revealed that there were not significant differences (p-value > 0.05) in the abundance of MPs between oil samples or types.

Assessment of anthropogenic particles content in commercial beverages.

Microplastic (MPs) pollution is a current global concern that is affecting all environmental compartments and food sources. In this work, anthropogenic particles occurrence (MPs and natural and synthetic cellulosic particles), have been determined in 73 beverages packed in different containers. Overall, 1521 anthropogenic particles were found, being the lowest occurrence in water samples (7.2 ± 10.1 items·L-1) while beer had the highest (95.5 ± 91.8 items·L-1). Colourless/white particles were the most detected followed by blue and red colours. The highest mean size was 783 ± 715 µm in soft drinks. Cellulosic, both natural and semisynthetic particles, were the composition mostly found but regarding plastic polymers, it was polyester. Phenoxy resin particles from the can coatings were also identified in all metal containers which indicates that leaching from the packaging may be happening. The total estimated daily intake were 0.077 and 0.159 items·kg-1 body weight (b.w.)·day-1 for children and adult population, respectively.

Levels, patterns and risk assessment of PCDD/Fs and dl-PCBs through dietary exposure in the Valencian Region (Spain).

Polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) and dioxin like polychlorinated biphenyls (dl-PCBs) levels were measured in representative vegetable oils and animal origin foodstuffs collected in a Total Diet Study carried out in the Valencian Region (Spain). A total amount of 3,300 food samples were collected and grouped into 5 main food groups: vegetable oils, meat and meat products, eggs, milk and dairy products and fish and sea products. The samples were analysed using gas chromatography coupled to high-resolution mass spectrometry (GC-HRMS). The food group which presented the highest concentration in wet weight (ww) for the sum of PCDD/Fs and dl-PCBs was fish and sea products (0.5 pg TEQ g-1 ww), whereas meat and meat products (0.6 pg TEQ g-1 lipid) showed the highest levels expressed in lipid terms. Occurrence data of PCDD/F and dl-PCBs were combined with consumption data to estimate the dietary exposure of adults (>15 years) and children (6-15 years). Finally, the estimated weekly intake (EWI) was calculated using a deterministic approach and considering the food consumption of the population, with fish and sea products being the main food group contributor. Likewise, considering the worst-case scenario (Upper Bound, UB), average EWI were 1.8 and 3.4 pg TEQ kg-1 body weight (bw) week-1 for adults and children, respectively. For children, the average EWI was almost twice above the tolerable weekly intake (TWI) of 2 pg TEQ kg-1 bw week-1 set by EFSA in 2018. In terms of risk characterisation, the overall obtained results showed that 19 % of adults and 43 % of children may exceed the TWI when using UB.

Air quality of health facilities in Spain.

The present study examines indoor air pollution in health facilities, focusing on compounds from various sources, such as industrial products, healthcare activities and building materials. It assesses chemical and microbiological concentrations in two public hospitals, two public healthcare centres, and one public health laboratory in Spain. Measurements included indoor air quality, microbiological contaminants, ambient parameters and non-target analysis across ten different locations. Outdoor air quality was also assessed in the surroundings of the hospitals. The results showed that around 350 substances were tentatively identified at a high confidence level, with over 50 % of compounds classified as of high toxicological risk. Three indoor and 26 outdoor compounds were fully confirmed with standards. These confirmed substances were linked to medical, industrial and agricultural activities. Indoor Air Quality (IAQ) results revealed that CO, CO2, formaldehyde (HCHO), O3 and total volatile organic compounds (TVOCs) showed average values above the recommended guideline levels in at least one of the evaluated locations. Moreover, maximum concentrations detected for CO, HCHO, O3 and TVOCs in hospitals surpassed those previously reported in the literature. SARS-CoV-2 was detected in three air environments, corresponding to COVID-19 patient areas. Fungi and bacteria concentrations were acceptable in all assessed locations, identifying different fungi genera, such as Penicillium, Cladosporium, Aspergillus, Alternaria and Botrytis.

Indoor Air Quality and Bioaerosols in Spanish University Classrooms.

A comprehensive study assessed indoor air quality parameters, focusing on relevant air pollutants such as particulate matter (PM10 and PM2.5), gaseous compounds (CO, CO2, formaldehyde, NO2) and volatile/semi-volatile organic chemicals, as well as respiratory viruses (including SARS-CoV-2), fungi and bacteria in Spanish university classrooms. Non-target screening strategies evaluated the presence of organic pollutants inside and outside the classrooms. Saliva samples from teachers and students were collected to explore correlations between respiratory viruses in the air and biological samples. Indoor results revealed the punctual exceedance of recommended guidelines for CO2, formaldehyde (HCHO), volatile organic compounds (TVOCs) and PM in the least naturally ventilated classrooms. Significant differences occurred between the classes, with the least ventilated one showing higher average concentrations of CO2, HCHO, NO2, PM10 and PM2.5. A respiratory virus (rhinovirus/enterovirus) was detected in the medium naturally ventilated classroom, although saliva samples tested negative. Suspect screening tentatively identified 65 substances indoors and over 200 outdoors, with approximately half reporting a high toxicological risk based on the Cramer rules. The study provides a comprehensive overview of indoor air quality, respiratory viruses and organic pollutants in university classrooms, highlighting the variations and potential health risks associated with ventilation differences.

Determination of 21 per- and poly-fluoroalkyl substances in paper- and cardboard-based food contact materials by ultra-high-performance liquid chromatography coupled to high-resolution mass spectrometry.

In this work, a new analytical method was developed and validated for the determination of the total content of 21 per- and poly-fluoroalkyl substances (PFASs) in paper- and cardboard-based food contact materials (FCMs). This method is based on green ultrasound-assisted lixiviation followed by ultra-high-performance liquid chromatography coupled to high-resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS). The method was validated in various paper- and cardboard-based FCMs, showing good linearity (R2 ≥ 0.99), limits of quantification (1.7-10 µg kg-1), accuracy (74-115%), and precision (RSD < 20%). The eco-friendly characteristics of the proposed analytical method were assessed according to the analytical eco-scale, demonstrating that it can be considered an excellent green analytical method (EcoScale score > 75). Finally, 16 field samples of paper- and cardboard-based FCMs, including pizza boxes, popcorn boxes, paper bags and cardboard boxes for potato fries, ice cream tubs, pastry trays, and cardboard-based packaging for cooked Spanish omelets, fresh grapes, frozen fish, and salads were analyzed, showing that they comply with the current European regulations regarding the studied PFASs. The developed method is currently accredited according to UNE-EN ISO/IEC 17025 by the Spanish National Accreditation Body (ENAC) to be implemented for official control analysis of FCMs in the Public Health Laboratory of Valencia, Generalitat Valenciana (Valencia, Spain).

Untargeted analysis and tentative identification of unknown substances in human tears by ultra-high performance liquid chromatography-high resolution mass spectrometry: Pilot study.

In this work, a new approach for the identification of unknown compounds in human tears has been developed and validated using ultra-high performance liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) linked to an intelligent data acquisition mode (AcquireX DS-dd-MS2) coupled to an automated data processing software (Compound Discoverer™ 3.2). As a pilot research study, four human tear samples from volunteers were analyzed. Data were acquired in both positive and negative ionization modes and exact mass, isotope pattern, and MS2 spectra match were used for the tentative identification. Following this approach, 58 substances were identified, 47 in positive mode and 11 in negative mode, with an estimated concentration ranging from 0.1 to 9000 ng mL-1. Most of them were amino acids, hormones, metabolites, and pharmaceuticals. In order to validate the proposed method, the system suitability was evaluated and 29 commercial analytical standards of the tentatively identified substances were analyzed, of which 28 were confirmed obtaining a high identification accuracy (96.6 %). These results confirm that the screening tool presented in this work can facilitate the discovery of new metabolites, novel potential biomarkers, and substances to which the person is exposed, such as pollutants.

Non-target screening and human risk assessment for adult and child populations of semi-volatile organic compounds in residential indoor dust in Spain.

In this work, an analytical strategy based on non-target screening of semi-volatile organic compounds and subsequent risk assessment for adult and child populations has been conducted for the first time in household indoor dust samples in Spain. The methodology was based on a microwave-assisted extraction followed by gas chromatography coupled to high resolution mass spectrometry determination, using a hybrid quadrupole-orbitrap analyzer. The procedure was applied to 19 residential indoor dust samples, collected in different Spanish regions (namely Galicia, La Rioja, Catalunya, the Balearic Islands, and the Valencian Region). From the generated data, 4067 features were obtained, of which 474 compounds were tentatively identified with a high level of identification confidence (probable structure by library spectrum match or confirmed by reference standard), using a restrictive set of identification criteria. Most of the identified chemicals were natural products, metabolites, additives, and substances with industrial applications in the field of foods, cosmetics, pharmaceuticals, pesticides, and plastics. Finally, risk assessment was carried out by applying the threshold of toxicological concern approach, showing that risk to adult and child populations associated with the presence of the identified substances in the indoor dust was not expected, although the existence of indoor environments with conditions of potential risk cannot be discarded under a worst-case scenario approach.

Exposure and Risk Assessment to Airborne dl-PCBs and Dioxins in the Population Living in the Neighborhood of a Cement Plant: A Pilot Study in the Valencian Region of Spain.

Emissions from cement manufacturing facilities may increase health risks in nearby populations. For this reason, dioxin-like PCB (dl-PCB), polychlorinated dibenzo-p-dioxin (PCDD), and polychlorinated dibenzofuran (PCDF) concentrations in PM10 samples were assessed in the vicinity of a cement manufacturing plant located in the Valencian Region (eastern Spain). The total concentrations of the sum of dl-PCBs, PCDDs, and PCDFs ranged between 1.85 and 42.53 fg TEQ/m3 at the assessed stations. The average daily inhalation dose (DID) for the sum in adults ranged from 8.93 · 10-4 to 3.75 · 10-3 pg WHO TEQ kg-1 b.w. d-1, and, for children, the DID ranged from 2.01 · 10-3 to 8.44 · 10-3 pg WHO TEQ kg-1 b.w. d-1. Risk assessment for adults and children was performed using both daily and chronic exposure. The hazard quotient (HQ) was calculated considering 0.025 pg WHO TEQ kg-1 b.w. d-1 to be the acceptable maximum permitted inhalation exposure. The HQ obtained was slightly higher than 1 for PCDD/Fs at one of the stations (Chiva), indicating a possible health risk for the population under study due to inhalation exposure. In the case of chronic exposure, cancer risk (>10-6) was observed for some samples in one of the assessed sampling sites (Chiva).

Assessment of acrylamide exposure in Spain by human biomonitoring: Risk and predictors of exposure.

Acrylamide (AA), a chemical compound currently classified as "reasonably anticipated to be a human carcinogen", is formed through the Maillard reaction in processed carbohydrate-rich foods and is also present in tobacco smoke. The primary sources of AA exposure in the general population are dietary intake and inhalation. Within a 24-h period, humans eliminate approximately 50% of AA in the urine, predominantly in the form of mercapturic acid conjugates such as N-acetyl-S-(2-carbamoylethyl)-L-cysteine (AAMA), N-acetyl-S-(2-carbamoyl-2- hydroxyethyl)-L-cysteine (GAMA3), and N-acetyl-3-[(3-amino-3-oxopropyl)sulfinyl]-L-alanine (AAMA-Sul). These metabolites serve as short-term biomarkers for AA exposure in human biomonitoring studies. In this study, we analysed first-morning urine samples from the adult population (aged 18-65 years) residing in the Valencian Region, Spain, (n = 505). AAMA, GAMA-3 and AAMA-Sul were quantified in 100% of the analysed samples, with geometric means (GM) of 84, 11 and 26 µg L-1, respectively, while the estimated daily intake of AA in the studied population ranged from 1.33 to 2.13 µg·kg-bw-1·day-1 (GM). Statistical analysis of the data indicated that the most significant predictors of AA exposure were smoking and the amount of potato fried products and, biscuits and pastries consumed last 24 h. Based on risk assessment approaches conducted, the findings suggest that exposure to AA could pose a potential health risk. Therefore, it is crucial to closely monitor and continuously evaluate AA exposure to ensure the well-being of the population.

Assessment of exposure to pesticide mixtures in five European countries by a harmonized urinary suspect screening approach.

Humans are exposed to a mixture of pesticides through diet as well as through the environment. We conducted a suspect-screening based study to describe the probability of (concomitant) exposure to a set of pesticide profiles in five European countries (Latvia, Hungary, Czech Republic, Spain and the Netherlands). We explored whether living in an agricultural area (compared to living in a peri-urban area), being a a child (compared to being an adult), and the season in which the urine sample was collected had an impact on the probability of detection of pesticides (-metabolites). In total 2088 urine samples were collected from 1050 participants (525 parent-child pairs) and analyzed through harmonized suspect screening by five different laboratories. Fourty pesticide biomarkers (either pesticide metabolites or the parent pesticides as such) relating to 29 pesticides were identified at high levels of confidence in samples across all study sites. Most frequently detected were biomarkers related to the parent pesticides acetamiprid and chlorpropham. Other biomarkers with high detection rates in at least four countries related to the parent pesticides boscalid, fludioxonil, pirimiphos-methyl, pyrimethanil, clothianidin, fluazifop and propamocarb. In 84% of the samples at least two different pesticides were detected. The median number of detected pesticides in the urine samples was 3, and the maximum was 13 pesticides detected in a single sample. The most frequently co-occurring substances were acetamiprid with chlorpropham (in 62 urine samples), and acetamiprid with tebuconazole (30 samples). Some variation in the probability of detection of pesticides (-metabolites) was observed with living in an agricultural area or season of urine sampling, though no consistent patterns were observed. We did observe differences in the probability of detection of a pesticide (metabolite) among children compared to adults, suggesting a different exposure and/or elimination patterns between adults and children. This survey demonstrates the feasibility of conducting a harmonized pan-European sample collection, combined with suspect screening to provide insight in the presence of exposure to pesticide mixtures in the European population, including agricultural areas. Future improvements could come from improved (harmonized) quantification of pesticide levels.