Simultaneous detection of mycotoxins and pesticides in human urine samples: A 24-h diet intervention study comparing conventional and organic diets in Spain.

Pesticides and mycotoxins, prominent chemical hazards in the food chain, are commonly found in plant-based foods, contributing to their pervasive presence in the human body, as evidenced by biomonitoring programs. Despite this, there is limited knowledge about their co-occurrence patterns. While intervention studies have demonstrated that organic diets can significantly reduce pesticide levels, their impact on mycotoxin exposure has been overlooked. To address this gap, this study pursued two objectives: first, to characterize the simultaneous presence of mycotoxins and pesticides in human urine samples by means of the control of the biomarkers of exposure, and second, to investigate the influence of consuming organic foods on these co-exposure patterns. A pilot study involving 20 healthy volunteers was conducted, with participants consuming either exclusively organic or conventional foods during a 24-h diet intervention in autumn 2021 and spring 2022 to account for seasonal variability. Participants provided detailed 24-h dietary records, and their first-morning urine samples were collected, minimally treated and analysed using LC-Q-ToF-MS by means of a multitargeted method in order to detect the presence of these residues. Results indicated that among the 52 screened compounds, four mycotoxins and seven pesticides were detected in over 25% of the samples. Deoxynivalenol (DON) and the non-specific pesticide metabolite diethylphosphate (DEP) exhibited the highest frequency rates (100%) and concentration levels. Correlations were observed between urine levels of mycotoxins (DON, ochratoxin alpha [OTα], and enniatin B [ENNB]) and organophosphate pesticide metabolites DEP and 2-diethylamino-6-methyl-4-pyrimidinol (DEAMPY). The pilot intervention study suggested a reduction in ENNB and OTα levels and an increase in ß-zearalenol levels in urine after a short-term replacement with organic food. However, caution is advised due to the study's small sample size and short duration, emphasizing the need for further research to enhance understanding of the human chemical exposome and refine chemical risk assessment.

Assessing human exposure to pesticides and mycotoxins: optimization and validation of a method for multianalyte determination in urine samples.

Humans are exposed to an increasing number of contaminants, with diet being one of the most important exposure routes. In this framework, human biomonitoring is considered the gold standard for evaluating human exposure to chemicals. Pesticides and mycotoxins are chemicals of special concern due to their health implications. They constitute the predominant border rejection notifications for food and feed in Europe and the USA. However, current biomonitoring studies are focused on a limited number of compounds and do not evaluate mycotoxins and pesticides together. In this study, an analytical method has been developed for the determination of 30 pesticides and 23 mycotoxins of concern in urine samples. A salting-out liquid-liquid extraction (SALLE) procedure was optimized achieving recoveries between 70 and 120% for almost all the compounds and limits as lower as when QuEChERS was applied. The compounds were then determined by liquid chromatography coupled to triple quadrupole mass spectrometry. Different chromatographic conditions and analytical columns were tested, selecting a Hypersild gold aQ column as the best option. Finally, the method was applied to the analysis of 45 urine samples, in which organophosphate and pyrethroid pesticides (detection rates (DR) of 82% and 42%, respectively) and ochratoxin A and deoxynivalenol (DR of 51% and 33%, respectively) were the most detected compounds. The proposed analytical method involves the simultaneous determination of a diverse set of pesticides and mycotoxins, including their most relevant metabolites, in human urine. It serves as an essential tool for biomonitoring the presence of highly prevalent contaminants in modern society.

Deeper insights into the effects of low dietary levels of polychlorinated biphenyls on pig metabolism using gas chromatography-high resolution mass spectrometry metabolomics.

Polychlorinated biphenyls (PCBs) are a class of contaminants of great concern, linked to the development of many chronic diseases. Adverse effects of PCBs have been documented in humans after accidental and massive exposure. However, little is known about the effect of chronic exposure to low-dose PCB mixtures, and studies regarding scattered lifetime exposures to non-dioxin-like (NDL)-PCBs are especially missing. In this work, serum samples from pigs chronically exposed through their diet during 22 days to Aroclor 1260 (i.e. a commercially available mixture of NDL-PCBs) underwent a metabolomics analysis using gas chromatography-high resolution mass spectrometry (GC-HRMS), with the objective to investigate the effect of exposure to low doses of NDL-PCBs (few ng/kg body weight (b.w.) per day). The study showed that the serum profiles of 84 metabolites are significantly altered by the administration of Aroclor 1260, of which 40 could be identified at level 1. The aggregate interpretation of the results of this study, together with the outcome of a previous one involving LC-HRMS profiling, provided a substantial and concise overview of the effect of low dose exposure to NDL-PCBs, reflecting the hepatotoxic and neurotoxic effects already reported in literature at higher and longer exposures. These results are intended to contribute to the debate on the current toxicological reference values for these substances.

Determination of sulfonylurea pesticide residues in edible seeds used as nutraceuticals by QuEChERS in combination with ultra-high-performance liquid chromatography-tandem mass spectrometry.

This work proposes a novel Quick, Easy, Cheap, Effective, Rugged, and Safe (QuEChERS) method in combination with ultra-high performance liquid chromatography-tandem mass spectrometry for the determination of sulfonylurea residues in edible seeds. The chromatographic separation of nine sulfonylureas was accomplished in less than 5.5 min, using a Luna Omega C18 column (50 × 2.1 mm, 1.6 µm). Mobile phase was supplied at 0.55 mL min-1 and consisted of 0.01% (v/v) aqueous acetic acid as eluent A and a mixture methanol/acetonitrile (80/20, v/v) as eluent B. Column temperature was established at 25 °C. A QuEChERS procedure was investigated as sample treatment for sulfonylureas extraction and sample clean-up. Different clean-up agents (i.e. PSA, Z-Sep+, EMR-Lipid and C18) were evaluated, selecting Z-Sep+ (25 mg) as the best option. The proposed method provided an extraction efficiency greater than 86.2%, while absolute matrix effect was lower than 50.1%. Matrix-matched calibration curves were required for analyte quantification. The analytical method was characterized according to SANTE/11813/2017 guideline, and including limits of detection and quantification, precision, and trueness. Linear dynamic ranges were established from 5 to 150 µg kg-1 for all analytes. Linearity (R2 ≥ 0.9974) and precision in terms of repeatability and intermediate precision (relative standard deviation ≤ 14.7%) are reported. The reporting limit was established at 5 µg kg-1, which is above the limits of quantification of the proposed method (≤ 1.64 µg kg-1) and below the maximum residue levels currently established by European legislation. In general, trueness is within the range of 70-120%. Despite greater recoveries were obtained at the reporting limit (i.e. 120-138%), relative standard deviations lower than 20% were obtained at this concentration level, so fulfilling the requirements of SANTE/11813/2017 guideline. This work represents the first analytical method intended for the analysis of sulfonylureas in sunflower, pumpkin and chia seeds, which are complex matrices due to their high content of fat as well as of growing interest due to their current commercialization as nutraceuticals.

Development and validation of a QuEChERS method for the analysis of 5-nitroimidazole traces in infant milk-based samples by ultra-high performance liquid chromatography-tandem mass spectrometry.

QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) methodology in combination with UHPLC-MS/MS is proposed for the determination of 5-nitroimidazole (5-NDZ) residues in infant milk-based products. Chromatographic separation was accomplished in a C18 Zorbax Eclipse Plus RRHD (50 × 2.1 mm, 1.8 µm) column under gradient elution conditions. Mobile phase consisted of 0.025% (v/v) aqueous formic acid (eluent A) and MeOH (eluent B), and was supplied at a flow rate of 0.5 mL/min. Under these conditions, eleven 5-NDZs including three metabolites were separated in less than 4 min. A novel QuEChERS method was optimized, and primary-secondary amine (PSA) sorbent was selected as clean-up agent. In addition, the proposed QuEChERS procedure was compared with other sample preparation methodologies, which are usually used in the analysis of 5-NDZs, namely solid phase extraction (SPE) using mixed cation exchange (MCX) cartridges and molecularly imprinted solid phase extraction (MISPE). Method comparison was carried out in terms of process efficiency, which includes matrix effect and extraction recovery. Higher process efficiency was generally achieved for QuEChERS and MISPE than for SPE. However, matrix effect was more significant for the non-selective QuEChERS method than for the selective MISPE. Higher extraction recoveries (63.2-94.1%) were observed for QuEChERS. The studied methods were validated in terms of linearity, detection limits (CCα), detection capabilities (CCß) and precision, observing similar performance characteristics independently of the proposed sample treatment. CCα and CCß ranged between 0.05 and 1.69 µg/L for all analytes and extraction techniques. QuEChERS-UHPLC-MS/MS method was also validated in terms of precision (relative standard deviations <10.3%), trueness (recovery>70.2%) and selectivity according to Regulation 2002/657/EC. It is proposed as a good alternative for the monitoring of 5-NDZ residues in enriched infant-milk based products and other similar milk based-products.