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Environmental monitoring is crucial for assessing the overall state of the ecosystems in terms of contaminant impact and chemical landscape. The use of honey bee (Apis mellifera) colonies considerably eases the sampling activities, as honey bees are exposed to a wide range of substances that are transported and accumulated within the beehives. In this work, combining low-resolution and high-resolution mass spectrometry, the APIStrip passive sampler has been employed to evaluate the presence of pesticide residues and the overall characterization of beehive environments. A total of 180 APIStrips have been deployed in 10 Danish apiaries, located in different landscapes, during a five-month sampling period. The targeted methodology for pesticide analysis was based on gas and liquid chromatography coupled with triple quadrupole mass spectrometry, covering 430 pesticide residues. A total of 29 pesticide residues were identified (fluopyram and azoxystrobin being the most frequently detected), with remarkable differences in the pesticide load between apiaries. For its part, the use of non-targeted approaches through liquid chromatography coupled with an Orbitrap mass spectrometer allowed the detection of unknown compounds that were specific of certain environments. Natural products such as eupatilin and gnaphaliin, which are derived from plant sources, were present exclusively in one of the apiaries. Additionally, the detection of drimane sesquiterpenoids, including compounds potentially originating from the Aspergillus genus, suggests the capability of APIStrips to early detect fungal contamination within beehives. This dual approach of low- and high-resolution mass spectrometry maximizes the analytical potential of APIStrips as a tool capable of detecting a wide range of substances with implications for both agricultural practices and ecological health.

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Primary Processed Foods are a class of food items that are ready for consumption after minimal processing in the supply chain. These products are ubiquitous in our daily diet, but so far a limited number of studies dealt with the optimization of quality control methods to check their content of contaminants. Among primary processed foods, bee pollen is a nutritionally acclaimed food supplement, whose contamination with pesticides and mycotoxins has been largely proven. For this reason, the present study aimed at optimizing for the first time a comprehensive LC-MS/MS method capable of analyzing 282 pesticides and 8 mycotoxins in bee pollen. To obtain a suitable method, two extraction procedures (QuEChERS and Accelerated Solvent Extraction), as well as different chromatographic gradients and columns, were tested. The optimized methodology, comprehending an extraction based on semi-automated QuEChERS, and an analytical method including inert LC column technology, was validated and applied to a sample set of 34 bee pollens. The analyzed samples collectively showed the presence of 41 pesticides and 1 mycotoxin.

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The analysis of pesticide residues and mycotoxins in baby food demands exceptionally low limits of quantitation, necessitating the use of highly sensitive instruments capable of conducting trace analyses. High-resolution instruments typically fail to detect such low levels. However, the latest advancements in liquid time-of-flight technology, when coupled with ion trapping, enable ion enrichment, thereby improving detection levels. This allows for the analysis of these substances at low concentration levels, benefiting from enhanced mass accuracy. Additionally, the use of mass accuracy data helped eliminate matrix interferences, thereby enabling high-confidence identification. We developed a multi-residue method to analyse 219 pesticide residues and 9 mycotoxin residues in baby food matrices. Utilizing a QuEChERS-based extraction method, the samples were then analysed using an LC-ZenoTOF 7600 system with mass window screening acquisition. For pesticides, the limit of quantitation was 0.001-0.003 mg/kg for 81 % of the evaluated compounds, 0.005 mg/kg for 13 %, 0.010 mg/kg for 4 % and 0.020-0.030 for 2 %; good linearities were obtained at these levels. Apparent recoveries were evaluated at 0.003, 0.005, and 0.010 mg/kg. At the lowest recovery level, 93 % of compounds showed recoveries between 70 and 120 %. The rest of the compounds were in the range of 63-129 %, with relative standard deviation values below 20 %. For mycotoxins, the limits of quantitation ranged from 0.0001 to 0.100 mg/kg, with matrix-matched concentrations assessed within this range. Recoveries were evaluated at low concentration range (0.001-0.003 mg/kg) and high range (0.020-0.050) with apparent recoveries values between 92 and 140 %. Finally, a total of 31 commercial baby food samples were analysed using this method. The results indicated that 16 samples contained pesticide residues, while two samples were found to have mycotoxins.

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The performance of the QuEChERS method in this study, as indicated by a high percentage (>90%) of recovery observations falling within the range of 60-140% and a sample replicate deviation (% RSD) of <20%, for the routine analysis of isoprocarb and carbaryl pesticides, has been evaluated over a 14-month period for the export of Indonesian coffee. Following a seven-day observation of the stability of these pesticides in coffee extract, it was found that the added standard calibration solution remained stable and useable for seven days when stored at 4 °C and -20 °C. This validated method, with high sensitivity (a LOQ of 0.001 mg kg-1 for isoprocarb and carbaryl), has been employed to monitor residues in Indonesian coffee exports to comply with maximum residue limits (MRLs). The samples with higher contamination levels were predominantly from robusta coffee (57.76%), followed by arabica coffee (6.17%). The detection rates for residues decreased by more than 90% in the last two months of the method's application. In the observation of coffee processing, it was found that isoprocarb residues in contaminated samples could be transferred to the processed coffee (roasted and its infusion) to a limited extent, while residues from the carcinogenic carbaryl were not detected due to evaporation. Additionally, chronic dietary risk assessment showed that contaminated samples of robusta and arabica coffees should not be considered a significant public health concern (hazard index HI < 1). However, continuous monitoring of pesticide residues in Indonesian coffee is still recommended, not only to conform to the MRLs of importing countries but also to ensure food trade.

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Recent research has highlighted the potential of honeybees and bee products as biological samplers for monitoring xenobiotic pollutants. However, the effectiveness of these biological samplers in tracking microplastics (MPs) has not yet been explored. This study evaluates several methods of sampling MPs, using honeybees, pollen, and a novel in-hive passive sampler named the APITrap. The collected samples were characterized using a stereomicroscopy to count and categorise MPs by morphology, colour, and type. To chemical identification, a micro-Fourier transform-infrared (FTIR) spectroscopy was employed to determine the polymer types. The study was conducted across four consecutive surveillance programmes, in five different apiaries in Denmark. Our findings indicated that APITrap demonstrated better reproducibility, with a lower variation in results of 39%, compared to 111% for honeybee samples and 97% for pollen samples. Furthermore, the use of APITrap has no negative impact on bees and can be easily applied in successive samplings. The average number of MPs detected in the four monitoring studies ranged from 39 to 67 in the APITrap, 6 to 9 in honeybee samples, and 6 to 11 in pollen samples. Fibres were the most frequently found, accounting for an average of 91% of the total MPs detected in the APITrap, and similar values for fragments (5%) and films (4%). The MPs were predominantly coloured black, blue, green and red. Spectroscopy analysis confirmed the presence of up to five different synthetic polymers. Polyethylene terephthalate (PET) was the most common in case of fibres and similarly to polypropylene (PP), polyethylene (PE), polyacrylonitrile (PAN) and polyamide (PA) in non fibrous MPs. This study, based on citizen science and supported by beekeepers, highlights the potential of MPs to accumulate in beehives. It also shows that the APITrap provides a highly reliable and comprehensive approach for sampling in large-scale monitoring studies.

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This manuscript presents a comprehensive review of high-resolution mass spectrometry in the field of food analysis and metabolomics. We have followed the historical evolution of metabolomics, its associated techniques and technologies, and its increasing role in food science and research. The review provides a critical comparison and synthesis of tentative identification guidelines proposed for over 15 years, offering a condensed resource for researchers in the field. We have also examined a wide range of recent metabolomics studies, showcasing various methodologies and highlighting key findings as a testimony of the versatility of the field and the possibilities it offers. In doing so, we have also carefully provided a compilation of the software tools that may be employed in this type of studies. The manuscript also explores the prospects of high-resolution mass spectrometry and metabolomics in food science. By covering the history, guidelines, applications, and tools of metabolomics, this review attempts to become a comprehensive guide for researchers in a rapidly evolving field.

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In this comprehensive study, we evaluated the feasibility of using hydrogen instead of helium as a carrier gas in a GC-MS/MS system for pesticide residue analysis, spanning three matrices: pepper, tomato, and zucchini. Initial assessments focused on the ion source's chemical inertness, employing nitrobenzene as a benchmark to monitor the hydrogenation process. A method with a duration of less than 12 minutes was developed, achieving good chromatographic peak resolution attributable to the enhanced chromatographic performance of hydrogen as a carrier gas. The study emphasized the optimization of system parameters, testing various ion source temperatures, detector voltages, and injection volumes. Sensitivity assessments, based on the DG-SANTE criteria, indicated that the majority of compounds were identifiable at a concentration of 5 µg kg-1 (81% in tomato, 84% in pepper and 73% in zucchini). Detailed validation for reproducibility, matrix effects, and linearity across 150 pesticides unveiled generally favorable outcomes, with a notable majority of compounds displaying low matrix effects, satisfactory linearity ranges and good reproducibility with most compounds returning a relative standard deviation (RSD) below 10%. When applied to 15 real samples, the hydrogen-based system's performance was juxtaposed against a helium-based counterpart, revealing that results are very comparable between both systems. This comparative approach highlights hydrogen's potential as a reliable and efficient carrier gas in pesticide residue analysis for routine food control laboratories, overcoming difficulties resulting from the lack of helium supplies.

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Microplastics from mulch films can be a source of chemical contamination to agricultural soils. In this context, biodegradable films have been widely positioned as a greener choice. However, their sorption/desorption capabilities, in contrast to the conventional plastic types remain understudied. It is for this reason that objective evaluation of their interactions with residual agricultural contaminants becomes important. Our findings reveal that polyethylene (PE) mulch films retained lower amounts of pesticide residues and demonstrated a higher desorption/release [median desorption = 71.86 µg/L or about 50%], while polybutylene adipate terephthalate (PBAT) mulch films retained higher amounts of pesticide residues onto their surface and demonstrated a much lower desorption [median desorption = 24.27 µg/L or about 17%] after a spraying event. A higher ambient temperature had no significant effect on final desorption amounts in both PE [median = 65.27 µg/L at 20 °C and 74.23 µg/L at 40 °C] and PBAT [median = 24.26 µg/L at 20 °C and 24.78 µg/L at 40 °C] mulch films. However, it did favour a faster desorption pace in PE films. Desorption in PBAT and PE plastic types was correlated with the log Kow value [Spearman's correlation: 0.857 and 0.837 respectively, p < 0.05]. However, only a moderate correlation with pKa was observed in PBAT [Spearman's correlation: 0.478, p < 0.05], while none for PE plastic type. Sorption of pesticides onto biodegradable PBAT microplastics were best explained by Elovich [R2: 0.937-0.959] and pseudo-second order kinetics [R2: 0.942-0.987], suggesting the presence of chemisorption. Furthermore, Weber Morris plots suggested the presence of a multi-step process and Boyd plots indicated that film diffusion or chemical bond formation was the rate-limiting step governing this phenomenon.

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The analysis of highly polar pesticides is challenging due to their unique physicochemical properties, requiring specialized chromatographic techniques for their accurate and sensitive detection. Furthermore, the high level of co-extracted polar matrix components that can co-elute with the analytes can interfere with the analysis. Consequently, there is lack of pesticide monitoring data, as the European Food Safety Authority has pointed out. This article explores the overcoming of such difficulties in the analysis of these compounds. Analytical methodologies for the extraction, clean-up, and direct determination of 11 highly polar anionic pesticides, including glyphosate, glufosinate, ethephon, fosetyl-aluminium, and their related metabolites in complex food matrices such as honey and pollen by hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry were successfully developed and validated. Solid-phase extraction and micro-solid-phase extraction employing strong anion exchange (SAX) cartridges were implemented for clean-up. The automation and miniaturization of SAX clean-up for these compounds were achieved for the first time. For method validation, SANTE/11312/2021 guideline was followed. Recoveries were between 70 and 120%, with RSDs below 20%. Limits of quantitation ranged from 0.005 to 0.020 mg kg-1. Linearity was evaluated from 0.002 to 0.200 mg kg-1. Matrix effects were assessed, showing medium to low signal suppression for most compounds. AMPA and glufosinate presented the highest signal suppression, but it was reduced after SAX clean-up. Analysis of real honey and pollen samples revealed the occurrence of the studied compounds in beehive products and showed the applicability of the validated methodologies for routine control of these complex samples.

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This study confirms the uptake, translocation and bioaccumulation of 100 nm polystyrene nanoplastics in the root, stem and leaves of the plant Lepidum sativum at exposure concentrations ranging from environmentally realistic 10 µg/L up to a high of 100 mg/L. Accumulation in plant tissues was characterised by aggregation in the intercellular spaces and heterogeneous distribution. Nanoplastic presence was confirmed in the root tips, root surface and stele, lateral roots, root hairs, stem vascular bundles, leaf veins and mesophyll, as well as leaf epidermis including stomatal sites. Quantification results show that majority of the particles were retained in the root and accumulation in stem and leaves was only 13 to 18 % of the median value in roots. There was a reduction of 38.89 ± 9.62 % in the germination rate, 55 % in plant fresh weight, as well as in root weight (> 80 %), root length (> 60 %), shoot weight (51 to 78 %) and number of lateral roots (> 28 %) at exposure concentrations at and above 50 mg/L. However, lower, environmentally probable exposure concentrations did not affect the plant health significantly. Our results highlight the urgent need for further exploration of this issue from the point of view of food safety and security. STATEMENT OF ENVIRONMENTAL IMPLICATION: Micro and nanoplastics have been reported in agricultural environments across the globe and reports regarding their hazardous effects over agricultural and plant health call for an urgent exploration of this issue. This work demonstrates the uptake, bioaccumulation and distribution of nanoplastics in an edible plant at an environmentally realistic concentration and raises serious concerns regarding the possible implications for food safety and security. It presents a novel approach which addresses the quantification of nanoplastic accumulation in plant tissues and helps identify the mechanism and trends behind this phenomenon which has been a challenge up until now.

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Pesticide contamination in emerging foods and supplements is currently a topic of great interest. This study focused on the evaluation of pesticide residues in commercial bee pollen samples to evaluate the risk associated with their consumption. To this end, an automated clean-up method for the pesticide extracts of bee pollen was developed. An LC-MS/MS and a GC-MS/MS method were validated for the analysis of 353 pesticides in 80 bee pollen samples purchased from different countries. The results showed the presence of 77 different pesticide residues in bee pollen, including plant protection chemicals and veterinary treatments. 85 % of the samples were contaminated with pesticides and no relevant differences were found between conventional and organic samples. Pesticide concentrations exceeding the imposed MRL were found in 40 % of the samples, but the risk assessment showed that consumers are not exposed to an unacceptable risk when consuming the evaluated bee pollen.

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Spices such as paprika, curry, turmeric, dry chilli, and black pepper are grown in various geographic locations and widely used by consumers across the world. Pesticides applied during crop production practices could contaminate the produce, affecting the quality and posing a health risk for consumers. The complexity of the spice matrix and the wide range of target pesticides potentially present require special sample extraction and clean-up treatments to overcome matrix interference and ion suppression. In this study, sample extracts from spice matrices (paprika/curry/turmeric/dry chilli/black pepper) were cleaned up by an automated µSPE clean-up method for multi-residue analysis of pesticides using LC-MS/MS. The automated µSPE clean-up method involves pre-filled cartridges containing various sorbent materials suitable for numerous co-extractives and the automated clean-up process was carried out using an autosampler. The regulatory limit for pesticides in spices varies with type, with a low MRL of 0.05 mg kg-1 or higher for 99% of the analytes. At spiking concentrations of 0.05 and 0.1 mg kg-1, good recoveries between 70 and 120% with RSD values below 20% were achieved for more than 98% of the compounds. With automatic clean-up of samples that takes 5 min/sample, 20% increased output per day shows an important advantage achieved compared to manual clean-up.

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Western honey bees are very sensitive bioindicators for studying environmental conditions, hence frequently included in many investigations. However, it is very common in both research studies and health surveillance programs to sample different components of the colony, including adult bees, brood and their food reserves. These practices are undoubtedly aggressive for the colony as a whole, and may affect its normal functioning and even compromise its viability. APIStrip-based passive sampling allows long-term monitoring of residues without affecting the colony in any way. In this study, we compared the effectiveness in the control of acaricide residues by using passive and conventional sampling, where the residue levels of the acaricides coumaphos, tau-fluvalinate and amitraz were evaluated. Conventional and APIStrip-based sampling differ in methods for evaluating bee exposure to residues. APIStrip is less invasive than conventional sampling, offers more efficient measurement of environmental contaminants, and can be stored at room temperature, saving costs and minimizing operator error.

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Manuka honey is a valuable commodity produced by bees foraging the flowers of Leptospermum scoparium, a bush native to New Zealand and Australia. Due to its high value and proven health benefits, authenticity fraud in the sale of this food is a significant risk, as recounted in the literature. Four compulsory natural products must be present at minimum concentrations to authenticate manuka honey (3-phenyllactic acid, 2'-methoxyacetophenone, 2-methoxybenzoic acid, and 4-hydroxyphenyllactic acid). However, spiking other kinds of honey with these compounds and/or the dilution of manuka honey with other varieties may result in fraud going undetected. In this work, liquid chromatography coupled with high-resolution mass spectrometry and a metabolomics-based strategy has allowed us to tentatively identify 19 natural products -putative manuka honey markers-, nine of which are reported for the first time. Chemometric models applied to these markers allowed the detection of both spiking and dilution fraud attempts of manuka honey, even at 75% manuka honey purity. Thus, the herein-reported methodology can be employed in the prevention and detection of manuka honey adulteration even at low levels, and the tentatively identified markers presented in this work proved valuable for manuka honey authentication procedures.

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The use of apicultural matrices for the environmental monitoring of pesticides is a widely employed approach that facilitates to a great extent the sampling procedures. Honey bees are one of the most commonly employed matrices in these studies due to their abundance in the colonies and their direct contact with the beehive and the environment. However, the analysis of this matrix is associated to a lack of representativity of the contaminants accumulated within the beehive, due mainly to the limited number of honey bees that are sampled and analyzed compared to the population in a hive. This small proportion of organisms which are sampled from the colony may lead to underestimations or overestimations of the total pesticide load, depending on the specific individuals that are included in the analysis. In the present work, the passive, non-invasive APIStrip-based sampling approach is compared to active bee sampling with a total of 240 samples taken from 15 apiaries from Austria, Denmark and Greece over a two-month period in 2022. The APIStrips have been found to provide a more comprehensive image of the pesticide residues accumulated in the beehive in terms of number of identified residues and robustness of the results. A total of 74 different pesticide residues were detected: the use of APIStrips allowed to detect 66 pesticides in the three countries, compared to 38 residues in honey bees. The use of APIStrips also resulted in a higher percentage of positive samples (containing at least one pesticide residue). The results provided by the passive sampling approach were also more consistent among the replicates and over time, which reveals an increased sampling robustness.

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Microwavable plastic food containers can be a source of toxic substances. Plastic materials such as polypropylene polymers are typically employed as safe materials in food packaging, but recent research demonstrates the migration of plastic substances or their by-products to food simulants, to foodstuff, and, more recently, to the human body through food consumption. However, a thorough evaluation of foodstuff in food contact materials under cooking conditions has not yet been undertaken. Here we show for the first time that plastic migrants present in food contact materials can react with natural food components resulting in a compound that combines a UV-photoinitiator (2-hydroxy-2-methyl-1-phenylpropan-1-one) with maltose from potato starch; this has been identified after cooking potatoes in microwavable plastic food containers. Additionally, polypropylene glycol substances have been found to transfer into food through microwave cooking. Identifying these substances formed in situ requires state-of-the-art high-resolution mass spectrometry instrumentation and metabolomics-based strategies.

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Clean-up step is essential during the multiresidue sample preparation process to remove undesired matrix components that may cause analytical interferences or suppression effect. However, its application generally by specific sorbents entails time-consuming work producing low recoveries for some compounds. Moreover, it usually needs to be adapted to the different co-extractives from the matrix present in the samples by using different chemical sorbents increasing the number of validation procedures. Therefore, the development of a more efficient and automated and unified clean-up procedure means a significant time reduction and laboratory work with improved performance. In this study, extracts from different matrices (tomato, orange, rice, avocado and black tea) were purified by manual dispersive clean-up (different procedures according to the matrix group) in parallel with an automated µSPE clean-up workflow, in both cases based on QuEChERS extraction. The latter procedure employed clean-up cartridges containing a mixture of sorbent materials (anhydrous MgSO4/PSA/C18/CarbonX) suitable for multiple matrices. All the samples were analysed by liquid chromatography mass spectrometry and the results obtained from both procedures have been compared in terms of the extract cleanness, performance, interferences, and sample workflow. At the levels studied, similar recoveries were achieved by both techniques (manual and automated) except for reactive compounds when PSA was used as the sorbent material producing low recoveries. However, the µSPE recoveries were between 70-120%. Furthermore, closer calibration line slopes were provided when µSPE was applied to the different matrix groups studied. It is important to note that up to 30% more samples per day can be analysed using an automated µSPE compared to the manual method (which requires shaking, centrifuging, then taking the supernatant and adding formic acid in ACN); it also provides good repeatability - an RSD (%) < 10%. Consequently, this technique is a very useful option for routine analyses, greatly simplifying the work of muti-residue methods.

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This nationwide monitoring aimed to investigate the prevalence of residues of plant protection products (PPPs) and veterinary medicine products (VMPs) based on random selection of apiaries of Apis mellifera. For a three-year period (2012, 2013 and 2016), this study targeted 306 PPPs, VMPs and other active substances in 442 samples of bee bread honeycomb (BBHC) and 89 samples of honeybees collected from up to 177 apiaries. The results indicate that honeybees were most often exposed to residues of coumaphos, tau-fluvalinate, chlorfenvinphos, and acrinathrin, with a prevalence from a maximum of 98.8% to 49.4% in BBHC samples. Residues of coumaphos, tau-fluvalinate, amitraz (DMF + DMPF), carbendazim and orthophenylphenol were also frequently detected, from a maximum of 55.1% to 13.5% of the honeybee samples. Neonicotinoid residues, namely clothianidin and thiamethoxam, whose outdoor uses in crops are completely banned in EU, were not detected. Imidacloprid was found in 3.4% to 13.3% of samples during 2013 and 2016, respectively. Imidacloprid exceeded its acute toxicity (LD50) value for honey bees in two samples of BBHC. Fipronil was detected in 0.5% of the samples during 2013. The diversity of active substances found (% of different residues analyzed) ranged from 33.9% to 37.2% in BBHC from 2012, 2013 to 2016, and was of 26.5% in honeybees in 2016. In at least 54% of the samples, the total residue load was in the range of 200 to 1500 µg·kg-1. Up to 50% of BBHC samples were positive for one or two residues. No toxic residues for honeybees were detected in up to 88.8% of bee samples. This systematic surveillance of active substances assisted the evaluation of which target pesticides to look for and provided support to the competent authorities in the bee health decision-making.

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BACKGROUND: Pesticide quantitation in foods relies on the availability of matrix reference materials, which, however, are scarce because of the general instability of pesticides. In particular, no avocado reference material has been developed. OBJECTIVE: This research aimed to develop a reference material to support the determination of pesticide residues in avocado. METHOD: Avocado was spiked with 11 selected pesticides at levels of 0.01-0.35 mg/kg. The production process included the assessment of between-unit heterogeneity, stability during dispatch, and best storage conditions according to ISO 17034. Reference values were estimated through an interlaboratory comparison study involving laboratories of demonstrated competence and adhering to ISO/IEC 17025. The corresponding expanded uncertainties were calculated as 4-19% in compliance with the Guide to the Expression of Uncertainty in Measurement. RESULTS: The reference material was sufficiently homogeneous and stable at 4°C during the entire study period (365 days) for most of the pesticides, and at 40°C during 10 days, but it was unstable at 50 and 60°C during 10 days. Four pesticides showed downward trends; however, this behavior was considered in the uncertainty budget. As this material complied with all requirements of proficiency testing, it was used in an interlaboratory proficiency test designed to investigate analytical performance and assist laboratories in improving the quality of measurement results. CONCLUSIONS: The presented material can be used for the development of novel analytical methods or in-house reference materials and adds to the scarce supply of reference materials for the determination of pesticides in vegetable matrixes with high oil contents and intermediate water contents. HIGHLIGHTS: A novel reference material has been developed for the reliable and accurate quantitation of multiple pesticides in avocado, allowing researchers to avoid the preparation of in-house reference materials for this purpose.

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