Food waste reduction remains a major challenge in the European Union, where annual losses exceed 88 million tons. Active food packaging offers a sustainable strategy to extend shelf-life and maintain product quality. This study evaluated the antioxidant performance of natural extracts from olive by-products stone, leaf and pomace obtained through a green ethanol-based extraction. Polyphenolic profiles were determined by UHPLC-DAD, and antioxidant capacity was measured using DPPH assays, with olive stone extract showing the best activity (EC50 = (63.6 ± 4.6) mg/L). Total biophenol contents ranged from 10 g/kg to 130 g/kg, with high levels of gallic acid and related polyphenols. Among all, olive stone extracts exhibited the strongest performance in terms of antioxidant power percentage ((84.4 ± 1.6) %AP). Packaging films incorporating these extracts significantly delayed lipid oxidation in minced beef meat with 50% fat, reducing it to 18.4% after 10 days versus 46.6% in the control, demonstrating strong potential for sustainable industrial application.

Per- and polyfluoroalkyl substances (PFAS) extensively contaminate the environment and can enter the food chain. Fruits and vegetables represent a significant, yet under-regulated, human exposure pathway due to the uptake of PFAS by plants from contaminated soil/water. PFAS exposure is linked to adverse health effects. EFSA has established a strict group tolerable weekly intake of 4.4 ng/kgbw/week for PFOA, PFOS, PFHxS, and PFNA, leading the EU to introduce specific regulations. However, EU maximum levels in plant-derived foods are lacking because the achievability of limits of quantification (LOQ) of 0.001 mu g/kg poses significant analytical challenges. This study presents the results of an extensive interlaboratory validation study among European laboratories using various triple quadrupoles and high-resolution mass spectrometers. The aim was to assess method robustness and achievability of analytical LOQs required by EU guidance with a focus on the EFSA-4 PFAS. The interlaboratory validation study involved 240 food extracts prepared in two different laboratories by two operators each. The quantification was performed across four laboratories and seven analytical setups. The validated method showed robustness, with an apparent recovery between 65% and 135%, uncertainty and RSD below 25%, and consistency with current EU guidelines. The application of internal 13C-labeled standards and the use of very sensitive mass spectrometers proved to be a critical factor in achieving low LOQs.

To ensure a high level of food safety throughout production and distribution, it is crucial to improve and harmonize analytical techniques for contaminant quantification. This constitutes the basis for reliable data regarding food product compliance with regulations and for resolving disputes and minimizing financial losses within the food industry. In this framework, the ScreenFood project is developing reference methods and reference materials for quantifying contaminants in both food and food packaging, with a specific emphasis on recycled materials. The project aims: i. To improve protocols for the quantification of the mineral oil aromatic hydrocarbons fraction and for the quantification of the fraction consisting of substances with three or more aromatic ring systems (3+MOAH); ii. To develop sensitive analytical procedures for detecting and quantifying per- and polyfluoroalkyl substances (PFAS) in selected matrices, in line with the requirements of Commission Regulation (EU) 2023/915; iii. To develop traceable and highly accurate reference materials for quality control and quality assurance purposes. These materials will be designed for key contaminants like MOSH, MOAH, and PFAS in relevant food and food-related matrices, such as vegetable oil, infant formula, vegetables, and food packaging materials; iv. to develop screening methods, addressing new/existing organic and inorganic contaminants, in virgin and recycled packaging such as bio-based food contact materials and reusable materials, and to be applied also in migration studies. In this presentation, the project and the ongoing activities related to the 4 objectives will be described, and the experimental results related to the validation of analytical methods for the quantification of PFAS in different matrices will be presented. Cereals and foods of plant origin are the categories covered in the studies. In case of cereals, an UHPLC-HRMS/MS based method was validated, obtaining a LOQ ranging between 2 ng/kg and 32 ng/kg and robustness in line with EU guidelines and recommendation for PFAS in food. Additionally, a metrological approach was employed to estimate the uncertainty budget, utilizing modeling and experimental methods, and comparing the outcomes. A comparison of uncertainty of different approaches was conducted after applying the method to 30 real samples. In the case of fruit and vegetables, the work aimed to validate, according to EURL-POPs guidance, an analytical method for assessing PFAS levels in vegetable and fruit samples using an interlaboratory approach. The study was conducted involving 4 different laboratories to achieve interlaboratory validation, including all the steps from the sample preparation (exchanging extracts prepared in two laboratories) to the instrumental analysis by means of 4 different liquid chromatography coupled with mass spectrometry apparatus. The study has led to a robust, accurate, and validated method for quantifying PFAS that will be used within the project for the characterization of candidate reference materials for PFAS, one in a tomato-based matrix and one in a cereal-based baby food, and that can be applied in migration studies from packaging. The outcome of the work and the project in this area will support the development of EU legislation for PFAS in matrices that are not yet encompassed by existing regulations and recommendations.

Food contamination by per- and polyfluoroalkyl substances (PFAS), especially ultra-short-chain (USC) compounds, poses a growing concern due to their environmental persistence and potential health risks. Despite the developing regulatory framework, analytical challenges persist in quantifying polar USC-PFAS in complex content food matrices. This study presents the development and validation of a novel high-performance liquid chromatography coupled to a tandem mass spectrometer (HPLC-MS/MS) method for the accurate determination of USC-PFAS (carbon chain length from one to four, C1-C4) in tomato-based products (i.e. concentrate, puree, and pulp), that, due to their high water content, are prone to USC-PFAS contamination. Leveraging a polar analytical column and a delay column, the method effectively mitigates system-related interferences, especially for trifluoroacetic acid (TFA) and achieves enhanced retention and sensitivity. Target compounds included Difluoroacetic acid (DFA), TFA, Perfluoropropanoic acid (PFPrA), Perfluorobutanoic acid (PFBA), Perfluoromethanesulfonic acid (PFMeS), Perfluoroethanesulfonic acid (PFEtS), Perfluoropropanesulfonic acid (PFPrS), and Perfluorobutanesulfonic acid (PFBS). The quantification based on isotope dilution ensures high accuracy and precision. The method demonstrated excellent linearity (R² ≥ 0.99), recoveries within 65-135 %, and low relative standard deviation (RSD) values (<10 %). PFBA was detected across all tested tomato products, with concentrations ranging from 0.056 to 0.265 µg/kg, indicating potential endogenous contamination potentially linked to processing concentration effects. This study fills a critical methodological gap, offering a robust analytical tool for USC-PFAS monitoring in complex food matrices, and supporting improved food safety regulation.

Alpha‐gal syndrome (AGS) is a mammalian meat allergy associated with tick bites and specific IgE to the oligosaccharide galactose‐α‐1,3‐galactose (α‐gal). Recent studies have shown that 10–20% of AGS patients also react to the dairy proteins. Considering the already described role of the meat lipid fraction in AGS manifestations, the aim of this work has been to investigate whether the milk fat globule proteins (MFGPs) could be involved in AGS.The MFGPs are extracted and their recognition by the IgE of AGS patients is proved through immunoblotting experiments. The identification of the immunoreactive proteins by LC‐HRMS analysis allows to demonstrate for the first time that butyrophillin, lactadherin, and xanthine oxidase (XO) are α‐gal glycosylated. The role of xanthine oxidase seems to be prevalent since it is highly recognized by both the anti‐α‐gal antibody and AGS patient sera.The results obtained in this study provide novel insights in the characterization of α‐Gal carrying glycoproteins in bovine milk, supporting the possibility that milk, especially in its whole form, may give reactions in AGS patients. Although additional factors are probably associated with the clinical manifestations, the avoidance of milk and milk products should be considered in individuals with AGS showing symptoms related to milk consumption.

In the present work, an analytical method for the quantification of per and poly fluoroalkyl substances (PFAS) in rice and maize has been developed and then validated with a metrological approach. PFAS are a group of human-made chemicals used in a variety of industries and consumer products for their water- and grease-resistant properties. Studies have shown that PFAS can contaminate soil and water, and there is concern about their bioaccumulation in edible plants, fruits, and cereals. The presence of PFAS has been identified in rice and other food products, including maize, as indicated by studies and scientific literature. This is particularly alarming since some PFAS have been associated with adverse health effects and rice and maize account for over 20% of the annual food intake worldwide. Despite this evidence, the regulation currently in place is not covering cereal matrices and limits of quantification for matrices encompassed by the current legislation are defined for a small group of PFAS. In this study an UHPLC-HRMS/MS based method was validated, obtaining a LOQ (Limit Of Quantification) ranging between 2 ng/kg and 32 ng/kg and robustness in line with EU guidelines and recommendation for PFAS in food. Additionally, a metrological approach was employed to estimate the uncertainty budget, utilizing modeling and experimental methods, and comparing the outcomes, aiming to characterize with high accuracy PFAS in rice and maize and support control bodies to assess contamination in suspected areas. A comparison of uncertainty of different approaches was conducted after applying the method to 30 real samples.

Main text This Supplementary Comparison, EURAMET.QM-S15 "PAHs in Protein Matrix", was organized by the Federal Institute of Metrology METAS. The target analytes were four PAHs: benz[a]anthracene (BaA); benzo[a]pyrene (BaP); benzo[b]fluoranthene (BbF); and chrysene (Chr), for which maximum limits are set in the European and Swiss food legislation. The measurands were the mass fractions (µg/kg) of these PAHs in a protein-rich matrix. Six National Metrology Institutions (NMIs) and Designated Institutes (DIs) participated in EURAMET.QM-S15. The study required solvent extraction, separation of the target analytes from interfering matrix components (clean-up), analytical/chromatographic separation and selective detection of the target analytes. Solvent extraction (ASE, PLE, HUPsSE, QuEChERS) followed by SPE cleanup was applied by the participants in the sample pretreatment and GC-IDMS/MS, GC-HR-IDMS and LC-FLD were applied for separation and detection. The results for the determination of the four PAHs in EURAMET.QM-S15 ranged from 0.5 µg/kg to 5.3 µg/kg for BaA, 0.6 µg/kg to 13.3 µg/kg for BaP, 0.6 µg/kg to 7.0 µg/kg for BbF and 0.5 µg/kg to 5.0 µg/kg for Chr. Due to poor extraction efficiencies, as a result of unfavorable choice of solvent and insufficient demonstration of metrological traceability of the calibrant, only the results of two participants were included in the evaluation of the Supplementary Comparison Reference Value (SCRV). The SCRV was assigned using the weighted mean of these two participants which agreed well with gravimetrically prepared mass fractions and evidence from an ancillary study performed with external expert laboratories. Four participant results were excluded from the SCRV for the technical reasons mentioned above. The SCRV (X ± u(X)) was 3.291 µg/kg ± 0.079 µg/kg for BaA, 4.28 µg/kg ± 0.10 µg/kg for BaP, 4.77 µg/kg ± 0.12 µg/kg for BbF and 2.967 µg/kg ± 0.070 µg/kg for Chr. The two institutes that were included in the assignment of consensus SCRV agreed within their standard uncertainties. Successful participation in EURAMET.QM-S15 demonstrates the following measurement capabilities in determining the mass fractions of organic compounds, with a molecular mass between 150 g/mol and 500 g/mol, having a low polarity (pKow < -2), in a mass fraction range from 0.1 µg/kg to 100 µg/kg in a high protein food matrix. To reach the main text of this paper, click on Final Report. Note that this text is that which appears in Appendix B of the BIPM key comparison database https://www.bipm.org/kcdb/. The final report has been peer-reviewed and approved for publication by the CCQM, according to the provisions of the CIPM Mutual Recognition Arrangement (CIPM MRA).

More than 7000 synthetic compounds known as per- and poly-fluoroalkyl substances (PFAS) are applied to food packaging and other materials to provide fat, fire, and/or water resistance properties. These compounds have exceptional environmental stability and persistence due to the strong C-F chemical bond, earning them the moniker "forever chemicals". Emission of PFAS from industrial waste leads to water, air, and soil contamination. Due to this ubiquitous nature, combined with the fact that PFAS in humans are known to have carcinogenic and reprotoxic effects and to cause vaccine resistance and depression of the immunity system, PFAS may constitute a major threat to human health. For this reason, the attention of the scientific community and of control bodies is increasing and as a consequence legislation and the scientific literature on PFAS are constantly evolving. This review aims to provide a comprehensive overview of the state of the art about current legislation addressing PFAS; targeted and screening method for identification, detection and quantification of PFAS; toxicity of PFAS; and contamination of environmental and food matrices and from food contact matrices. A comprehensive review of the latest scientific research and recent developments in the legislation of PFAS will provide insights into the current understanding of PFAS and its health implications. Moreover, it will serve as a valuable reference for further studies related to PFAS and could help in informing future policy decisions.

An innovative spectroscopic method that allows to chemically and structurally characterize viruses directly in suspension within few minutes was developed. A library of five different plant viruses was obtained combining dielectrophoresis (DEP), performed with a device specifically designed to capture and agglomerate virus particles, and Raman spectroscopy to provide a chemical fingerprint of virions. The tested viruses, purified from infected plants, were chosen for their economic impact on horticultural crops and for their different morphological and structural features. Using the Raman-DEP device, specific profiles for each virus were successfully obtained, relying on chemical differences occurring even with genetically similar viruses belonging to the same taxonomic species and morphologically indiscernible by transmission electron microscopy (TEM). Moreover, we investigated the potentiality of Raman-DEP to follow dynamic changes occurring upon heat treatment of tobacco mosaic virus (TMV) particles. Raman peak deviations linked to TMV coat protein conformation were observed upon treatment at temperatures equal or higher than 85°C, substantiating the rod-to-spherical shape transitions observed by TEM and the concomitant drastic loss of infectivity following plant inoculation. Overall, the Raman-DEP method can be useful for the characterization of virus (nano)particles, setting the basis to create a database suitable for the study of viruses or virus derived-nanoparticles relevant for the agricultural, medical, or biotechnological fields.

This article presents an interlaboratory comparison (ILC) on Raman spectroscopy as a technique for relative quantification of the two most common polymorphs of titanium dioxide (TiO2)—anatase and rutile—in binary mixtures. Some standard methods are currently employed internationally for the determination of TiO2 content in samples (ISO 591-1, ASTM D3720-90), but require extensive sample preparation, do not distinguish between the two polymorphs or are accurate only for small fractions of either polymorph. Raman spectroscopy is a well-suited characterization technique for measuring and differentiating TiO2 in a fast, non-invasive way, while requiring no particular reagent or sample preparation. Eleven international participants conducted the study under the framework of Versailles Project on Advanced Materials and Standards. The collected data was analyzed by means of partial least squares regression after spectral preprocessing. The resulting models all show discrepancies of lower than 2% from the nominal values in the quantitative analysis over the concentration range of 5%–95% mixture fractions, with many datasets showing substantial improvement margins on this figure. The results of this ILC provide validation of Raman spectroscopy as a reliable method for quantification of TiO2 phases.

Walnut is considered the healthiest of all nuts, mainly because of its polyunsaturated fatty acid content. As far as its protein component is concerned, a rather interesting class of oil-body (OB) associated proteins, which, together with lipids, generates the "oleosome", still needs to be explored in detail. This study aimed to characterize the walnut oleosome proteins by a double off-gel and in-gel approach, with a focus on the effect of processing on protein profile and oleosin immune recognition. The off-gel approach (label free quantification, LFQ) allowed the identification of all the three oleosome specific associated class of proteins: oleosins, caleosins, and steroleosins. The in-gel separation coupled with immunoblotting experiments proved that walnut oleosins were immune recognized by walnut allergic patients' IgEs, providing first evidence of their potential role in walnut allergenicity. Oleosin immune recognition seemed to be increased by boiling procedure compared to raw and roasted walnut, probably due to the increased solubility of oleosome associated proteins in this condition.

The use of oak barrel alternatives, including oak chips, oak staves and oak powder, is quite common in the production of spirits obtained from the distillation of vegetal fermented products such as grape pomace. This work explored the use of unconventional wood formats such as peeled and sliced wood. The use of poplar wood was also evaluated to verify its technological uses to produce aged spirits. To this aim, GC-MS analyses were carried out to obtain an aromatic characterisation of experimental distillates treated with these products. Moreover, the same spirits were studied for classification purposes using NMR, NIR and e-nose. A significant change in the original composition of grape pomace distillate due to sorption phenomena was observed; the intensity of this effect was greater for poplar wood. The release of aroma compounds from wood depended both on the toasting level and wood assortment. Higher levels of xylovolatiles, namely, whisky lactone, were measured in samples aged using sliced woods. Both the NIR and NMR analyses highlighted similarities among samples refined with oak tablets, differentiating them from the other wood types. Finally, E-nose seemed to be a promising alternative to spectroscopic methods both for the simplicity of sample preparation and method portability.

In this work, a layer of graphene was used as a standard material for the measurement of the dimensions of Raman microscopes focal volumes of different confocal Raman spectrometers equipped with different objectives and excitation laser wavelengths. This method consists in probing the volume near the focal point of the system by using a flat graphene monolayer sheet with a straight edge. Graphene was selected because of its high Raman cross section and mechanically and chemically stability, allowing fast measurements and easy manipulation. In this paper, a method to employ graphene to accurately and precisely measure the three dimensions of the focal volume of a Raman microscope is presented; scanning along the axial and lateral directions, it is possible to reconstruct the three dimensions of the focal volume. Furthermore, these operations can be combined in a single procedure which allows the measurement of projections of the volume on planes parallel to the optical axis. Knowledge of these parameters enable absolute quantification of Raman-active molecules and support high-resolution Raman imaging.

In this work, biomolecules, such as membrane proteins, lipids, and DNA, were identified and their spatial distribution was mapped within a single Escherichia coli cell by Raman hyperspectral imaging. Raman spectroscopy allows direct, nondestructive, rapid, and cost-effective analysis of biological samples, minimizing the sample preparation and without the need of chemical label or immunological staining. Firstly, a comparison between an air-dried and a freeze-dried cell was made, and the principal vibrational modes associated to the membrane and nucleic acids were identified by the bacterium’s Raman chemical fingerprint. Then, analyzing the Raman hyperspectral images by multivariate statistical analysis, the bacterium biological status was investigated at a subcellular level. Principal components analysis (PCA) was applied for dimensionality reduction of the spectral data, then spectral unmixing was performed by multivariate curve resolution–alternating least squares (MCR-ALS). Thanks to multivariate data analysis, the DNA segregation and the Z-ring formation of a replicating bacterial cell were detected at a sub-micrometer level, opening the way to real-time molecular analysis that could be easily applied on in vivo or ex vivo biological samples, avoiding long preparation and analysis process.

In this work the quantification of antimicrobial properties of differently sized AgNPs immobilized on a surface was studied. Three different sizes of spheroidal AgNPs with a diameter of (6, 30 and 52) nm were synthetized and characterized with UV–vis, SEM, TEM and ICP-MS. The MIC (Minimal Inhibitory Concentration) and MBC (Minimal Bactericidal Concentration) against Escherichia coli were investigated. Then, the antibacterial efficacy (R) of amino-silanized glasses coated with different amounts of the three sizes of AgNPs were quantified by international standard ISO 22196 adapted protocol against E. coli, clarifying the relationship between size and antibacterial properties of immobilized AgNPs on a surface. The total amount of silver present on glasses with an R ∼ 6 for each AgNPs size was quantified with ICP-MS and this was considered the Surface MBC (SMBC), which were found to be (0.023, 0.026 and 0.034) μg/cm2 for (6, 30 and 52) nm AgNPs, respectively. Thus, this study demonstrates that active surfaces with a bactericidal effect at least ≥ 99.9999 % could be obtained using an amount of silver almost 100 times lower than the MBC found for colloidal AgNPs. The immobilization reduces the aggregation phenomena normally occuring in liquid media, maximizing the exposed specific superficial area of the AgNPs and their direct contact with bacterial cells. Starting from this glass model system, our work could broaden the way to the development of a wide range of antibacterial materials with very low amount of silver that can be safely applied in biomedical and food packaging fields.