Non-intentionally Added Substances Research Articles

Hazard assessment of fenozan, a released non-intentionally added substance from polyester-based can coating

Since the safety of new-generation polyester-based internal coatings regarding the migration of non-intentionally added substances (NIAS) is poorly documented, studies are needed to identify NIAS originating from these food-contact materials (FCM). The aim of this study was to identify volatile and semi-volatile NIAS from polyester-based coatings in order to assess their hazard and ensure consumers’ safety with regard to exposure from canned food. Extraction and migration tests were carried out on a single polyester-coated tin plate (5 batches) using two solvents: acetonitrile and ethanol 95%, then FCM’s extracts and migrates were analysed by GC-MS. An antioxidant degradation (hydrolysis) product, 3-(3,5-di-tert-butyl-4-hydroxyphenyl)propionic acid or fenozan (CAS RN: 20170-32-5), was identified and confirmed by reference standard in all migrates. To assess fenozan’s toxicity, several in vitro bioassays, such as the Ames test (to assess point mutation), the micronucleus assay (to detect chromosomal aberrations), and the iodide uptake assay (to study one mode of action for thyroid disruption) were conducted. Fenozan was negative in the Ames test on three strains of S. typhimurium (TA98, TA100, and TA1535) and on one strain of E.coli (WP2), with and without metabolic activation system (S9 mix) using direct incorporation and pre-incubation methods. The in vitro micronucleus assay conducted on HepG2 cells also exhibited a negative response following a 4-hour treatment with the S9 mix, and a 48-hour treatment without the S9 mix. A weak inhibitory effect was obtained when testing fenozan in the iodide uptake assay using rat thyroid FRTL-5 cells. Significant inhibition started from 800 µM of fenozan, with a maximal inhibition of almost 47% at 1000 µM. The findings indicate that fenozan exhibits an anti-thyroid activity in vitro.

Advancing Analytical Techniques in PET and rPET: Development of an ICP-MS Method for the Analysis of Trace Metals and Rare Earth Elements.

Despite the extensive use of recycled polyethylene terephthalate (rPET) in food contact materials (FCMs), research on the presence of heavy metals (HMs) and rare earth elements (REEs) during various recycling stages (e.g., flakes, granules, and preforms) remains limited. This study aimed to address these gaps by validating a rapid and sensitive analytical method to quantify 26 HMs and 4 REEs in PET and rPET matrices. An ICP-MS method was validated per EURACHEM guidelines, assessing linearity, limits of detection (LOD), limits of quantification (LOQ), accuracy, and repeatability. The method was employed for initial screening of HMs and REEs classified as non-intentionally added substances (NIASs) in PET and rPET samples. The findings showed high accuracy and reliability, with recovery rates between 80% and 120%. Analysis revealed varying concentrations of HMs and REEs, with the highest levels in 100% rPET preforms, notably Zn, Cu, and Al among HMs, and La among REEs. The study identified critical contamination points during the recycling process, highlighting the need for targeted interventions. This research provides a crucial analytical framework for assessing HMs and REEs in PET and rPET, ensuring FCM safety compliance and supporting efforts to enhance rPET product safety, promoting public health protection and advancing the circular economy.

Novel selection of recycled PET surrogates based on non-targeted screening of non-intentionally added substances and chemometrics

Waste PET is utilized to produce recycled PET (rPET) through bottle-to-bottle recycling in order to reduce environmental pollution. However, non-intentionally added substances (NIAS) may be generated during rPET production, potentially posing a threat to food safety. In this study, a non-target screening of 13 batches with a total of 39 rPET samples was performed using GC-QTOF-MS and 240 compounds were tentatively identified. It was worth noting that the detection of phthalate esters in rPET from China, such as bis(2-ethylhexyl) phthalate, exhibits a higher diversity (17 different types) and frequency (7.8 %–100 %) in comparison with related studies. Subsequently, based on the principles for selection of surrogates used in challenge test, seven molecular descriptors were selected as variables for compound characterization, followed by principal component analysis (PCA) for representation. Based on this analysis and referencing the EFSA and FDA surrogate suggestion lists, eight surrogates were proposed for China, namely chlorobenzene, toluene, benzophenone, bis(2-ethylhexyl) terephthalate, methyl salicylate, methyl stearate, 1-phenyldecane and phenylcyclohexane. High-throughput detection methods were developed for each surrogate with the regression correlation coefficients over 0.999, spiked recoveries ranging from 99.3-107.5 %, LODs of 0.003–2.192 ng/mL, LOQs of 0.009–7.306 ng/mL, RSD of 0.71–4.23 %, respectively. This research provides foundational data and methodological support for the evaluation of rPET challenge testing in China, offering technical support for subsequent safety assessments of rPET.

Designing safe recycled high-density polyethylene (HDPE) for child toys

New high-density polyethylene (HDPE) manufactured from different percentage of post-consumer recycled HDPE milk bottles was studied through two static and dynamic migration tests using saliva simulant to assess the potential hazard to children. Sixty-nine compounds were identified, including several additives used in PE synthesis such as alkanes, alkenes, antioxidants and plasticizers as well as non-intentionally added substances (NIAS) like degradation products such as 2,6-di-tert-butyl-1,4-benzoquinone, 2,4-di-tert-butylphenol, phenol, 2,5-bis(1,1-dimethylethyl)-, 3,5-di-tert-butyl-4-hydroxybenzaldehyde, and 3,5-di-tert-butyl-4-hydroxyacetophenone, or various residues from flavoring agents, cleaning products and essential oils. Some of these compounds as the isomers p and o t-butylcyclohexyl acetate, 3-Octanol, 3,7-dimethyl- and thujanol acetate (3-) pose a potential risk to children, as their concentrations exceed the recommended Cramer values for high percentages of recycling. This suggests improving recycling processes by incorporating advanced cleaning to remove residual products and contaminants.

Mass spectral characterisation of cyclic oligoesters in a biodegradable mulch film.

Plastic mulch film manages weed growth and moisture loss on the surface of cropping beds. The chemical components of such plastics include polymer(s), additives and non-intentionally added substances (NIASs). The unknown chemical nature and behaviours of these constituents require investigation due to their potential to add to the anthropogenic chemical burden in the agrifood system. Solvent extracts of a commercial 15% polylactic acid (PLA)/85% poly(butylene adipate-co-terephthalate) mulch film were investigated using gas chromatography-mass spectrometry (GC-MS) with electron ionisation to characterise the additive and NIAS components. The obscurity of some of the NIASs meant their identification was not readily achieved through routine MS library comparisons. As such, the identification of several polymer-derived compounds required interpretation of the MS data and re-application of the derived fragmentation patterns with reference to the wider literature. Unknowns were confirmed using commercially available compounds. Unknown NIASs were identified as cyclic oligoesters comprised of the monomeric building blocks of the polymer system. Cyclic structures derived from the monomers of polybutylene adipate (PBA) and polybutylene terephthalate (PBT) fragmented through a primary pathway involving 1,5- and 1,3-H transfers at ester linkages. Characteristic ions at m/z 111, 129, 183 and 201 for PBA-derived cyclic oligoesters and m/z 104, 132, 149 and 221 for PBT-derived cyclic oligoesters were assigned in the mass spectra of unknowns. Cyclic oligoesters containing sebacate moieties were also identified, indicating the presence of polybutylene sebacate as an unexpected component of the mulch. Systematic analyses of the sort reported here are valuable for providing alternative approaches for the identification of plastic-related chemicals. Open publication of MS spectral data is required to build a greater understanding of the mulch film chemical components contributing to the environmental chemical load introduced to agroecosystems.

Strategy proposal using QSAR models to approach mutagenicity assessment of non intentionally added substances in recycled plastic resins

ContextTransition to the use of recycled plastics raises an issue concerning safety assessment of Non Intentionally Added Substances (NIAS). To assess the mutagenic potential of the recycled polyethylene impurities and to evaluate the need to perform in vitro assays on recycled resins, this study lies in identifying existing NIAS associated with recycled Low/High Density Polyethylene and assessing the mutagenicity data-gaps by employing in silico tools. MethodsQuantitative Structure-Activity Relationship (QSAR) models predicting Ames mutagenicity were selected from literature, then NIAS were run to 1/evaluate performances of each model, 2/apply a QSAR strategy on the NIAS molecular space and address data-gaps. ResultsAmong the 165 NIAS identified, experimental Ames results were not found for 50 substances while the substances with experimental data were predominantly negatives. No individual model was able to predict all NIAS due to applicability domain limitations. Taking into account 1/calculated performances, 2/availability of applicability domain, 3/description of the Training Set, an Integrated Strategy was founded including Sarpy, Consensus and Protox to extend the applicability domain. Conclusion & perspectivesExisting data and predictions generated by this strategy suggest a low mutagenic potential of NIAS. Further investigation is needed to explore other genotoxicity mechanisms.

Assessing the Conformity of Plasticizer-Free Polymers for Foodstuff Packaging Using Solid Phase Microextraction Coupled to Gas Chromatography/Mass Spectrometry

Phthalates are the synthetic chemical plasticizers with the most varied uses and are a source of concern due to their toxicity and ubiquity, so much so that even plasticizer-free polymers can contain them as non-intentionally added substances (NIAS). Food packaging is among the materials with the greatest impact. In this study, a simple protocol is proposed for the location and identification of dimethyl phthalate, diethyl phthalate, dipropyl phthalate, and dibutyl phthalate which is applicable to compliance studies of food packaging materials and for the associated risk assessment. Solid phase microextraction gas chromatography/mass spectrometry was used to evaluate the migration of four NIAS from food packaging to release media simulating food substrates. Three plasticizer-free polymers were used: two that were lab-made and based on sodium alginate and a commercial polyethylene film. Linearity ranged from the LOQ to 10 µg/mL; within-day and between-day precision values were between 12.3–25.7% and 21.9–35.8%, respectively; the LOD and LOQ were in the range 0.029–0.073 µg/mL and 0.122–0.970 µg/mL. Migration tests were conducted for different periods of time at room temperature and at 8 °C. Exposure to microwaves (MW) was also evaluated. All packaging materials tested had global migration limits lower than 10 mg/dm2 of material surface.

Composition and release rates of chemicals in inkjet fabrics determined by non-targeted screening and targeted analysis

Unveiling composition and release rates of chemicals in chemical-intensive products (CIPs) such as inkjet fabrics that are applied extensively in advertising and publicizing industries, is of importance to sound management of chemicals. This study tentatively identified 212 compounds from 69 inkjet fabric samples using gas chromatograph coupled with quadrupole time-of-flight mass spectrometry (GC-QTOF-MS). Contents of six phthalate esters (PAEs) were quantified to range from 3.0 × 102 mg/kg to 3.1 × 105 mg/kg with GC-MS. Bis(2-ethylhexyl) phthalate was predominantly detected to average 96 g/kg. The inkjet fabrics collected from southern China contained fewer non-intentionally added substances (NIASs) than from northern China. Annual mass release rates (RM) of the 6 PAEs from inkjet fabrics to air were estimated to range from 1.4 × 10−2 kg/year to 2.8 × 104 kg/year in China in 2020, and the mean indoor RM was comparable with the outdoor one. Equilibrium partition coefficients of the compounds between the product and air, ambient temperature, and concentrations of chemicals in the product, are key factors leading to RM with the high variance. The findings indicate that contents of the NIASs in the CIPs should be minimized, and the refining concept should be adopted in design of the CIPs, so as to control the release of chemicals from the CIPs.

Untargeted screening of NIAS and cyclic oligomers migrating from virgin and recycled polyethylene terephthalate (PET) food trays

Polyethylene terephthalate is considered one of the most used and approved recycled plastic polymer as food contact materials (FCM); however, the fundamental details arising from the recycled content of PET trays in relation to processing conditions or extent of recycling are not well understood. In the present work, the presence and migration of different series of PET cyclic oligomers from industrially recycled trays were investigated using official food simulant D1 at 40 °C for 10 days. The trays were produced in two different facilities, using different extrusion processing technology, namely single and twin-screw extruder. Tested food packaging trays were produced from different ratios of virgin (vPET) and recycled PET (rPET), varying from 0% up to 100% w/w content. Analysis of the trays using MALDI-TOF-MS revealed a decrease in the molecular weight distribution (Mw) in rPET compared to virgin PET plastic. Untargeted screening with UHPLC-QTOF-MS, revealed the presence and migration of several intentionally added substances (IAS) as well as non-intentionally added substances (NIAS). The results demonstrated differences of the cyclic oligomers migration into the selected food simulant, depending on the rPET/vPET ratios as well as among the applied extrusion technology applied. Following multivariate analysis, marker compounds found to be benzyl butyl phthalate (BBP), butyl lactate, dodecenamide, 2-methyl-1,3-dioxolane, PET 2nd series cyclic dimer as well as PET 1st series cyclic dimer can be considered as marker compounds.

Understanding intentionally and non-intentionally added substances and associated threshold of toxicological concern in post-consumer polyolefin for use as food packaging materials

The use of post-consumer recycled (PCR) polymers in food contact materials (FCMs) can facilitate achieving a circular economy by reducing environmental waste and landfill accumulation. This study aimed to identify potentially harmful substances, including non-intentionally added substances (NIAS) and unapproved intentionally added substances (IAS), in polyolefin samples from material recovery facilities using gas-chromatography mass-spectrometry. Selected phthalates and bisphenols were quantified by targeted gas-chromatography tandem mass-spectrometry. The analysis detected 9 compounds in virgin polymers and 52 different compounds including alcohols, hydrocarbons, phenols in virgin and hydrocarbons, aromatic, phthalates, organic acids, per- and polyfluoroalkyl substances (PFAS) in PCR polymers. The Cramer classification system was used to assesses the Threshold of Toxicological Concern associated with the detected compounds. The PCR sample showed a slightly higher proportion of Cramer Class III compounds (48.08 %) than the virgin sample (44.44 %), indicating higher toxicity potential. Quantification detected bisphenols only in PCR material including BPA (2.88 ± 0.53 μg/g), BPS (5.12 ± 0.003 μg/g), BPF (3.42 ± 0.01 μg/g), and BADGE (4.638 μg/g). Phthalate concentrations were higher in PCR than virgin samples, with the highest levels detected as DIDP, at 6.18 ± 0.31 μg/g for PCR and 6.04 ± 0.02 for virgin. This study provides critical understanding of the safety and potential risks associated with using PCR polyolefins from different sources in food contact applications.

Determination of volatile migrants from breast milk storage bags

Breast milk storage bags are commonly used to facilitate extended periods of breastfeeding. These bags must meet the necessary safety standards for infant food and comply with current regulations. As they are typically made from polyethylene and feature printing inks on their surface, there is a potential for the transfer of polymer and ink additives, as well as non-intentionally added substances (NIAS), into the milk. An initial study was conducted using two breast milk bags to identify the primary migrants that could be released from this particular food packaging. Migration tests were conducted in both milk and the corresponding food simulant D1 (50% ethanol/water, v/v) under conditions mimicking typical breast milk storage conditions (4–5ºC, 7 days) and defrosting conditions (40ºC, 2 h). The results showed the presence of volatile migrants in both situations, whether long storage periods at refrigerated conditions or brief periods if the temperature is not under control. The outcomes also highlighted that the primary migrants were breakdown products from Irgafos 168, a widely used trisarylphosphite-based antioxidant. These included 2,4-ditertbutylphenol (2,4-dtBP), 1,3-di-tert-butylbenzene (1,3-dtBB), and 2,6-di-tert-butyl-1,4-benzoquinone (2,6-dtBBQ). A subsequent study was carried out involving eight breast milk bags to evaluate the migration of these compounds and to conduct a comprehensive risk assessment of the materials used, ensuring the safety of infants' health is not compromised. Results showed that 1,3-dtBB exceeded the maximum recommended migration values in two of the studied bags. Nevertheless, further toxicity tests are necessary to ensure the safety of these materials in relation to food.

A vision for safer food contact materials: Public health concerns as drivers for improved testing

Food contact materials (FCMs) and food contact articles are ubiquitous in today’s globalized food system. Chemicals migrate from FCMs into foodstuffs, so called food contact chemicals (FCCs), but current regulatory requirements do not sufficiently protect public health from hazardous FCCs because only individual substances used to make FCMs are tested and mostly only for genotoxicity while endocrine disruption and other hazard properties are disregarded. Indeed, FCMs are a known source of a wide range of hazardous chemicals, and they likely contribute to highly prevalent non-communicable diseases. FCMs can also include non-intentionally added substances (NIAS), which often are unknown and therefore not subject to risk assessment. To address these important shortcomings, we outline how the safety of FCMs may be improved by (1) testing the overall migrate, including (unknown) NIAS, of finished food contact articles, and (2) expanding toxicological testing beyond genotoxicity to multiple endpoints associated with non-communicable diseases relevant to human health. To identify mechanistic endpoints for testing, we group chronic health outcomes associated with chemical exposure into Six Clusters of Disease (SCOD) and we propose that finished food contact articles should be tested for their impacts on these SCOD. Research should focus on developing robust, relevant, and sensitive in-vitro assays based on mechanistic information linked to the SCOD, e.g., through Adverse Outcome Pathways (AOPs) or Key Characteristics of Toxicants. Implementing this vision will improve prevention of chronic diseases that are associated with hazardous chemical exposures, including from FCMs.

Evaluation and identification of chemical migrants leached from baby food pouch packaging

Multilayer plastic pouch (MLPP) has an estimated 9% annual increase of market growth. However, the migrants it can introduce to food has not yet been systematically studied. A total of 79 MLPPs for baby food were purchased from major retail outlets in Australia. The methodology for testing chemical migration followed the design of previous studies using four types of selected simulants according to the European Committee Regulation No. 10/2011 method. Four bisphenols and five phthalic acid diesters (PAEs) were detected, including the ones known for endocrine disrupting effect in human. Three intentionally added and 23 non-intentionally added substances (NIAS) were tentatively identified through a suspect screening procedure. Out of the 23 NIAS, neopentyl glycol – phthalic acid – 1,6-hexanediol – phthalic acid oligomer was identified for the first time with MLPP. A further two NIAS were detected for the first time in baby food related products. For 40% of the pouches where adipic acid - diethylene glycol was detected, the estimated exposure from consuming one pouch of food per day may exceed the threshold of toxicological concern established based on the Cramer classification.

Analysis of Migrant Cyclic PET Oligomers in Olive Oil and Food Simulants Using UHPLC-qTOF-MS.

Oligomers are a particular category of non-intentionally added substances (NIAS) that may be present in food contact materials (FCMs), such as polyethylene terephthalate (PET), and consequently migrate into foods. Here, an ultra-high-pressure liquid chromatography quadruple time-of-flight mass spectrometry (UHPLC-qTOF-MS) method was developed for the analysis of 1st series cyclic PET oligomers in virgin olive oil (VOO) following a QuEChERS clean-up protocol. Oligomer migration was evaluated with two different migration experiments using bottles from virgin and recycled PET: one with VOO samples stored in household conditions for a year and one using the food simulant D2 (95% v/v ethanol in water) at 60 °C for 10 days. Calibration curves were constructed with fortified VOO samples, with the LOQs ranging from 10 to 50 µg L-1 and the recoveries ranging from 86.6 to 113.0%. Results showed no migration of PET oligomers in VOO. However, in the simulated study, significant amounts of all oligomers were detected, with the migration of cyclic PET trimers from recycled bottles being the most abundant. Additional substances were tentatively identified as linear derivatives of PET oligomers. Again, open trimer structures in recycled bottles gave the most significant signals.

Authentication of recycled and virgin polyethylene terephthalate based on UPLC-Q-TOF-MS using non-volatile organic compounds and chemometrics

Plastic packaging waste, such as polyethylene terephthalate (PET) has increased significantly in recent decades, arousing a considerable and serious public concern regarding the environment, economy, and policy. Plastic recycling is a useful tool to mitigate this issue. Here, a feasible study was performed to investigate the potential of a novel method for identifying virgin and recycled PET. Ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) was combined with various chemometrics, as a simple and reliable method that achieved a high discrimination rate for 105 batches of virgin PET (v-PET) and recycled PET (r-PET) based on 202 non-volatile organic compounds (NVOCs). Making use of orthogonal partial least-squares discrimination analysis (OPLS-DA) together with non-parametric tests, 26 marker compounds (i.e. 12 intentionally added substances (IAS) and 14 non-intentionally added substances (NIAS) as well as 31 marker compounds (i.e. 11 IAS and 20 NIAS) obtained from positive and combination of positive and negative ionization modes of UPLC-Q-TOF-MS, respectively, were successfully identified. Moreover, 100% accuracy was obtained using a decision tree (DT). Cross-discrimination based on misclassified samples using various chemometrics allowed the prediction accuracy to be improved and to identify a large sample set, thus greatly enhancing the application scope of this method. The possible origins of these detected compounds can be the plastic itself, as well as contamination from food, medicine, pesticides, industry-related substances, and degradation and polymerization products. As many of these compounds are toxic, especially those pesticide related, this indicates an urgent requirement for closed loop recycling. Overall, this analytical method provides a quick, accurate, and robust way to distinguish virgin from recycled PET and thus addresses the issue of potential virgin PET adulteration thereby detecting fraud in the area of PET recycling.

Mass Spectrometry-Based Techniques for the Detection of Non-Intentionally Added Substances in Bioplastics

The safety of food contact materials is a hot topic since chemicals can migrate from packaging into food, thus raising health concerns about and/or producing changes in the organoleptic properties of foodstuffs. Migration tests are required to demonstrate the compliance with current regulations and to investigate the transferred compounds. In this context, mass spectrometry is the analytical technique of choice for the detection and quantitation of both intentionally added substances, such as antioxidants, stabilizers, processing aids, and non-intentionally added substances (NIAS). Untargeted strategies represent a major analytical challenge, providing a comprehensive fingerprinting of the packaging material and migrating components, allowing for NIAS identification. Hyphenated mass spectrometry-based techniques have been devised for screening the presence of migrating contaminants and for quantitation purposes. Both low-resolution (LRMS) and high-resolution (HRMS) methods were screened, with a special emphasis on the latter because of its capability to directly characterize food contact materials with minimal/no sample preparation, avoiding chromatographic separation, and reducing sample handling, analysis costs, and time. Examples related to the migration of contaminants from existing or newly developed bioplastic materials will be discussed, providing an overview of the most used MS-based methods, covering the state-of-the-art approaches from 2012 up to 2022.

Identification of potential migrants from epoxy and organosol coatings used in metal food cans

The coatings of metal cans may release complex mixtures of migrants into the contained foods, including non-intentionally added substances (NIAS), such as reaction products. All migrating substances should be studied to demonstrate their safety. In this work, the characterisation of two epoxy and organosol coatings was performed using several techniques. Firstly, the type of coating was identified using FTIR-ATR. Screening techniques based on purge and trap (P&T) and solid-phase microextraction (SPME) coupled to GC-MS were used to investigate volatiles from coatings. For the identification of semi-volatile compounds, an appropriate extraction was performed before analysis by GC-MS. The most abundant substances were compounds with at least one benzene ring and an aldehyde or alcohol group in their structures. Furthermore, a method to quantify some of the identified volatiles was explored. Secondly, HPLC with fluorescence detection (HPLC-FLD) was used to determine non-volatile compounds such as bisphenol analogues and bisphenol A diglycidyl ethers (BADGEs), with subsequent confirmation by LC-MS/MS. Additionally, migration assays were performed by this technique to determine non-volatile compounds migrating into food simulants. Bisphenol A (BPA) and all BADGE derivatives except BADGE.HCl were detected in the migration extracts. Moreover, BADGE-solvent complexes such as BADGE.H2O.BuEtOH, BADGE.2BuEtOH, etc. were also tentatively identified using the accurate mass provided by time-of-flight mass spectrometry (TOF-MS).

Characterization of a new polymeric food contact coating with emphasis on the chemical analysis and safety assessment of non-intentionally added substances (NIAS).

Regulators have established safety requirements for food packaging raw materials and finished products, including by-products of polymer synthesis known as non-intentionally added substances (NIAS). However, there are no official guidance or regulations for best practices to evaluate the safety of NIAS. Here we described the process we followed to identify, characterize, and prioritize for safety assessment low molecular weight NIAS from an epoxy coating (V70) made with tetramethyl bisphenol F-based diglycidyl ether resin (TMBPF-DGE). We assembled a database of 15000 potential oligomers with masses up to 1000Da and conducted extraction and migration testing of V70 coating. Acetonitrile extract contained higher number and concentration of substances compared to ethanolic-based food simulants. The extract contained 16 substances with matches in the database with estimated concentration of 18.27 μg/6dm2; seven of these substances have potentially genotoxic oxirane functionality. TMBPF-DGE+hydroquinone (TMBPF-DGE+HQ) was most abundant (55% of total concentration) and was synthesized and prioritized for safety assessment. TMBPF-DGE+HQ exposure from can beverage was estimated at 5.2 μg/person/day, and it was not mutagenic or genotoxic in in vitro assays. The overall mixture of substances that migrated into ethanolic simulant was also negative in the mutagenicity bioassay. Our findings suggest that exposure to TMBPF-DGE+HQ from the V70 coating is exceedingly small and that the coating migrates are not genotoxic.

Cytotoxicity, endocrine disrupting activity, and chemical analysis of 42 food contact silicone rubber products

After migration in 95 % ethanol (food simulant) at 70 °C for 2 h (accelerated conditions), the cytotoxicity and endocrine-disruption activity of 42 food contact silicone products (FCSPs) obtained from the Chinese market were studied. Of 31 kitchenwares, 96 % showed mild or above cytotoxicity (relative growth rate < 80 %) using the HeLa neutral red uptake test; and 84 % showed estrogenic (64 %), anti-estrogenic (19 %), androgenic (42 %), and anti-androgenic (39 %) activities by the Dual-luciferase reporter gene assay. The mold sample induced late phase HeLa apoptosis as detected by Annexin V-FITC/PI double staining flow cytometry, in addition, the migration of mold sample has a higher risk of endocrine disruption at high temperature usage. Encouragingly, 11 bottle nipples had neither cytotoxic nor hormonal activity. Utilizing multiple mass spectrometry techniques, non-intentionally added substances (NIASs) in 31 kitchenwares were analyzed, and the migration levels of 26 organic compounds and 21 metals were quantified, furthermore, the safe risk of single migrant was evaluated through their special migration limit (SML) or threshold of toxicological concern (TTC). Using “nchoosek” statement and Spearman's correlation analysis in MATLAB, the migration of 38 compounds or combinations including metals, plasticizers, methylsiloxanes, and lubricants, had strong correlation with cytotoxicity or hormonal activity. The coexistence of various chemical substances in migrants leads to complex biological toxicity of FCSPs, so it is very important to detect the toxicity of the final products. The combination of bioassays and chemical analyses are valuable tools to facilitate the identification and analyses of FCSPs and migrants that have potential safety risks.

Interlaboratory Study to Evaluate a Testing Protocol for the Safety of Food Packaging Coatings.

According to European regulations, migration from food packaging must be safe. However, currently, there is no consensus on how to evaluate its safety, especially for non-intentionally added substances (NIAS). The intensive and laborious approach, involving identification and then quantification of all migrating substances followed by a toxicological evaluation, is not practical or feasible. In alignment with the International Life Sciences Institute (ILSI) and the European Union (EU) guidelines on packaging materials, efforts are focused on combining data from analytics, bioassays and in silico toxicology approaches for the risk assessment of packaging materials. Advancement of non-targeted screening approaches using both analytical methods and in vitro bioassays is key. A protocol was developed for the chemical and biological screening of migrants from coated metal packaging materials. This protocol includes guidance on sample preparation, migrant simulation, chemical analysis using liquid chromatography (LC-MS) and validated bioassays covering endocrine activity, genotoxicity and metabolism-related targets. An inter-laboratory study was set-up to evaluate the consistency in biological activity and analytical results generated between three independent laboratories applying the developed protocol and guidance. Coated packaging metal panels were used in this case study. In general, the inter-laboratory chemical analysis and bioassay results displayed acceptable consistency between laboratories, but technical differences led to different data interpretations (e.g., cytotoxicity, cell passages, chemical analysis). The study observations with the greatest impact on the quality of the data and ultimately resulting in discrepancies in the results are given and suggestions for improvement of the protocol are made (e.g., sample preparation, chemical analysis approaches). Finally, there was agreement on the need for an aligned protocol to be utilized by qualified laboratories for chemical and biological analyses, following best practices and guidance for packaging safety assessment of intentionally added substances (IAS) and NIAS to avoid inconsistency in data and the final interpretation.