Results of multi-actor collaboration in risk analysis: a simplified risk assessment toolkit for rapid detection of emerging risks.

The dynamic field of food safety faces continuous challenges, prompting stakeholders to develop collaborative actions for improved food safety systems. As part of these actions, the EU-FORA fellowship programme was dedicated to a multi-actor collaboration addressing risks of the unregulated mycotoxins T-2 and HT-2 toxins in oats. Critical gaps in risk assessment procedures were identified, leading to a joint effort to develop a strategy for rapid data collection and risk assessment, including the development of a risk assessment toolkit comprising of a training manual and two intuitive Microsoft® Excel files. The toolkit enables efficient data collection and processing, facilitating risk assessment calculations and rapid risk detection. Applying the toolkit to assess T-2 and HT-2 toxin risks in Belgian oats revealed minimal concerns, except for children aged 3-9 years, likely due to an overestimation. The toolkit is available on the FoodSafety4EU Platform and will be refined based on user feedback, promoting better risk assessment practices. This approach empowers stakeholders, from professionals to policymakers, fostering collaboration and enhancing food safety practices.

Polar pesticides in food of animal origin: interlaboratory validation to evaluate method fitness-for-purpose of official control.

The present work reports on the design, execution and evaluation of results of an interlaboratory validation study aimed at verifying the fitness-for-purpose of a LC-MS/MS method for the detection of polar pesticides in food of animal origin in official control and monitoring programmes. To this scope, five participant laboratories, with relevant expertise, were recruited. After passing a pre-trial test, the participants were asked to analyse test samples of bovine fat, chicken eggs and cow's milk, contaminated with 11 polar pesticides (group A: Aminomethyl phosphonic acid (AMPA), cyanuric acid, ethephon, glyphosate, fosetyl aluminium, 2-hydroxyethyphosphonic acid (HEPA), maleic hydrazide, N-acetyl-glyphosate, group B: N-acetyl glufosinate (NAG), 3-methylphosphinicopropionic acid (MPP) and glufosinate ammonium) at two different levels (0.05 and 0.25 mg/kg-1 and 0.01 and 0.05 mg/kg-1 for group A and B respectively. The method was based on acidified methanol/water extraction followed by dSPE clean up with C18 sorbent. For LC-MS/MS analysis isotopically labelled standards were used for all targeted analytes. With a couple of exceptions, average recoveries ranged from 85% to 110%, with repeatability (RSDr) ranging from 3% to 25%, and reproducibility (RSDR) from 4% to 26%. The assessment by different laboratories provided also insights on key factors impacting method performance characteristics and its implementation by new users.

Aflatoxin B1 Degradation by Ery4 Laccase: From In Vitro to Contaminated Corn.

Aflatoxins (AFs) are toxic secondary metabolites produced by Aspergillus spp. and are found in food and feed as contaminants worldwide. Due to climate change, AFs occurrence is expected to increase also in western Europe. Therefore, to ensure food and feed safety, it is mandatory to develop green technologies for AFs reduction in contaminated matrices. With this regard, enzymatic degradation is an effective and environmentally friendly approach under mild operational conditions and with minor impact on the food and feed matrix. In this work, Ery4 laccase, acetosyringone, ascorbic acid, and dehydroascorbic acid were investigated in vitro, then applied in artificially contaminated corn for AFB1 reduction. AFB1 (0.1 µg/mL) was completely removed in vitro and reduced by 26% in corn. Several degradation products were detected in vitro by UHPLC-HRMS and likely corresponded to AFQ1, epi-AFQ1, AFB1-diol, or AFB1dialehyde, AFB2a, and AFM1. Protein content was not altered by the enzymatic treatment, while slightly higher levels of lipid peroxidation and H2O2 were detected. Although further studies are needed to improve AFB1 reduction and reduce the impact of this treatment in corn, the results of this study are promising and suggest that Ery4 laccase can be effectively applied for the reduction in AFB1 in corn.

Industrial-Scale Cleaning Solutions for the Reduction of Fusarium Toxins in Maize.

Grain cleaning is the most effective non-destructive post-harvest mitigation strategy to reduce high levels of mycotoxins on account of the removal of mold-infected grains and grain fractions with high mycotoxin content. In this study, the reduction in the concentration of some co-occurring Fusarium toxins in maize, namely deoxynivalenol (DON), zearalenone (ZEA) and fumonisins B1 and B2 (FBs), was evaluated at an industrial-scale level by mechanical removal (sieving and density separation) of dust, coarse, small, broken, shriveled and low-density kernels and/or optical sorting of defected kernels. Samples were dynamically collected according to the Commission Regulation No. 401/2006 along the entire process line. Mycotoxin analyses of water-slurry aggregate samples were performed by validated LC methods. Depending on the contamination levels in raw incoming maize, the overall reduction rates ranged from 36 to 67% for DON, from 67 to 87% for ZEA and from 27 to 67% for FBs. High levels of DON, ZEA and FBs were found in all rejected fractions with values, respectively, up to 3030%, 1510% and 2680%, compared to their content in uncleaned maize. Results showed that grain cleaning equipment based on mechanical and or optical sorting technologies can provide a significant reduction in Fusarium toxin contamination in maize.

Mycotoxin Analysis of Grain via Dust Sampling: Review, Recent Advances and the Way Forward: The Contribution of the MycoKey Project.

The sampling protocols for the official control of the levels of mycotoxins in foodstuffs are very costly and time-consuming. More efforts are needed to implement alternative sampling plans able to support official control, or to adapt the current ones. The aim of the research carried out within the European Horizon 2020 MycoKey project was to evaluate the applicability at industrial scale of the dust sampling approach to detect multiple mycotoxins in grains. To this end, two trials were performed on an EU industrial site: (i) control of the unloading of wheat from train wagons; (ii) control of the unloading of wheat from trucks. In line with previous studies, the MycoKey results indicated that dust sampling and mycotoxin analysis represent a fitness for purpose approach for non-destructive and rapid identification of wheat commodities compliant to the maximum permitted levels. Based on reviewed and newly generated results, this article discusses potential applications and limits of the dust sampling methodology, identifying future research needs.

Introduction to This Special Issue of Toxins: Application of Novel Methods for Mycotoxin Analysis.

Crop contamination by mycotoxins is a global problem that poses significant economic burdens due to the food/feed losses that are caused by reduced production rates; the resulting adverse effects on human and animal health and productivity; and the trade losses associated with the costs incurred by inspection, sampling, and analysis before and after shipments [...].

Monitoring Alternaria toxins in Italian food to support upcoming regulation.

The collection of occurrence data on Alternaria toxins in food and feed across the European countries is required since 2012 by the European Commission, endorsing the relevant scientific opinion by the EFSA CONTAM Panel. Within this framework, occurrence data for Alternaria toxins (Alternariol, Alternariol monomethyl ether, Tenuazonic acid, Tentoxin, and Altenuene) in 97 samples of cereal foods, tomato products, and sunflower seeds have been provided as requested by the Italian national monitoring programme (years 2017-2020). To this purpose, an LC-MS/MS method was set up and validated, obtaining fit for purpose sensitivity, recoveries (70-120%), repeatability (≤20%) and within laboratory reproducibility (≤26%). Occurrence data showed that oilseeds were the most contaminated food group with levels of Tenuazonic acid up to 16752 µg/kg and Tentoxin up to 570 µg/kg, whereas for the other mycotoxin/commodities combinations, the percentage of left censored data (below the limit of quantification) ranged from 74 to 100%.

Undertaking a New Regulatory Challenge: Monitoring of Ergot Alkaloids in Italian Food Commodities.

The present manuscript reports on monitoring data of 12 ergot alkaloids (EAs) in cereal and cereal-derived products, collected in Italy over the period 2017-2020, for official control purposes under the edge of the Commission Recommendation 2012/154/EU on the monitoring of the presence of EAs in feed and food. To these purposes, an LC-MS/MS method was set up and applied, after in-house verification of its analytical performance. Besides satisfactory recoveries and precision, the method's quantification limits proved suitable to assess the compliance of cereals and cereal-based foods with the recently issued EU maximum permitted levels (Commission Regulation 2021/1399/EU). The validity of the generated data was also evaluated through the adoption of four proficiency tests, from which acceptable z-score values (-2 ≤ z ≤ 2) were obtained. The method was then applied to analyse a total of 67 samples, collected in Italy over the period 2017-2020. The samples consisted of 18 cereal grains, 16 flours (14 of wheat and 2 of spelt) and 31 other types of cereals derivatives (including 9 for infants). Overall, the EAs analysis returned a high percentage of left-censored data (>86%). Among the positive samples, the highest contamination levels, up to 94.2 µg/kg, were found for ergocristine (12% incidence), followed by ergocristinine (7% incidence) with levels of up to 48.3 µg/kg.

Determination of Zearalenone and Trichothecenes, Including Deoxynivalenol and Its Acetylated Derivatives, Nivalenol, T-2 and HT-2 Toxins, in Wheat and Wheat Products by LC-MS/MS: A Collaborative Study.

An analytical method for the simultaneous determination of trichothecenes-namely, nivalenol (NIV), deoxynivalenol (DON) and its acetylated derivatives (3- and 15-acetyl-DON), T-2 and HT-2 toxins-and zearalenone (ZEN) in wheat, wheat flour, and wheat crackers was validated through a collaborative study involving 15 participants from 10 countries. The validation study, performed within the M/520 standardization mandate of the European Commission, was carried out according to the IUPAC (International Union of Pure and Applied Chemistry) International Harmonized Protocol. The method was based on mycotoxin extraction from the homogenized sample material with a mixture of acetonitrile-water followed by purification and concentration on a solid phase extraction column. High-performance liquid chromatography coupled with tandem mass spectrometry was used for mycotoxin detection, using isotopically labelled mycotoxins as internal standards. The tested contamination ranges were from 27.7 to 378 µg/kg for NIV, from 234 to 2420 µg/kg for DON, from 18.5 to 137 µg/kg for 3-acetyl-DON, from 11.4 to 142 µg/kg for 15-acetyl-DON, from 2.1 to 37.6 µg/kg for T-2 toxin, from 6.6 to 134 µg/kg for HT-2 toxin, and from 31.6 to 230 µg/kg for ZEN. Recoveries were in the range 71-97% with the lowest values for NIV, the most polar mycotoxin. The relative standard deviation for repeatability (RSDr) was in the range of 2.2-34%, while the relative standard deviation for reproducibility (RSDR) was between 6.4% and 45%. The HorRat values ranged from 0.4 to 2.0. The results of the collaborative study showed that the candidate method is fit for the purpose of enforcing the legislative limits of the major Fusarium toxins in wheat and wheat-based products.

Application of an Integrated and Open Source Workflow for LC-HRMS Plant Metabolomics Studies. Case-Control Study: Metabolic Changes of Maize in Response to Fusarium verticillioides Infection.

Liquid chromatography coupled with high-resolution mass spectrometry (LC-HRMS) represents the most powerful metabolomics platform to investigate biological systems. Reproducible and standardized workflows allow obtaining a meaningful biological interpretation. The purpose of this study was to set up and apply an open-source workflow for LC-HRMS plant metabolomics studies. Key steps of the proposed workflow were as follows: (1) experimental design, (2) sample preparation, (3) LC-HRMS analysis, (4) data processing, (5) custom database search, (6) statistical analysis, (7) compound identification, and (8) biochemical interpretation. Its applicability was evaluated through the study of metabolomics changes of two maize recombinant inbred lines with contrasting phenotypes with respect to disease severity after Fusarium verticillioides infection of seedlings. Analysis of data from the case-control study revealed abundance change in metabolites belonging to different metabolic pathways, including two amino acids (L-tryptophan and tyrosine), five flavonoids, and three N-hydroxynnamic acid amides.

Critical Comparison of Analytical Performances of Two Immunoassay Methods for Rapid Detection of Aflatoxin M1 in Milk.

Aflatoxin B1 (AFB1) is a secondary metabolite produced by some Aspergillus spp. fungi affecting many crops and feed materials. Aflatoxin M1 (AFM1), the 4-hydroxylated metabolite of AFB1, is the main AFB1-related compound present in milk, and it is categorized by the International Agency for Research on Cancer (IARC) as a "group 1 human carcinogen". The aim of this work was to evaluate and compare the analytical performances of two commercial immunoassays widely applied for the detection of AFM1 in milk, namely strip test immunoassay and enzyme linked immunosorbent assay (ELISA). Assay validation included samples at AFM1 levels of 25, 50, 75 ng/kg and blank samples (AFM1 < 0.5 ng/kg). With respect to a screening target concentration (STC) of 50 ng/kg the two assays showed cut-off values of 37.7 ng/kg and 47.5 ng/kg for strip test and ELISA, respectively, a false suspect rate for blanks <0.1% (for both assays) and a false negative rate for samples containing AFM1 at levels higher than STC, of 0.4% (for both assays). The intermediate precision (RSDip) was <32% for the strip test and <15% for the ELISA. Method verification through long-term intra-laboratory quality control (QC) measurements confirmed the results from the validation study. Furthermore, a satisfactory correlation of the results obtained with both immunoassays and the AOAC Official Method 2000.08 was obtained for the analysis of cow milk samples naturally contaminated with AFM1 at levels within "not detected" (< 0.5 ng/kg) and 50 ng/kg. Finally, the extension of the scope of the strip test method to goat and sheep milk was evaluated by applying the experimental design foreseen in the EU regulation.

In Vitro Fumonisin Biosynthesis and Genetic Structure of Fusarium verticillioides Strains from Five Mediterranean Countries.

Investigating the in vitro fumonisin biosynthesis and the genetic structure of Fusarium verticillioides populations can provide important insights into the relationships between strains originating from various world regions. In this study, 90 F. verticillioides strains isolated from maize in five Mediterranean countries (Italy, Spain, Tunisia, Egypt and Iran) were analyzed to investigate their ability to in vitro biosynthesize fumonisin B1, fumonisin B2 and fumonisin B3 and to characterize their genetic profile. In general, 80% of the analyzed strains were able to biosynthesize fumonisins (range 0.03-69.84 µg/g). Populations from Italy, Spain, Tunisia and Iran showed a similar percentage of fumonisin producing strains (>90%); conversely, the Egyptian population showed a lower level of producing strains (46%). Significant differences in fumonisin biosynthesis were detected among strains isolated in the same country and among strains isolated from different countries. A portion of the divergent FUM1 gene and of intergenic regions FUM6-FUM7 and FUM7-FUM8 were sequenced to evaluate strain diversity among populations. A high level of genetic uniformity inside the populations analyzed was detected. Apparently, neither geographical origin nor fumonisin production ability were correlated to the genetic diversity of the strain set. However, four strains from Egypt differed from the remaining strains.

Rapid and reliable detection of glyphosate in pome fruits, berries, pulses and cereals by flow injection - Mass spectrometry.

A flow injection - mass spectrometry method for rapid glyphosate detection in food commodities was developed and validated. The sample preparation protocol included a simple and rapid extract purification step through polymeric solid phase extraction cartridges followed by addition of isotopically labeled glyphosate to the final test sample. The optimized method was subjected to intra-laboratory validation (spiking range 0.5-100 mg/kg) in chickpeas, grapes and apples, as representatives of three different commodity groups as defined in SANTE/11813/2017 guidelines. Recoveries were in the range 60-111%, repeatability and within laboratory reproducibility were ≤17%.The trueness of the results generated with the developed method was evaluated by analysis of a set of incurred chickpea and wheat samples (glyphosate range 0.5-36 mg/kg) and comparison with the reference method (Quick Polar Pesticides Method), confirming the method fitness-for-purpose of rapid compliance testing.

Fluorescence Polarization Immunoassay for the Determination of T-2 and HT-2 Toxins and Their Glucosides in Wheat.

T-2 and HT-2 toxins and their main modified forms (T-2 glucoside and HT-2 glucoside) may co-occur in cereals and cereal-based products. A fluorescence polarization immunoassay (FPIA) was developed for the simultaneous determination of T-2 toxin, HT-2 toxin and relevant glucosides, expressed as sum. The developed FPIA, using a HT-2-specific antibody, showed high sensitivity (IC50 = 2.0 ng/mL) and high cross-reactivity (100% for T-2 toxin and 80% for T-2 and HT-2 glucosides). The FPIA has been used to develop two rapid and easy-to-use methods using two different extraction protocols, based on the use of organic (methanol/water, 90:10, v/v) and non-organic (water) solvents, for the determination of these toxins in wheat. The two proposed methods showed analytical performances in terms of sensitivity (LOD 10 µg/kg) recovery (92-97%) and precision (relative standard deviations ≤13%), fulfilling the criteria for acceptability of an analytical method for the quantitative determination of T-2 and HT-2 toxins established by the European Union. Furthermore, the methods were then validated in accordance with the harmonized guidelines for the validation of screening methods included in the Regulation (EU) No. 519/2014. The satisfactory analytical performances, in terms of intermediate precision (≤25%), cut-off level (80 and 96 µg/kg for the two methods) and rate of false positives (<0.1%) confirmed the applicability of the proposed methods as screening method for assessing the content of these toxins in wheat at the EU indicative levels reported for T-2 and HT-2 toxins.

In-house validation and small-scale collaborative study to evaluate analytical performances of multimycotoxin screening methods based on liquid chromatography-high-resolution mass spectrometry: Case study on Fusarium toxins in wheat.

A strong trend toward using highly selective mass spectrometry technologies for screening of multiple mycotoxins has been observed in recent years. In the present study, the process of validation of a multimycotoxin screening method based on liquid chromatography-high-resolution mass spectrometry method is presented. The method was intended for the simultaneous screening of the major Fusarium toxins (deoxynivalenol, 3- and 15-acetyl deoxynivalenol, T-2 and HT-2 toxins, zearalenone, enniatins A, A1, B, and B1, and beauvericin) in wheat. The sample preparation protocol was based on a double extraction (methanol followed by acetonitrile/water mixture) and purification through solid-phase extraction C18 column. To provide insights for full exploitation of the potential of the double-stage high-resolution mass spectrometry detection, a full-scan acquisition event followed by a sequence of 5 fragmentation events (variable data-independent acquisition) was set for mycotoxin detection, the latter to be exploited for confirmatory purposes. Method analytical performances were evaluated through in-house validation and small-scale interlaboratory study, designed according to Commission Regulation 519/2014/EU, setting performance requirements for screening methods for mycotoxins. Screening target concentrations were close to European Union maximum permitted or indicative levels. The in-house validation provided the precision of the response under repeatability conditions and the intermediate precision (both resulting lower than 30%), the cutoff value, and the rate of false suspect results for negative (free of the mycotoxin of interest) samples, which resulted lower than 0.1% in all cases. The collaborative study provided reproducibility and laboratory independent cutoff values. Analysis of reference materials proved method trueness and suitability for screening of the major Fusarium mycotoxins in wheat. Finally, the applicability of the full-scan/variable data-independent acquisition detection approach was successfully tested on a set of naturally contaminated wheat samples, where 2 characteristic product ions could be detected for all identified mycotoxins even at levels in the low µg/kg range.

Occurrence of deoxynivalenol and deoxynivalenol-3-glucoside in durum wheat from Argentina.

The occurrence of deoxynivalenol, 3- and 15-deoxynivalenol and deoxynivalenol-3-glucoside in 84 durum wheat samples, from the Argentinean main growing area, was investigated during 2012/13 and 2013/14 using LC-MS/MS. Deoxynivalenol was found in all samples at concentrations varying between

Biophenols from Table Olive cv Bella di Cerignola: Chemical Characterization, Bioaccessibility, and Intestinal Absorption.

In this study, the naturally debittered table olives cv Bella di Cerignola were studied in order to (i) characterize their phenolic composition; (ii) evaluate the polyphenols bioaccessibility; (iii) assess their absorption and transport, across Caco2/TC7. LC-MS/MS analysis has confirmed the presence of hydroxytyrosol acetate, caffeoyl-6'-secologanoside, and comselogoside. In vitro bioaccessibility ranged from 7% of luteolin to 100% of tyrosol, highlighting the flavonoids sensitivity to the digestive conditions. The Caco2/TC7 polyphenols accumulation was rapid (60 min) with an efficiency of 0.89%; the overall bioavailability was 1.86% (120 min), with hydroxytyrosol and tyrosol the highest bioavailables, followed by verbascoside and luteolin. In the cells and basolateral side, caffeic and coumaric acids metabolites, probably derived from esterase activities, were detected. In conclusion, the naturally debittered table olives cv Bella di Cerignola can be considered as a source of bioaccessible, absorbable, and bioavailable polyphenols that, for their potential health promoting effect, permit inclusion of table olives as a functional food suitable for a balanced diet.

Parasitic weed management by using strigolactone-degrading fungi.

BACKGROUND: Seed germination is a key phase of the parasitic plant life cycle that is stimulated by the secondary metabolites, mainly strigolactones (SLs), secreted by the host roots. Interventions during this stage would be particularly suitable for parasitic weed management practices, as blocking these chemical signals would prevent seed germination and thus parasite attack. Four fungal strains with different ecological functions were considered for their possible ability to metabolise SLs: Fusarium oxysporum and F. solani, biocontrol agents of Phelipanche ramosa; Trichoderma harzianum, a potential biopesticide; Botrytis cinerea, a phytopathogenic fungus. Four different SLs [the natural strigol, 5-deoxystrigol (5DS) and 4-deoxyorobanchol (4DO), and the synthetic analogue GR24] were added to fungal cultures, followed by determination of the SL content by liquid chromatography-tandem mass spectrometry. RESULTS: Differences were observed among microorganisms, treatments and SLs used. T. harzianum and F. oxysporum were the most capable of reducing the SL content; considering the whole set of fungi used, 5DS and 4DO proved to be the most degradable SLs. CONCLUSIONS: Beneficial microscopic fungi could differently be used for biocontrolling parasitic weeds, acting as a 'physiological' barrier, by preventing the germination of their seeds through the ability to biotransform the stimulatory signals. © 2016 Society of Chemical Industry.

Study of the natural occurrence of T-2 and HT-2 toxins and their glucosyl derivatives from field barley to malt by high-resolution Orbitrap mass spectrometry.

This paper reports a new method for the determination of T-2 and HT-2 toxins and their glucosylated derivatives in cereals, and some survey data aimed at obtaining more comprehensive information on the co-occurrence of T-2 and HT-2 toxins and their glucosylated derivatives in naturally contaminated cereal samples. For these purposes, barley samples originating from a Northern Italian area were analysed by LC-HRMS for the presence of T-2, HT-2 and relevant glucosyl derivatives. Quantitative analysis of T-2 and HT-2 glucosides was performed for the first time using a recently made available standard of T-2 glucoside. The glucosyl derivative of HT-2 was detected at levels up to 163 µg kg(-1) in 17 of the 18 analysed unprocessed barley grains, whereas the monoglucosyl derivative of T-2 toxin was detected in only a few samples and at low µg kg(-1) levels. The ratio between glucosylated toxins (sum of T-2 and HT-2 glucosides) and native toxins (sum of T-2 and HT-2) ranged from 2% to 283%. Moreover, taking advantage of the possibility of retrospective analysis of full-scan HRMS chromatograms, samples were also screened for the presence of other type-A trichothecenes, namely neosolaniol, diacetoxyscirpenol and their monoglucosyl derivatives, which were detected at trace levels. A subset of nine different samples was subjected to micro-maltation in order to carry out a preliminary investigation on the fate of T-2, HT-2 and relevant glucosides along the malting process. Mycotoxin reduction from cleaned barley to malt was observed at rates ranging from 4% to 87%.

Improved method for the simultaneous determination of aflatoxins, ochratoxin A and Fusarium toxins in cereals and derived products by liquid chromatography-tandem mass spectrometry after multi-toxin immunoaffinity clean up.

An improved method for the quantitative determination of aflatoxins (B1, B2, G1, G2), ochratoxin A, fumonisins (B1, B2), zearalenone, deoxynivalenol, nivalenol, T-2 and HT-2 toxins in cereals and derived products, at levels comparable with EU maximum permitted levels, was developed. The effective co-extraction of the mycotoxins under investigation was achieved in 4min by a double extraction approach, using water followed by methanol. Clean up of the extract was performed by a new multi-toxin immunoaffinity column. Analytical performance characteristics were evaluated through single laboratory validation. Raw wheat and maize, corn flakes and maize snacks were chosen as representative matrices for method validation. The validation assay was carried out at 50, 100 and 150% of EU maximum permitted levels for each mycotoxin. Statistical analysis of the results (ANOVA) provided the within laboratory reproducibility and the error contributions from repeatability, between day effects, and influences from different matrix composition. Recoveries generally higher than 70% were obtained for all tested mycotoxins with relative standard deviation (within laboratory reproducibility) lesser than 37%. Limits of quantification (calculated as the lowest amount of each analyte which could be determined with a precision of 10%) ranged from 1µg/kg to 30µg/kg. The trueness of generated data was assessed by analysis of reference materials. The proposed method was proven to be suitable to assess, with a single analysis, compliance of the selected cereal based foods with the EU maximum permitted or recommended levels for all regulated mycotoxins.