Alternaria toxins (ATs) are a group of toxins produced by Alternaria fungi that frequently contaminate tomatoes and tomato products. Recently, the European Food Safety Authority evaluated ATs for their genotoxic and carcinogenic properties. Alternaria infestation is often controlled using ad hoc treatment strategies (fungicides). In this study, two analytical methods were developed, validated and applied for the determination of five ATs and 195 pesticides in tomato products collected from the Italian market. Two distinct groups, organic (n = 20) and conventional (n = 20) Italian tomato sauces, were characterized in depth. Tenuazonic acid, alternariol and alternariol monomethyl ether were found up to 517, 27 and 7.1 µg/kg, respectively, while pesticides were detected between 0.0026 and 0.0421 mg/kg in conventional products, and, interestingly, up to 0.0130 mg/kg in organic products. No correlation emerged between the detected levels of ATs and pesticides and the type of tomato cultivation, but the probability of pesticide contamination in conventional products was eight times higher than in organics. Some considerations about exposure assessment and risk characterization for ATs were also proposed in the overall population and in more sensitive and/or exposed subgroups, underlining the need for new focused toxicological and monitoring studies to establish reliable reference values. Moreover, these data highlight that fungicide treatments may not protect tomatoes from ATs contamination, although it may remove fungi infestation. As organic product consumption is increasing, it is important to lay down dedicated regulations for maximum permitted levels to ensure the food safety of these products that are often perceived by consumers as a healthier and environmentally friendlier choice.

Mycotoxin contamination of feed and feed materials represent a serious health hazard. This study details the occurrence of aflatoxin B1 (AFB1), zearalenone (ZEN) and ochratoxin A (OTA) in 826 feed and 617 feed material samples, collected in two Italian Regions (Umbria and Marche) from 2018 to 2022 analyzed using a UPLC-FLD platform. The developed method was validated and accredited (ISO/IEC 17025) with satisfactory accuracy and precision data obtained in repeatability and intralaboratory reproducibility conditions. Feed had a higher incidence of contaminated samples (26%) with respect to feed materials (6%). AFB1 was found up to 0.1045 mg/kg in cattle feeds and 0.1234 mg/kg in maize; ZEN was detected up to 6.420 mg/kg in sheep feed while OTA was rarely reported and in lower concentrations (up to 0.085 mg/kg). Co-contamination of at least two mycotoxins was reported in 0.8% of the analyzed samples. The incidence of above maximum content/guidance level samples was 2% for feed and feed materials while almost 3-fold-higher for maize (5.8%) suggesting how mycotoxin contamination can affect some matrices more than others. Obtained data can be useful to improve official monitoring plans and therefore further raise awareness of this issue between agriculture stakeholders, healthcare entities and non-professionals.

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 mu g/kg and Tentoxin up to 570 mu g/kg, whereas for the other mycotoxin/commodities combinations, the percentage of left censored data (below the limit of quantification) ranged from 74 to 100%.

Aflatoxin M-1 (AFM(1)) is a hepatocarcinogenic and genotoxic derivative of aflatoxin B-1 excreted into milk after ingestion of feed contaminated by Aspergillus genus fungi. Because of the important role of dairy products, especially cow cheese, in the human diet, there is great concern about the presence of AFM(1) in this food category. EC Regulation No. 1881/2006 establishes the importance of the enrichment factor (EF), an essential parameter that must be defined in order to evaluate the maximum level of the toxin in cheese aiming to ensure that cheese has been produced from compliant milk. The Italian Ministry of Health has established two provisional AFM(1) EFs (5.5 and 3.0) to be applied to as many cheese categories (hard and soft), defined according to the moisture content on a fat free basis (MFFB) classification. Two experimental productions of Primosale and Fior di Latte cheese, both belonging to the soft cheese category, showed an EF of 4.1 and 2.9 respectively. Data in literature also suggest that the EF attribution based on the current categorization may need reconsideration.

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.

Twelve different approaches commonly used for the simultaneous LC tandem MS (MS/MS) determination of mycotoxins (deoxynivalenol, aflatoxins, ochratoxin A, T-2 and HT-2 toxins, fumonisins, and zearalenone) were tested in cereals and feed materials. They comprised different extraction solvents, types of cleanup [solid-phase extraction, QuEChERS, and immunoaffinity (IMA)], and calibration approaches (external or matrix-matched). The percentage of mycotoxins with acceptable recovery, according to Regulation (EC) No. 401/2006, ranged from 9 to 100%. The approach giving the highest percentage of acceptable results was selected and further tested for corn, rice, and feed spiked at three different mycotoxin levels (low, medium, and high). The method is based on extraction with MeOH-water (70 + 30, v/v) and cleanup with two multiantibody IMA columns. For corn and rice spiked at low mycotoxin levels, a significant matrix effect was observed and was compensated by using 13C calibration. At higher mycotoxin levels (medium and high), matrix effects were negligible as no significant differences were observed for the majority of recovery results calculated by 13C calibration and external calibration. Although the proposed method still needs improvement in terms of accuracy and, to a lesser extent, precision, it was successfully tested with four proficiency tests in buckwheat, corn, rice, and feed, giving acceptable z-scores for 97% (34 out of 35) of results.

The objective of this study was to develop a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the determination of Ochratoxin A (OTA) and Ochratoxin α (OTα) in poultry tissues and eggs. The two toxins were extracted by a mixture of acetonitrile/water, purified with a reversed phase C18 solid phase extraction column (SPE) and determined by LC-MS/MS. The LC-MS/MS method performances were evaluated in terms of linearity in solvent and in matrix (ranged from 0.5 to 15.10 µg L-1 for OTA and from 0.60 to 17.85 µg L-1 for OTα), limit of detection (LOD), limit of quantitation (LOQ), specificity, accuracy and precision in repeatability conditions. Recovery experiments were performed by spiking poultry liver, kidney, muscle and eggs around 1 µg kg-1 and 10 µg kg-1. LODs were 0.27 and 0.26 µg kg-1 while LOQs were fixed at 1.0 and 1.2 µg kg-1 for OTA and OTα, respectively. Main recoveries for OTA ranged from 82 to 109% and for OTα ranged from 55 to 89%. The values of within-laboratory relative standard deviation (RSDr) were equal to or below 20%. Considering the results obtained and that all analytical performance criteria were fulfilled, the new extraction and purification method developed for OTA and OTα determination in animal tissues and eggs was found appropriate for control laboratories and research activities designed to ensure food safety.

A simple multiresidue method is described for assaying 10 sulphonamides (SAs) (sulfadiazine, sulfathiazole, sulfapyridine, sulfamerazine, sulfamethazine, sulfamonomethoxine, sulfachlorpyridazine, sulfamethoxazole, sulfaquinoxaline and sulfadimethoxine) in muscle samples. Samples were prepared by homogenizing the tissue, extracting with ethyl acetate and cleaning up with a cation-exchange solid-phase extraction (SPE) column. The detection of analytes was achieved by HPLC-diode array detection (DAD) at 270 nm. The procedure was validated according to the European Union regulation 2002/657/EC determining specificity, decision limit, detection capability, trueness and precision. The results of validation process demonstrate that the method is suitable for application in European Union statutory veterinary drug residue surveillance programmes.

It is impossible to determine whether an analytical procedure is suitable to the purposes without some knowledge of its uncertainty. Several approaches for its estimation have already been proposed in the past years. Discussion on the estimation of uncertainty of chemical measurement has also influenced the ISO 17025 which explicitly refers to the bottom-up approach.On the other hand, the European Commission Decision 2002/657/EC concerning the performance of methods and the interpretation of results in the official control of residues in products of animal origin imposes the determination of some new parameters, such as decision limit (CCalpha) which represents an index of results dispersion. This decision also indicates that official laboratories must be accredited according to ISO 17025.In this work, the bottom-up approach together with in-house validation data is applied for the evaluation of measurement uncertainty associated with residue determination of 10 sulfonamides in muscle and, at the same time, the decision limit was calculated following the 2002/657/EC criteria. (C) 2004 Elsevier B.V. All rights reserved.

A simple multi-residue method is described for assaying 13 quinolones (QNs; pipemidic acid, rufloxacin, enoxacin, ofloxacin, norfloxacin, ciprofloxacin, danofloxacin, enrofloxacin, difloxacin, cinoxacin, oxolinic acid, nalidixic acid and flumequine) in feeds. The samples were extracted by a metaphosphoric acid/acetonitrile mixture at pH 2.6 (ASE 200) and automatically purified onto OASIS HLB cartridges (ASPEC XL). The determination was achieved by liquid chromatography (LC) using a C5 analytical column (150×4.6mm, 5μm) either with photodiode-array or fluorescence detection. The procedure was validated by spiking a feed sample at three different levels (5, 10 and 25mgkg−1) and linearity, detection limit, quantification limit, accuracy and precision were checked. The automated preparation of the sample permits a very fast analysis which is an important goal for routine purposes.