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Mycotoxins' exposure by inhalation and/or dermal contact can occur in different branches of industry especially where heavily dusty settings are present and the handling of dusty commodities is performed. This study aims to explore the possible contribution of the occupational exposure to aflatoxins by analysing urine samples for the presence of aflatoxins B 1 and M 1 and aflatoxin B 1 -N 7 -guanine adduct. The study was conducted in 2017 on two groups of volunteers, the workers group, composed by personnel employed in an Italian feed plant (n = 32), and a control group (n = 29), composed by the administrative employees of the same feed plant; a total of 120 urine samples were collected and analysed. A screening method and a quantitative method with high-resolution mass spectrometry determination were developed and fully validated. Limits of detections were 0.8 and 1.5 pg/mL urine for aflatoxin B 1 and M 1 , respectively. No quantitative determination was possible for the adduct aflatoxin B 1 -N 7 -guanine. Aflatoxin B 1 and its adduct were not detected in the analysed samples, and aflatoxin M 1 , instead, was found in 14 samples (12%) within the range 1.9-10.5 pg/mL urine . Only one sample showed a value above the limit of quantification (10.5 pg/mL urine ). The absence of a statistical difference between the mean values for workers and the control group which were compared suggests that in this specific setting, no professional exposure occurs. Furthermore, considering the very low level of aflatoxin M 1 in the collected urine samples, the contribution from the diet to the overall exposure is to be considered negligible.

Ochratoxin A is one of the most diffused mycotoxin present in a large spectrum of food commodities, mainly produced by Aspergillus ochraceus, Aspergillus carbonarius, Aspergillus niger and Penicillium verrucosum. EU has set maximum limits for a number of matrices such as cereals, wine, spices and liquorice, whilst other commodities such as beer and meat products that are susceptible of OTA contamination and are largely consumed are not included. In 2013, within the framework of the Regulation (EC) 882/2004 on official controls, the European Commission issued the mandate M/520 regarding the standardisation for methods of analysis for mycotoxins in food to the European Committee for Standardisation. Of the 11 priorities of the mandate, the one on "HPLC determination of OTA in meat, meat products and edible offal" was assigned to the Italian National Reference Laboratory for feed and food. The method was single-laboratory validated, and all the performance characteristics of the method were compliant with the corresponding reference values indicated in Regulation (EC) n. 401/2006. The method was applied to characterise a set of 5 pork-based materials (ham, kidney, liver and canned chopped pork) to be used for an inter-laboratory method validation study. Three ham materials (levels of contamination of 0.77, 2.22 and 12.3 μg/kg, respectively), one liver material (contamination level of 2.80 μg/kg) and one chopped pork meat (contamination level of 0.66 μg/kg) were tested for homogeneity and stability.

Mycotoxins exposure by inhalation and/or dermal contact is possible in different branches of industry especially where heavily dusty settings are present and the handling of dusty commodities is performed. This study aims to explore the validity of the biomonitoring as a tool to investigate the intake of mycotoxins in a population of workers operating in an Italian feed plant. Serum samples were collected for the determination of aflatoxins B1 (AFB1), AFB1-Lysine adduct and ochratoxin A (OTA). A method based on liquid-liquid extraction coupled with high resolution mass spectrometry determination was developed and fully validated. For AFB1, a high number of non-detected samples (90%) was found and no statistical difference was observed comparing workers and control group. None of the analyzed samples showed the presence of AFB1-Lysine adduct. For OTA, the 100% of the analyzed samples was positive with a 33% of the samples showing a concentration higher than the limit of quantification (LOQ), but no statistical difference was highlighted between the average levels of exposed and control groups. In conclusion, the presence of AFB1 and OTA in serum cannot be attributable to occupational exposure.

Objective: To assess the risk of mycotoxin exposure (aflatoxin M1, ochratoxin A, and zearalenone) in celiac disease (CD) breastfeeding mothers and healthy control mothers, as well as in their offspring, by quantifying these contaminants in breast milk., Study Design: Thirty-five breastfeeding women with CD on a gluten-free diet and 30 healthy breastfeeding controls were recruited. Milk sampling was performed three times per day for three consecutive days. Mycotoxin content was investigated by an analytical method using immunoaffinity column clean-up and high-performance liquid chromatography (HPLC) with fluorometric detection., Results: Aflatoxin M1 (AFM1) was detected in 37% of CD group samples (mean ± SD = 0.012 ± 0.011 ng/mL; range = 0.003-0.340 ng/mL). The control group showed lower mean AFM1 concentration levels in 24% of the analyzed samples (0.009 ± 0.007 ng/mL; range = 0.003-0.067 ng/mL, ANOVA on ranks, p -value < 0.01). Ochratoxin A and zearalenone did not differ in both groups., Conclusion: Breast milk AFM1 contamination for both groups is lower than the European safety threshold. However, the estimated exposures of infants from CD mothers and control mothers was much higher (≃15 times and ≃11 times, respectively) than the threshold set by the joint FAO/WHO Expert Committee on Food Additives (JECFA). Since incongruities exist between JECFA and the European Union standard, a novel regulatory review of the available data on this topic is desirable. Protecting babies from a neglected risk of high AFM1 exposure requires prompt regulatory and food-control policies., Competing Interests: C.C. receives a consultancy honorary from Schär and Menarini Diagnostics. The other authors have no conflicts of interest relevant to this article to disclose.

The analytical scenario for determining contaminants in the food and feed sector is constantly prompted by the progress and improvement of knowledge and expertise of researchers and by the technical innovation of the instrumentation available. Mycotoxins are agricultural contaminants of fungal origin occurring at all latitudes worldwide and being characterized by acute and chronic effects on human health and animal wellness, depending on the species sensitivity. The major mycotoxins of food concern are aflatoxin B1 and ochratoxin A, the first for its toxicity, and the second for its recurrent occurrence. However, the European legislation sets maximum limits for mycotoxins, such as aflatoxin B1, ochratoxin A, deoxynivalenol, fumonisins, and zearalenone, and indicative limits for T-2 and HT-2 toxins. Due to the actual probability that co-occurring mycotoxins are present in a food or feed product, nowadays, the availability of reliable, sensitive, and versatile multi-mycotoxin methods is assuming a relevant importance. Due to the wide range of matrices susceptible to mycotoxin contamination and the possible co-occurrence, a multi-mycotoxin and multi-matrix method was validated in liquid chromatography-tandem mass spectrometry (LC-MS/MS) with the purpose to overcome specific matrix effects and analyze complex cereal-based samples within the Italian Total Diet Study project., Competing Interests: The authors declare no conflict of interest.

Mycotoxins are low molecular weight secondary metabolites produced by certain strains of filamentous fungi such as Aspergillus, Penicillium and Fusarium, which attack crops in the field, and grow on foods also during storage under favorable conditions of temperature and humidity. Foods mainly contributing to the intake of mycotoxins with diet are cereals, maize being the most risky commodity due to the potential co-occurrence of more than one mycotoxin, this can be of particular concern especially for vulnerable group of population such as celiac patients that show increased maize-based products consumption. In this study the exposure of celiac patients to fumonisins (FBs) and zearalenone (ZON) has been assessed. The higher exposures, for all the matrices and for both the selected mycotoxins, were for children age group. The lower and upper bound exposure ranged between 348-582 ng/kg bw/day for FBs and 22-83 ng/kg bw/day for ZON; these values result well below the TDI for the selected mycotoxins, representing the 17-29% and 9-33% of the TDI set for FBs and ZON, respectively. Even considering the worst scenario the exposure values reported for children were lower, namely 1385 ng/kg bw/day for FBs and 237 ng/kg bw/day for ZON, than the corresponding toxicological thresholds., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Four hundred and seventy-two pasta samples were collected from long retail distribution chain sales points located in North, Central and South Italy. Representative criteria in the sample collection were followed in terms of number of samples collected, market share, and types of pasta. Samples were analysed by an accredited HPLC-UV method of analysis. The mean contamination level (64.8 μg/kg) of deoxynivalenol (DON) was  in the 95th percentile (239 μg/kg) and 99th percentile (337 μg/kg), far below the legal limit (750 μg/kg) set by Regulation EC/1126/2007, accounting for about one tenth, one third and half the legal limit, respectively. Ninety-nine percent of samples fell below half the legal limit. On the basis of the obtained occurrence levels and considering the consumption rates reported by the Italian official database, no health concern was assessed for all consumer groups, being that exposure was far below the Tolerable Daily Intake (TDI) of 1000 ng/kg b.w/day. Nevertheless, despite this, particular attention should be devoted to the exposure to DON by high consumers, such as children aged 3-5 years, who could reach the TDI even with very low levels of DON contamination.

Use of proper sampling methods throughout the agri-food chain is crucial when it comes to effectively detecting contaminants in foods and feeds. The objective of the study was to estimate the performance of sampling plan designs to determine aflatoxin B(1) (AFB(1)) contamination in corn fields. A total of 840 ears were selected from a corn field suspected of being contaminated with aflatoxin. The mean and variance among the aflatoxin values for each ear were 10.6 mug/kg and 2233.3, respectively. The variability and confidence intervals associated with sample means of a given size could be predicted using an equation associated with the normal distribution. Sample sizes of 248 and 674 ears would be required to estimate the true field concentration of 10.6 mug/kg within +/-50 and +/-30%, respectively. Using the distribution information from the study, operating characteristic curves were developed to show the performance of various sampling plan designs.

According to general consensus, the global climate is changing, which may also affect agricultural and livestock production. The potential impact of climate change on food security is a widely debated and investigated issue. Nonetheless, the specific impact on safety of food and feed for consumers has remained a less studied topic. This review therefore identifies the various food safety issues that are likely to be affected by changes in climate, particularly in Europe. Amongst the issues identified are mycotoxins formed on plant products in the field or during storage; residues of pesticides in plant products affected by changes in pest pressure; trace elements and/or heavy metals in plant products depending on changes in their abundance and availability in soils; polycyclic aromatic hydrocarbons in foods following changes in long-range atmospheric transport and deposition into the environment; marine biotoxins in seafood following production of phycotoxins by harmful algal blooms; and the presence of pathogenic bacteria in foods following more frequent extreme weather conditions, such as flooding and heat waves. Research topics that are amenable to further research are highlighted.

The scope of this study was to evaluate the exposure of the Italian population to ochratoxin A (OTA) attributable to wine consumption. With this aim 1166 wine samples (773 red wines, 290 white, 75 rose, and 28 dessert wines), collected in 19 different Italian regions and mostly produced between 1988 and 2004, were analyzed for OTA content. The obtained results are reported by year of harvest, geographical area of production, and type of wine. Red wine showed the highest maximum level of contamination (7.50 ng/mL), even though rose wines were characterized by a higher mean value (0.01 ng/mL). A gradually increasing mean concentration was also observed from the north (0.05 ng/mL) to south of Italy (0.54 ng/mL). Exposure calculations, performed using two different consumption databases, indicate a daily intake for consumer only of 0.59 up to 1.24 ng/(kg of b.w.)/day and of 0.33 up to 0.90 ng/(kg of b.w.)/day for the total population. Even in the worst case, corresponding to the calculation of the intake for consumers only in southern Italy and Islands and considering the mean consumption data increased by 1 standard deviation, a quite low exposure (1.68 ng/(kg of b.w.)/day, accounting for 9.8% of TDI) was obtained. Considering the overall OTA dietary exposure, obtained exposure rates indicate that wine did not pose a risk to the Italian population health.

An interlaboratory study was conducted to evaluate the effectiveness of an immunoaffinity column cleanup liquid chromatography (LC) method for the determination of aflatoxin B1 levels in corn samples, enforced by European Union legislation. A test portion was extracted with methanol-water (80 + 20); the extract was filtered, diluted with phosphate-buffered saline solution, filtered on a microfiber glass filter, and applied to an immunoaffinity column. The column was washed with deionized water to remove interfering compounds, and the purified aflatoxin B1 was eluted with methanol. Aflatoxin B1 was separated and determined by reversed-phase LC with fluorescence detection after either pre- or postcolumn derivatization. Precolumn derivatization was achieved by generating the trifluoroacetic acid derivative, used by 8 laboratories. The postcolumn derivatization was achieved either with pyridinium hydrobromide perbromide, used by 16 laboratories, or with an electrochemical cell by the addition of bromide to the mobile phase, used by 5 laboratories. The derivatization techniques used were not significantly different when compared by the Student's t-test; the method was statistically evaluated for all the laboratories. Five corn sample materials, both spiked and naturally contaminated, were sent to 29 laboratories (22 Italian and 7 European). Test portions were spiked with aflatoxin B1 at levels of 2.00 and 5.00 ng/g. The mean values for recovery were 82% for the low level and 84% for the high contamination level. Based on results for spiked samples (blind pairs at 2 levels) as well as naturally contaminated samples (blind pairs at 3 levels), the values for relative standard deviation for repeatability (RSDr) ranged from 9.9 to 28.7%. The values for relative standard deviation for reproducibility (RSDR) ranged from 18.6 to 36.8%. The method demonstrated acceptable within- and between-laboratory precision for this matrix, as evidenced by the HorRat values.

The need to obtain a representative sample deserves particular consideration since a wrong sampling plan can greatly affect the reliability of the measured levels of mycotoxins. This can even result in legal disputes and barriers to trade. Reported here is an holistic view for an ideal sampling plan, which is based on two consecutive steps: (i) To establish 'why, where and when' sampling has to be performed by assessing the purpose, the appropriate time and the site for collecting the samples; (ii) To establish 'how' to draw samples by assessing practical ad hoc guidelines, considering that, for bulk goods in particular, mycotoxins are not at all homogeneously distributed in a lot. So far, step 1 is not yet covered by specific guidelines while for step 2, European regulations establish the procedures for the sampling of bulk and retail products potentially contaminated by mycotoxins.

The aim of this study was to investigate the distribution of fumonisin B1 in various corn milling fractions processed by an industrial plant. Corn kernels and six derived milling fractions (germ, bran, large and small grits, animal feed flour, and flour) were sampled. In addition, in order to evaluate the effect of cooking, samples of polenta were prepared starting from naturally contaminated flour obtained from the industrial processing cycle. The industrial plant worked continuously at a rate of 60 tons per day. Two sublots of 5 tons each were investigated with samples of derived products taken at regular time intervals. Due to a similar heterogeneous distribution of fumonisin B1 with other mycotoxins, such as aflatoxins, the sampling scheme was derived from the European Directive 98/53 for aflatoxins. Both lots of kernels showed fumonisin contamination at 4.54 and 5.09 mg/kg, respectively. Germ, bran, and animal feed flour showed contamination levels, namely 8.92 mg/kg (lot 1) and 9.56 mg/kg (lot 2), 7.08 mg/kg (lot 1) and 8.08 mg/kg (lot 2), and 9.36 mg/kg (lot 1) and 6.86 mg/kg (lot 2) higher than large and small grits and flour (0.39 mg/kg [lot 1] and 0.42 mg/kg [lot 2], 0.60 mg/kg [lot 1] and 1.01 mg/kg [lot 2], and 0.40 mg/kg [lot 1] and 0.45 mg/kg [lot 2], respectively). These results seem to account both for the industrial yields of the derived products and the distribution of fumonisin contamination in a kernel. The cooking of polenta in a domestic pressure cooker did not affect fumonisin contamination because the mycotoxin concentrations were similar to those of the starting flour (0.40 and 0.45 mg/kg).

In order to estimate the incidence of ochratoxin A (OA) in biological fluids, a study was carried out to determine the concentration of OA in breast milk of donor mothers in Italy. Out of 111 samples, 22 were contaminated in the range 0.1-12 micrograms/kg.

A study was undertaken to evaluate the total aflatoxin content in naturally contaminated eggs. Two pools of eggs from laying hens were collected after 2 and 7 days of treatment with aflatoxin B1 (AFB1). An HPLC method has been developed for the determination of AFB1, aflatoxin M1 (AFM1), aflatoxin B2a (AFB2a) and aflatoxicol (Ro) both in free form and after release from water-soluble conjugates. The bound form was cleaned up after acid hydrolysis of the aqueous phase. Eggs collected after 2 days of treatment revealed residues of AFB1, AFB2a and Ro in the organic phase, but none in the aqueous portion. After 7 days of treatment both AFB1 and its hydroxy derivative were found in the organic phase but the aqueous portion showed only hydroxylated metabolites accounting for 35% of the total aflatoxin content.

A study was performed to evaluate the contamination by ochratoxin A in coffee beans. Twenty-nine samples of green coffee were collected from large lots of material by representative sampling. The analyses of green coffee samples showed a significantly high contamination percentage (58%) ranging from 0.2 to 15 micrograms/kg. Naturally and artificially contaminated samples were roasted at different operation times (5-6 min) to verify the percentage of destruction of the mycotoxin. The percentage ranged from 48% to 87% and from 90% to 100% in artificially and naturally contaminated samples respectively. The beverages prepared from artificially contaminated coffee using the most common types of coffee makers showed no residues of ochratoxin A.

Abstract: The influence of the durum wheat milling process (pre-cleaning, cleaning, hulling, different fineness milling steps) on the deoxynivalenol (DON) content was evaluated, by establishing its contamination distributed in the different products and byproducts (bran, shorts, flour, semolina, and pasta). In order to include also the DON distribution within the pasta manufacturing process, four samples of semolina were worked to obtain four different lots of dried pasta. The results obtained in this study show that, with respect to wheat grain (100%), a reduction of DON contamination of 30%, 66% and 63% in clean wheat, peeled wheat and semolina was observed respectively. On the other hand, an increase of DON contamination was observed in the byproducts, especially in the foliage waste and in the wheat bran fractions, which are intended for feed production. However, at the present time, there is no legal limit set for DON content in feed. As far as the second-stage processing is concerned, with respect to semolina (100%), the observed DON reduction in dry and cooked pasta was estimated 8% and 41% respectively. The overall process, from wheat grain to cooked pasta, leads to a mean DON contamination decrease of 78%. The outputs of this study, confirming the trend of previous works, are of crucial importance in order to early evaluate the safety of the finished products, consistently with their final destination, once the contamination of the raw material is established. [Copyright &y& Elsevier]

Abstract: Three hundred samples of cocoa and chocolate-based products were randomly collected in South, Center and North Italy. One hundred and seventy nine, out of the 300 samples analyzed, were positive for ochratoxin A (OTA) contamination, representing 60% of the purchased products. All 40 cocoa samples showed OTA contamination, while grouping all the chocolate products the positive samples accounted for 53%. The mean OTA concentration levels for cocoa and chocolate products were well below the corresponding Italian legal limits. Teenagers’ male age group resulted as the highest consumers for cocoa powder (2.9 g per day), while the OTA highest weekly intake was reported for the younger age groups (0.10 and 0.38 ng/kg bw/week, for infant and children respectively). With respect to chocolate products, the highest daily consumption and the highest intake are reported for children (7.3 g and 0.57 ng/kg bw/week, respectively). As far as single chocolate products are concerned, the highest weekly intake is referred to the consumption of Easter chocolate eggs by the infant/children age group (0.10 ng/kg bw/week). The overall analysis of the exposure assessment suggests that the very low value of body weight for infant and children categories drives the exposure calculations. On the basis of the risk assessment carried out in this study, the exposure to OTA, due to the consumption of cocoa and chocolate products, is to be considered not a major concern. Therefore, the Italian Superior Council of Health considered the data arising from this study, adequate to repeal the existing Italian legal limits and to align national legislation with the EU provisions. [Copyright &y& Elsevier]