Chiara Dall'Asta

Masked mycotoxins: A review

The aim of this review is to give a comprehensive overview of the current knowledge on plant metabolites of mycotoxins, also called masked mycotoxins. Mycotoxins are secondary fungal metabolites, toxic to human and animals. Toxigenic fungi often grow on edible plants, thus contaminating food and feed. Plants, as living organisms, can alter the chemical structure of mycotoxins as part of their defence against xenobiotics. The extractable conjugated or non-extractable bound mycotoxins formed remain present in the plant tissue but are currently neither routinely screened for in food nor regulated by legislation, thus they may be considered masked. Fusarium mycotoxins (deoxynivalenol, zearalenone, fumonisins, nivalenol, fusarenon-X, T-2 toxin, HT-2 toxin, fusaric acid) are prone to metabolisation or binding by plants, but transformation of other mycotoxins by plants (ochratoxin A, patulin, destruxins) has also been described. Toxicological data are scarce, but several studies highlight the potential threat to consumer safety from these substances. In particular, the possible hydrolysis of masked mycotoxins back to their toxic parents during mammalian digestion raises concerns. Dedicated chapters of this article address plant metabolism as well as the occurrence of masked mycotoxins in food, analytical aspects for their determination, toxicology and their impact on stakeholders.

Occurrence of deoxynivalenol and its 3-β-D-glucoside in wheat and maize

Deoxynivalenol-3-β-D-glucoside (D3G), a phase II plant metabolite of the mycotoxin deoxynivalenol (DON), occurs in naturally Fusarium-contaminated cereals. In order to investigate the frequency of occurrence as well as the relative and absolute concentrations of D3G in naturally infected cereals, 23 wheat samples originating from fields in Austria, Germany and Slovakia as well as 54 maize samples from Austrian fields were analysed for DON and D3G by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Both analytes were detected in all the 77 field samples. DON was found at levels from 42 to 4130 ng g−1 (977 ± 1000 ng g−1 on average). The D3G concentrations in all cereal samples were in the range 10–1070 ng g−1 (216 ± 253 ng g−1 on average), corresponding to about 5–46 mol% of their DON concentrations (15 ± 8 mol% on average).

Rapid and Comprehensive Evaluation of (Poly)phenolic Compounds in Pomegranate (Punica granatum L.) Juice by UHPLC-MSn

The comprehensive identification of phenolic compounds in food and beverages is a crucial starting point for assessing their biological, nutritional, and technological properties. Pomegranate (Punica granatum L.) has been described as a rich source of (poly)phenolic components, with a broad array of different structures (phenolic acids, flavonoids, and hydrolyzable tannins) and a quick, high throughput, and accurate screening of its complete profile is still lacking. In the present work, a method for UHPLC separation and linear ion trap mass spectrometric (MSn) characterization of pomegranate juice phenolic fraction was optimized by comparing several different analytical conditions. The best solutions for phenolic acids, anthocyanins, flavonoids, and ellagitannins have been delineated and more than 70 compounds have been identified and fully characterized in less than one hour total analysis time. Twenty-one compounds were tentatively detected for the first time in pomegranate juice. The proposed fingerprinting approach could be easily translated to other plant derived food extracts and beverages containing a wide array of phytochemical compounds.

Difficulties in fumonisin determination: the issue of hidden fumonisins

In this paper, the results obtained by five independent methods for the quantification of fumonisins B1, B2, and B3 in raw maize are reported. Five naturally contaminated maize samples and a reference material were analyzed in three different laboratories. Although each method was validated and common calibrants were used, a poor agreement about fumonisin contamination levels was obtained. In order to investigate the interactions among analyte and matrix leading to this lack of consistency, the occurrence of fumonisin derivatives was checked. Significant amounts of hidden fumonisins were detected for all the considered samples. Furthermore, the application of an in vitro digestion protocol to raw maize allowed for a higher recovery of native fumonisins, suggesting that the interaction occurring among analytes and matrix macromolecules is associative rather than covalent. Depending on the analytical method as well as the maize sample, only 37–68% of the total fumonisin concentrations were found to be extractable from the samples. These results are particularly impressive and significant in the case of the certified reference material, underlying the actual difficulties in ascertaining the trueness of a method for fumonisin determination, opening thus an important issue for risk assessment.

In Vitro Digestion Assay for Determination of Hidden Fumonisins in Maize

Hidden fumonisins have received great attention in the last years as they have been frequently found in maize products in addition to the free forms. Several papers have shown that interaction with macromolecular components such as protein and starch is at the base of the phenomenon: although the nature of the interaction (covalent or not) is still not clarified, the occurrence of hidden forms is generally revealed by the application of an alkaline hydrolysis procedure. In this study, an in vitro digestion model has been applied to raw maize to evaluate the possible release of hidden fumonisins under gastrointestinal conditions. Upon digestion of the food matrix, an increased amount of total detectable fumonisins was observed in comparison with the analysis on the nondigested matrix, an amount even higher than that calculated through the application of the hydrolysis procedure. Besides the analytical issues, our data have serious implications, since consumers may be exposed to a systematic higher risk than that estimated by conventional techniques.

Free and bound fumonisins in gluten-free food products

In this work a multiresidual LC-ESI-MS/MS method for the simultaneous detection of free and bound fumonisins is described, which allowed for a very low LOD and a very good recovery for all the analytes. The method was applied to the determination of free and bound fumonisins in several gluten-free products from the Italian market. Free fumonisins were found to occur in 90% of the samples: the overall median value was below the EU legal limit for foods for human consumption (800 microg/kg). Nonetheless, fumonisins occurred in several samples at concentrations above the legal limit, reaching also very strong contamination levels (maximum concentration level: 3310 microg/kg). Anyway, considering the limited diet of people suffering of the celiac disease or allergic to other wheat proteins, the incidence of fumonisin contamination may be envisaged as problematic. Furthermore, bound fumonisins were found to be present in all the analysed samples at similar or even higher amounts than the free forms. In many cases the sum of free and bound fumonisins exceeded the EU legal limit for total fumonisins also for those samples characterized by a low contamination of free fumonisins, thus opening a new important task to be addressed for the risk assessment in this field.

Fate of Fusarium mycotoxins in the cereal product supply chain: the deoxynivalenol (DON) case within industrial bread-making technology.

Fusarium mycotoxins are a relevant problem in the cereal supply chain at a worldwide level, with wheat, maize and barley being the main contaminated crops. Mould growth can happen in the pre-harvest phase and also during transport and storage due to ineffective drying conditions. Among Fusarium toxins, deoxynivalenol (DON) is considered the most important contaminant in wheat due to its widespread occurrence. In the last years the European Food Safety Authority (EFSA) and the European Commission have frequently expressed opinions on Fusarium toxins, setting limits, regulations and guidelines in order to reduce their levels in raw materials and food commodities. In particular, European legislation (Reg. 1881/2006) sets the maximum limit for DON in flour and bread as 750 and 500 µg kg−1 respectively. Relatively few studies have taken into account the loss of trichothecenes during processing, focusing on how processing factors may influence their degradation. In particular, the description of DON behaviour during bread-making is very difficult, since complex physico-chemical modifications occur during the transformation of the raw ingredients into the final product. In the present study, we studied how DON concentration may be influenced by modifying bread-making parameters, with a special emphasis on the fermentation and baking stages, starting from a naturally contaminated flour at both pilot and industrial scales. Exploiting the power of a Design of Experiments (DoE) approach to consider the great complexity of the studied system, the obtained model shows satisfying goodness-of-fit and prediction, suggesting that the baking step (time/temperature ranges) is crucial for minimizing native DON level in bread.

Dietary exposure to fumonisins and evaluation of nutrient intake in a group of adult celiac patients on a gluten-free diet

SCOPE The main objectives of this study were to estimate dietary fumonisin exposure and nutrient intake in a group of patients diagnosed with celiac disease compared to non-celiac subjects. METHODS AND RESULTS The fumonisin level in 118 frequently consumed corn-based products was determined and dietary habits were recorded using a 7-day weighed food record. Data were then compared to those obtained for a control group. The fumonisin intake in the celiac patients was significantly higher than in controls, with mean values (± SE) of 0.395 ± 0.049 and 0.029 ± 0.006 μg/kg body weight per day, respectively. With regard to nutritional habits, celiac patients showed a preference for a high fat diet, coupled with a high intake of sweets and soft drinks and a low intake of vegetables, iron, calcium and folate. CONCLUSION These findings may have serious health implications for the celiac population due to the widespread occurrence of fumonisins in most of the widely consumed gluten-free products, leading to continuous exposure to this particular mycotoxin. Moreover, the recorded nutritional quality of the celiac patients diet raises concerns regarding its long-term adequacy and its potential impact on chronic conditions such as type 2 diabetes and cardiovascular diseases.

Role of maize hybrids and their chemical composition in Fusarium infection and fumonisin production

This study was designed to investigate the role of hybrids in maize Fusarium section Liseola interaction and fumonisin production, with particular emphasis on the occurrence and accumulation of hidden fumonisins in maize (masking phenomenon). In this 2 year study, naturally infected field crops were chosen with 10 maize hybrids, six of them grown in both years. Maize samples collected in 2010 showed a higher incidence of fungal infection as well as higher fumonisin contamination than those obtained in 2009 but a very similar incidence of F. section Liseola. Fumonisin masking was confirmed in raw maize, with a lower amount of hidden forms as compared to free fumonisins detected in the year with higher contamination. The chemical composition of the different hybrids was determined and correlated with the contamination data: the results obtained highlight the main role of fatty acids, with a higher fumonisin contamination in hybrids showing a higher linoleic acid content and a higher masking action in hybrids with higher oleic to linoleic ratio. These results represent a good basis to explain maize hybrid susceptibility to F. section Liseola infection, fumonisin contamination, and masking not related to a specific commercial hybrid but extendable to all hybrids.

Fate of Fusarium mycotoxins in cereal product supply chain: the DON case within industrial bread-making technology

Fusarium mycotoxins are a relevant problem in the cereal supply chain at a worldwide level, with wheat, maize and barley being the main contaminated crops. Mould growth can happen in the pre-harvest phase and also during transport and storage due to ineffective drying conditions. Among Fusarium toxins, deoxynivalenol (DON) is considered the most important contaminant in wheat due to its widespread occurrence. In the last years the European Food Safety Authority (EFSA) and the European Commission have frequently expressed opinions on Fusarium toxins, setting limits, regulations and guidelines in order to reduce their levels in raw materials and food commodities. In particular, European legislation (Reg. 1881/2006) sets the maximum limit for DON in flour and bread as 750 and 500 µg kg−1 respectively. Relatively few studies have taken into account the loss of trichothecenes during processing, focusing on how processing factors may influence their degradation. In particular, the description of DON behaviour during bread-making is very difficult, since complex physico-chemical modifications occur during the transformation of the raw ingredients into the final product. In the present study, we studied how DON concentration may be influenced by modifying bread-making parameters, with a special emphasis on the fermentation and baking stages, starting from a naturally contaminated flour at both pilot and industrial scales. Exploiting the power of a Design of Experiments (DoE) approach to consider the great complexity of the studied system, the obtained model shows satisfying goodness-of-fit and prediction, suggesting that the baking step (time/temperature ranges) is crucial for minimizing native DON level in bread.