Food and feed contamination with mycotoxins is a major public health concern. Humans and animals are exposed to these toxins by consuming contaminated products throughout their lives. In this study, a method based on dispersive liquid–liquid microextraction (DLLME), followed by liquid chromatography with fluorescence detection (LC-FLD), was validated for the determination of aflatoxins (AFs) M1, B1, B2, G1, G2, zearalenone (ZEN), and ochratoxin A (OTA). The method was applied to 150 raw cow milk samples and 90 market durum wheat samples from two Tunisian climatic regions: the littoral region (Mahdia) and the continental region (Béja). This work was carried out to obtain more surveillance data to support rapid initiatives to assure safe foods and protect consumer health and to estimate the daily exposure of the Tunisian population consuming those products. AFG2 and OTA were found in wheat with incidences of 54.4 and 11.1%, respectively. On the other side, milk samples were contaminated by AFG2, AFB1, and AFB2 with incidences of 8.7%, 2.0%, and 0.67%, respectively. Some of the samples showed OTA concentrations above the maximum limit allowed by the European Union, which represents a health risk for consumers in Tunisia, where no legislation exists about the maximum content of mycotoxins in food.

Milk is a staple food that is essential for human nutrition because of its high nutrient content and health benefits. However, it is susceptible to being contaminated by Aflatoxin M1 (AFM1), which is a toxic metabolite of Aflatoxin B1 (AFB1) presented in cow feeds. This research investigated AFM1 in Tunisian raw cow milk samples. A total of 122 samples were collected at random from two different regions in 2022 (Beja and Mahdia). AFM1 was extracted from milk using the QuEChERS method, and contamination amounts were determined using liquid chromatography (HPLC) coupled with fluorescence detection (FD). Good recoveries were shown with intra-day and inter-day precisions of 97 and 103%, respectively, and detection and quantification levels of 0.003 and 0.01 µg/L, respectively. AFM1 was found in 97.54% of the samples, with amounts varying from values below the LOQ to 197.37 µg/L. Lower AFM1 was observed in Mahdia (mean: 39.37 µg/L), respectively. In positive samples, all AFM1 concentrations exceeded the EU maximum permitted level (0.050 µg/L) for AFM1 in milk. In Tunisia, a maximum permitted level for AFM1 in milk and milk products has not been established. The risk assessment of AFM1 was also determined. Briefly, the estimated intake amount of AFM1 by Tunisian adults through raw cow milk consumption was 0.032 µg/kg body weight/day. The Margin of Exposure (MOE) values obtained were lower than 10,000. According to the findings, controls as well as the establishment of regulations for AFM1 in milk are required in Tunisia.

Mycotoxin contamination in wheat, a staple food critical to human nutrition, poses significant public health concerns. This study investigated the natural occurrence of 17 mycotoxins in Tunisian durum wheat, assessed the influence of soil tillage practices on mycotoxin contamination, and performed an associated exposure risk assessment. A total of 167 wheat samples were randomly collected over two years (2021 and 2022) from fields managed under conventional tillage (CT) and no-tillage (NT) systems during both pre- and post-harvest periods. Mycotoxins were extracted using the QuEChERS method and quantified via UHPLC-MS/MS. The results demonstrated contamination by ZEN, DON, OTA, ENA1, ENB, and ENB1. Among regulated mycotoxins, OTA was the most prevalent, detected in 68 out of 167 samples with a mean concentration of 1.85 µg/kg. ZEN was the most abundant, detected in 65 samples with a mean concentration of 26.85 µg/kg, while DON was less frequently detected in 62 samples with a mean concentration of 0.68 µg/kg. Regarding emerging mycotoxins, ENB was the most prevalent and abundant, found in 51 samples with a mean concentration of 10.13 µg/kg; ENB1 and ENA1 were detected in 20 and 10 samples, with mean concentrations of 3.38 µg/kg and 1.69 µg/kg, respectively. Furthermore, mycotoxin concentrations varied according to agricultural practices. DON, ZEN, ENA1, ENB, and ENB1 showed higher frequencies and concentrations (ranging from 0.08 to 210.11 µg/kg) in samples collected during the 2021 pre-harvest period from NT fields. In contrast, OTA exhibited greater prevalence and higher concentrations (ranging from 2.33 to 9.78 µ/kg) in samples collected during the 2022 post-harvest period from CT fields. The Estimated Daily Intake (EDI) of mycotoxins by Tunisian adults was calculated based on contamination levels in raw durum wheat from fields under NT and CT practices, resulting in the following values (ng/kg bw/day), with the first value corresponding to NT samples and the second to CT samples: OTA (17.3; 20.8), ZEN (466.3; 194.0), DON (8.0; 7.56), ENA1 (4.30; 18.85), ENB (105.17; 121.08), and ENB1 (49.91; 40.91). Both the Margin of Exposure (MOE) values for OTA and the Hazard Quotients (HQ) for ZEN and DON exceeded established safety thresholds, indicating potential health risks for Tunisian adults. These findings highlight the urgent need to implement stricter mycotoxin regulations in Tunisia and enhance surveillance systems. Further research is warranted to elucidate the mechanisms by which soil tillage practices influence mycotoxin contamination and to develop targeted mitigation strategies to ensure food safety.

The contamination of cereals by mycotoxins represents a major concern due to their harmful effects on human health and food quality. The current study investigated the occurrence of major mycotoxins (AFB1, AFB2, AFG1, AFG2, OTA, ENA, ENA1, ENB, and ENB1) in 158 raw cereal samples (durum wheat, barley, and maize) collected from two Tunisian regions: Beja (continental region) and Mahdia (coastal region). Mycotoxins were extracted using the QuEChERS method and quantified by UHPLC–MS/MS. Several mycotoxins were detected at high levels across all the cereals. In the Beja region, durum wheat was contaminated with AFB1, AFG1, ENA, ENA1, ENB, and ENB1, with ENB being the most frequent (70%). Mahdia durum wheat was contaminated only with ENA, ENA1, ENB, and ENB1, with ENB1 being the most prevalent (22.6%). Barley from both regions was contaminated only with ENs. The ENB was the most frequent (Beja 66%, Mahdia 28.6%). Maize from Beja was contaminated by OTA and ENs, with ENA1 being the most frequent (22.5%), while maize from Mahdia was contaminated by AFB1, AFB2, AFG2, and ENs (AFB1 was the most frequent, 35%). All wheat samples contaminated with AFG1 (6.55%) exceeded the European Union maximum limit for AFs in cereals (4 µg/kg). Similarly, maize samples contaminated with AFB1 (17.5%), AFB2 (10%), and AFG2 (2.5%) exceeded the EU maximum limit for aflatoxins in maize (10 µg/kg). Additionally, maize samples contaminated with OTA (5%) exceeded the EU maximum limit for OTA in unprocessed cereals (5 µg/kg). The co-occurrence of multiple mycotoxins was observed in all cereal types, with up to six different mycotoxins detected in a single sample. Dietary risk assessment revealed high EDIs of AFB1, AFG1, and ENs through the consumption of wheat and barley by Tunisian adults. The calculated MOE values for AFB1 and AFG1 in wheat were below 10,000 (MOE = 1190 for AFB1 and 2.5 for AFG1), suggesting a potential health concern associated with dietary exposure. Despite this potential risk, AFB1 and AFG1 were detected in only 3% and 7% of the analyzed samples, respectively. These results highlight the need for regular monitoring and the establishment of regulations to control mycotoxins in Tunisian cereals.