Exposure to Mycotoxin-Mixtures via Breast Milk: An Ultra-Sensitive LC-MS/MS Biomonitoring Approach

Abstract

Exposure to natural food contaminants during infancy may influence health consequences later in life. Hence, breast milk may serve as a vehicle to transport these contaminants, including mycotoxins, from mothers to their infants. Analytical methods mostly focused on single exposures in the past, thus neglecting co-occurrences and mixture effects. Here, we present a highly sensitive multi-biomarker approach by a sophisticated combination of steps during sample preparation including a Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) extraction followed by a solid phase extraction (SPE) cleanup and utilizing stable isotopes for compensating challenging matrix effects. The assay was validated in-house, reaching limits of detection (LOD) for all 34 analytes in the range of 0.1 to 300 ng/L with satisfying extraction efficiencies (75-109%) and stable intermediate precisions (1-18%) for most analytes. Compared to a similar multi-mycotoxin assay for breast milk, LOD values were decreased by a factor of 2-60x enabling the assessment of chronic low-dose exposures. The new method was applied to a small set of Nigerian breast milk samples (n = 3) to compare results with already published data. Concentration levels of samples that were found to be contaminated before could be confirmed. In addition, other mycotoxins were determined in all three samples, for example the newly investigated alternariol monomethyl ether (AME) was found for the first time in this biological fluid at concentrations up to 25 ng/L. Moreover, in a pooled Austrian sample obtained from a milk bank, trace amounts of multiple mycotoxins including AME (1.9 ng/L), beauvericin (5.4 ng/L), enniatin B (4.7 ng/L), enniatin B 1 (<LOQ), ochratoxin A (<LOQ) and the estrogenic zearalenone (<LOQ) confirmed co-occurrence and exposure even in a country with high food safety standards. In conclusion, the method facilitates the determination of mycotoxins at ultra-trace levels in breast milk, enabling the generation of occurrence data necessary for comprehensive co-exposure assessment.

Beauvericin and Enniatins: In Vitro Intestinal Effects

“…Another interesting research was conducted on pooled breast milk samples to assess possible contaminations by BEA and ENNs. The highest concentration of 6.2 ng/L was found for BEA, while ENN B was present at a level of 4.7 ng/L and ENN B1 was detected below its respective sample LOQ value [166].…”

Alternaria alternata Toxins Synergistically Activate the Aryl Hydrocarbon Receptor Pathway In Vitro

“…These uncovered co-contaminations of various Alternaria toxins in sunflower seed oil, tomato products, infant foods, and diverse fruit and vegetable juices [10][11][12]. Recently, a low abundance of AOH and AME was reported even in breast milk samples [13]. The two dibenzo-α-pyrones AOH and AME (Figure 1c,d) were previously reported to affect cell viability [14] and to moderately induce DNA strand breaks [15]; the latter occurring upon poisoning of topoisomerase II, as demonstrated for AOH [16].…”

The Occurrence of Aflatoxins in Nuts and Dry Nuts Packed in Four Different Plastic Packaging from the Romanian Market

“…As such, young children at the weaning age may be exposed through consumption of both household-formulated and industrially processed contaminated complementary foods [ 27 , 28 ]. Human exposure to aflatoxins from foods naturally contaminated was assessed by a biomonitoring point of view through biomarkers present in urine and breast milk samples [ 29 , 30 ]. Although low in concentration, aflatoxin traces were discovered even in blood samples of women and children.…”