There is a critical need to better understand the patterns, levels and combinatory effects of exposures we are facing through our diet and environment. Mycotoxin mixtures are of particular concern due to chronic low dose exposures caused by naturally contaminated food. To facilitate new insights into their role in chronic disease, mycotoxins and their metabolites are quantified in bio-fluids as biomarkers of exposure. Here, we describe a highly sensitive urinary assay based on ultra-high performance liquid chromatography - tandem mass spectrometer (UHPLC-MS/MS) and 13C-labelled or deuterated internal standards covering the most relevant regulated and emerging mycotoxins. Utilizing enzymatic pre-treatment, solid phase extraction and UHPLC separation, the sensitivity of the method was significantly higher (10-160x lower LODs) than in a previously described method used for comparison purpose, and stable isotopes provided compensation for challenging matrix effects. This method was in-house validated and applied to re-assess mycotoxin exposure in urine samples obtained from Nigerian children, adolescent and adults, naturally exposed through their regular diet. Owing to the methods high sensitivity, biomarkers were detected in all samples. The mycoestrogen zearalenone was the most frequently detected contaminant (82%) but also ochratoxin A (76%), aflatoxin M1 (73%) and fumonisin B1 (71%) were quantified in a large share of urines. Overall, 57% of 120 urines were contaminated with both, aflatoxin M1 and fumonisin B1, and other co-exposures were frequent. These results clearly demonstrate the advanced performance of the method to assess lowest background exposures (pg mL-1 range) using a single, highly robust assay that will allow for the systematic investigation of low dose effects on human health.

中文翻译：

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多种尿液霉菌毒素暴露生物标志物的超灵敏、稳定同位素辅助定量

迫切需要更好地了解我们通过饮食和环境所面临的暴露的模式、水平和组合影响。由于自然污染的食物引起的慢性低剂量暴露，霉菌毒素混合物尤其受到关注。为了促进对其在慢性疾病中的作用的新见解，真菌毒素及其代谢物在生物流体中被量化为暴露的生物标志物。在这里，我们描述了一种基于超高效液相色谱 - 串联质谱仪 (UHPLC-MS/MS) 和 13C 标记或氘代内标的高灵敏度尿液检测，涵盖了最相关的监管和新兴真菌毒素。利用酶法预处理、固相萃取和UHPLC分离，该方法的灵敏度明显高于之前描述的用于比较目的的方法（LOD 低 10-160 倍），并且稳定同位素为具有挑战性的基质效应提供了补偿。该方法经过内部验证并应用于重新评估从尼日利亚儿童、青少年和成人获得的尿液样本中的霉菌毒素暴露情况，这些样本通过日常饮食自然暴露。由于该方法灵敏度高，在所有样品中均检测到生物标志物。霉菌雌激素玉米赤霉烯酮是最常检测到的污染物 (82%)，但在大部分尿液中也检测到赭曲霉毒素 A (76%)、黄曲霉毒素 M1 (73%) 和伏马菌素 B1 (71%)。总体而言，120 份尿液中有 57% 被黄曲霉毒素 M1 和伏马菌素 B1 污染，并且其他共同暴露也很常见。