Entering the ppq level in mycotoxin biomarker analysis

There is increasing awareness of the need to understand the patterns and levels of dietary mycotoxins mixtures. To achieve this goal, mycotoxins and their metabolites are being measured in urine. We previously published a multi-mycotoxin LC-MS/MS survey of 120 Nigerian urine samples, however we were aware that in measuring the mixture a compromise on assay sensitivity was incurred compared to some established single-analyte methods. In order to allow more accurate and comparative dietary mycotoxin exposure determination, a highly sensitive urinary assay, including 13C-labelled or deuterated internal standard, was developed and validated for seven important mycotoxins (and several metabolites) including AFM1, FB1, OTA, DON, DOM-1, NIV, ZEN, CIT, dihydro-CIT, α- and β-ZEL, and AOH. The assay includes β- glucuronidase pre-treatment, SPE enrichment, UHPLC separation and high precision triple quadrupole MS/MS detection. Analytical sensitivity was significantly improved, providing parity or better compared to existing single analyte methods, but in a single more robust multi-mycotoxin assay. On re-analysis of the Nigerian urine samples we observed an increased number of positive samples for all tested mycotoxins: from 50.8% to 100%, with highest occurrence of ZEN (82%), OTA (76%), AFM1 (73%), and FB1 (71%). Overall, a significant share of samples (68 out of 120 ; 57%) were contaminated with both AFM1 and FB1.