*2.1. LC-MS*/*MS Optimisation*

The optimisation of LC-MS/MS parameters was accomplished by directly applying tuning solutions of the selected mycotoxins at concentrations of 1 µg/mL each, using 0.1% CH3COOH and MeOH as a mobile phase. The MRM mode was used and the analytes were ionized in both positive (ESI+) and negative modes (ESI−) (Table S1). In the case of positional isomers, proper quantification of 3Ac-DON and 15Ac-DON cannot be achieved through the difference of the product ion in MRM mode. In positive mode (339 *m*/*z*) the most abundant parent ion was the same for both analytes, but different product ions were chosen (339/231.2 and 339/279.1 for 3Ac-DON and 339/321.2 and 339/261.2 for 15Ac-DON) [27,28]. However, other studies show higher ionisation in positive mode with NH4<sup>+</sup> adducts for 3Ac-DON and 15-AcDON [13,23] or in negative modes [14,16]. If two different daughter ions for both isomeric forms are be chosen, positive false results could sometimes occur. For proper qualification and quantification it is therefore essential to have successful baseline separation of analytes.
