*4.3. Samples and Reference Materials*

Poultry and swine feedstuff samples (total *n* = 99) were collected in 2017 and 2018 by Veterinary Inspectorate officers working with feed manufacturers, in accordance to Commission Regulation (EC) NO. 2009/152 [48]. Delivered samples were milled using a ZM 200 ultra-centrifugal high-speed instrument with 1 mm sieve (Retsch GmbH, Haan, Germany;) and stored in a dark place at room temperature until analysis. A validation study was conducted using low contamination feed samples (pseudo-blanks) [49] with DON concentration of 50 ± 13 µg/kg (Figure 6).

**Figure 6.** Chromatogram of pseudo-blank sample with DON concentration of 50 ± 13 µg/kg.

For confirmation of the method trueness, three reference materials (RMs) were tested: maize (TET030RM; Fapas, Fera Science, York, UK), maize (12199.15-G; Chiron AS, Trondheim, Norway) and wheat (TET007RM; Fapas) (Table 2). Moreover, the method was verified in the "Deoxynivalenol and related compounds in food and feed matrices"; EURLPT-MP01 proficiency test on DON, 3Ac-DON, 15Ac-DON and DON-3Glc in wheat and maize carried out by the European Union Reference Laboratory mycotoxins and plant toxins.
