Tolerable limits set for deoxynivalenol (DON) do not consider DON conjugates such as DON-3-glucoside. Conjugates may be metabolized in vivo to DON. Such masked mycotoxins and the potentially toxic Fusarium pigment are not routinely analyzed in cereals. We quantified DON, DON-3-glucoside, and a red Fusarium pigment in hard red spring wheat, using a new liquid chromatography–mass spectrometry method. Extraction protocols using centrifugation and shaking, and methanol–methylene chloride (50:50 [vol/vol]) or acetonitrile-water (84:16 [vol/vol]) were assessed. Purposively and randomly selected hard spring wheat samples were extracted with solvent filtered through a C18 column and analyzed using liquid chromatography–UV–mass spectrometry. Isocratic mobile phase (70% methanol) was used. Recoveries were 96.4% (DON) and 70.0% (DON-3-glucoside), while limits of detection were 1 μg/kg (MS) and 10 μg/kg (UV), and limits of quantification were 1 μg/kg (UV) and 0.5 μg/kg (MS), respectively. The pigment limits of quanti...