Simultaneous Determination of Deoxynivalenol,Deoxynivalenol-3-Glucoside and Nivalenol in Wheat Grains by HPLC-PDA with Immunoaffinity Column Cleanup

Deoxynivalenol-3-glucoside (D3G) is a modified mycotoxin formed by the metabolism of plants through the conjugation of deoxynivalenol (DON) with glucose. Toxicology studies of D3G for human and animal health are still under investigation, and the development of practical and reliable methods for its direct determination, especially in cereal matrices, is of great importance. In the present study, a methodology for simultaneous determination of D3G, DON, and nivalenol (NIV) in wheat grains, using immunoaffinity column (IAC) cleanup, separation by C18 column and detection by ultraviolet (UV) absorption, was optimized and in-house validated. The results demonstrated adequate values of D3G recovery from IAC and spiked samples. Intraday precision, linearity, limit of detection and limit of quantification (LOQ) were also adequate for the determination of these mycotoxins. Range of applicability varied from 47.1 to 1000 μg/kg for D3G and from 31.3 to 1000 μg/kg for DON and NIV, with recovery ranging from 84.7?±?7.2 % to 112.3?±?8.1 %. A high incidence of D3G (41.2 %, all samples <LOQ) was verified in commercial samples of wheat grains and whole wheat-flour (n?=?17). Also, the presence of D3G occurred simultaneously with DON in 100 % of the D3G-positive samples. DON levels varied from <LOQ to 325.8 μg/kg, and NIV was detected in only 29.5 % (from <LOQ to 140.6 μg/kg). To the best of our knowledge, this is the first method of simultaneous determination of NIV, D3G, and DON by high-performance liquid chromatography with photodiode array detector (HPLC-PDA) reported until now.