Development of a simultaneous extraction and UHPLC-HESI-Q-Orbitrap-MS workflow for the determination of 10 pro-carcinogenic N-nitrosamines in animal derived foods

A sensitive and selective analytical method was developed for the simultaneous determination of 10 pro-carcinogenic N-nitrosamines (NAs), namely N-nitrosodimethylamine (NDMA), N-nitroso-N-methylethylamine (NMEA), N-nitrososarcosine (NSAR), N-nitrosomorpholine (NMOR), N-nitrosopyrrolidine (NPYR), N-nitrosodiethylamine (NDEA), N-nitrosopiperidine (NPIP), N-nitrosodi- n -propylamine (NDPA), N-nitrosomethylphenylamine (NMPhA), N-nitrosodi- n -butylamine (NDBA), in different processed food matrices including meat, fish and dairy products. A green liquid-solid extraction with ethyl acetate acidified (0.2% formic acid) was optimized by experimental design. UHPLC-Q-Orbitrap-MS with a heated electrospray ionization source was used for analysis, enabling accurate mass identification. Matrix-matched calibration curves were used for quantification, ensuring reliable results despite pronounced matrix effects in such complex foods. The method validation demonstrated good linearity (R² = 0.9750–0.9987), satisfactory recoveries (60–110%), low detection (0.348–0.848 µg/kg), and quantification (1.06–2.57 µg/kg) limits, and high reproducibility across all matrices. Application to commercial samples revealed trace levels of NPYR, NPIP and NDBA with significantly elevated amounts detected in some smoked, lipid-rich samples containing high nitrate and nitrite levels, suggesting a potential combination among these factors.