Development, Validation, and Application of a Cold-Induced Phase Separation Method for 10 Nitroimidazole Residues in Meats and Egg

This study developed a novel analytical method based on cold-induced phase separation (CIPS) coupled with ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) for simultaneous determination of 10 nitroimidazoles (NIMs) in various animal-derived food matrices, such as pork, chicken, egg, beef, and mutton. Systematic optimization of critical parameters (acetonitrile/water ratio 8:2 v/v, neutral pH) yielded excellent analytical performance, with quantification limits (LOQs) of 0.1–0.5 μg/kg, recoveries of 78.0–106.5%, and relative standard deviations below 20%, fully compliant with the Codex Alimentarius standards. The CIPS technique effectively precipitated lipids and polar interferents through freezing, significantly reducing co-extractive interference and maintaining matrix effects (MEs) within − 13 to 15.6%, thereby enabling accurate quantification without matrix-matched calibration while improving throughput and reducing costs. Method validation demonstrated excellent applicability and reliability across different matrices. The simple, low-cost method provides an efficient solution for monitoring trace NIMs in animal-derived foods, demonstrating potential for food safety applications.