Development and Validation of a Rapid UPLC-MS/MS Determination of Cereulide in ARA Oil and ARA Powder

Arachidonic acid (ARA) is an essential long-chain polyunsaturated fatty acid for infant growth and development, commonly added to infant formula and supplementary foods as ARA oil or ARA powder. Since December 2025, infant formula products have been recalled in several countries due to cereulide contamination in ARA ingredients. Cereulide, a heat-resistant emetic toxin produced by Bacillus cereus, presents a serious threat to infant health. However, specific analytical methods for cereulide detection in ARA raw materials remain scarce, hindering effective quality control and source prevention of food safety hazards. This study developed and validated a rapid ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the quantitative determination of cereulide in ARA oil and powder. Sample pretreatment was optimized according to matrix characteristics: for microencapsulated ARA powder, water dissolution was performed to release cereulide before acetonitrile extraction, whereas direct extraction was applied to ARA oil. Subsequent HLB-P pass-through solid-phase extraction effectively eliminated matrix interference in both matrices. The method exhibited excellent linearity (R² > 0.999) from 0.1 to 20 μg·L^-1, with an LOD of 0.03 μg·kg^-1 and an LOQ of 0.1 μg·kg^-1, respectively. Spike recoveries reached 90.7-107.0% for ARA oil (RSDs ≤ 6.5%) and 88.5-107.9% for ARA powder (RSDs ≤ 7.0%). This rapid and reliable method provides a critical analytical tool to monitor cereulide in ARA raw materials and mitigate infant safety hazards at the source.