Journal of Chromatography A (2010), 1217, 48, 7579--7585 DOI: 10.1016/j.chroma.2010.10.037
A bicarbonate buffer-based extraction method for the simultaneous analysis of five nut allergens (Ana o 2, cashew-nut; Cor a 9, hazelnut; Pru 1, almond; Ara h3/4, peanut; Jug r 4, walnut) in cereals and biscuits using liquid chromatography–electrospray-linear ion trap-tandem mass spectrometry (LC–ESI-LIT-MS2) was developed and validated. The method was based on our earlier published LC–MS2-based method in a research frame aimed at the identification and determination of hidden allergens in foods by using selective biomarker peptides. A C18 particle-packed column and a silica-based C18 monolithic column were compared in terms of chromatographic performances, such as peak shape, resolution, analysis time and selectivity. The C18 particle-packed column exhibited better performances and was further used for method development and validation. By operating under MS2 selected reaction monitoring (SRM) acquisition mode, linearity, limits of detection (LOD) and quantitation, trueness and precision were evaluated on breakfast samples enriched with a mix of the five nuts. Good linearity of the matrix matched-calibration curves was obtained and detection limit values generally varied from 14 to 55 mg nut/kg matrix. Recoveries were in the 76 ± 4% to 94 ± 3% range with RSD <15%. The capabilities of LIT to perform MSn fragmentation was exploited to improve selectivity of the analysis, and the LC–(SRM) MS2 method was compared in terms of LOD, linearity, precision and accuracy with a LC–(SRM) MS3 method. Finally, both the LC–MS2 and LC–MS3 methods were successfully applied to the analysis of nut traces in commercially available breakfast cereals and biscuits.