Simultaneous determination of major peanut allergens Ara h1 and Ara h2 in baked foodstuffs based on their signature peptides using ultra-performance liquid chromatography coupled to tandem mass spectrometry

Abstract

A robust ultra-high performance liquid chromatography method with tandem mass spectrometry was established for the quantitative analysis of the major peanut allergens Ara h1 and Ara h2 in baked foodstuffs based on their signature peptides. Tryptic peptides DLAFPGSGEQVEK and NLPQQCGLR were selected as the target analytes for Ara h1 and Ara h2, respectively. Their corresponding isotope-labeled peptides DL*AFPGSGEQV*EK and NL*PQQCGL*R were synthesized and used as internal standards to overcome matrix effects during sample analysis. The Ara h1 or Ara h2 content values in samples were calculated according to the measured signature peptide of Ara h1 or Ara h2 and the calibration curve established using the hydrolysates of their corresponding proteins. The limits of quantification for Ara h1 and Ara h2 were 0.30 and 0.13 mg kg⁻¹, respectively. The developed method had satisfactory accuracy, precision, and sensitivity according to in-house validation. The current method was successfully employed to measure the peanut allergens Ara h1 and Ara h2 in commercial baked foods, and could be considered a robust candidate method for promoting the quality control of commercial baked foods and providing accurate allergen warning information.