Multianalytical Approach to Understand Polyphenol-Mal d 1 Interactions to Predict Their Impact on the Allergenic Potential of Apples.

Interactions between phenolic compounds and the allergen Mal d 1 are discussed to be the reason for better tolerance of apple cultivars, which are rich in polyphenols. Because Mal d 1 is susceptible to proteolytic digestion and allergenic symptoms are usually restricted to the mouth and throat area, the release of native Mal d 1 during the oral phase is of particular interest. Therefore, we studied the release of Mal d 1 under different in vitro oral digestion conditions and revealed that only 6-15% of the total Mal d 1 present in apples is released. To investigate proposed polyphenol-Mal d 1 interactions, various analytical methods, e.g., isothermal titration calorimetry, ¹H-¹⁵N-HSQC NMR, and untargeted mass spectrometry, were applied. For monomeric polyphenols, only limited noncovalent interactions were observed, whereas oligomeric polyphenols and browning products caused aggregation. While covalent modifications were not detectable in apple samples, a Michael addition of epicatechin at cysteine 107 in r-Mal d 1.01 was observed.