Effect of roasting and creaming processes on hazelnut allergenicity: a proteomic and immunological investigation

Hazelnuts are among the most common causes of food-induced allergic reactions [1,2], and thermal or mechanical processing may influence their allergenic potential [3]. This study aimed to investigate the impact of roasting and creaming on hazelnut protein profile and allergenic potential using a multidisciplinary approach that combines electrophoresis, immunoblot analysis, and high-resolution mass spectrometry-based proteomics. Protein extracts were prepared from raw and roasted hazelnuts, and from two hazelnut pastes (raw and roasted), following a defatting step. After protein quantification, SDS-PAGE experiments were performed to assess differences in protein profiles across the various samples. Distinct patterns were observed between raw and processed samples, particularly in the 40–65 kDa region, where roasting appeared to affect the integrity and intensity of specific protein bands. Western blot analyses were carried out using sera from hazelnut-allergic individuals to detect IgE-binding proteins. Several reactive bands were identified, notably in the 60, 45, 30, and 23 kDa regions. Each protein band was excised and subjected to in-gel tryptic digestion followed by LC-HRMS/MS analysis. Bioinformatic analysis revealed the consistent presence of key hazelnut allergens, including Cor a 9, Cor a 11, and Cor a 14, across all samples. Notably, Cor a 9 and Cor a 11 were detected even in samples undergoing roasting and creaming, suggesting their stability and persistence after processing. Additionally, proteins from Carpinus fangiana with high sequence similarity to known hazelnut allergens (particularly Cor a 9) were identified, possibly reflecting isoforms not yet annotated in current hazelnut-specific databases. Complementary bottom-up proteomics of total protein digests confirmed the presence of allergenic proteins and highlighted the detection of several peptide modifications. Immunoblotting results showed that processed samples retained IgE-reactive bands, indicating a retained allergenicity. Interestingly, the roasted hazelnut and both hazelnut pastes displayed similar IgE-binding profiles, whereas raw hazelnuts exhibited some distinct reactive bands, suggesting potential structural changes induced by thermal or mechanical treatment. In conclusion, the results highlight that both roasting and creaming do not significantly reduce the allergenicity of hazelnuts. The persistence of key allergens even after processing underlines the need to have at disposal reliable analytical methods for allergen detection and risk assessment plan to protect allergic consumers.