Impact of boiling on the allergens in fish bone samples identified by microfluidic chips and MALDI-TOF MS

Fish bones hold significant potential in the food industry. Investigating the allergenic characteristics of fish bone food products can enhance our understanding of fish allergies. In this study, the allergenic proteins in aqueous extracts from large yellow croaker (Larimichthys crocea) bones boiled for various durations were analyzed and identified using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and protein database searches. Tryptic products of potential allergens were detected using MALDI-TOF MS and further identified through analysis with Bruker's FlexAnalysis software and the FindMod tool available on the ExPASy proteomics server of the SIB (http://expasy.org/). 25 proteins related to allergy were identified. Two proteins reported as allergens were identified, along with twenty-three (23) proteins that, to the best knowledge of the authors, have not been reported as allergens. At least one aligned tryptic peptide were detected for 17 of the 25 proteins. The 17 potentially allergenic proteins exhibit peptide coverage ranging from 6.72 % to as high as 80 %. The results indicate that boiling the bones for 10 min releases many potentially allergenic proteins. However, the sensitization of most proteins diminishes when the bones are boiled for 30 to 120 min. Despite this, boiling does not completely eliminate the allergenicity of proteins in large yellow croaker bone samples. It is recommended to boil large yellow croaker fish bones for 30 min or longer to reduce most of the protein allergenicity when processing fish bones. Boiling may affect the allergenicity of proteins in fish bones by modifying their structure. These findings provide valuable guidance for monitoring allergens in aquatic food by-products, promisingly assisting to ensure the safety of allergy sufferers. Additionally, this research offers a reference for allergy management and the development of diagnostic reagents derived from aquatic food by-products.