Seafood fraud has become a global issue, threatening food security and safety. Adulteration, substitution, dilution, and incorrect labeling of seafood products are fraudulent practices that violate consumer safety. In this context, developing sensitive, robust, and high-throughput molecular tools for food and feed authentication is becoming crucial for regulatory purposes. Analytical approaches such as proteomics mass spectrometry have shown promise in detecting incorrectly labeled products. For the application of these tools, genome information is crucial, but currently, for many marine species of commercial importance, such information is unavailable. However, when combining proteomic analysis with spectral library matching, commercially important fish species were successfully identified, differentiated, and quantified in pure muscle samples and mixtures, even when genome information was scarce. This study further tested the previously developed spectral library matching approach to differentiate between 29 fish species from the North Sea and examined samples including individual fish, laboratory-prepared mixtures and commercial products. For authenticating libraries generated from 29 fish species, fresh muscle samples from the fish samples were matched against the reference spectral libraries. Species of the fresh fish samples were correctly authenticated using the spectral library approach. The same result was obtained when evaluating the laboratory-prepared mixtures. Furthermore, processed commercial products containing mixtures of two or three fish species were matched against these reference spectral libraries to test the accuracy and robustness of this method for authentication of fish species. The results indicated that the method is suitable for the authentication of fish species from highly processed samples such as fish cakes and burgers. The study shows that current and future challenges in food and feed authentication can efficiently be tackled by reference spectral libraries method when prospecting new resources in the Arctic.
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