Based on a non-targeted metabolomic approach, a comprehensive analysis of the metabolic profiles of two raspberry cultivars, Selina (TSM) and Tulameen (CSM), was conducted to explore the potential for further processing of these two varieties. A total of 752 metabolites were identified in both raspberry varieties, with 375 metabolites detected in negative ion mode and additional metabolites identified in positive ion mode. The total number of differential metabolites between TSM and CSM was 62, with orthogonal partial least squares discriminant analysis (OPLS-DA) revealing that the main differential metabolites between the two varieties were phenolic compounds, organic acids, and alkaloids. KEGG pathway analysis identified the top three significantly different metabolic pathways as biosynthesis of amino acids
valine, leucine, and isoleucine biosynthesis
and alanine, aspartate, and glutamate metabolism. In conclusion, this study comprehensively analyzed the metabolic characteristics of TSM and CSM raspberry varieties, providing valuable insights and revealing their potential application prospects. Compared to previous studies, this research provides new insights into the metabolic differences between TSM and CSM cultivars, specifically highlighting newly identified metabolites that were not reported before. These findings include novel differences in phenolic compounds and alkaloids, which play important roles in plant stress response and have potential health benefits. This enhances our understanding of the unique properties of each cultivar and underscores the biological functions of the newly identified metabolites, contributing to the broader field of functional food development.