Structural similarity analysis and AI-predicted binding mode of bitter flavonoids in pomelo (Citrus maxima) using HRMS-based metabolomics and molecular fingerprinting.

The bitterness of pomelo is primarily attributed to flavonoids, which significantly impacts consumer acceptance. However, the molecular basis by which specific structural features of flavonoids interact with bitter taste receptors remains unclear. This study combined UHPLC-Orbitrap-MS, electronic tongue, and in silico prediction to analyze flavonoids in eight pomelo cultivars. A total of 35 flavonoids were identified, among which 25 were classified as potentially bitter. The key findings revealed that the neohesperidosyl group and the 4'/5'-hydroxyls groups on the B-ring are crucial for bitterness, while methoxylation mitigates bitterness. Analysis of the flavonoid-TAS2R14 complex showed that a key hydrogen bond between the B-ring hydroxyl group and the SER65 residue governs bitterness intensity. These findings elucidate how flavonoid structures modulate bitter receptor interactions, providing important theoretical foundations for flavor-guided breeding and the development of natural bitterness modulators.