Sulfation Rewrites the Biological Profile of Dietary Flavonoids: Implications for Stability, Redox Signaling, and...

Dietary flavonoids undergo extensive Phase II metabolism, yet the functional consequences of sulfation for their biological activity remain poorly understood. In this study, we compared the physicochemical properties and cellular effects of nine structurally diverse flavonoids and their sulfated derivatives. Flavonoid 3'-O- or 4'-O-sulfates were prepared enzymatically using bacterial aryl sulfotransferase, structurally confirmed, and evaluated as potential metabolites that can be formed by human cytosolic sulfotransferases (SULTs). The results showed that sulfation significantly increased the stability of otherwise labile flavonoids under physiologically relevant conditions, while generally reducing their antiradical capacity and cytotoxicity toward murine RAW264.7 macrophage cells. In contrast to nonsulfated flavonoids, particularly fisetin and quercetin, all tested flavonoid sulfates failed to activate the NRF2 (nuclear factor erythroid 2 p45-related factor 2) pathway in RAW264.7 cells. On the other hand, some of the parent flavonoids as well as flavonoid sulfates partially attenuated lipopolysaccharide-stimulated expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS), indicating that sulfated metabolites retain modulatory effects on inflammatory responses. Using recombinant human sulfotransferases, we identified multiple enzymes responsible for the formation of the tested sulfates, confirming their relevance as plausible metabolites. Overall, our findings demonstrate that metabolic sulfation profoundly reshapes flavonoid stability and biological activity, underscoring the importance of considering conjugated metabolites when evaluating the health effects of dietary flavonoids.