Polyphenols exert beneficial effects in type 2 diabetes mellitus (T2DM). However, their mechanism of action remains largely unknown. Endothelial Akt-kinase plays a key role in the pathogenesis of cardiovascular complications in T2DM and therefore the modulation of its activity is of interest. This work aimed to characterize effects of structurally different polyphenols on Akt-phosphorylation (pAkt) in endothelial cells (Ea.hy926) and to describe structure-activity features. A comprehensive screening via ELISA quantified the effects of 44 polyphenols (10 µM) on pAkt Ser473. The most pronounced inhibitors were luteolin (44 ± 18%), quercetin (36 ± 8%), urolithin A (35 ± 12%), apigenin, fisetin, and resveratrol; (p < 0.01). The results were confirmed by Western blotting and complemented with corresponding experiments in HUVEC cells. A strong positive and statistically significant correlation between the mean inhibitory effects of the tested polyphenols on both Akt-residues Ser473 and Thr308 (r = 0.9478, p = 0.0003) was determined by immunoblotting. Interestingly, the structural characteristics favoring pAkt inhibition partially differed from structural features enhancing the compounds’ antioxidant activity. The present study is the first to quantitatively compare the influence of polyphenols from nine different structural subclasses on pAkt in endothelial cells. These effects might be advantageous in certain T2DM-complications involving over-activation of the Akt-pathway. The suggested molecular mode of action of polyphenols involving Akt-inhibition contributes to understanding their effects on the cellular level.

Polyphenols are plant‐derived natural compounds, which are attributed to various beneficial effects on human health. Therefore, they are extensively investigated in different experimental setups. These compounds exhibit favorable antidiabetic effects in various clinical trials. However, their mode of action is not completely elucidated. A potential molecular target of certain polyphenols could be the dipeptidyl peptidase IV (DPP IV). This enzyme is abundant in human serum and represents a serine exopeptidase that cleaves and thus rapidly deactivates incretin hormones which are responsible for adequate postprandial insulin secretion from the pancreas. Multiple inhibitors of DPP IV have already been approved as antidiabetic drugs. In the course of screening assays exploring in vitro DPP IV inhibitory activities of polyphenols, some implausible observations prompted a detailed analysis of the interaction of polyphenols with fluorescence signals. The results revealed that quercetin quenched the fluorescence intensity, while 6‐methoxyflavone and urolithin B enhanced fluorescence signals depending on particular experimental conditions such as solvent composition. Thus, individual polyphenols might conflict with the experimental setup of fluorescence‐based DPP IV inhibitor screening assays. To avoid false positive or false negative results, polyphenols under investigations should be carefully controlled for potential assay interferences.