||Polyphenols (PPH) represent a group of chemical substances found in plants, characterized by the
presence of more than one phenol unit. The largest and best studied polyphenols are the flavonoids.
Polyphenols contained in red wine have beneficial effects on cardiovascular health. The aim of our
research was a comparative and interactive study of four red wine polyphenols - resveratrol (R),
quercetin (Q), kaempferol (K) and isorhamnetin (IH) which may have a long term properties as the
increase of (NO) synthase expression, acting on the prometer activity when used alone or in
equimolar mixtures. To determine their activity, we performed a luciferase reporter gene assay on
EA.hy926 cells stably transfected with a luciferase reporter gene construct containing eNOS promoter.
The Bradford assay was also performed to evaluate the cytotoxicity and the differences in cell number.
The median effect equation, as an interaction analysis evaluating synergy or antagonism of the
combinations was done according to the principle of mass-action law and the dose reduction index
(DRI) was calculated for all mixtures. All single polyphenols activated eNOS promoter. The EC50 values
were in micromolar concentrations ranging from 3.44 μM (R2 = 0.96) for kaempferol to 9.89 μM for
isorhamnetin (R2 = 0.94). All mixtures activated eNOS promoter, but their interactions varied from
synergy (Q+R, Q+IH+K, Q+R+K and Q+R+IH+K), through additive (R+IH+K) to antagonistic interaction
(R+IH, R+K, Q+IH, Q+K, IH+K and R+Q+IH). In this study, we show for the first time that red wine
polyphenols activated eNOS promoter when used alone and in mixtures with different types of