The objective of the study was a comparative analysis of the antihemolytic activity against two Staphylococcus aureus strains (8325-4 and NCTC 5655) as well as α-hemolysin and of the membrane modifying action of four hydrolysable tannins with different molecular mass and flexibility: 3,6-bis-O-di-O-galloyl-1,2,4-triO -galloyl-β-d-glucose (T1), 1,2,3,4,5-penta-O-galloyl-β-d-glucose (T2), 3-O-galloyl-1,2-valoneoyl-β-d-glucose (T3) and 1,2-di-O-galloyl-4,6-valoneoyl-β-d-glucose (T4). We showed that all the compounds studied manifested antihemolytic effects in the range of 5-50 µM concentrations. However, the degree of the reduction of hemolysis by the investigated tannins was not uniform. A valoneoyl group-containing compounds (T3 and T4) were less active. Inhibition of the hemolysis induced by α-hemolysin was also noticed on preincubated with the tannins and subsequently washed erythrocytes. In this case the efficiency again depended on the tannin structure and could be represented by the following order: T1 > T2 > T4 > T3. We also found a relationship between the degree of antihemolytic activity of the tannins studied and their capacity to increase the ordering parameter of the erythrocyte membrane outer layer and to change zeta potential. Overall, our study showed a potential of the T1 and T2 tannins as anti-virulence agents. The results of this study using tannins with different combinations of molecular mass and flexibility shed additional light on the role of tannin structure in activity manifestation. Over the past few years, a significant increase in bacterial resistance to antibiotics and transference of resistance genes from animal to human strains has become a global medical problem. Therefore, constant search for new antimicrobial agents among them being compounds of plant origin, including polyphenols, is ongoing 1,2. In addition to the antibiotic approaches of combating bacteria, anti-virulence strategies have also been considered recently. An anti-virulent strategy assumes a direct effect of compounds on virulent factors by reducing

Erythrocytes are constantly exposed to ROS due to their function in the organism. High tension of oxygen, presence of hemoglobin iron and high concentration of polyunsaturated fatty acids in membrane make erythrocytes especially susceptible to oxidative stress. A comparison of the antioxidant activities of polyphenol-rich plant extracts containing hydrolysable tannins from sumac leaves (Rhus typhina L.) and condensed tannins from grape seeds (Vitis vinifera L.) showed that at the 5-50 µg/ml concentration range they reduced to the same extent hemolysis and glutathione, lipid and hemoglobin oxidation induced by erythrocyte treatment with 400 µM ONOO -or 1 mM HClO. However, extract (condensed tannins) from grape seeds in comparison with extract (hydrolysable tannins) from sumac leaves stabilized erythrocytes in hypotonic NaCl solutions weakly. Our data indicate that both hydrolysable and condensed tannins significantly decrease the fluidity of the surface of erythrocyte membranes but the effect of hydrolysable ones was more profound. In conclusion, our results indicate that extracts from sumac leaves (hydrolysable tannins) and grape seeds (condensed tannins) are very effective protectors against oxidative damage in erythrocytes.

Polyphenols, including tannins, are phytochemicals with pronounced antimicrobial properties. We studied the activity of two hydrolysable tannins, (i) gallotannin—1,2,3,4,5-penta-O-galloyl-β-D-glucose (PGG) and (ii) ellagitannin—1,2-di-O-galloyl-4,6-valoneoyl-β-D-glucose (dGVG), applied alone and in combination with antibiotics against Staphylococcus aureus strain 8324-4. We also evaluated the effect of these tannins on bacterial membrane integrity and fluidity and studied their interaction with membrane proteins and lipids. A correlation between the antimicrobial activity of the tannins and their membranotropic action depending on the tannin molecular structure has been demonstrated. We found that the antibacterial activity of PGG was stronger than dGVG, which can be associated with its larger flexibility, dipole moment, and hydrophobicity. In addition, we also noted the membrane effects of the tannins observed as an increase in the size of released bacterial membrane vesicles.

Platelet activation is closely associated with an increase in intracellular Ca(2+) concentration. Various compounds including Ca(2+) ionophores are able to trigger platelet aggregation by increasing intracellular Ca(2+) concentration in platelets. In the present study, we monitored the effect of the phytoestrogen ferutinin, which acts as a Ca(2+) ionophore in human blood platelets; its ionophore-like properties include upregulation of [Ca(2+)](in), activation of fibrinogen receptors and increased fibrinogen binding. Using spectrofluorometry and triple-color flow cytometry, we demonstrate that ferutinin increases [Ca(2+)](in) in both isolated platelets and platelets in whole blood from humans. This effect was almost completely blocked by the Ca(2+) chelator EGTA and was not sensitive to either Gd(3+) or econazole, which inhibit VOC and SOC channels, respectively. Nor was the effect sensitive to thapsigargin, an inhibitor of endoplasmic reticulum Ca(2+) ATPases. Ferutinin stimulated the expression of the active form of the GPIIb-IIIa complex and whole blood platelet aggregation only weakly and had no statistically significant effect on the binding of fibrinogen. These results demonstrate apparently inconsistent effects of ferutinin, which raises intraplatelet Ca(2+) concentration but fails to have an effect on spontaneous blood platelet aggregation. This pattern of responses may be caused by the combination of ferutinin's Ca(2+) ionophoric and estrogenic properties.

In this study, we found that the sumac tannins (Rhus typhina L.) exert to a various extent antihemolytic effects and antibacterial activity against Bacillus cereus and Pseudomonas aeruginosa depending on structural specificity of bacteria and different mechanisms of their toxic action. The sumac tannins exert the most expressed activity against B. cereus. The antihemolytic effect of the sumac tannins seems to be connected to a greater extent with their modifying action on the erythrocyte membrane structure. It was found that the sumac tannins are incorporated into the erythrocyte membrane, causing transformation of discocytes into echinocytes and enhancing the rigidity of the hydrophilic region of the lipid bilayer. We suggest that the embedding of sumac tannins into the membrane of erythrocytes alters their physical properties and, as a consequence, can limit their interaction with bacterial toxins.