Hippophae rhamnoides L. Fruits Reduce the Oxidative Stress in Human Blood Platelets and Plasma

Abstract

Effects of the phenolic fraction from Hippophae rhamnoides fruits on the production of thiobarbituric acid reactive substances (TBARS, a marker of lipid peroxidation) and the generation of superoxide anion (O2 (-∙)) in human blood platelets (resting platelets and platelets stimulated by a strong physiological agonist, thrombin) were studied in vitro. We also examined antioxidant properties of this fraction against human plasma lipid peroxidation and protein carbonylation induced by a strong biological oxidant, hydrogen peroxide (H2O2) or H2O2/Fe (a donor of hydroxyl radicals). The tested fraction of H. rhamnoides (0.5- 50 µg/mL; the incubation time: 15 and 60 min) inhibited lipid peroxidation induced by H2O2 or H2O2/Fe. The H. rhamnoides phenolic fraction inhibited not only plasma lipid peroxidation, but also plasma protein carbonylation stimulated by H2O2 or H2O2/Fe. Moreover, the level of O2 (-∙) in platelets significantly decreased. In comparative experiments, the H. rhamnoides fraction was a more effective antioxidant than aronia extract or grape seed extract (at the highest tested concentration, 50 µg/mL). The obtained results suggest that H. rhamnoides fruits may be a new, promising source of natural compounds with antioxidant and antiplatelet activity beneficial not only for healthy people, but also for those with oxidative stress-associated diseases.

Figure 1

Effects of the phenolic fraction from fruits of H. rhamnoides (0.5–50 µg/mL; 15 and 60 min) on plasma lipid peroxidation induced by H₂O₂ (a) and plasma lipid peroxidation induced by H₂O₂/Fe (b). Data represent means ± standard error (SE) of 4-5. The effect of five different concentrations of the tested fraction (0.5, 1, 5, 10, and 50 μg/mL) was statistically significant according to ANOVA I test, p < 0.05 for concentrations 0.5, 5, 10, and 50 μg/mL (for 15 and 60 min) (a); p > 0.05 for concentration 1 μg/mL (for 15 and 60 min) (a); p < 0.05 for concentrations 0.5, 1, and 5 μg/mL (for 15 min) (b); p < 0.02 for concentrations 10 and 50 μg/mL (for 15 min) (b); p < 0.02 for concentration 50 μg/mL (for 60 min) (b); p > 0.05 for concentrations 0.5, 1, 5, and 10 μg/mL (for 60 min) (b).

Figure 2

Effects of the phenolic fraction from fruits of H. rhamnoides (0.5–50 µg/mL; 15 and 60 min) on plasma protein carbonylation induced by H₂O₂ (a) and plasma protein carbonylation induced by H₂O₂/Fe (b). Data represent means ± standard error (SE) of 4. The effect of five different concentrations of the tested fraction (0.5, 1, 5, 10, and 50 μg/mL) was statistically significant according to ANOVA I test, p < 0.05 for concentration 50 μg/mL (for 15 min) (a); p < 0.05 for concentrations 10 and 50 μg/mL (for 60 min) (a); p > 0.05 for concentrations 0.5, 1, 5, and 10 μg/mL (for 15 min) (a); p > 0.05 for concentrations 0.5, 1, and 5 μg/mL (for 60 min) (a); p < 0.05 for concentration 50 μg/mL (for 15 min) (b); p < 0.05 for concentrations 5, 10, and 50 μg/mL (for 60 min) (b); p > 0.05 for concentrations 0.5, 1, 5, and 10 μg/mL (for 15 min) (b); p > 0.05 for concentrations 0.5 and 1 μg/mL (for 60 min) (b).

Figure 3

Effects of the phenolic fraction from fruits of H. rhamnoides (0.5–50 µg/mL; 15 min) on lipid peroxidation in resting blood platelets and blood platelets activated by thrombin. Data represent means of 3-4 donors ± standard error (SE). The effect of five different concentrations of the tested fraction (0.5, 1, 5, 10, and 50 μg/mL) was statistically significant according to ANOVA I test, p < 0.05 for concentrations 1, 5, 10, and 50 μg/mL; p > 0.05 for concentration 0.5 μg/mL.

Figure 4

Effects of the phenolic fraction from fruits of H. rhamnoides (0.5–50 µg/mL; 15 min) on O₂ ^−∙ generation in resting blood platelets and blood platelets activated by thrombin. Data represent means of 4–6 donors ± standard error (SE). The effect of five different concentrations of the tested fraction (0.5, 1, 5, 10, and 50 μg/mL) was statistically significant according to ANOVA I test, p < 0.05 for concentrations 5, 10, and 50 μg/mL (resting platelets); p < 0.05 for concentrations 1, 5, 10, and 50 μg/mL (platelets activated by thrombin); p > 0.05 for concentration 0.5 μg/mL (resting platelets and platelets activated by thrombin); p > 0.05 for concentration 1 μg/mL (resting platelets).

Figure 5

Effects of the phenolic fraction from fruits of H. rhamnoides (50 µg/mL; 15 min), A. melanocarpa extract (50 µg/mL, 15 min), and grape seed extract (50 µg/mL, 15 min) on plasma lipid peroxidation induced by H₂O₂ or H₂O₂/Fe. The results represent 3–8 independent experiments and are expressed as means ± SE. The effects were significant according to ANOVA I test, for peroxidation induced by H₂O₂: H. rhamnoides fraction-treated plasma versus A. melanocarpa extract-treated plasma (p < 0.02); H. rhamnoides fraction-treated plasma versus grape seed extract-treated plasma (p < 0.05); for peroxidation induced by H₂O₂/Fe: H. rhamnoides fraction-treated plasma versus A. melanocarpa extract-treated plasma (p < 0.01); H. rhamnoides fraction-treated plasma versus grape seed extract-treated plasma (p < 0.01).

Figure 6

Effects of the phenolic fraction from fruits of H. rhamnoides (50 µg/mL; 15 min), A. melanocarpa extract (50 µg/mL, 15 min), and grape seed extract (50 µg/mL, 15 min) on autoperoxidation of blood platelets and platelet lipid peroxidation induced by thrombin. The results represent 3-4 independent experiments and are expressed as means ± SE. The effects were significant according to ANOVA I test, for autoperoxidation: H. rhamnoides fraction-treated platelets versus A. melanocarpa extract-treated platelets (p < 0.01); H. rhamnoides fraction-treated platelets versus grape seed extract-treated platelets (p < 0.01); for peroxidation induced by thrombin: H. rhamnoides fraction-treated platelets versus A. melanocarpa extract-treated platelets (p < 0.05); H. rhamnoides fraction-treated platelets versus grape seed extract-treated platelets (p < 0.05).

Figure 7

Effects of the phenolic fraction from fruits of H. rhamnoides (50 µg/mL; 15 min), A. melanocarpa extract (50 µg/mL, 15 min), and grape seed extract (50 µg/mL, 15 min) on O₂ ^−∙ generation in resting blood platelets and blood platelets activated by thrombin. The results represent 3–6 independent experiments and are expressed as means ± SE. The effects were significant according to ANOVA I test, for resting platelets: H. rhamnoides fraction-treated platelets versus A. melanocarpa extract-treated platelets (p > 0.05); H. rhamnoides fraction-treated platelets versus grape seed extract-treated platelets (p > 0.05); for peroxidation induced by thrombin: H. rhamnoides fraction-treated platelets versus A. melanocarpa extract-treated platelets (p < 0.05); H. rhamnoides fraction-treated platelets versus grape seed extract-treated platelets (p < 0.05).