Interactions Of Procyanidins From Nelumbo Nucifera Gaertn. Combined With Four Kind Of Antioxidants

There is an increasing interest in the use of combined antioxidants, because not only can they play synergistic antioxidant effects in vitro, but also synergistically inhibit oxidative damage induced by free radical in vivo. The purpose of this study is to evaluate the interactions of LSPC combined with four kinds of antioxidants in vitro, and select the combination with better synergism to research their amelioration effect on scopolamine-induced memory impairment mice, providing theoretical basis and reference for research and development combined antioxidants and anti-memory impairment functional foods. The important results were obtained as follows:1. The interaction of LSPC combined with four kinds of antioxidants in the DPPH free radical scavenging experimentThe antioxidant capacity of different antioxidants was determined by DPPH free radical scavenging ability in vitro. The result indicated that the IC50 of EGCG, LSPC, VC, Cys and NAC increased significantly, which suggested that their antioxidant capacity were significantly decreased.Evaluate the interactions by using Isobologram analysis method and response surface method. The results indicated that when the dose ratio of LSPC and Cys at 2:3, 4:3 and 1:3, the interaction index (γ) of the combination was significantly less than other dose ratios, namely two substances had equivalent dose or one substance slight more than the other, they had obvious synergistic effect, and when concentration range of two substances were 1-4 μg/mL and 1.5-6 μg/mL respectively, they also showed synergism. When the dose ratio of LSPC and NAC at 1:3 and 2:3, the interaction index (y) of the combination was significantly less than other dose ratios, namely two substances had equivalent dose or NAC slight more than LSPC, they had obvious synergistic effect, and when concentration range of two substances were 1-4 μg/mL and 1.5-6 μg/mL respectively, they also showed synergism. When the dose ratio of LSPC and VC at 1:8 and 1:2, the interaction index (y) of the combination was significantly less than other dose ratios, namely the dose of VC were much more than LSPC, they had obvious synergistic effect, but when concentration range of two substances were 1.5-6 μg/mL and 1.5-6 μg/mL respectively, they showed additive action. When the dose ratio of LSPC and EGCG at 3:16 and 3:8, the interaction index (y) of the combination was significantly less than other dose ratios, namely the dose of EGCG were much more than LSPC, they had obvious synergistic effect, but when concentration range of two substances were 1.5-6 μg/mL and 1-4 μg/mL respectively, they showed antagonism.To sum up, in the selected ratio and concentration range, both LSPC combined with Cys and LSPC combined with NAC had synergistic antioxidant effect, especially when the dose ratio of LSPC and NAC at 1:3 and 2:3, the interaction index (y) of the combination was the least, showing the best synergism.2. Amelioration effect of LSPC combined with NAC on scopolamine-induced memory impairment miceComparing to model group, both sole substance and LSPC combined with NAC can significantly decrease number of errors in Y-maze test, reverse the increment of AchE activity in brain, increase the level of T-AOC, SOD and GPx activity, the ratio of GPx/SOD both in brain and serum, and inhibit the enhancement of tNOS and nNOS activity and the mRNA expression of nNOS in brain. Compared with sole LSPC and NAC, high dose LSPC combined with NAC can much more significantly increase the antioxidant system activity of brain and serum, and inhibit the NO system activity of brain.LSPC combined with NAC can significantly ameliorate the memory impairment induced by scopolamine in mice by synergistic improving brain cholinergic activity, increasing antioxidant activity of brain and serum, and inhibiting brain NO system activity.3. Xanthine oxidase inhibition capacity of LSPC and other natural productsThe result of xanthine oxidase inhibition assay indicated that the IC50 of allopurinol, EGCG, green tea extract and LSPC increased significantly, and the IC50 of them were lower than 100 μg/mL, while the IC50 of procyanidins from Litchi chinensis and cranberry extract were higher than 200 μg/mL, which suggested that their inhibition capacity were significantly decreased.Evaluate the interactions by using Isobologram analysis method and response surface method. The results indicated that when the dose ratio of LSPC and EGCG at 1:1 and 1:2, the interaction index (γ) of the combination was significantly less than other dose ratios, namely two substances had equivalent dose or EGCG slight more than LSPC, they had obvious synergistic effect, but when concentration range of two substances were 10-40 μg/mL and 10-40 μg/mL respectively, they showed additive action.