小球藻水萃物對細胞與老化小鼠氧化損傷之影響

Abstract

本研究旨在探討小球藻水萃物(Chlorella vulgaris extract, CE)對 D-半乳糖誘導氧化壓力小鼠之抗氧化影響。細胞試驗利用過氧化氫誘導 MRC-5 細胞氧化壓力作為細胞模式,觀察小球藻水萃物對細胞存活率及過氧化氫誘導細胞損傷之影響。動物試驗分為六組,分別為未誘導之對照組(Control)、D-半乳糖誘導之誘導組(D-gal)、維生素 C 之正對照組(Vitamin C)及小球藻水萃物之低劑量(CE- L)、中劑量(CE- M)、高劑量(CE- H)處理組,除對照組外,以皮下注射 D-半乳糖誘導氧化壓力老化小鼠為試驗動物。實驗為期八周,進行新物體辨識試驗與水迷宮游泳行為試驗,並於試驗結束時採血分析血液抗氧化酵素活性及脂質過氧化物含量。研究結果顯示經破壁處理取得之小球藻水萃物具有顯著較高的萃出產率與水萃物之含量(P<0.05)。此外,在處理濃度 2mg/ml 以下時,對細胞不具有細胞毒性,但對於 200 μM 及 250 μM 過氧化氫誘導之細胞損傷沒有顯著的保護作用。在動物試驗中,小球藻水萃物能顯著改善由 D-半乳糖誘導引起小鼠體重下降、脾臟腫大、抗氧化酵素穀胱甘肽氧化酶活性降低與脂質過氧化物丙二醛含量增加(P<0.05)之情形,並在中劑量組中有顯著較佳的識別記憶、高劑量組中有顯著較佳的空間記憶表現(P<0.05)。綜而言之,小球藻水萃物在細胞試驗中無法顯著有效預防由 200 μM 以上過氧化氫所造成之細胞損傷,但具有良好的體外清除自由基能力及增強 D-半乳糖誘導老化小鼠體內抗氧化之潛力。

The objective of this study was to investigate the antioxidant effect of Chlorella extract on D-galactose induced oxidative stress in mice. The study was divided into three parts which includes extraction of Chlorella, in vitro cell test and in vivo animal test. First, the Chlorella cells were broken by high-pressure, then extracted with hot water followed by centrifugation. The supernatant was lyophilized using a freeze dryer to obtain the Chlorella water extract powder (CE). The extraction results showed that CE obtained from the broken cell wall had significantly higher yield and water extractive contents (P<0.05). In vitro study aimed to evaluate the possible protective effects of CE against H2O2-induced cell damage in MRC-5 cells. Treatment of CE on MRC-5 cell for 24 hours resulted in no changes (P>0.05) in cell viability compared to control, indicating that the concentrations (<2 mg/ml) used in this study did not induce damage to cell. However, the cell viability of the cells pretreated with CE and further exposure to 200 μM or 250 μM H2O2 for 4 hours, showed no significant differences between treatment groups and induced group. The result indicated that pretreatment of cells with CE had no significantly protected the cells against the damage induced by H2O2. In vivo study was performed to investigate the effect of CE on learning, memory and oxidative stress in D-galactose (D-gal) induced aging mice. Mice were injected subcutaneously with D-gal daily for 8 weeks and administered simultaneously with CE or vitamin C. The results showed that CE significantly improved body weight loss and memory and learning ability in Novel object recognition (NOR) and Morris water maze test (MWM) compared to the D-gal induced mice. Furthermore, CE increased the glutathione peroxidases activity and decreased the malondialdehyde level (P<0.05). To sum up, although CE could not significantly prevent cell damage induced by H2O2 in the in vitro study, it had good ability to scavenge free radicals and enhance the antioxidant potential of D-gal induced aging mice.