【作者】 汪求真；

【导师】 马爱国；

【作者基本信息】 青岛大学 ， 营养与食品卫生学， 2004， 硕士

【摘要】 目的 维生素E（VE）是人体所需的重要脂溶性维生素和营养性抗氧化剂，但大剂量补充是否具有不良影响尚未见到明确报道，本研究拟通过大剂量维生素E补充从生物大分子氧化损伤及细胞功能方面探讨大剂量维生素E的影响。 方法 实验动物选用Wistar大鼠48只按体重随机分成4组，对照组给予普通大鼠饲料，三个干预组均喂饲添加了大剂量α-生育酚乙酸酯的饲料，其中维生素E的含量分别为500 IU／kg、2000 IU／kg、7500 IU／kg，补充时间为8周。实验结束时将大鼠安乐处死，收集血样和尿样进行抗氧化能力及细胞活性分析。抗氧化能力分析主要包括DNA氧化（H₂O₂诱导）损伤、血清超氧化物歧化酶（SOD）、谷胱甘肽过氧化物酶（GSH-Px）和丙二醛（MDA）含量分析及尿中O⁶-甲基鸟嘌呤含量分析。DNA氧化损伤采用“彗星”电泳技术进行分析，尿O⁶-甲基鸟嘌呤含量分析采用高效毛细管电泳法，SOD等抗氧化酶及MDA用生化试剂盒进行测定。细胞功能分析主要包括红细胞膜流动性和外周血淋巴细胞转化率测定。红细胞膜流动性测定采用荧光偏振法，应用四甲基偶氮唑盐（MTT）法培养大鼠外周血淋巴细胞并测定淋巴细胞转化率，观察大剂量维生素E对免疫细胞增殖活性的影响。 结果 补充较大剂量VE能显著增强大鼠机体抗氧化能力及细胞增殖和膜流动的功能活性。研究结果显示：当VE剂量为500IU／kg时，与对照组（75IU／kgVE）相比，大鼠血清VE水平提高了69.2％，SOD活性显著升高（P＜0.05），提高了11％；血清和红细胞膜中GSH-Px活性均明显升高，分别提高了5.4％（P＜0.05）和101.5％（P＜0.01）；血清MDA水平明显降低，比对照组下降了34％（P＜0.01）。DNA损伤分析结果显示由10μmol／L过氧化氢诱发的DNA氧化损伤均显著降低（P＜0.05）以及尿中O⁶-甲基鸟嘌呤排出量也呈现明显下降（P＜0.05）。细胞功能分析结果显示：补充500IU／kg VE 8周后，大鼠红细胞膜流动性和淋巴细胞转化率均比对照组显著提高（P＜0.01，P＜0.05）。 但是，长期补充过大剂量VE可引起生物大分子损伤和细胞功能降低。当饲料中维生素E的剂量进一步增加为2000 IU／kg、7500 IU／kg（分别为大鼠需要量的27倍、100倍）时，与对照组（75IU／kgVE）相比，大鼠血清VE浓度分别 中文摘要提高了93.7%、n7.4%，而SOD活性却分别下降了17%、18%（尸<0.05，尸<0.01）;彗星电泳结果显示，10拜mol/L过氧化氢诱发的DNA氧化损伤明显加重（尸<0.05，尸<0.01）。细胞功能分析结果显示:补充75。。IU/kgVES周后，大鼠淋巴细胞转化率比对照组显著降低（尸<0.05），而2000 IU/kgVE对细胞转化率没有影响（尸>.05），对细胞增殖功能既没有产生保护作用，也没有产生损伤作用。红细胞膜流动性在这两个过大剂量补充组均未见改善。 结论①so0IU/kg维生素E可以增强血清、细胞膜GSH一Px、SOD等抗氧化酶的活性，降低血清中脂质过氧化终产物MDA水平;同时，补充该剂量维生素E可降低DNA诱发氧化损伤及自发烷化损伤水平;提高大鼠红细胞膜流动性和淋巴细胞转化率。②饲料中维生素E进一步增高为Zoo0lu/kg、7500IU/kg时，可降低血清SOD活性，并且加重较大剂量过氧化氢诱发的DNA氧化损伤;7500IU/kgVE可降低淋巴细胞转化率。而GSH一Px、MDA、红细胞膜流动性等则未见改善。 总之，较大剂量维生素E可提高机体的抗氧化能力，并且改善细胞增殖及膜流动等细胞活性。但是，过大剂量维生素E则会引起生物大分子损伤和细胞功能下降，并且随着剂量增大，这种损伤作用可能更加严重。更多还原

【Abstract】 Objectives Vitamin E is an essential fat-soluble vitamin that has antioxidant activities. But it had not been reported definitely whether supplementation of high dose vitamin E would cause adverse effects. In this in vivo study we investigated the influence of different levels of high dose vitamin E on oxidative damage of macromolecule and cell function to determine its effect when used excessively.Methods 48 Wistar rats , 60~90g body weight(half male and half female) were randomly divided into 4 groups based on body weight. The control group was fed with normal rat diet which contented vitamin E 75 IU/kg and the other three interfered groups were supplemented with different levels of high dose all-rac-a-tocopheryl acetate enriched diets which contented vitamin E 500 IU/kg,2000 IU/kg,7500IU/kg,respectively. The period of the trial was about 8 weeks. Urine of each rat was collected one week before the end of the trail. At the end of the trial, the rat will be died without pain by drug anesthesia, and samples of the whole blood were collected for analysis. Antioxidation activity analysis included DNA oxidative damage induced by H2O2, activities of serum superoxide dismutase (SOD), glutatathione peroxidase (GSH-Px) content of malanydiadehyde(MDA) and the concerntration of O6-methylguanine in urine. The comet assay or the alkaline single cell gel electrophoresis (SCGE) assay was used to measure the DNA oxidative damage. High performance capillary zone electrophoresis was used to determine O6-methylguanine. SOD, GSH-Px and MDA were measured by biochemical methods with the kits. Cell function a-nalysis included the erythrocyte membrane fluidity and the lymphocyte transformation rate . The method of fluorescence polarization was used to measure membrane fluidity and MTT was used to detected the lymphocyte transformation rate which reflected proliferation activity of immune cell.Results Relatively high dose vitamin E could significantly increase antioxidation activity and cell function detected by lymphocyte proliferation and membrane fluidity . Compared with the control group, 500IU/kg VE increased serum VE by 69. 2%, serum SOD activity by 11% , serum and membrane GSH-Px activities by 5. 4% and 101. 5% while serum MDA decreased by 34%. 500IU/kg VE significantly lowered DNA oxidative damageinduced by 10 tmol/L H2O2 and urine O6-methylguanine was also decreased. After 500IU/ kg VE supplementation with 8 weeks , the erythrocyte membrane fluidity and lymphocyte transformation rate were significantly increased.It worth notion that long term supplementation with too excessive vitamin E would damage macrobiomolecule and decrease cell function. 2000 IU/kg,7500 IU/kg vitamin E increased serum VE by 93. 7%, 117. 4% respectively. But serum SOD activities were decreased by 17% and 18% respectively and DNA oxidative damages induced by 10 mol/L H2O2 were both aggravated in this two groups. At the same time, lymphocyte transformation rate of the rat supplemented with 7500 lU/kg VE for 8 weeks was significantly decreased.Conclusion (1)500 lU/kg vitamin E could increase the activities of antioxidant enzymes such as SOD,GSH-Px and decrease the formation of serum MDA. DNA oxidative damage induced by H2O2 and DNA alkyl damage were decreased by 500 lU/kg VE . As to cell function, 500 lU/kg VE significantly improved membrane fluidity and promoted lymphocyte transformation. (2)2000 IU/kg,7500 IU/kg vitamin E decreased serum SOD activities and aggravated DNA oxidative damages induced by H2O2. Lymphocyte transformation rate was decreased by 7500 IU/kg VE.Above all, relatively high dose vitamin E could increase antioxidation activity of the body and increase cell function indexes such as membrane fluidity and lymphocyte proliferation rate. However, too excessive vitamin E would induce macrobiomolecule damage and decrease cell function.更多还原