【作者】 孙永叶；

【导师】 马爱国；

【作者基本信息】 青岛大学 ， 营养与食品卫生， 2003， 硕士

【摘要】 目的，通过对实验动物(豚鼠／大鼠)进行不同剂量的维生素C(以下简称VC)或维生素E(以下简称VE)的干预，观察不同剂量的VC、VE对实验动物抗氧化能力、细胞功能及淋巴细胞DNA氧化损伤的影响，并探讨其机理， 方法：1．VC干预实验 采用幼年健康荷兰种豚鼠48只，体重180～240克，雌雄各半，按体重、性别随机分为四组，每组12只。各组豚鼠每天每千克体重VC的添加剂量分别为0(缺乏组)、7.5mg(对照组)、125mg(中高剂量组)、250mg(高剂量组)，其中7.5mg／kg／d为豚鼠VC的适宜摄入量，作为正常对照组。实验期为20天，期间饲料和饮水不限。实验10天时留取各组豚鼠尿液。 2．VE干预实验 采用健康初断乳Wistar大鼠39只，体重65～85克，雌雄各半，按体重、性别随机分为三组，每组12-14只。各组VE的补充剂量分别为24.18mg／kg饲料(对照组)、70.83mg／kg饲料(中高剂量组)、114.04mg／kg饲料(高剂量组)，其中第1组饲料VE的含量为大鼠适宜摄入量(约35IU)，作为对照组。实验期为8周，饲料及饮水不限。实验过程中，每周称大鼠体重，第8周留取尿液。 3．两组实验结束后，腹主动脉取血，分离获取血浆，收集淋巴细胞、红细胞等。采用单细胞凝胶电泳法检测DNA自发及诱导氧化损伤；采用荧光偏振法测定红细胞膜流动性；采用毛细管电泳法测定尿中O~6-甲基鸟嘌呤的含量；采用荧光法测定大鼠血浆VE；用2、4-二硝基苯肼法测定豚鼠血浆VC；采用试剂盒测定血浆中SOD、MDA以及GSH-Px。 结果： 一、维生素C干预实验研究结果显示： 1．经过20天的干预，各组豚鼠血浆VC含量间差别较大(F=107.595，P＜0.001)，缺乏组血浆VC含量为6.26μmol／L，而对照组、中高剂量组和高剂量组分别升高2.91倍(24.46μmol／L)、8.36倍(58.62μmol／L)和9.30倍(64.45μmol／L)。 2．血浆抗氧化指标分析结果显示：各组血浆SOD差别无统计学意义(F=1.151，p＞0.05)；血浆MDA水平：缺乏组为2.20nmol／ml，明显高于其余VC干预组中文摘要 (F=4 .60，P<0 .01)。GSH-Px活力缺乏组为265.93酶活力单位，明显低于对照组 321 .20酶活力单位(P<0 .05);而VC补充中高剂童、高剂量组GSH-Px含量与缺 乏组和对照组相比无明显差异(P>0.05)，研究结果未发现在补充VC后血浆 GSH-Px水平有明显提高。3.红细胞膜流动性分析结果显示:VC缺乏组的荧光偏振度(P)和微粘滞度(n) 明显高于VC干预组(F值分别为4.241、4.747，P值分别小于 0.05和0.01)。其 中，缺乏组P值为0.3049，对照组0.2817，中高剂t组和高剂盘组分别为0.2791、 0.2770。说明缺乏组豚鼠红细胞膜流动性较其余三组均降低，差别有统计学意义 (P<0 .05);而补充较高剂量VC的两组及对照组红细胞膜流动性差异无显著性 (P>0.05)。4.淋巴细胞DNA损伤情况:各组自发损伤差别无统计学意义(F句.453，P>0.05)。 10pmo比的HZ仇诱发DNA损伤时，各组间差异有显著性(F==2 .700，P<0 .05); 对照组损伤最轻(AU=72.56)，而缺乏组(AU== 1 2729)及中高剂量组(AU== 121)、 高剂量组(AU=133)DNA损伤均较对照组加重(P值均小于0.05)，补充VC没 有表现出对DNA损伤的保护作用。而25 p inoUL及50pmo比的玩伍诱发DNA 损伤时，各组间差异均无统计学意义(F值分别为0.542、0.746，P值均>0 .05)。 尿中06一甲基鸟嗦吟含t分析，各组06一甲基鸟嗦吟的含t差异有统计学意义 (F== 3.028，P<0.05)，缺乏组较高，为3.科叫九肌醉，与对照组(l .48m眺肌 配)相比差异有统计学意义(P<0 .05)，补充组尿06.甲基鸟嗦吟的含盘较缺乏组 偏低，稍高于对照组，但各组间差别无显著性。二、维生素E干预实验研究结果显示:1.各组血浆vE含童分别为8.“叹办，56.94m留L，64.34m目工(F值为74.76，p<0 .01). 中高剂量组及高剂!组血浆VE含量均明显高于对照组，其含盘分别为对照组的 6.6倍、7.4倍(p值均小于0.01);补充VE的两组间差别无统计学意义(P沁.05).2.DNA损伤研究结果显示:VE干预各组淋巴细胞DNA自发损伤率均较低，各组 l’ed差别无统计学意义(F值为0.555，P>0.05)。当5 pmo比、10 pmo比及25p mo讥的H刃:诱发DNA损伤时，各组损伤均明显加重，但组间差异无显著性 (P>0.05)，随着玩仇剂t的增加，高剂量VE没有表现出对DNA损伤的保护 作用。3.DNA烷化产物护一甲基鸟嗦吟分析结果显示:留取的24小时尿液各组尔甲基 鸟嗓吟的含量差异无显著性(F=2 .493，P>0.05)，但中高剂童组及高剂t组较对 照组含量有逐渐增高的趋势。 结论:VC干预实验组中文摘要1.豚鼠缺乏VC将会出现相应的缺乏症状，如:倦怠、出血、关节肿胀、瘫痪等; 而补充较高剂量的VC对豚鼠的生长发育没有明显作用。2.维生素C缺乏可导致豚鼠体内抗氧化物酶GSH一Px含量降低，M[DA含量增高， 但对SOD活性变化不明显，甚至有升高的趋势:VC缺乏可致红细胞膜流动性降 低。补充较高剂量的VC对其无影响。3.VC缺乏及补充较高剂t的VC对DNA自发损伤无改变:而在低剂l的HZOZ诱 导损伤时，VC缺乏及更多还原

【Abstract】 Objective: To investigate the antioxidative activities of vitamin C, E at different levels of dosage in guinea pigs and rats.Methods: 1. 48 healthy young guinea pigs (weighed from 180g to 240g) were selected and randomized into 4 groups with 12 hi each group. Each of mem hi different group took different dose of VC (0, 7.5, 125, 250mg/kg wt) for 20 days. The second group was the control group because 7.5mg/kg wt was near to the appropriate dose for human RDA. Urine was collected hi the mid of the trial and whole blood was collected by the end of the trial. 2. 39 healthy weaning Wistar rats (weighed from 65g to 85g) were randomly allocated to 3 treatment groups with 12-14 hi each group. Those rats were fed on fully compounded diet for 2 months. The content of vitamin E hi each group diet was separately 24.18,70.83,114.04mg/kg fodder. The first group was the control group. Urine and whole blood were collected after experiment. 3. Serous vitamin C % serous vitamin E and SOD, GSH, MDA were examined; DNA damage were analyzed by SCGE; O6-Meg was analyzed by high performance capillary zone electrophoresis; erythrocyte membrane fluidity was detected by fluorescence polarization method. Results Vitamin C experiment:1. After 20 days’ intervention, there were great differences hi serous vitamin C among those groups (F=107.595, p<0.001). The deficient group was 6.26Mmol/L, the control group (24.46Mmol/L) and the two supplemented groups were separately 2.91 folds, 8.36 folds, 9.30 folds higher than the deficient group.2. The difference of serous SOD among these groups had no significance (F=1.151, p>0.05). MDA of the deficient group was 2.20nmol/ml which was higher than the other three groups (0.70nmol/mLx 1.27nmol/mL% 0.87nmol/mL) while there were no differences among the three groups. GSH-Px was reduced hi the deficient group (p<0.05). But in the supplemented group, there was no significant change (p>0.05).3. The fluorescence polarization of erythrocyte membrane of the four groups were 0.3049^ 0.2817, 0.2791, 0.2770, the deficient group was the lowest and there were no differences between the supplemented groups and the control group (p>0.05).4. After the experiment, intrinsic DNA damage had no differences hi 4 groups ( F=0.453, p>0.05) , but DNA damage induced by 10Mmol/L H202 had significant differences (F=2.709, p<0.05):the control group ( 72.56 AU ) was less DNA damage than those ofother three groupsC 127.29> 121 <, 133 AU respectively, p<0.05). DNA damage induced by 25rtnol/L and 50Mmol/L H202 had no significant differences (F=0.542, 0.746, P >0.05).There were different levels O6-Meg in four groups (F=3.028, p<0.05). The level in the deficient group was higher than the control group (p<0.05). Vitamin ?experiment:1. The serous vitamin E levels were 8.64mg/L, 56.94mg/L and 64.34mg/L respectively in three groups (F=74.76, p<0.01). The values in two supplemented groups were greatly higher than the control group (6.6-fold, 7.4-fold of the control group).2. No any differences of intrinsic DNA damage in three groups were detected ( F=0.555, p>0.05). When DNA damage were induced by 5Mmol/L, 10 umol/L and 25umol/L H202, there were also no differences among those groups (p>0.05).3. O6-Meg levels in three groups were detected and had no significant differences (F=2.493, p>0.05).Conclusion1. ’Vitamin C is necessary for the growth of cavy and a high dose supplementation has no additional effects on the growth of cavy.2. Deficiency of vitamin C could lead to a reduction of GSH-Px, increasing fluorescence polarization of erythrocyte membrane and MDA.3. Different doses of vitamin C did not affect the intrinsic DNA damage. HjOi could induce DNA oxidative damage. But VC supplementation could not show the protective action.4. Supplementation of vitamin E has no effects on the growth of young rats and did not increase their serous SOD, GSH-Px and decrease MDA, fluorescence polarization of erythrocyte membrane.5. Vitamin E could not protect DNA from oxidative damage更多还原