抗氧化剂的体外评价及在运输应激大鼠中的抗应激效果研究

发布时间：2018-06-23 23:50

本文选题：运输应激 + 大鼠　；参考：《华中农业大学》2014年硕士论文

【摘要】：动物的运输是畜牧生产中一个重要的环节,运输应激对动物生产性能、健康状况等产生一系列负面影响。家畜在运输应激后使机体内氧化和抗氧化失衡,导致机体内发生氧化应激。本研究旨在评价抗氧化剂维生素C、维生素E、α-硫辛酸、槲皮素和止痢草油的抗氧化能力和特性,并通过建立大鼠运输应激模型,比较这5种抗氧化剂缓解大鼠运输应激的效果,为家畜运输应激的营养调控提供理论依据。主要研究内容和结果如下： 1、不同抗氧化剂体外抗氧化效果评价及特性研究。利用DPPH法测定维生素C、维生素E、槲皮素和止痢草油清除DPPH自由基的能力,结果表明抗氧化剂清除DPPH自由基的IC50如下：维生素C为0.0059mg/ml,维生素E为0.0249mg/ml,槲皮素为0.0051mg/ml,止痢草油为0.0687mg/ml,这4种抗氧化剂对DPPH自由基的清除能力大小顺序为：槲皮素维生素C维生素E止痢草油。通过建立大鼠肝匀浆脂质自氧化模型,测定维生素C、a-硫辛酸、维生素E、槲皮素和止痢草油的脂质自氧化抑制率,各抗氧化剂的IC50如下：维生素C为0.255mg/ml,维生素E为0.0052mg/ml,α-硫辛酸为2.06mg/ml,槲皮素为0.0058mg/ml,止痢草油为0.067mg/ml,抑制脂质自氧化的高低顺序为：维生素E槲皮素止痢草油维生素Cα-硫辛酸。 2、抗氧化剂对运输应激大鼠的抗应激效果的研究。试验选择7-8周龄SPF级雄性Wistar大鼠70只,随机分成7组,每组10个重复,每个重复1只大鼠。其中非应激对照组和应激对照组均饲喂基础日粮,5个日粮处理组在基础日粮中分别添加1.5g/kg维生素C、2g/kg α-硫辛酸、1g/kg维生素E、1g/kg槲皮素、100mg/kg止痢草油。试验期28d,试验期结束后,应激组大鼠根据运输应激模拟条件(大鼠置于35℃恒温摇床中,60r/min,2h)进行处理,运输应激处理结束后乙醚麻醉并屠宰大鼠,收集样品。测定血清激素水平、血清和肝脏中抗氧化系统指标、空肠组织形态结构及空肠mRNA表达量。结果显示： 1)添加止痢草油能显著增加大鼠体重、日增重和日采食量(P0.05),添加维生素E显著增加大鼠体重(P0.05),添加槲皮素可以显著增加大鼠的平均日采食量(P0.05)。 2)运输导致大鼠体重显著下降,体温显著升高(P0.05)。添加各抗氧化剂后,体重失重与应激对照组无显著差异(P0.05),运输后维生素C组大鼠体温显著高于其他各组(P0.05)。 3)运输应激处理后应激对照组大鼠血清中皮质醇和去甲肾上腺素水平均显著升高(P0.05)。与应激对照组相比,日粮添加维生素E显著降低了去甲肾上腺素水平(P0.05)。 4)与非应激组相比,应激对照组的血清T-AOC显著降低(P0.05),MDA水平显著升高(P0.05),SOD和GSH-Px活力均显著降低(P0.05)；添加抗氧化剂后,各抗氧化剂组与应激对照组相比,MDA水平显著降低(P0.05),SOD活力显著升高(P0.05),维生素C、维生素E与止痢草油组T-AOC显著升高,维生素E、槲皮素、a-硫辛酸和止痢草油组GSH-Px活力均显著升高(P0.05)。 5)抗氧化剂对运输应激大鼠肝脏抗氧化功能的影响：应激对照组的肝脏T-AOC显著下降,脂质过氧化物MDA水平显著升高(P0.05)。而添加维生素C、维生素E、槲皮素和止痢草油均能提高T-AOC,降低MDA水平(P0.05)。与非应激对照组相比,应激对照组均显著降低了肝脏的SOD和GSH-Px活力(P0.05),添加槲皮素和止痢草油组可以提高SOD和GSH-Px活力(P0.05),而添加α-硫辛酸组肝脏的SOD和GSH-Px活力与应激对照组没有显著差异(P0.05)。 6)大鼠空肠组织结构形态分析：应激对照组显著提高了隐窝深度,显著降低了空肠绒毛高度和绒毛高度与隐窝深度比(P0.05)。添加维生素E、a-硫辛酸、槲皮素和止痢草油均能显著提高绒毛高度(P0.05)；添加维生素C、维生素E、槲皮素均能降低隐窝深度(P0.05)；添加这5种抗氧化剂均能显著提高绒毛高度与隐窝深度比(P0.05)。各日粮处理组的空肠绒毛高度、隐窝深度、绒毛高度与隐窝深度比,与非应激对照组相比差异不显著(P0.05)。 7)大鼠空肠HSps mRNA表达量的变化：应激对照组与非应激对照组相比,大鼠空肠HSp27、Hsp70、Hsp90mRNA表达量均显著升高(P0.05)；日粮添加维生素C、α-硫辛酸、维生素E、槲皮素和止痢草油均能显著降低大鼠空肠Hsp27、Hsp70、 Hsp90mRNA表达量(P0.05)。综上所述,本研究的结论是：1)由体外试验可得,槲皮素和维生素E清除DPPH自由基和抑制脂质自氧化的效果较好。2)日粮中添加维生素E、槲皮素和止痢草油均能增加动物采食量和体重,改善动物的生长性能。3)由大鼠运输应激模型可知,补充抗氧化剂均能降低大鼠血清和肝脏中脂质过氧化,提高抗氧化酶SOD和GSH-Px活力,提高机体抗氧化能力。其中α-硫辛酸在动物体内发挥自由基清除能力、抑制脂质自氧化的作用最强,止痢草油对机体总抗氧化能力增强作用较明显,槲皮素、维生素C、维生素E在抑制脂质自氧化和增强抗氧化酶活力均有效果,这5种抗氧化剂在运输应激大鼠中均具有一定的抗运输应激作用。综合指标的测定可知,维生素E、槲皮素和止痢草油在运输应激大鼠中抗运输应激效果较好。
[Abstract]:The transport of animals is an important link in animal production. Transportation stress has a series of negative effects on animal production and health. The oxidative stress in the body is caused by the imbalance of oxidation and antioxidation in the body after transportation stress. The aim of this study is to evaluate the antioxidant vitamin C, vitamin E, alpha lipoic acid and mistletoe. The antioxidative ability and characteristics of PEP and SCR were compared, and the effect of these 5 antioxidants on transport stress was compared by establishing a rat model of transport stress, which provided a theoretical basis for the nutrition regulation of livestock stress. The main contents and results were as follows:
1, evaluation and characterization of antioxidant effects of different antioxidants in vitro. Using DPPH to determine the ability of vitamin C, vitamin E, quercetin and dysentery oil to remove DPPH free radicals, the results showed that the antioxidants removed the DPPH free radical IC50 as follows: vitamin C 0.0059mg/ml, vitamine E 0.0249mg/ml, quercetin as 0.0051mg/ml, dysentery The order of the 4 antioxidants is 0.0687mg/ml, the order of the scavenging ability of these 4 antioxidants to DPPH radicals is quercetin vitamin E, vitamin E, antidysentery oil. The lipid self oxidation inhibition rate of vitamin C, a- lipoic acid, vitamin E, quercetin and antidysentery oil is determined by establishing rat liver homogenate lipid autoxidation model, and the IC50 of each antioxidant is as follows. Vitamin C is 0.255mg/ml, vitamin E is 0.0052mg/ml, alpha lipoic acid is 2.06mg/ml, quercetin is 0.0058mg/ml, and dysentery oil is 0.067mg/ml, and the order of inhibiting lipid self oxidation is: vitamin E quercetin vitamin C alpha - lipoic acid.
2, the anti stress effect of antioxidant on stress rats was studied. 70 male Wistar rats of SPF grade 7-8 weeks old were randomly divided into 7 groups, with 10 repetitions in each group and 1 rats each. The non stress control group and the stress control group were fed with basal diet, and 5 diet treated groups added 1.5g/kg to the basal diet. C, 2g/kg alpha lipoic acid, 1g/kg vitamin E, 1g/kg quercetin, 100mg/kg dysentery oil. Test period 28d. After the test period, the stress group rats were treated according to the simulation conditions of transport stress (rats were placed in the constant temperature rocking bed at 35 degrees C, 60r/min, 2h). After the transport of stress treatment, the rats were anaesthetized and slaughtered to collect samples and determine serum hormones. Levels, serum and liver antioxidant index, jejunal morphology and jejunal mRNA expression.
1) adding dysentery oil could significantly increase the weight of rats, daily gain and daily intake (P0.05), and vitamin E significantly increased the weight of rats (P0.05), and the addition of quercetin could significantly increase the average daily feed intake (P0.05) of rats.
2) transport resulted in a significant decrease in body weight and a significant increase in body temperature (P0.05). There was no significant difference between weight loss and stress control group (P0.05) after the addition of all antioxidants (P0.05). The body temperature of rats in group C after transportation was significantly higher than that of other groups (P0.05).
3) the levels of cortisol and norepinephrine in the serum of stress control group were significantly increased after transportation stress treatment (P0.05). Compared with the stress control group, dietary vitamin E significantly decreased norepinephrine level (P0.05).
4) compared with the non stress group, the serum T-AOC of the stress control group was significantly lower (P0.05), the level of MDA increased significantly (P0.05), and the activity of SOD and GSH-Px decreased significantly (P0.05). After the addition of antioxidants, the level of MDA decreased significantly (P0.05), SOD activity increased significantly (P0.05), vitamin C, vitamins and dysentery. T-AOC increased significantly in grass oil group, while GSH-Px activity in vitamin E, quercetin, a- lipoic acid and Zhili grass oil group increased significantly (P0.05).
5) the effect of antioxidant on the antioxidant function of liver in stress rats: the liver T-AOC in stress control group decreased significantly and the level of lipid peroxide MDA increased significantly (P0.05). The addition of vitamin C, vitamin E, quercetin and antidysentery oil could increase T-AOC and decrease MDA level (P0.05). Compared with non stress control group, the stress control group was all compared with the control group. The activity of SOD and GSH-Px in the liver (P0.05) was significantly reduced. The activity of SOD and GSH-Px (P0.05) was increased by the addition of quercetin and antidysentery oil, while the activity of SOD and GSH-Px in the liver of the group adding alpha lipoic acid was not significantly different from that of the stress control group (P0.05).
6) the morphological analysis of the jejunum tissue: the stress control group significantly increased the depth of the recess, significantly reduced the height of the jejunum and the height of the villi and the depth of the recess (P0.05). The addition of vitamin E, a- lipoic acid, quercetin and antidysentery oil significantly increased the height of the villi (P0.05), and vitamin C, vitamin E and quercetin were all reduced. Low recess depth (P0.05); adding these 5 antioxidants could significantly increase the ratio of the height of the villi to the depth of the recess (P0.05). The height of jejunum villus, the depth of the recess, the height of the villi and the depth of the recess were not significant (P0.05).
7) the changes in the expression of HSps mRNA in the jejunum of rats: the expression of HSp27, Hsp70 and Hsp90mRNA in the jejunum of the rats increased significantly (P0.05) in the stress control group compared with the non stress control group. The diet supplemented with vitamin C, alpha lipoic acid, vitamin E, quercetin and antidysentery oil significantly reduced the expression of Hsp70 and Hsp90mRNA (P0.05) in the rat jejunum.
To sum up, the conclusions of this study are as follows: 1) in vitro tests can be obtained, quercetin and vitamin E can remove DPPH free radicals and inhibit lipid autoxidation, the effect of vitamin E, quercetin and antidysentery oil can increase animal food intake and weight, improve animal growth performance.3) by rat model of transport stress, supplemented by rat transport stress model. Antioxidants can reduce lipid peroxidation in serum and liver of rats, increase the activity of antioxidant enzyme SOD and GSH-Px, and improve the antioxidant capacity of the body. Among them, alpha lipoic acid plays the free radical scavenging ability in animals and inhibits lipid self oxidation. Vitamin C and vitamin E have effect on inhibiting lipid autoxidation and enhancing antioxidant enzyme activity. These 5 kinds of antioxidants have a certain anti transport stress in transport stress rats. The comprehensive index shows that vitamin E, quercetin and antidysentery oil have better effect on transportation stress in the transport stress rats.
【学位授予单位】：华中农业大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：S852.2

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