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首页> 外文期刊>Animal Reproduction Science >Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen cryopreserved in extenders with antioxidants.
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Lipid peroxidation and generation of hydrogen peroxide in frozen-thawed ram semen cryopreserved in extenders with antioxidants.

机译：冷冻解冻的公羊精液中的脂质过氧化和过氧化氢的生成在抗氧化剂增量剂中冷冻保存。

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摘要

The objective of this study was to evaluate the effect of addition of the antioxidants Trolox and catalase to a ram semen cryopreservation extender on lipid peroxidation and hydrogen peroxide generation on the extender and in the thawed semen. Semen was collected from 23 Santa Ines rams (one ejaculate per ram) and diluted at 32 degrees C to a concentration of 400x106 cells/ml in one of the following solution: Tris-egg yolk extender (control), or the same extender supplemented with either 50 micro M Trolox/108 sperm (Trolox), 50 micro g catalase/ml (Catalase) or a combination of Trolox and catalase (Tro+cat, 50 micro M Trolox/108 sperm and 50 micro g catalase/ml). The semen was loaded into 0.25 ml straws, cooled and frozen in a programmable freezer and subsequently stored in liquid nitrogen. Prior to evaluation, frozen straws were thawed in a water bath (42 degrees C for 20 s). Lipid peroxidation (LPO), both spontaneous and catalyzed, on the semen and the extender were measured using the thiobarbituric acid (TBA) assay in accordance with the method described by Buege and Aust (1978). Hydrogen peroxide (H2O2) generation was measured using the horseradish peroxidase-dependent oxidation of phenol red to a derivative with absorbance at 610 nm, according to the method described by Pick and Keisari (1980). Spontaneous LPO resulted in the least production of thiobarbituric acid-reactive substances (TBARS) in the Tro+cat (1.37+or-0.02 nMol/108 sperm), compared to amounts in the other treatments groups. In the catalyzed LPO experiments, the least (P<0.05) amounts of TBARS were observed in Trolox (2.52+or-0.02 nMol/108 sperm) and Tro+cat (2.54+or-0.02 nMol/108 sperm) groups, compared to the control (3.81+or-0.02 nMol/108 sperm) and catalase (3.83+or-0.02 nMol/108 sperm) groups. Hydrogen peroxide generation was less (P<0.05) in the Trolox (6.00+or-0.18 nMol/40x106 sperm/+or-40 min) and Tro+cat (6.08+or-0.18 nMol/40x106 sperm/+or-40 min) groups than in the control (6.97+or-0.18 nMol/40x106 sperm/+or-40 min) and catalase (6.53+or-0.18 nMol/40x106 sperm/+or-40 min) groups. Compared to the control group, Trolox and catalase treatment significantly reduced TBARS in catalyzed LPO and hydrogen peroxide concentrations in the samples (P<0.05). ROS (reactive oxygen species) generation occurred in all extenders, without sperm cells. The data presented provide evidence that ROS are produced in ram semen, both in the extender and during the freezing and thawing process. In addition, the data suggest that the antioxidants Trolox and catalase may be used to control the oxidative stress imposed on ram spermatozoa by the cryopreservation process.
机译：这项研究的目的是评估在冷冻精液中添加抗氧化剂Trolox和过氧化氢酶对脂质和过氧化氢的作用。从23个Santa Ines公羊（每个公羊射精一次）中收集精液，并在以下一种溶液中于32摄氏度稀释至400x10 6 细胞/ ml的浓度：Tris-卵黄蛋黄质补充剂（对照），或添加了50 micro M Trolox / 10 8 精子（Trolox），50 micro g过氧化氢酶/ ml（Catalase）或Trolox和过氧化氢酶的组合（Tro + cat，50 micro M Trolox / 10 8 精子和50 micro g过氧化氢酶/ ml）。将精液装入0.25 ml的吸管中，在可编程的冷冻机中冷却并冷冻，然后储存在液氮中。在评估之前，将冷冻的稻草在水浴中融化（42摄氏度，持续20 s）。根据Buege and Aust（1978）描述的方法，使用硫代巴比妥酸（TBA）测定法测量精液和增量剂上自发和催化的脂质过氧化（LPO）。使用辣根过氧化物酶依赖性的酚红氧化成衍生物并在610 nm处吸收的方法测量了过氧化氢（H 2 O 2 ）的生成。皮克和凯萨里（1980）。与其他治疗组相比，自发LPO导致Tro + cat（1.37 + or-0.02 nMol / 10 8 精子）中硫代巴比妥酸反应性物质（TBARS）的产生最少。在催化的LPO实验中，在Trolox（2.52 + or-0.02 nMol / 10 8 精子）和Tro + cat中观察到最少（ P <0.05）的TBARS量（2.54 + or-0.02 nMol / 10 8 精子）组与对照组（3.81 + or-0.02 nMol / 10 8 精子）和过氧化氢酶（3.83 + or -0.02 nMol / 10 8 精子）组。 Trolox（6.00+或-0.18 nMol / 40x10 6 精子/ +或-40分钟）中的过氧化氢生成量较少（ P <0.05）比对照组（6.97+或-0.18 nMol / 40x10 6 精子/ +或6.08+或-0.18 nMol / 40x10 6 精子/ +或-40 min -40分钟）和过氧化氢酶（6.53+或-0.18 nMol / 40x10 6 精子/ +或-40分钟）组。与对照组相比，Trolox和过氧化氢酶处理显着降低了样品中LPO和过氧化氢的催化TBARS浓度（ P <0.05）。 ROS（活性氧物质）的产生发生在所有没有精子细胞的补充剂中。所提供的数据提供了证据，证明在精液中，膨胀剂中以及冷冻和解冻过程中都产生了ROS。此外，数据表明抗氧化剂Trolox和过氧化氢酶可用于控制通过低温保存过程施加在精子上的氧化应激。

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