原花青素和乳酸链球菌素对猪精液常温保存效果的研究

发布时间：2018-07-26 20:09

【摘要】：随着规模化养猪业发展,猪人工授精技术的普及面不断扩大,而精液保存质量的优劣决定生产效率高低,其中稀释剂发挥着至关重要的作用。精液常温保存过程中需要进行稀释,会减弱精液原有抗氧化作用,使过多活性氧无法及时清除,质膜易受被损伤,导致精子质量下降。同时,微生物的存在也会影响精液保存质量。为同时解决抗菌和抗氧化的问题,本研究在稀释液中添加原花青素(Oligomeric Proantho Cyanidins,OPC)和乳酸链球菌素(Nisin),检测精液保存质量、渗透压、总抗氧化能力、丙二醛含量、酸性磷酸酶活性和抑菌效果等,以期为OPC和Nisin在精液稀释液中的联合添加提供一些理论参考。本研究主要获得以下结果:1.猪精液稀释液中添加不同浓度(10、20、30、40、50、60、70μg/mL)OPC溶液,可优化精液保存效果。其中,稀释液中添加50μg/mL OPC效果最好,常温保存第5d的精子活率61.63%、质膜完整率41.88%、顶体完整率75.88%,显著高于对照组(P0.05)。常温保存第5d时精子T-AOC为1.43 U/mL,高于对照组,二者差异不显著(P0.05);MDA含量为2.37 nmol/mL,显著低于对照组(P0.05)。另外,稀释液中添加OPC对稀释液渗透压无显著影响(P0.05)。2.稀释液中添加不同浓度(10、20、30、40、50μg/mL)Nisin到稀释液中,能够在提高精液保存质量同时抑制细菌繁殖。添加40μg/mL Nisin组的常温保存效果最好,保存到第5d精子活率能达到60.12%,精子质膜完整率为44.98%,顶体完整率为71.78%,显著高于对照组(P0.05)。稀释液中添加30μg/mL Nisin抑制金黄色葡萄球菌效果显著优于庆大霉素(P0.05),抑制鲁菲不动杆菌效果与乳酸链球菌素相当(P0.05);稀释液中添加40μg/mL Nisin抑制大肠杆菌效果显著低于庆大霉素(P0.05)。3.猪精液稀释液中联合添加50μg/mL OPC和20μg/mL Nisin的常温保存效果优于单独添加两种物质组和对照组(P0.05),常温保存到第5 d精子活率为62.07%,精子质膜、顶体完整率分别为56.00%和78.64%,精子线粒体活性为60.46%。第5 dOPC:Nisin=50:20组精子T-AOC为1.32 U/mL,酸性磷酸酶活性为573.34 U/L,均显著高于对照组和单独添加药物组(P0.05);MDA含量最少,为2.36 nmol/mL,显著低于其余各组(P0.05)。4.稀释液中添加OPC-Nisin可显著抑制金黄色葡萄球菌生长(P0.05),且作用效果优于庆大霉素,但对大肠杆菌和鲁菲不动杆菌的抑制效果欠佳(P0.05)。综上所述,添加原花青素和乳酸链球菌素能提高精液保存质量并延长保存时间。原花青素最适添加浓度为50μg/mL,乳酸链球菌素最适添加浓度为40μg/mL,联合添加最适浓度为50μg/mL原花青素和20μg/mL乳酸链球菌素。
[Abstract]:With the development of large-scale pig industry, the popularization of artificial insemination technology is expanding, and the quality of semen preservation determines the production efficiency, among which diluents play a crucial role. In the process of semen preservation at room temperature, it is necessary to dilute, which will weaken the original antioxidant effect of semen, make too much reactive oxygen species can not be cleared in time, and the plasma membrane will be easily damaged, which will lead to the decline of sperm quality. At the same time, the existence of microbes will also affect the quality of semen preservation. In order to solve the problem of anti-bacterial and anti-oxidation simultaneously, the semen quality, osmotic pressure, total antioxidant capacity and malondialdehyde (MDA) content were determined by adding proanthocyanidin (Oligomeric Proantho Cyanidins (OPC) and Lactic streptococcin (Nisin),) to the diluent. The activity of acid phosphatase and bacteriostatic effect are expected to provide some theoretical reference for the combined addition of OPC and Nisin in semen diluent. In this study, the following results were obtained: 1: 1. The preservation effect of porcine semen was optimized by adding different concentrations of OPC solution (10 ~ (20) C ~ (30) C ~ (40) C ~ (50) C ~ (60) ~ (70) 渭 g/mL) to porcine semen diluent. The sperm motility, plasma membrane integrity and acrosome integrity were 61.63, 41.88 and 75.88, respectively, which were significantly higher than those in the control group (P0.05). At the 5th day of normal temperature preservation, the T-AOC of sperm was 1.43 U / mL, which was higher than that of the control group (P0.05), and the content of MDA was 2.37 nmol / mL, which was significantly lower than that of the control group (P0.05). In addition, the addition of OPC to the diluent had no significant effect on the osmotic pressure of the diluent (P0.05). Adding different concentrations of Nisin (10 ~ 20 ~ 30 ~ 40 ~ (50 渭 g/mL) to the diluent could improve the quality of semen preservation and inhibit the propagation of bacteria. The sperm motility was 60.12%, the sperm plasma membrane integrity rate was 44.98 and the acrosome integrity rate was 71.78, which was significantly higher than that of the control group (P0.05). The inhibitory effect of 30 渭 g/mL Nisin on Staphylococcus aureus was significantly better than that of gentamicin (P0.05), and the inhibitory effect on Acinetobacter rufii was similar to that on Streptococcus lactis (P0.05), and the inhibitory effect of 40 渭 g/mL Nisin on Escherichia coli was significantly lower than that on gentamicin (P0.05). The effect of adding 50 渭 g/mL OPC and 20 渭 g/mL Nisin to porcine semen diluent at room temperature was better than that of the control group (P0.05). The sperm motility was 62.07 and the plasma membrane of sperm was 62.07 on the 5th day after preservation at room temperature. The acrosome integrity rates were 56.00% and 78.64%, respectively. The sperm mitochondrial activity was 60.46%. The sperm T-AOC and acid phosphatase activity in the fifth dOPC:Nisin=50:20 group were 1.32 U / mL and 573.34 U / L, respectively, which were significantly higher than those in the control group and the control group (P0.05), the lowest (2.36nmol / mL), and significantly lower than those in the other groups (P0.05) .4. The addition of OPC-Nisin in diluent significantly inhibited the growth of Staphylococcus aureus (P0.05), and the effect was better than gentamicin, but the inhibitory effect on Escherichia coli and Acinetobacter rufii was not good (P0.05). In conclusion, the addition of proanthocyanidin and lactic acid streptococcin can improve the quality of semen preservation and prolong the storage time. The optimal concentration of proanthocyanidins was 50 渭 g / mL, the optimal concentration of nisin was 40 渭 g / mL, the optimal concentration of proanthocyanidins and 20 渭 g/mL nisin was 50 渭 g/mL and 20 渭 g/mL respectively.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S828

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