本文關(guān)鍵詞： 豬精液 常溫保存 D-異抗壞血酸鈉 溶菌酶 精液保存效果　出處：《西北農(nóng)林科技大學(xué)》2017年碩士論文　論文類(lèi)型：學(xué)位論文

【摘要】：精液稀釋和保存是人工授精技術(shù)的關(guān)鍵步驟,在豬精液常溫保存過(guò)程中,隨著保存時(shí)間的延長(zhǎng),過(guò)量活性氧(ROS)在精液中開(kāi)始積累,而豬精子的特殊膜結(jié)構(gòu)使其更容易受到自由基的氧化損傷,最終導(dǎo)致精子活率下降、頂體受損甚至受精能力喪失。此外,精液中的細(xì)菌和一些病原微生物不僅會(huì)直接和精子競(jìng)爭(zhēng)營(yíng)養(yǎng)物質(zhì),其代謝活動(dòng)的產(chǎn)物還會(huì)損傷精子的正常生理功能,這是精液質(zhì)量下降、人工授精效果降低的重要原因。本試驗(yàn)通過(guò)測(cè)定精子活率、質(zhì)膜完整率、頂體完整率、總抗氧化能力(T-AOC)、丙二醛(MDA)含量、活性氧(ROS)水平、精子運(yùn)動(dòng)性能和精液中細(xì)菌含量等指標(biāo),探究了D-異抗壞血酸鈉和溶菌酶對(duì)豬精液常溫保存效果的影響,并分析篩選了兩種物質(zhì)在豬精液常溫保存基礎(chǔ)稀釋粉中的最適添加濃度,為豬精液常溫保存稀釋粉配方的改良提供了參考。以下為本研究主要結(jié)果:1.將不同濃度(0、0.002、0.004、0.006、0.008、0.010 mg/mL)的D-異抗壞血酸鈉添加到豬精液常溫保存基礎(chǔ)稀釋液中可以不同程度地改善精液保存效果。隨著D-異抗壞血酸鈉濃度的增加,精液的保存效果呈現(xiàn)先增后減的趨勢(shì)。當(dāng)D-異抗壞血酸鈉的濃度為0.006 mg/mL時(shí),精液有效保存時(shí)間為5d,且精子活率、質(zhì)膜完整率、頂體完整率和精液總抗氧化能力(T-AOC)顯著高于對(duì)照組和其他試驗(yàn)組(P0.05),丙二醛(MDA)含量和活性氧(ROS)水平顯著低于其他組。保存第3d時(shí),0.006 mg/mL試驗(yàn)組的精子運(yùn)動(dòng)性能參數(shù)最好,且顯著優(yōu)于對(duì)照組和低濃度組(0.002 mg/mL)。當(dāng)D-異抗壞血酸鈉的濃度為0.008 mg/mL時(shí),精液各項(xiàng)指標(biāo)雖然顯著高于對(duì)照組(P0.05),但相比于0.006mg/mL試驗(yàn)組卻有明顯的下降趨勢(shì),所以,添加0.006 mg/mL D-異抗壞血酸鈉時(shí),精液常溫保存效果最好。2.在豬精液常溫保存基礎(chǔ)稀釋粉中添加不同濃度(0、0.5、1.0、1.5、2.0、2.5 mg/mL)的溶菌酶,對(duì)于豬精液的常溫保存有一定的改善作用。當(dāng)溶菌酶的濃度為2.0 mg/mL時(shí),保存各天的精子活率、質(zhì)膜完整率和頂體完整率都顯著高于其他各組(P0.05)。保存的第1 d、第3 d和第5 d,同一天的精液中的細(xì)菌含量隨著溶菌酶濃度的增加而減少,到第5d時(shí),各組間細(xì)菌含量差異顯著,當(dāng)溶菌酶的濃度為2.5 mg/mL時(shí),精液中的細(xì)菌含量顯著低于其他各組(P0.05)。保存第3d,當(dāng)溶菌酶濃度為2.0 mg/mL時(shí),VSL(直線速度)、VAP(路徑速度)和ALH(側(cè)擺幅度)3項(xiàng)運(yùn)動(dòng)性能參數(shù)顯著高于對(duì)照組(P0.05),CMS(曲線運(yùn)動(dòng))和BCF(鞭打頻率)則顯著低于對(duì)照組(P0.05),其他各試驗(yàn)組間無(wú)顯著性差異。綜合對(duì)比結(jié)果表明,2.0 mg/mL為溶菌酶在豬精液常溫保存稀釋粉中的最適添加濃度。3.在交叉混合試驗(yàn)中,將0.006 mg/mL的D-異抗壞血酸鈉和1.5 mg/mL的溶菌酶聯(lián)合添加到豬精液常溫保存基礎(chǔ)稀釋粉中能夠顯著提升保存后豬精子的活率和質(zhì)膜完整率,且效果顯著好于對(duì)照組和兩種物質(zhì)單獨(dú)使用的試驗(yàn)組(P0.05)。綜上,D-異抗壞血酸鈉和溶菌酶對(duì)于豬精液常溫保存的效果有一定的提高作用,能夠有效保護(hù)精子性能,提升精液保存質(zhì)量,其最適添加濃度分別為0.006 mg/mL和2.0mg/mL,聯(lián)合添加的最適濃度組合為0.006 mg/mL的D-異抗壞血酸鈉和1.5 mg/mL的溶菌酶。
[Abstract]:Semen dilution and preservation is a key step in artificial insemination, pig semen preservation at room temperature in the process, with the increase of the storage time, the excess reactive oxygen species (ROS) accumulation in semen, and the special membrane structure of pig sperm to make it easier to oxidative damage by free radicals, resulting in decreased sperm motility, top body damage or even loss of ability to fertilization. In addition, semen bacteria and some pathogenic microorganisms will not only directly and sperm competition of nutrients, the normal physiological function of the product will damage the sperm metabolic activities, the semen quality decline, the important reason to reduce the effects of artificial insemination. In this experiment, through the determination of sperm motility, membrane integrity rate of acrosome integrity rate, total antioxidant capacity (T-AOC), malondialdehyde (MDA) content, reactive oxygen species (ROS) level, bacteria content in semen and sperm motility index performance, explores the D- and sodium erythorbate Effect of preservation effect of lysozyme on boar semen of normal temperature, and the analysis of the screening of two kinds of material stored in the powder based diluted the optimum concentration in normal porcine semen, provides a reference for improvement of preservation of diluted powder formula for boar semen of normal temperature. The following is the main results of this study: 1. different concentrations (0,0.002,0.004,0.006,0.008,0.010 mg/mL) D- ISO sodium ascorbate was added to the normal pig semen preservation based diluent can improve the semen preservation effect. With the increase of D- concentration of sodium erythorbate, preservation of semen showed the trend of first increase and then decrease. When D- concentration of sodium D-isoascorbate was 0.006 mg/mL, semen effective preservation time is 5D, and sperm the survival rate, plasma membrane integrity, acrosome integrity and sperm total antioxidant capacity (T-AOC) was significantly higher than the control group and the other experimental groups (P0.05), malondialdehyde (MDA) content and reactive oxygen species (ROS) The level is significantly lower than other groups. The retention of article 3D, the best sperm motion parameters of 0.006 mg/mL in the experimental group, and significantly better than the control group and low concentration group (0.002 mg/mL). When the D- concentration of sodium D-isoascorbate was 0.008 mg/mL, while the semen indexes were significantly higher than the control group (P0.05), but compared to 0.006mg/mL the test group was significantly decreased, so, adding 0.006 mg/mL D- sodium erythorbate, adding different concentrations of semen stored at room temperature the best preserved in.2. Based Diluted pig semen powder at room temperature (0,0.5,1.0,1.5,2.0,2.5 mg/mL) of lysozyme, for boar semen stored at room temperature to a certain extent. When the concentration of lysozyme was 2 mg/mL when the day of the preservation of sperm motility, plasma membrane integrity and acrosome integrity rate were significantly higher than other groups (P0.05). The preservation of first D, third D and fifth D, the same day in semen containing bacteria The amount decreases with the increasing of lysozyme concentration at 5D, the difference between groups was significant when the bacterial content, lysozyme concentration was 2.5 mg/mL, the bacterial content in semen was significantly lower than that of the other groups (P0.05). The preservation of 3D, when the concentration of lysozyme was 2 mg/mL, VSL (straight line speed (VAP). The path and ALH (speed) side swing) 3 sports performance parameters were significantly higher than the control group (P0.05), CMS (curve movement) and BCF (BCF) is significantly lower than the control group (P0.05), the other was no significant difference between each experimental group. Comparison results show that 2 mg/mL lysozyme in room temperature preservation of boar sperm dilution powder in the optimum concentration of.3. in the cross hybrid test, 0.006 mg/mL D- d-vic sodium and 1.5 mg/mL lysozyme and added to the room temperature preservation of boar sperm dilution based powder can significantly improve the preservation of pig sperm motility and membrane integrity Rate, and the effect was significantly better than the control group and the experimental group of two substances used alone (P0.05). In conclusion, D- of sodium erythorbate and lysozyme has a certain improving effect on pig semen of normal temperature preservation effect, can effectively protect the sperm performance, enhance the quality of semen preservation, the optimal concentration was 0.006 mg/mL and 2.0mg/mL, in combination with the optimal concentration combination of 0.006 mg/mL D- sodium erythorbate and 1.5 mg/mL respectively.

【學(xué)位授予單位】：西北農(nóng)林科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：S828

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