| Summary: | The dilution method and ratio were tested to assess their effects on the <i>Hu</i> ram semen after cryopreservation. Experiment I aimed to explore the effect of various dilution ratios (1:1, 1:2, 1:3, 1:4) of diluent I (Tris-based and egg yolk) under the condition of 1:1 dilution of diluent II (diluent I and glycerol) on the <i>Hu</i> ram semen preserved in liquid nitrogen regarding spermatozoa motility and kinetic parameters. Experiment II aimed to investigate the effect of various dilution ratios (1:1, 1:2, 1:3, 1:4) of diluent I under the condition of 1:2 dilution of diluent II to the <i>Hu</i> ram semen for cryopreservation on spermatozoa motility and kinetic parameters. The purpose of experiment III is to assess the effect of various dilution methods and ratios on the cryopreservation of <i>Hu</i> ram semen by detecting spermatozoa motility, kinetic parameters, plasma membrane integrity, acrosome integrity and reactive oxygen species (ROS) level. Experiment III includes four groups: one-step dilution method and two-step dilution method. The two-step dilution method includes two groups: 1:2, 1:1 and 1:3, 1:2, and the one-step dilution method includes two groups: 1:5 and 1:11. The results indicated that the post-thawed spermatozoa total motility (TM), progressive motility (PM) and average motion degree (MAD) were highest in the 1:2 group and significantly higher (<i>p</i> < 0.05) than those in the 1:1 and 1:4 groups under the condition of 1:1 dilution of diluent II. The post-thawed spermatozoa TM and PM of the 1:3 group were significantly higher (<i>p</i> < 0.05) than those of the other groups under the condition of 1:2 dilution of diluent II. The post-thawed spermatozoa TM, PM, plasma membrane integrity and acrosome integrity of the two-step group (1:3, 1:2) were the highest and significantly higher (<i>p</i> < 0.05) than those in the other groups. Additionally, the post-thawed spermatozoa ROS level of the two-step group (1:3, 1:2) was significantly lower (<i>p</i> < 0.05) than that in the one-step groups (1:5 and 1:11). Therefore, a two-step dilution (1:3, 1:2) was found to be the most suitable method and ratio for diluting the <i>Hu</i> ram semen after cryopreservation.
|
|---|