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In order to improve the quality of frozen spermatozoa of Yunnan semi-fine wool sheep, 1, 4-cyclohexanediol (1, 4-CHD) as a synthetic ice blocker was used for cryopreservation of ram spermatozoa in this study. Briefly, following collection by electric stimulation, equilibration at 5℃ following dilution with the freezing extender, and pre-freezing in liquid nitrogen vapor, the ram spermatozoa were preserved in liquid nitrogen for one month. In addition, the effects of osmolarity of the diluting extenders used for evaluation of frozen spermatozoa quality were also assessed. The results indicated addition of 1, 4-CHD could not increase the motility of ram spermatozoa after cryopreservation and thawing. With the elevation of the concentrations of 1, 4-CHD, the motility and moving velocity of frozen ram spermatozoa showed a steady decrease. Additionally, the presence of 1, 4-CHD cannot increase the percentage of frozen spermatozoa with intact acrosome and membrane. When the isotonic binding buffer was used to dilute the thawed spermatozoa, the percentage of cells labeled with propidium iodide (PI) after cryopreservation in the presence of 1, 4-CHD was significantly higher than that of spermatozoa frozen in the absence of 1, 4-CHD (P0.05). However, the percentage of frozen-thawed spermatozoa with exposed PS in the presence of 1, 4-CHD was significantly less than that of spermatozoa frozen in the absence of 1, 4-CHD (P0.01). When the basic extenders with an osmolarity of 404mOsm, 528mOsm, 648mOsm, or 853mOsm were used to dilute the frozen-thawed spermatozoa respectively, there is no significant difference between the four groups with respect to the moving velocity and membrane integrity (P0.05). In conclusion, the presence of 1, 4-CHD cannot improve the motility, moving velocity, acrosome staus, and membrane integrity of frozen ram spermatozoa. However, 1, 4-CHD may inhibit apoptosis caused by freezing and thawing.