Prolong semen quality following the chilled process is important for assistant reproductive techniques such as artificial insemination. The objective of the current study is to evaluate the effect of adding different concentrations of vitamin D3 to the Tris-egg yolk extender on some semen parameters stored at 5ºC. Three Awassi rams were used in this study. Semen samples were collected pooled and diluted with an egg-yolk extender (1:10). Four Aliquots were divided into three treatments with vitamin D3 (T1= 0.02, T2=0.004, T3= 0.002 gm) and one control (T0). All samples were chilled to 5ºC. Samples were estimated at time h0 and after 24, 48,72, and 96 hours following treatment. Individual motility, dead sperm, abnormal sperm, and hypo-osmotic swelling test (HOST) was used in this study. The individual motility significantly decreased in T1 compared to T0, T2, and T3 in all studded hours. Also, the individual motility in T3 was significantly greater at h48, h72, and h96, compared to T0, T1. The dead sperm was significantly greater in T1 on h0 compared to T0, T2, and T3. The HOST test was significantly greater in T0, T2, and T3 on h0 compared to T1. In conclusion, low vitamin D3 supplementation to the Awassi semen might potentiate the individual motility for up to 72 hours and trend to enhance some semen parameters.