This study was conducted to investigate the effect of activating solutions containing different concentrations of calcium, magnesium and pH change in the Billard standard fertilization solution on sperm motility of brown trout. Sperm catching from the male fish was done after anesthetizing. Sperm samples were then mixed together and density (87.0±30.10 million per milliliter) and spermatocrit (23.0±0.58%) of the pooled sample were calculated. Experimental activating solutions were prepared by addition of bivalent ions of calcium and magnesium with a concentration of 1, 2, and 3 mmol L-1 in the Billard solutions. Three other experimental groups were prepared by making changes in the PH of Billrd solution (8, 8.3, and 9). Then, Sperm motility in each experimental solutions and standard solution Billard (control) were compared. The highest Sperm motility of brown trout was found in the solution containing 1 mM calcium ion with the 64.7±8.6 s, which did not show significant difference with the solution containing pH 8 (p>0.01).The minimum duration of sperm motility was also recorded in pH 9.5 fertilization solution with the time of 24.3±2.6 s compared to the other treatments (p<0.01). Also, no significant difference was found in duration of sperm motility between Billard standard solution (control treatment) with treatments containing 1 mM magnesium, 2 mM magnesium and pH 8.5 (p>0.01). Among bivalent ions the most motility of brown trout sperm was observed in the solution containing 1mM calcium. Therefore, the type and concentration of bivalent ions followed by the pH in activating solutions could put the sperm motility under control