The effect of yttrium addition on the grain refinement, formation of long period stacking ordered (LPSO) structures, and improved hardness in hot extruded Mg99.5-xNi0.5Yx (x = 0, 0.5, and 1 at%) alloys was investigated. It was revealed that with increasing Y content, the volume fraction of the LPSO phase (VLPSO) increased, and the average dynamic recrystallization (DRX) grain size (D) was refined. It was shown and quantified that both VLPSO and D contribute to the improvement of hardness, where the deviation from the Hall-Petch relationship was ascribed to the strengthening effect of VLPSO. The equation of H = 36.7 + 0.64 VLPSO + 35/√D was proposed to quantify the synergistic effects of hardening mechanisms to predict the hardness of Mg99.5-xNi0.5Yx alloys after deformation by hot extrusion.