To understand whether genotypic variation in acid phosphatase (APase) activity in rapeseed (Brassica napus L.) induced by phosphorus (P) deficiency has impact on P efficiency, soil APase activity in the rhizosphere for rapeseed P-efficient genotype 102 and P-inefficient genotype 105 was measured against organic and inorganic P sources in the pot experiment, and the activities of root-secreted APase and leaf intracellular APase were investigated in different P-starvation periods in the nutrient solution. Higher activity of root-secreted APase in B. napus was induced under low P conditions. However, P nutrition and P uptake efficiency of the plants supplied with organic P were not directly related to the activity of root-secreted APase due to several confounding factors affecting APase availability. The higher activity of leaf APase improved P remobilization in plants and played important roles in enhancing P use efficiency, shown by the significant correlation between leaf APase activity and P use efficiency in a rapeseed recombinant inbred population of 135 lines.