The fatigue life of bone illustrates a large degree of scatter that is likely related to underlying differences in composition and microarchitecture. Vascular canals act as stress concentrations, the magnitude and volume of which may depend on the size and spatial distribution of canals. The purpose of this study was to establish the relationship between vascular canal microarchitecture, stressed volume and the fatigue life of both secondary osteonal and plexiform bovine bone. Twenty-one cortical bone samples were prepared from bovine femora and tibiae and imaged using micro-computed tomography (μCT) to quantify canal diameter, canal separation and canal number. Samples were cyclically loaded in zero-compression to a peak magnitude of 95 MPa, and fatigue life was defined as the number of cycles until fracture. Finite element models were created from μCT images and used to quantify the stressed volume, i.e., the volume of bone stressed higher than a yield stress of 108 MPa. Fatigue life ranged from 162-633,437 cycles with the fatigue life of plexiform bone (n = 15) being more than 4.5 times longer than secondary bone (n = 6). The fatigue life of secondary bone was negatively correlated with canal diameter (r² = 0.73) and canal separation (r² = 0.56), while the fatigue life of plexiform bone was negatively correlated with canal separation (r² = 0.41), but positively correlated with canal number (r² = 0.36). Stressed volume was related to canal microarchitecture in secondary bone only, where canal diameters and canal separation were larger than approximately 50 μm and 200 μm, respectively. Consequently, stressed volume explained 89% of the fatigue life variance in secondary bone but was not related to the fatigue life of plexiform bone. These findings suggest that the volume of the stress concentration surrounding vascular canals is dictated by canal size and spacing and may play an important role in the fatigue failure of osteonal bone. We suspect that a larger stressed volume is more likely to encounter and facilitate the propagation of pre-existing microcracks, thereby leading to a reduction in fatigue life.