The primary objective of this work was to analyse the adequacy of the Double Cantilever Beam (DCB) test in determining fracture toughness under pure mode I loading of cortical bone tissue. A new data reduction scheme based on specimen compliance and the crack equivalent concept was used to overcome the difficulties inherent in crack monitoring during its growth. It provides a complete resistance curve, which is fundamental in estimating the fracture energy. A cohesive zone model was used to simulate damage initiation and propagation, thus assessing the efficacy of the proposed testing method and data reduction scheme. Subsequently, the DCB test was applied to evaluate the mode I fracture energy of hydrated and thermally dehydrated cortical bone tissue from young bovine femur, in the tangential-longitudinal propagation system. The results obtained demonstrate the efficacy of the DCB test and the proposed data reduction scheme on the bone fracture characterization under mode I loading.