Objective Establish and validate a biomechanical modeling method based on micro-magnetic resonance imaging (μMRI) and microstructure segmentation to noninvasively assess microstructural behavior of the proximal femur. Methods Firstly, μMRI images were obtained for the femoral samples, and bone microstructures were segmented through regionized image processing to create the μMRI finite element model (μMRI-FEM). Finite element analysis was performed utilizing a lateral fall posture simulation, and stress and strain results were calculated. Secondly, the accuracy of μMRI image segmentation of bone microstructure was verified using micro-computed tomography (μCT), and the accuracy of μMRI FEM calculation results was verified using a finite element model constructed based on μCT (μCT-FEM). Finally, simulated lateral fall posture, the accuracy of bone surface strain calculated by μMRI-FEM was verified through strain gauge measurements in vitro mechanical loading experiments. Results The bone microstructure parameters BV/TV calculated by μMRI and μCT were significantly correlated (r=0.89, p<0.05). The maximum/minimum principal stress/principal strain percentile results calculated by μMRI-FEM and μCT-FEM were highly correlated (R2>0.9). Moreover, the strain results calculated by μMRI-FEM were highly correlated with the strain results measured by mechanical experiments (R2=0.82). Conclusions The micro finite element model based on μMRI segmentation of bone microstructure can accurately characterize the micro mechanical behavior of the proximal femur, which provided an important tool for non-invasive assessment of hip femur microstructure degeneration and osteoporosis fracture risk in vivo.