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The equine hoof wall has outstanding impact resistance, which enables high-velocity gallop over hard terrain with minimum damage. To better understand its viscoelastic behavior, complex moduli were de- termined using two complementary techniques: conventional ( ∼5 mm length scale) and nano ( ∼1 μm length scale) dynamic mechanical analysis (DMA). The evolution of their magnitudes was measured for two hydration conditions: fully hydrated and ambient. The storage modulus of the ambient hoof wall was approximately 400 MPa in macro-scale experiments, decreasing to ∼250 MPa with hydration. In contrast, the loss tangent decreased for both hydrated ( ∼0.1–0.07) and ambient ( ∼0.04–0.01) conditions, over the frequency range of 1–10 Hz. Nano-DMA indentation tests conducted up to 200 Hz showed little frequency dependence beyond 10 Hz. The loss tangent of tubular regions showed more hydration sensitivity than in intertubular regions, but no significant difference in storage modulus was observed. Loss tangent and effective stiffness were higher in indentations for both hydration levels. This behavior is attributed to the hoof wall’s hierarchical structure, which has porosity, functionally graded aspects, and material interfaces that are not captured at the scale of indentation. The hoof wall’s viscoelasticity characterized in this work has implications for the design of bioinspired impact-resistant materials and structures. 

The equine hoof wall has a complex, hierarchical structure that can inspire designs of impact-resistant materials. In this study, we utilized micro-computed tomography (micro-CT) and serial block-face scanning electron microscopy (SBF-SEM) to image the microstructure and nanostructure of the hoof wall. We quantified the morphology of tubular medullary cavities by measuring equivalent diameter, surface area, volume, and sphericity. Highresolution micro-CT revealed that tubules are partially or fully filled with tissue near the exterior surface and become progressively empty towards the inner part of the hoof wall. Thin bridges were detected within the medullary cavity, starting in the middle section of the hoof wall and increasing in density and thickness towards the inner part. Porosity was measured using three-dimensional (3D) micro-CT, two-dimensional (2D) micro-CT, and a helium pycnometer. The highest porosity was obtained using the helium pycnometer (8.07%), followed by 3D (3.47%) and 2D (2.98%) micro-CT. SBF-SEM captured the 3D structure of the hoof wall at the nanoscale, showing that the tubule wall is not solid, but has nano-sized pores, which explains the higher porosity obtained using the helium pycnometer. The results of this investigation provide morphological information on the hoof wall for the future development of hoof-inspired materials and offer a novel perspective on how various measurement methods can influence the quantification of porosity.