Biomechanics and composition of fixed neurocranial tessellated cartilage from the neurocranium of four mobulid ray species with different depth ranges

This study investigated how the mechanical and biochemical properties of neurocranial tessellated cartilage vary among four mobulid ray species occupying progressively greater depth ranges: Mobula eregoodoo (0-50 m), Mobula thurstoni (0-100 m), Mobula mobular (0-1112 m), and Mobula tarapacana (0-1896 m). Because specimens from these highly protected pelagic species are often preserved before analysis, the authors first evaluated the influence of neutral buffered formalin fixation using neurocranial cartilage from a ray, shark, and chimaera. Fixation generally reduced mechanical properties, with M. eregoodoo showing substantial decreases in high-strain Young’s modulus, ultimate strength, toughness, and stress-relaxation behavior compared with frozen tissue. The four fixed Mobula species were then characterized across multiple mechanical scales using bulk unconfined compression, MicroTester micro-indentation, and nanoindentation together with measurements of glycosaminoglycan content, water content, and tissue mineral density. Bulk testing showed a general decline in high-strain stiffness, ultimate strength, and toughness with increasing species depth range, while stress-relaxation time generally increased. Micro-indentation similarly identified significant interspecies differences in local stiffness: full-layered M. eregoodoo cartilage was significantly stiffer than M. tarapacana cartilage, while the unmineralized cartilage of M. thurstoni was significantly stiffer than M. tarapacana. Total water content increased in the deepest-ranging species and was negatively correlated with high-strain stiffness, toughness, and ultimate strength, whereas glycosaminoglycan content and tissue mineral density did not significantly predict these bulk mechanical properties. Overall, the findings suggest that mobulid species occupying greater depth ranges possess a more hydrated and mechanically compliant neurocranial cartilage matrix and demonstrate that tissue fixation can substantially reduce the measured mechanical properties of tessellated cartilage while generally preserving interspecies trends.