Elucidating the role of nanostructured pigment granules in the dynamic coloration of cephalopods

Cephalopods such as squid, octopus, and cuttlefish can quickly and accurately modulate their coloration in order to camouflage themselves to avoid predators, hunt prey, or communicate within their natural environment. This ability is made possible, in part, by chromatophores: pigmentary organs that can expand or relax to impart a global color change to the animal. While it is known that visible color originates from the nanostructured pigment granules that populate the chromatophore, their compositional and structural contributions to coloration remain largely unknown. The overarching goal of this dissertation is to elucidate the structure and function of the cephalopod chromatophore pigment granules as a means to inspire the design of new advanced optical materials. First, the presence of ommochrome pigments within the chromatophore granules was confirmed, and these pigments were identified as xanthommatin and decarboxylated xanthommatin. In this process, the pigments were extracted from the granules of isolated squid Doryteuthis pealeii chromatophores, resulting in a 70.8% decrease in the diameter of the granules, as well as a loss of absorbance, suggesting that non-pigmentary structural components, likely proteins, contribute to granule structure as well. In the same study, we examined the effect of granule structure on the optical properties of the pigments, finding that the extracted pigment features a blue shift in maximum absorbance as well as an altered fluorescence profile compared to the intact granules, suggesting that the optical properties of chromatophore pigment granules arise from an interaction between the pigmentary and structural components. Next, we performed a proteomic study of chromatophores and pigment granules isolated from D. pealeii dermal tissue to determine what proteins are in the granules that may also contribute to coloration. Lens crystallin proteins, specifically Ω- and S-crystallin isoforms, were found in the pigment granules, suggesting that the interaction between the pigment and these high-refractive index proteins may enhance absorbance through scattering effects. Reflectin, a cephalopod-specific protein responsible for structural coloration, was also identified in chromatophores, but not within the pigment granules. This finding inspired a re-examination of the chromatophore, leading to the conclusion that these organs incorporate elements of protein-based structural coloration, likely through thin-film interference that may amplify the colors displayed by the animal, originating from reflectin in sheath cells in the periphery of the chromatophore. Finally, a new, scalable method for the synthesis of xanthommatin, the primary pigment confined to the chromatophore granules, was developed and characterized. By replacing the traditionally used oxidizing agent with an applied electrochemical potential, the reaction becomes simpler while retaining its kinetics, specificity, and yield. Computational chemistry, informed by previous literature, was used to determine the mechanisms underlying the reaction so that it can more easily be applied to the synthesis of xanthommatin derivatives. Together, these results indicate a possible pathway through which the production of asymmetric phenoxazinones might be scaled up for future materials applications.