Structure and composition of the tunic in the sea pineapple Halocynthia roretzi: A complex cellulosic composite...

Biological organisms produce high-performance composite materials, such as bone, wood and insect cuticle, which provide inspiration for the design of novel materials. Ascidians (sea squirts) produce an organic exoskeleton, known as a tunic, which has been studied quite extensively in several species. However, currently, there are still gaps in our knowledge about the detailed structure and composition of this cellulosic biocomposite. Here, we investigate the composition and hierarchical structure of the tough tunic from the species Halocynthia roretzi, through a cross-disciplinary approach combining traditional histology, immunohistochemistry, vibrational spectroscopy, X-ray diffraction, and atomic force and electron microscopies. The picture emerging is that the tunic of H. roretzi is a hierarchically-structured composite of cellulose and proteins with several compositionally and structurally distinct zones. At the surface is a thin sclerotized cuticular layer with elevated composition of protein containing halogenated amino acids and cross-linked via dityrosine linkages. The fibrous layer makes up the bulk of the tunic and is comprised primarily of helicoidally-ordered crystalline cellulose fibres with a lower protein content. The subcuticular zone directly beneath the surface contains much less organized cellulose fibres. Given current efforts to utilize biorenewable cellulose sources for the sustainable production of bio-inspired composites, these insights establish the tunic of H. roretzi as an exciting new archetype for extracting relevant design principles. STATEMENT OF SIGNIFICANCE: Tunicates are the only animals able to produce cellulose. They use this structural polysaccharide to build an exoskeleton called a tunic. Here, we investigate the composition and hierarchical structure of the tough tunic from the sea pineapple Halocynthia roretzi through a multiscale cross-disciplinary approach. The tunic of this species is a composite of cellulose and proteins with two distinct layers. At the surface is a thin sclerotized cuticular layer with a higher protein content containing halogenated amino acids and cross-linked via dityrosine linkages. The fibrous layer makes up the bulk of the tunic and is comprised of well-ordered cellulose fibres with a lower protein content. Given current efforts to utilize cellulose to produce advanced materials, the tunic of the sea pineapple provides a striking model for the design of bio-inspired cellulosic composites.