How to Produce a Chemical Defense: Structural Elucidation and Anatomical Distribution of Aplysioviolin and Phycoerythrobilin in the Sea Hare Aplysia californica

We previously used bioassay‐guided fractionation to identify phycoerythrobilin ( 1 ) and its monomethyl ester, aplysioviolin ( 2 ), as components in the ink secretion of a marine gastropod, the sea hare Aplysia californica , that act as chemical deterrents against predatory blue crabs. This was the first report of 1 as a natural product. Compound 2 was previously reported as a natural product from three species of Aplysia ( A. fasciata, A. dactylomela , and A. parvula ), but the reported structure and composition of stereoisomers of 2 are different among these species. Sea hares are thought to produce 2 from phycoerythrin, a photosynthetic pigment in their red‐algal diet composed of a phycobiliprotein covalently linked to the chromophore 1 , by cleavage of the covalent bond and methylation of 1 , but neither the sequence nor the anatomical location of the cleavage and methylation is known. In this study, we clarify the structure of 1 and 2 in ink secretion of A. californica , and describe the distribution of 1 and 2 in the tissues of sea hares. We conclude that cleavage of the covalent bond in phycoerythrin occurs first, forming 1 in the digestive gland, followed by methylation of 1 to yield 2 in the ink gland.