The purple pigment aplysioviolin in sea hare ink deters predatory blue crabs through their chemical senses

Sea hares release an ink secretion composed of purple ink and white opaline as a potential chemical defence against predators. The aim of our study was to identify deterrent molecules in the ink of Aplysia californica against an allopatric generalist crustacean predator, the blue crab Callinectes sapidus, and to define the mechanisms of action of the deterrents against crabs. We used two behavioural assays, a squirting assay and an ingestion assay, to show that ink is highly effective and that opaline is moderately effective in suppressing feeding of crabs. Results with reversibly blinded crabs demonstrate that the deterrence is mediated through the crabs’ chemical senses. We used bioassay-guided fractionation to identify the purple molecules aplysioviolin and phycoerythrobilin as a major and minor deterrent, respectively, in ink against crabs. These molecules derive from a light-harvesting protein in the photosynthetic system of dietary algae. This is the first demonstration of an animal converting a photosynthetic pigment into a chemical deterrent. Mixing opaline and ink enzymatically produces hydrogen peroxide, which also functions as a chemical deterrent against crabs. Our results and those of other studies show that sea hares use a diversity of molecules in their skin, mucus and ink secretion to chemically defend themselves against their potential predators. Aplysioviolin, phycoerythrobilin and hydrogen peroxide also exist in ink secretion of Aplysia dactylomela, a sea hare sympatric to blue crabs, and thus we posit that these molecules are potentially effective in ecologically relevant predator–prey interactions and need to be scrutinized in more ecologically relevant experiments.