Physicochemical characterisation of bile pigments in butterflies

Abstract

The blue bottle butterfly, Graphium weiskei, possesses pigmentary blue colouration which is rare in nature, as well as green and purple, on its wings. Previous work claimed that the wing colourations are caused by the combination of bile pigments; sarpedobilin and phorcabilin with other pigments like papillochromes and ommochromes. Here, efforts are made to identify if the colouration is caused by a combination of different pigments, or one pigment that isomerizes into other forms to produce the three different colours.To uncover the chemical nature, Raman spectroscopy is used to ascertain the functional groups of the three colourations. UV-Vis spectroscopy is used to show the absorbance spectrum of the different colours on the wings and in the extracts from the wings, and density functional theory (DFT) is used to compute the various isomeric forms of bile pigments in lepidopterans using pterobilin (biliverdin IXγ) as a starting compound.The results indicate that the three colourations are identical to one another and to G. sarpedon wings which contains sarpedobilin. The wings also different from commercial bile pigment (biliverdin IXα). Extraction using acidifed methanol allows a removal of the green-causing pigment while the pigments causing blue and purple largely remain in the wing. UV-Vis signified that all the colours have similar absorbance peaks around 393 nm and around 674 nm. However, the blue and purple wing portions both have a peak around 580 nm albeit in varying quantities. This pinpoints the 580 nm peak to responsible for the purple colouration and the blue (which leaves behind a purple colouration after extraction). The extraction only successfully worked on the 674 nm peak resulting in a 664 nm peak in acidic methanol confirming sarpedobilin‘s presence. Simulating pterobilin isomers indicates the ability to form isomers that gives green, blue, and purple colourations and vary slightly in Raman spectra.In conclusion, the blue, green, and purple colourations on the wings of Graphium weiskei all contain sarpedobilin and another unknown substance with 393 nm absorbance that is inactive in Raman. Blue and purple wings additionally contain a 579 nm substance which if Raman active may be another sarpedobilin or if not Raman active, may be unknown or a sarpedobilin with a protein attached. The wings may also contain ommochromes or precursor molecules like kynurenines and 3OH-kynurenine in all the portions. Further evidence was given to prove that the 393 nm and 579 nm substances are not carotenoids.Kiyesola Kolewole, B.Eng.Literaturverzeichnis: Seite [44]-[57]Masterarbeit University of Salzburg 202