Contents lists available at ScienceDirect Biochemical Systematics and Ecology journal homepage: www.elsevier.com/locate/biochemsyseco Identifcation of anthocyanins in the corona of two species of Passiflora and their hybrid by ultra-high performance chromatography with electrospray ionization tandem mass spectrometry (UHPLC-ESI-MS/MS) Lilian Cristina Baldon Aizza, Alexandra Christine Helena Frankland Sawaya, Marcelo Carnier Dornelas ∗ Universidade Estadual de Campinas, Brazil ARTICLEINFO Keywords: Anthocyanins Corona Flower color Passiflora Ultra-high performance chromatography electrospray ionization tandem mass spectrometry ABSTRACT Plantsbelongingtothegenus Passiflora produceexuberantfowerswithcharacteristiccorona flamentsthatvary widely in size, morphology and color. This variability is thought to be an adaptation to diferent animal polli- nators.Ingeneral, Passiflora fowersarepollinatedbybees,hummingbirdsandbats.Theaimofthisstudywasto identifytheanthocyaninspresentinthecorona flamentsoftwowildparentspecies, P. incarnata and P. coccinea, that are pollinated by bees and hummingbirds, respectively, and their artifcial commercial hybrid P. ‘Lady Margaret’. The anthocyanin profles of the corona flaments were characterized by ultra-high performance chromatography with electrospray ionization and tandem mass spectrometry (UHPLC-ESI-MS/MS). Monoglucosides of two anthocyanidins (cyanidin and pelargonidin) were found in P. coccinea corona flaments, whereas diglucosides of four anthocyanidins (cyanidin, petunidin, peonidin and malvidin) were identifed in P. incarnata corona. The P. ‘Lady Margaret’ hybrid contained both mono and diglucosides of the anthocyanidins identifed in the parent species and also novel anthocyanin molecules not found in the parents. The use of UHPLC-ESI-MS/MS was an efcient method to identify the anthocyanins present in the corona of Passiflora species. P. incarnata acquired its red fowers by suppression of the delphinidin pathway. The wide variety of anthocyanins found in the corona flaments of the P. ‘Lady Margaret’ interspecifc hybrid indicated that it in- herited the anthocyanidin pathways as well as the glycosylation pattern of both parent species. 1. Introduction An ecological question of great interest is the relationship between foral diversity and pollinators, responsible for the reproductive success of species. Among all foral traits, color provides a visual signal to at- tract preferentially one or another pollinator and has been interpreted as an evolutionary adaptation associated with the attraction of specifc animals (Faegri and van der Piel, 1997; Waser and Price, 1981; Fenster et al., 2004; Thomson and Wilson, 2008; Miller et al., 2009). Usually, anthocyanins play a major role in the pigmentation of fowers and many studies have focused on the importance of anthocyanin produc- tion, especially in petal cells, for pollinator preference (Waser and Price, 1981; Fenster et al., 2004; Thomson and Wilson, 2008). Anthocyanins are favonoid pigments frequently responsible for the bright color of many angiosperm fowers and fruits. The basic antho- cyanidin structure is composed of two aromatic rings (A and B) linked by a central heterocyclic ring (C; Fig.1A)(Wu and Prior, 2005; Tanaka etal.,2008; Hichirietal.,2011).Colorsrangingfromredtopurpleand blue, as well as diverse intermediate hues naturally occurring in plants, are produced by diferent patterns of hydroxylation, methylation, gly- cosylationandpossibleacylationoftheanthocyanidins(Fig.1B, Holton andCornish,1995; Moletal.,1998; Koesetal.,2005; Grotewold, 2006; Halbwirth, 2010). Anthocyanin biosynthesis begins through the condensation of three malonyl-CoA molecules to 4-p-coumaroyl-CoA to form dihy- drokaempferol (DHK) as a result of chalcone synthase, chalcone-fa- vonone isomerase and favonone 3-hydroxylase catalysis (Fig. 2). Di- hydroquercetin and dihydromyricetin are from the DHK B-ring hydroxylation. In this context, favonoid 3′-hydroxylase (F3′H) and favonoid 3′5′-hydroxylase (F3′5′H) are the key enzymes (Holton and https://doi.org/10.1016/j.bse.2019.05.003 Received 14 February 2019; Received in revised form 3 May 2019; Accepted 11 May 2019 ∗ Corresponding author. UNICAMP, Instituto de Biologia, Depto de Biologia Vegetal, R. Monteiro Lobato, 255, 13083-862, Campinas, SP, Brazil. E-mail address: dornelas@unicamp.br (M.C. Dornelas). Biochemical Systematics and Ecology 85 (2019) 60–67 0305-1978/ © 2019 Elsevier Ltd. All rights reserved. T