title: Esculetin and esculin (esculetin 6-O-glucoside) occur as inclusions and are differentially distributed in the vacuole of palisade cells in Fraxinus ornus leaves: a fluorescence microscopy analysis.
abstract: The location of individual coumarins in leaves of Fraxinus ornus acclimated at full solar irradiance was estimated using their specific UV- and fluorescence spectral features. Using a combination of UV-induced fluorescence and blue light-induced fluorescence of tissues stained with diphenylborinic acid 2-amino-ethylester, in wide field or confocal laser scanning microscopy, we were able to visualize the distribution of esculetin and esculetin 6-O-glucoside (esculin) in palisade cells. Coumarins are not uniformly distributed in the cell vacuole, but accumulate mostly in the adaxial portion of palisade cells. Our study indeed shows, for the first time, that coumarins in palisade cells accumulate as vacuolar inclusions, as previously reported in the pertinent literature only for anthocyanins. Furthermore, esculetin and esculin have a different vacuolar distribution: esculetin largely predominates in the first 15 μm from the adaxial epidermis. This leads to hypothesize for esculetin and esculin different transport mechanisms from the endoplasmic reticulum to the vacuole as well as potentially different roles in photoprotection. Our study open to new experiments aimed at exploring the mechanisms that deliver coumarins to the vacuole using different fluorescence signatures of coumarin aglycones and coumarin glycosides.
authors: Istituto per la Protezione delle Piante, Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, I-50019, Sesto. F. no, Firenze, Italy.
authors: Massimiliano
authors: M
authors: Tattini
authors: Dipartimento di Scienze delle Produzioni Agroalimentari e dell'Ambiente, sez. Colture Arboree, Università di Firenze, Viale delle Idee 30, I-50019, Sesto. F. no, Firenze, Italy.
authors: Martina
authors: M
authors: Di Ferdinando
authors: Istituto per la Protezione delle Piante, Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, I-50019, Sesto. F. no, Firenze, Italy; Dipartimento di Scienze delle Produzioni Agroalimentari e dell'Ambiente, sez. Colture Arboree, Università di Firenze, Viale delle Idee 30, I-50019, Sesto. F. no, Firenze, Italy.
authors: Cecilia
authors: C
authors: Brunetti
authors: Dipartimento di Chimica 'Ugo Schiff', Università di Firenze, Via della Lastruccia 3, I-50019, Sesto. F. no, Firenze, Italy.
authors: Andrea
authors: A
authors: Goti
authors: Istituto per la Protezione delle Piante, Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, I-50019, Sesto. F. no, Firenze, Italy.
authors: Susanna
authors: S
authors: Pollastri
authors: Dipartimento di Scienze delle Produzioni Agroalimentari e dell'Ambiente, sez. Colture Arboree, Università di Firenze, Viale delle Idee 30, I-50019, Sesto. F. no, Firenze, Italy; Department of Animal and Plant Sciences, University of Sheffield, Western Bank, Sheffield S10 2TN, UK.
authors: Chandra
authors: C
authors: Bellasio
authors: Centro di Microscopie Elettroniche (Ce.M.E.), Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, I-50019, Sesto. F. no, Firenze, Italy.
authors: Cristiana
authors: C
authors: Giordano
authors: Dipartimento di Scienze delle Produzioni Agroalimentari e dell'Ambiente, sez. Colture Arboree, Università di Firenze, Viale delle Idee 30, I-50019, Sesto. F. no, Firenze, Italy.
authors: Alessio
authors: A
authors: Fini
authors: Istituto di Fisica Applicata 'Carrara' (IFAC), Consiglio Nazionale delle Ricerche, Via Madonna del Piano 10, I-50019, Sesto. F. no, Firenze, Italy. Electronic address: g.agati@ifac.cnr.it.
authors: Giovanni
authors: G
authors: Agati
mesh_terms: D004929:Esculin / Q000032:analysis* / Q000378:metabolism; D031661:Fraxinus / Q000737:chemistry* / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D058503:Mesophyll Cells / Q000737:chemistry / Q000378:metabolism; D008856:Microscopy, Fluorescence; D018515:Plant Leaves / Q000737:chemistry / Q000378:metabolism / Q000528:radiation effects; D013056:Spectrophotometry, Ultraviolet; D014466:Ultraviolet Rays; D014468:Umbelliferones / Q000032:analysis / Q000378:metabolism; D014617:Vacuoles / Q000737:chemistry / Q000378:metabolism
chemical_list: D014468:Umbelliferones; D004929:Esculin; C007628:esculetin
keywords: Confocal laser scanning microscopy (CLSM); Coumarins; Fluorescence micro-imaging; Fluorescence micro-spectroscopy; Naturstoff reagent; UV-induced auto-fluorescence; Vacuolar inclusions