title: A fast and precise method to identify indolic glucosinolates and camalexin in plants by combining mass spectrometric and biological information.
abstract: In this manuscript, a fast and accurate identification and quantitation by mass spectrometry of indolic glucosinolates and camalexin involved in defense in Arabidopsis thaliana are described. Two elicitation systems, inoculation with Botrytis cinerea and treatment with AgNO(3), were used in Col-0 wild-type and mutant genotypes impaired in the biosynthesis of the selected metabolites. Identification of analytes was carried out by nontargeted LC/ESI-QTOF-MS profiling. Confirmation of indolic glucosinolates and camalexin was achieved by their absence in the cyp79B2/B3 and pad3 mutants as well as their respective fragmentation upon collision-induced dissociation. Camalexin accumulation was induced only after AgNO(3) treatment, whereas all indolic glucosinolates were constitutively present. Inoculation with Botrytis did not influence camalexin concentration but caused most aliphatic and indolic glucosinolates contents to decrease. Only the pen 3.1 mutant showed increased indolic glucosinolate levels after Botrytis or AgNO(3) treatments. In addition, profiles of secondary metabolite in nontreated Col-0 and mutant plants were analyzed by means of partial least squares coupled to discriminant analysis (PLS-DA), and differences in the basal levels of indolic glucosinolates and tryptophan between cyp79B2/B3 plants and the rest of genotypes, including Col-0, were found. This probably has to be taken into consideration when comparing stress responses of Col-0 and cyp79B2/B3. The use of mutants carrying alterations in biosynthetic pathways is proposed as a useful strategy to identify secondary metabolites.
authors.affiliation: Departament de Ciències Agràries i del Medi Natural, Universitat Jaume I, Castelló de la Plana, Spain.
authors.forename: Sara Izquierdo
authors.initials: SI
authors.lastname: Zandalinas
authors.forename: Vicente
authors.initials: V
authors.lastname: Vives-Peris
authors.forename: Aurelio
authors.initials: A
authors.lastname: Gómez-Cadenas
authors.forename: Vicent
authors.initials: V
authors.lastname: Arbona
mesh_terms: D017360:Arabidopsis / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D020171:Botrytis; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D005961:Glucosinolates / Q000032:analysis* / Q000096:biosynthesis; D007211:Indoles / Q000032:analysis* / Q000378:metabolism; D013058:Mass Spectrometry; D009154:Mutation; D012835:Silver Nitrate / Q000494:pharmacology; D013844:Thiazoles / Q000032:analysis* / Q000378:metabolism; D000094162:Phytoalexins
chemical_list: D029681:Arabidopsis Proteins; D003577:Cytochrome P-450 Enzyme System; D005961:Glucosinolates; D007211:Indoles; D012835:Silver Nitrate; D013844:Thiazoles; C102405:camalexin; D000094162:Phytoalexins; C530432:cytochrome P-450 79B2 protein, Arabidopsis; C530433:cytochrome P-450 79B3, Arabidopsis