Quantification and localization of hesperidin and rutin in Citrus sinensis grafted on C-limonia after Xylella fastidiosa infection by HPLC-UV and MALDI imaging mass spectrometry

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Soares, Mamba Santos ^[1] ; da Silva, Danielle Fernandes ^[1] ; Forim, Moacir Rossi ^[1] ; das Gracas Fernandes da Silva, Maria Fatima ^[1] ; Fernandes, Joao Batista ^[1] ; Vieira, Paulo Cezar ^[1] ; Silva, Denise Brentan ^[2] ; Lopes, Norberto Peporine ^[2] ; de Carvalho, Sergio Alves ^[3] ; de Souza, Alessandra Alves ^[3] ; Machado, Marcos Antonio ^[3]

Afiliação do(s) autor(es)

^[1] Univ Fed Sao Carlos, Dept Quim, BR-13565905 Sao Carlos, SP - Brazil

^[2] Univ Sao Paulo, Fac Ciencias Farmaceut Ribeirao Preto, Nucleo Pesquisas Prod Nat & Sintet, BR-14040903 Ribeirao Preto, SP - Brazil

^[3] Inst Agron, Ctr APTA Citros Sylvio Moreira, BR-13490970 Cordeiropolis, SP - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Phytochemistry; v. 115, p. 161-170, JUL 2015.
• Citações Web of Science: 24

Resumo

A high performance liquid chromatography-ultraviolet (HPLC-UV) method was developed for quantifying hesperidin and rutin levels in leaves and stems of Citrus limonia, with a good linearity over a range of 1.0-80.0 and 1.0-50.0 mu g mL(-1) respectively, with r(2) >0.999 for all curves. The limits of detection (LOD) for both flavonoids were 0.6 and 0.5 mu g mL(-1), respectively, with quantification (LOQ) being 2.0 and 1.0 mu g mL(-1), respectively. The quantification method was applied to Citrus sinensis grafted onto C. limonia with and without CVC (citrus variegated chlorosis) symptoms after Xylella fastidiosa infection. The total content of rutin was low and practically constant in all analyses in comparison with hesperidin, which showed a significant increase in its amount in symptomatic leaves. Scanning electron microscopy studies on leaves with CVC symptoms showed vessel occlusion by biofilm, and a crystallized material was noted. Considering the difficulty in isolating these crystals for analysis, tissue sections were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) to confirm the presence of hesperidin at the site of infection. The images constructed from MS/MS data with a specific diagnostic fragment ion (m/z 483) also showed higher ion intensities for it in infected plants than in healthy ones, mainly in the vessel regions. These data suggest that hesperidin plays a role in the plant-pathogen interaction, probably as a phytoanticipin. This method was also applied to C sinensis and C limonia seedlings, and comparison with the graft results showed that the rootstock had an increased hesperidin content similar to 3.6 fold greater in the graft stem than in the stem of C sinensis seedlings. Increase in hesperidin content by rootstock can be related to induced internal defense mechanisms. (C) 2015 Elsevier Ltd. All rights reserved. (AU)

• Processo FAPESP: 08/57859-5 - Controle biorracional de insetos pragas
• Beneficiário: Maria Fátima das Graças Fernandes da Silva
• Modalidade de apoio: Auxílio à Pesquisa - Temático