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Optimization of sample preparation for intact cell mass spectrometry (matrix-assisted laser desorption/ionization linear time-of-flight mass spectrometry) of endophytic Xylaria

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Autor(es):
Amaral, Luciana da Silva [1, 2] ; Rodrigues-Filho, Edson [1] ; Kubicek, Christian P. [3, 4] ; Herwig, Christoph [3] ; Marchetti-Deschmann, Martina [2] ; Allmaier, Guenter [2]
Número total de Autores: 6
Afiliação do(s) autor(es):
[1] Univ Fed Sao Carlos, Dept Quim, CP 676, BR-13565905 Sao Carlos, SP - Brazil
[2] Vienna Univ Technol, TU Wien, Inst Chem Technol & Analyt, Getreidemarkt 9-164, A-1060 Vienna - Austria
[3] Vienna Univ Technol, TU Wien, Inst Chem Environm & Biosci Engn, Getreidemarkt 9-166, A-1060 Vienna - Austria
[4] Steinschotelgasse 7, A-1100 Vienna - Austria
Número total de Afiliações: 4
Tipo de documento: Artigo Científico
Fonte: RAPID COMMUNICATIONS IN MASS SPECTROMETRY; v. 32, n. 10, p. 815-823, MAY 30 2018.
Citações Web of Science: 0
Resumo

RationaleAlthough the fruiting-body of the fungi of the genus Xylaria shows a great variety of morphological characteristics, their mycelial forms are always very similar, imposing difficulties for their identification. Intact cell mass spectrometry (ICMS) using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) can be a fast and reliable strategy to support the differentiation/identification of Xylaria species in those cases where fruit-bodies are not available. MethodsMany experimental parameters such as sample preparation and culture media are crucial for filamentous fungi analysis by MALDI-TOFMS. For the purposes of this study, we used four matrices (CHCA, DHB, FA and SA) with five different concentrations (0.1, 0.3, 0.5, 1.0 and 2.5%) of TFA in the matrix, the influence of six different culture media (solid and liquid), and three mycelium peptide/protein extraction protocols (acid, basic and thymol-supported solution) to optimize the sample preparation of the endophytic fungus X. arbuscula. ResultsIt was observed that sinapinic acid (30mg/mL) dissolved in acetonitrile/0.1% TFA and PDA were the best matrix solution and culture medium, respectively, for the ICMS of X. arbuscula. The formic acid and ammonium bicarbonate (AB) protocols provided similar mass spectra; however, a higher number of peaks were observed using AB extraction. Mass spectra obtained from different thymol-containing solutions (EtOH/aqueous 0.1% TFA and ACN/aqueous 0.1% TFA) show increasing peak abundances at m/z 3000-6500. ConclusionsX. arbuscula could be analyzed by ICMS. However, an extraction step was required to provide suitable MALDI mass spectra. Formic acid-, AB- and thymol-containing solutions were demonstrated to be good cocktails for the extraction of peptide/protein biomarkers from these fungi. (AU)

Processo FAPESP: 13/18603-3 - Estudos metabolômicos e biossintéticos visando contribuições Pará a química e taxonomia dè micro-organismos da família Xylariaceae
Beneficiário:Luciana da Silva Amaral
Linha de fomento: Bolsas no Brasil - Doutorado
Processo FAPESP: 10/52312-8 - Rede de estudos em biodiversidade microbiana: isolamento, desenvolvimento de estratégias de identificação rápida, conservação, interações entre espécies e explorações biotecnológicas
Beneficiário:Edson Rodrigues Filho
Linha de fomento: Auxílio à Pesquisa - Programa BIOTA - Regular