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dc.contributor.authorAlves, Elianapt_PT
dc.contributor.authorMelo, Tâniapt_PT
dc.contributor.authorSimões, Cláudiapt_PT
dc.contributor.authorFaustino, Maria A. F.pt_PT
dc.contributor.authorTomé, João P. C.pt_PT
dc.contributor.authorNeves, Maria G. P. M. S.pt_PT
dc.contributor.authorCavaleiro, José A. S.pt_PT
dc.contributor.authorCunha, Ângelapt_PT
dc.contributor.authorGomes, Newton C. M.pt_PT
dc.contributor.authorDomingues, Pedropt_PT
dc.contributor.authorDomingues, M. Rosário M.pt_PT
dc.contributor.authorAlmeida, Adelaidept_PT
dc.description.abstractRATIONALE The photodynamic process involves the combined use of light and a photosensitizer, which, in the presence of oxygen, originates cytotoxic species capable of oxidizing biological molecules, such as lipids. However, the effect of the photodynamic process in the bacterial phospholipid profile by a photosensitizer has never been reported. A lipidomic approach was used to study the photodynamic oxidation of membrane phospholipids of Staphylococcus warneri by a tricationic porphyrin [5,10,15‐tris(1‐methylpyridinium‐4‐yl)‐20‐(pentafluorophenyl)porphyrin triiodide, Tri‐Py+‐Me‐PF]. METHODS S. warneri (108 colony forming units mL–1) was irradiated with white light (4 mW cm–2, 21.6 J cm–2) in the presence of Tri‐Py+‐Me‐PF (5.0 μM). Non‐photosensitized bacteria were used as control (irradiated without porphyrin). After irradiation, total lipids were extracted and separated by thin‐layer chromatography (TLC). Isolated fractions of lipid classes were quantified by phosphorus assay and analyzed by mass spectrometry (MS): off‐line TLC/ESI‐MS, hydrophilic interaction (HILIC)‐LC/MS and MS/MS. RESULTS The most representative classes of S. warneri phospholipids were identified as phosphatidylglycerols (PGs) and cardiolipins (CLs). Lysyl‐phosphatidylglycerols (LPGs), phosphatidylethanolamines (PEs), phosphatidylcholines (PCs) and phosphatidic acids (PAs) were also identified. After photodynamic treatment, an overall increase in the relative abundance of PGs was observed as well as the appearance of new oxidized species from CLs, including hydroxy and hydroperoxy derivatives. Formation of high amounts of lipid hydroperoxides was confirmed by FOX2 assay. Photodynamic oxidation of phospholipid standards revealed the formation of hydroperoxy and dihydroperoxy derivatives, confirming the observed CL oxidized species in S. warneri. CONCLUSIONS Membrane phospholipids of S. warneri are molecular targets of the photoinactivation process induced by Tri‐Py+‐Me‐PF. The overall modification in the relative amount of phospholipids and the formation of lipid hydroxides and hydroperoxides indicate the lethal damage caused to photosensitized bacterial cells.pt_PT
dc.description.sponsorshipThe authors thank Vanessa Oliveira MSc. for her help in bacteria identification by PCR testing. Thanks are due to Fundação para a Ciência e a Tecnologia (FCT, Portugal), European Union, QREN, FEDER and COMPETE for funding the QOPNA research unit (project PEst‐C/QUI/UI0062/2011 and the project PTDC/QUI‐BIQ/104968/2008), to the Portuguese National NMR Network that was also supported by funds from FCT, and to the RNEM (REDE/1504/REM/2005) for the Portuguese Mass Spectrometry Network. Thanks are also due to the Centre for Environmental and Marine Studies for funding the Microbiology Research Group (project Pest‐C/MAR/LA0017/2011). Eliana Alves (SFRH/BD/41806/2007), Tânia Melo (SFRH/BD/84691/2012) and Cláudia Simões (SFRH/BD/46293/2008) are grateful to FCT for their PhD grants.pt_PT
dc.titlePhotodynamic oxidation of Staphylococcus warneri membrane phospholipids: new insights based on lipidomicspt_PT
degois.publication.titleRapid Communications in Mass Spectrometrypt_PT
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