title: Photodynamic oxidation of Escherichia coli membrane phospholipids: new insights based on lipidomics.
abstract: BACKGROUND
  The irreversible oxidation of biological molecules, such as lipids, can be achieved with a photosensitizing agent and subsequent exposure to light, in the presence of molecular oxygen. Although lipid peroxidation is an important toxicity mechanism in bacteria, the alterations caused by the photodynamic therapy on bacterial phospholipids are still unknown. In this work, we studied the photodynamic oxidation of Escherichia coli membrane phospholipids using a lipidomic approach.
  
  
  METHODS
  E. coli ATCC 25922 were irradiated for 90 min with white light (4 mW cm(-2), 21.6 J cm(-2)) in the presence of a tricationic porphyrin [(5,10,15-tris(1-methylpyridinium-4-yl)-20-(pentafluorophenyl)porphyrin triiodide, Tri-Py(+)-Me-PF]. Lipids were extracted and separated by thin-layer chromatography. Phospholipid classes were quantified by phosphorus assay and analyzed by electrospray ionization tandem mass spectrometry. Fatty acids were analyzed by gas chromatography. Quantification of lipid hydroperoxides was performed by FOX2 assay. Analysis of the photodynamic oxidation of a phospholipid standard was also performed.
  
  
  RESULTS
  Our approach allowed us to see that the photodynamic treatment induced the formation of a high amount of lipid hydroperoxides in the E. coli lipid extract. Quantification of fatty acids revealed a decrease in the unsaturated C16:1 and C18:1 species suggesting that oxidative modifications were responsible for their variation. It was also observed that photosensitization induced the oxidation of phosphatidylethanolamines with C16:1, C18:1 and C18:2 fatty acyl chains, with formation of hydroxy and hydroperoxy derivatives.
  
  
  CONCLUSIONS
  Membrane phospholipids of E. coli are molecular targets of the photodynamic effect induced by Tri-Py(+) -Me-PF. The overall change in the relative amount of unsaturated fatty acids and the formation of PE hydroxides and hydroperoxides evidence the damages in bacterial phospholipids caused by this lethal treatment.
authors.affiliation: Centre for Environmental and Marine Studies, Department of Biology, University of Aveiro, Aveiro, Portugal.
authors.forename: Eliana
authors.initials: E
authors.lastname: Alves
authors.forename: Nuno
authors.initials: N
authors.lastname: Santos
authors.forename: Tânia
authors.initials: T
authors.lastname: Melo
authors.forename: Elisabete
authors.initials: E
authors.lastname: Maciel
authors.forename: M Luísa
authors.initials: ML
authors.lastname: Dória
authors.forename: Maria A F
authors.initials: MA
authors.lastname: Faustino
authors.forename: João P C
authors.initials: JP
authors.lastname: Tomé
authors.forename: Maria G P M S
authors.initials: MG
authors.lastname: Neves
authors.forename: José A S
authors.initials: JA
authors.lastname: Cavaleiro
authors.forename: Ângela
authors.initials: Â
authors.lastname: Cunha
authors.forename: Luisa A
authors.initials: LA
authors.lastname: Helguero
authors.forename: Pedro
authors.initials: P
authors.lastname: Domingues
authors.forename: Adelaide
authors.initials: A
authors.lastname: Almeida
authors.forename: M Rosário M
authors.initials: MR
authors.lastname: Domingues
mesh_terms: D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects*; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism* / Q000528:radiation effects; D005227:Fatty Acids / Q000737:chemistry / Q000378:metabolism; D008027:Light; D013058:Mass Spectrometry; D010084:Oxidation-Reduction; D010743:Phospholipids / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D005227:Fatty Acids; D010743:Phospholipids