title: Antimicrobial photodynamic effect of phenothiazinic photosensitizers in formulations with ethanol on Pseudomonas aeruginosa biofilms.
abstract: BACKGROUND
  Methylene blue (MB) and toluidine blue (TB) are recognized as safe photosensitizers (Ps) for use in humans. The clinical effectiveness of the antimicrobial photodynamic therapy with MB and TB needs to be optimized, and ethanol can increase their antimicrobial effect. Formulations of MB and TB containing ethanol were evaluated for their ability to produce singlet oxygen and their antibacterial effect on Pseudomonas aeruginosa biofilms.
  
  
  METHODS
  Photoactivated formulations were prepared by diluting the Ps (250 μM) in buffered water (pH 5.6, sodium acetate/acetic acid), 10% ethanol (buffer: ethanol, 90:10), or 20% ethanol (buffer: ethanol, 80:20). Biofilms also were exposed to the buffer, 10% ethanol, or 20% ethanol without photoactivation. Untreated biofilm was considered the control group. The production of singlet oxygen in the formulations was measured based on the photo-oxidation of 1,3-diphenylisobenzofuran. The photo-oxidation and CFU (log10) data were evaluated by two-way ANOVA and post-hoc Tukey's tests.
  
  
  RESULTS
  In all the formulations, compared to TB, MB showed higher production of singlet oxygen. In the absence of photoactivation, neither the buffer nor the 10% ethanol solution showed any antimicrobial effect, while the 20% ethanol solution significantly reduced bacterial viability (P=0.009). With photoactivation, only the formulations containing MB and both 10% and 20% ethanol solutions significantly reduced the viability of P. aeruginosa biofilms when compared with the control.
  
  
  CONCLUSIONS
  MB formulations containing ethanol enhanced the antimicrobial effect of the photodynamic therapy against P. aeruginosa biofilms in vitro.
authors: Graduate Program in Oral Science, Periodontology Unit, School of Dentistry, Federal University of Santa Maria, Santa Maria, Rio Grande do Sul State, Brazil. Electronic address: emiproch@hotmail.com.
authors: Emilia Pithan
authors: EP
authors: Prochnow
authors: Graduate Program in Oral Science, Periodontology Unit, School of Dentistry, Federal University of Santa Maria, Santa Maria, Rio Grande do Sul State, Brazil. Electronic address: maritieli_rm@hotmail.com.
authors: Maritieli Righi
authors: MR
authors: Martins
authors: Graduate Program in Oral Science, Periodontology Unit, School of Dentistry, Federal University of Santa Maria, Santa Maria, Rio Grande do Sul State, Brazil. Electronic address: cibelecampgnolo@hotmail.com.
authors: Cibele Bruno
authors: CB
authors: Campagnolo
authors: Graduate Program in Nanoscience, Franciscan University Center, Santa Maria, Rio Grande do Sul, Brazil. Electronic address: robertochrist@gmail.com.
authors: Roberto Christ
authors: RC
authors: Santos
authors: Graduate Program in Chemistry, Chemical-Physical Unit, Federal University of Santa Maria, Santa Maria, Rio Grande do Sul State, Brazil. Electronic address: mvilletti@hotmail.com.
authors: Marcos Antonio
authors: MA
authors: Villetti
authors: Graduate Program in Oral Science, Periodontology Unit, Stomatology Department, School of Dentistry, Federal University of Santa Maria, Santa Maria, Rio Grande do Sul State, Brazil. Electronic address: kzkantorski@hotmail.com.
authors: Karla Zanini
authors: KZ
authors: Kantorski
mesh_terms: D018441:Biofilms / Q000187:drug effects* / Q000254:growth & development; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D004339:Drug Compounding / Q000379:methods; D000431:Ethanol / Q000737:chemistry*; D010640:Phenothiazines / Q000008:administration & dosage* / Q000737:chemistry; D010778:Photochemotherapy / Q000379:methods*; D017319:Photosensitizing Agents / Q000008:administration & dosage / Q000737:chemistry; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000502:physiology*; D026082:Singlet Oxygen / Q000737:chemistry / Q000528:radiation effects; D013242:Sterilization / Q000379:methods
chemical_list: D010640:Phenothiazines; D017319:Photosensitizing Agents; D026082:Singlet Oxygen; D000431:Ethanol
keywords: Laser in dentistry; Microbiology; Photodynamic inactivation; Photodynamic therapy (PDT)