title: Gallesia integrifolia (Spreng.) Harms: In vitro and in vivo antibacterial activities and mode of action.
abstract: BACKGROUND
  Gallesia integrifolia (Phytolaccaceae) is commonly known as "pau-d'alho" in Brazil or "garlic plant" due to the strong scent of garlic peculiar to all parts of the plant. The bark decoction is used for the treatment of microbial infections among other diseases by different ethnic groups in Brazil, Peruvian Amazonians, Bolivia and Mosetene Indians. This study aimed to advance in the antibacterial activity and characterize the mode of action of the hydroethanolic extract of the inner stem bark of G. integrifolia (HEGi) using in vivo and in vitro experimental models.
  
  
  METHODS
  The qualitative and quantitative phytochemical analyzes of HEGi were carried out using colorimetric and HPLC technique. The cytotoxic potential of HEGi was evaluated against CHO-K1 cells by Alamar blue assay and its acute toxicity was assessed by the Hippocratic screening test using Swiss-Webster mice. The antibacterial activity was evaluated by micro- dilution method against ten strains of Gram-positive and Gram-negative bacteria. The mode of action of HEGi was investigated by outer membrane permeability, nucleotide leakage and potassium efflux assays. In vivo infection model was established by using Staphylococcus aureus infection model Wistar rats.
  
  
  RESULTS
  Qualitative phytochemical analysis of HEGi revealed the presence of saponins, alkaloids, phenolic compounds and flavonoids. Phytochemical quantification of HEGi showed that higher total phenolic (80.10±0.62mg GAE/g) and flavonoid (16.10±0.03mg RE/g) contents. HPLC fingerprint analysis revealed the presence of gallic acid, rutin, and morin. In the Alamar blue assay no cytotoxic effect of HEGi in CHO-K1 cells was observed up to 200µg/mL, and no signs or symptoms of acute toxicity were observed in mice of both sexes at higher doses of up to 2000mg/kg, p.o. HEGi demonstrated bacteriostatic effect against selected Gram positive and Gram negative bacterial pathogens. Its mode of action is associated, at least partly, with changes in the permeability of bacterial membranes, evidenced by the increased entry of hydrophobic antibiotic in Pseudomonas aeruginosa, intense K(+) efflux and nucleotides leakage in Shigella flexneri, Streptococcus pyogenes and S. aureus. HEGi attenuated the experimental blood borne S. aureus infection in rats at all the tested doses levels (10, 50 and 250mg/kg).
  
  
  CONCLUSIONS
  HEGi is safe at the dose tested when used acutely, and it presented broad antibacterial effect, which support its traditional use in the treatment of bacterial infections. It contains well known important phytochemicals, recognized to be active against bacterial pathogens in vitro and might be collectively responsible for the antibacterial activity of HEGi. It is bacteriostatic in nature, with membrane perturbation being one of it mode of action. HEGi represent a potential phytotherapic antibacterial agent.
authors.affiliation: Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), Av. Fernando Correa da Costa, no. 2367, Coxipó, Cuiabá, Mato Grosso 78060-900, Brazil.
authors.forename: Karuppusamy
authors.initials: K
authors.lastname: Arunachalam
authors.affiliation: Research Laboratory of Natural Products, Federal University of Tocantins, Faculty of Medicine, Palmas, Tocantins 77020-210, Brazil.
authors.forename: Sérgio Donizeti
authors.initials: SD
authors.lastname: Ascêncio
authors.affiliation: Research Laboratory of Natural Products, Federal University of Tocantins, Faculty of Medicine, Palmas, Tocantins 77020-210, Brazil.
authors.forename: Ilsamar Mendes
authors.initials: IM
authors.lastname: Soares
authors.affiliation: Department of Biotechnology, Federal University of Tocantins, Gurupi, Tocantins 77404-970, Brazil.
authors.forename: Raimundo Wagner
authors.initials: RW
authors.lastname: Souza Aguiar
authors.affiliation: Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), Av. Fernando Correa da Costa, no. 2367, Coxipó, Cuiabá, Mato Grosso 78060-900, Brazil.
authors.forename: Larissa Irene
authors.initials: LI
authors.lastname: da Silva
authors.affiliation: Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), Av. Fernando Correa da Costa, no. 2367, Coxipó, Cuiabá, Mato Grosso 78060-900, Brazil.
authors.forename: Ruberlei Godinho
authors.initials: RG
authors.lastname: de Oliveira
authors.affiliation: Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), Av. Fernando Correa da Costa, no. 2367, Coxipó, Cuiabá, Mato Grosso 78060-900, Brazil.
authors.forename: Sikiru Olaitan
authors.initials: SO
authors.lastname: Balogun
authors.affiliation: Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), Av. Fernando Correa da Costa, no. 2367, Coxipó, Cuiabá, Mato Grosso 78060-900, Brazil. Electronic address: taba@terra.com.br.
authors.forename: Domingos Tabajara
authors.initials: DT
authors.lastname: de Oliveira Martins
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology* / Q000627:therapeutic use*; D001522:Behavior, Animal / Q000187:drug effects; D016466:CHO Cells; D002470:Cell Survival / Q000187:drug effects; D003412:Cricetulus; D005260:Female; D006090:Gram-Negative Bacteria / Q000187:drug effects / Q000254:growth & development; D006094:Gram-Positive Bacteria / Q000187:drug effects / Q000254:growth & development; D008297:Male; D051379:Mice; D008826:Microbial Sensitivity Tests; D029602:Phytolaccaceae; D008517:Phytotherapy; D024301:Plant Bark; D010936:Plant Extracts / Q000494:pharmacology* / Q000627:therapeutic use*; D017208:Rats, Wistar; D013203:Staphylococcal Infections / Q000188:drug therapy* / Q000382:microbiology
chemical_list: D000900:Anti-Bacterial Agents; D010936:Plant Extracts
keywords: Antimicrobial; Cytotoxicity; Flavonoids; Gallesia integrifolia; Polyphenols; Traditional medicine