title: Antibacterial mode of action of the hydroethanolic extract of Leonotis nepetifolia (L.) R. Br. involves bacterial membrane perturbations.
abstract: BACKGROUND
  Leonotis nepetifolia (L) R. Br., Lamiaceae, a pantropical shrub, popularly known in Brazil as "cordão-de-frade", "rubim", is reportedly used in Brazilian ethnomedicine as well as in different countries in the treatments of ailments such as infections, inflammations, wounds, stomach disorders, among others.
  
  
  OBJECTIVE
  To evaluate its potential cytotoxicity and antibacterial mode of action of the hydroethanolic extract of L. nepetifolia (HELn) leaves, including phytochemical analysis.
  
  
  METHODS
  The cytotoxicity of HELn was investigated by Alamar blue assay, using CHO-K1 cells. Antibacterial activity of HELn was tested by broth microdilution methods against a panel of bacteria of clinical interest. The mode of action of L. nepetifolia was studied by targeting bacterial membranes. Phytochemical analysis was performed by determining total secondary metabolites with spectrophotometric assays and HPLC.
  
  
  RESULTS
  HELn is not cytotoxic in the in vitro evaluation (IC50>200 μg/mL). It demonstrated a good spectrum of antibacterial activity with major activity against Shigella flexneri, Enterococcus faecalis, Staphylococcus aureus and Bacillus subtilis with MIC=6.25 µg/mL, Helicobacter pylori with MIC of 25 µg/mL and Streptococcus pyogenes with MIC of 50 µg/mL. Its mode of action is associated, at least partly, with changes in the permeability of bacterial membranes, as evidenced by the increased entry of hydrophobic antibiotics in Shigella flexneri and intense efflux of K(+) and nucleotide leakage in E. faecalis and Shigella flexneri. In addition, the presence of phenols, flavonoids and carotenoids, described in the literature to possess antibacterial effects, were detected in the composition of HELn, with high phenol content (11.55%), especially the flavonoids (6.47%).
  
  
  CONCLUSIONS
  The results indicate that HELn has low cytotoxicity and potent antibacterial activity. It is bacteriostatic in nature, possibly acting at the level of bacterial membranes, especially on the cytoplasmic membrane and outer membrane, thus supporting its popular use in infectious processes. In addition, the presence of phenols, flavonoids, carotenoids, fatty acids and steroids, described in the literature as possessing antimicrobial activity, were detected in the composition of HELn.
authors.affiliation: Area of Pharmacology, Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), 78060-900 Cuiabá, MT, Brazil.
authors.forename: Darley Maria
authors.initials: DM
authors.lastname: Oliveira
authors.affiliation: Research Laboratory of Natural Products, Federal University of Tocantins, Faculty of Medicine, 7020-210 Palmas, TO Brazil.
authors.forename: Fernanda Germano
authors.initials: FG
authors.lastname: Melo
authors.affiliation: Area of Pharmacology, Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), 78060-900 Cuiabá, MT, Brazil.
authors.forename: Sikiru Olaitan
authors.initials: SO
authors.lastname: Balogun
authors.affiliation: Department of Chemistry, Federal University of Roraima, 69304-000 Boa Vista, RR, Brazil.
authors.forename: Adriana
authors.initials: A
authors.lastname: Flach
authors.affiliation: Department of Chemistry, Federal University of Roraima, 69304-000 Boa Vista, RR, Brazil.
authors.forename: Edineide Cristina Alexandre
authors.initials: EC
authors.lastname: de Souza
authors.affiliation: Department of Chemistry, Federal University of Roraima, 69304-000 Boa Vista, RR, Brazil.
authors.forename: Gilmar Prado
authors.initials: GP
authors.lastname: de Souza
authors.affiliation: Department of Chemistry, Federal University of Roraima, 69304-000 Boa Vista, RR, Brazil.
authors.forename: Iolanda do Nascimento Araújo
authors.initials: Ido N
authors.lastname: Rocha
authors.affiliation: Area of Pharmacology, Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), 78060-900 Cuiabá, MT, Brazil.
authors.forename: Luiz Antonio Mendonça Alves
authors.initials: LA
authors.lastname: da Costa
authors.affiliation: Research Laboratory of Natural Products, Federal University of Tocantins, Faculty of Medicine, 7020-210 Palmas, TO Brazil.
authors.forename: Ilsamar Mendes
authors.initials: IM
authors.lastname: Soares
authors.affiliation: Area of Pharmacology, Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), 78060-900 Cuiabá, MT, Brazil.
authors.forename: Larissa Irene
authors.initials: LI
authors.lastname: da Silva
authors.affiliation: Research Laboratory of Natural Products, Federal University of Tocantins, Faculty of Medicine, 7020-210 Palmas, TO Brazil.
authors.forename: Sérgio Donizeti
authors.initials: SD
authors.lastname: Ascêncio
authors.affiliation: Area of Pharmacology, Department of Basic Sciences in Health, Faculty of Medicine, Federal University of Mato Grosso (UFMT), 78060-900 Cuiabá, MT, 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 / Q000008:administration & dosage / Q000302:isolation & purification / Q000494:pharmacology*; D001419:Bacteria / Q000187:drug effects*; D001938:Brazil; D016466:CHO Cells; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D003412:Cricetulus; D020128:Inhibitory Concentration 50; D019686:Lamiaceae / Q000737:chemistry*; D008519:Medicine, Traditional; D008826:Microbial Sensitivity Tests; D010936:Plant Extracts / Q000008:administration & dosage / Q000494:pharmacology*; D018515:Plant Leaves
chemical_list: D000900:Anti-Bacterial Agents; D010936:Plant Extracts
keywords: Antimicrobial; Flavonoids; Lamiaceae; Leonotis nepetifolia; Phenolics; Phytochemistry