title: Cleistochlamys kirkii chemical constituents: Antibacterial activity and synergistic effects against resistant Staphylococcus aureus strains.
abstract: BACKGROUND
  Cleistochlamys kirkii (Benth) Oliv., (Annonaceae) is a medicinal plant traditionally used in Mozambique to treat infectious diseases.
  
  
  OBJECTIVE
  To find antibacterial lead compounds from C. kirkii and provide scientific validation for its use in traditional medicine.
  
  
  METHODS
  Through bioassay-guided fractionation, nine compounds (1-9), with different scaffolds, were isolated from the methanol extract of C. kirkii whose structures were identified by spectroscopic methods. Compounds 1-9 were evaluated for their in vitro antibacterial activity against a panel of eight Gram-positive, including five drug-resistant strains of Staphylococcus aureus, vancomycin-resistant Enterococcus faecalis, and two Gram-negative bacteria strains. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined. A chemosensitization assay, using the checkerboard method, was also performed in order to evaluate the type of interaction of compounds with antibiotics/compounds against two S. aureus resistant strains (ATCC 9144 and CIP 106760) and a susceptible strain (ATCC 6538).
  
  
  RESULTS
  Dichamanetin (3), a rare C-benzylated flavanone, was very active against all the Gram-positive strains tested, displaying MIC values in the range of 1-7.5 μg/mL. The C-benzylated flavanones chamanetin (1), isochamanetin (2), and the α,β-unsaturated lactone (-)-cleistenolide (6) also showed relevant antibacterial activity against some of the Gram-positive strains assayed. Compounds 4, 5, and 7-9 have shown no significant activity at the concentration ranges tested. In the combination with antibiotics, polycarpol (8) (MIC 125 μg/mL) showed a strong synergistic effect against the methicillin-resistant S. aureus ATCC 9144. When combined with oxacillin (MIC 125 μg/mL), compound 8 reduced the MIC to 1.5 μg/mL (FICI=0.11). Similarly, it reduced the MIC of amoxicillin (MIC 250 μg/mL) to 7.5 μg/mL (FICI=0.18). Synergy was also obtained when this compound was combined with both β-lactam antibiotics (FICI=0.30) and with vancomycin (FICI=0.24) against vancomycin-intermediate S. aureus (VISA) CIP 106760. Remarkable, compound 8 was also able to reduce synergistically the MIC value of dichamanetin (3) (FICI=0.18) against this strain.
  
  
  CONCLUSIONS
  These results suggested that C. kirkii constituents may be valuable as a leads for restoring antibiotic activity against resistant S. aureus strains.
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal.
authors.forename: Filipe
authors.initials: F
authors.lastname: Pereira
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal.
authors.forename: Ana Margarida
authors.initials: AM
authors.lastname: Madureira
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal.
authors.forename: Shirley
authors.initials: S
authors.lastname: Sancha
authors.affiliation: Centro de Estudos Moçambicanos e de Etnociências (CEMEC), Faculdade de Ciências Naturais e Matemática, Universidade Pedagógica, Campus de Lhanguene, Av. de Moçambique, 21402161 Maputo, Mozambique.
authors.forename: Silva
authors.initials: S
authors.lastname: Mulhovo
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal.
authors.forename: Xuan
authors.initials: X
authors.lastname: Luo
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal.
authors.forename: Aida
authors.initials: A
authors.lastname: Duarte
authors.affiliation: Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Av. Prof. Gama Pinto, 1649-003 Lisbon, Portugal. Electronic address: mjuferreira@ff.ulisboa.pt.
authors.forename: Maria-José U
authors.initials: MJ
authors.lastname: Ferreira
mesh_terms: D029061:Annonaceae / Q000737:chemistry*; D000900:Anti-Bacterial Agents / Q000737:chemistry* / Q000494:pharmacology*; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects*; D008826:Microbial Sensitivity Tests / Q000379:methods; D010936:Plant Extracts / Q000737:chemistry / Q000494:pharmacology; D010946:Plants, Medicinal / Q000737:chemistry*; D013211:Staphylococcus aureus / Q000187:drug effects*; D014640:Vancomycin / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D010936:Plant Extracts; D014640:Vancomycin
keywords: Annonaceae; Antibacterial; C-benzylated flavanones; Cleistochlamys kirkii; Dug-resistant Staphylococcus aureus; Synergy