title: Synergistic mutual potentiation of antifungal activity of Zuccagnia punctata Cav. and Larrea nitida Cav. extracts in clinical isolates of Candida albicans and Candida glabrata.
abstract: BACKGROUND
  Zuccagnia punctata Cav. (Fabaceae) and Larrea nitida Cav. (Zygophyllaceae) are indistinctly or jointly used in traditional medicine for the treatment of fungal-related infections. Although their dichloromethane (DCM) extract have demonstrated moderate antifungal activities when tested on their own, antifungal properties of combinations of both plants have not been assessed previously.
  
  
  OBJECTIVE
  The aim of this study was to establish with statistical rigor whether Z. punctata (ZpE) and L. nitida DCM extract (LnE) interact synergistically against the clinically important fungi Candida albicans and Candida glabrata and to characterize the most synergistic combinations.
  
  
  METHODS
  For synergism assessment, the statistical-based Boik's design was applied. Eight ZpE-LnE fixed-ratio mixtures were prepared from four different months of 1 year and tested against Candida strains. Lϕ (Loewe index) of each mixture at different fractions affected (ϕ) allowed for the finding of the most synergistic combinations, which were characterized by HPLC fingerprint and by the quantitation of the selected marker compounds.
  
  
  METHODS
  Lϕ and confidence intervals were determined in vitro with the MixLow method, once the estimated parameters from the dose-response curves of independent extracts and mixtures, were obtained. Markers (four flavonoids for ZpE and three lignans for LnE) were quantified in each extract and their combinations, with a valid HPLC-UV method. The 3D-HPLC profiles of the most synergistic mixtures were obtained by HPLC-DAD.
  
  
  RESULTS
  Three over four IC50ZpE/IC50LnE fixed-ratio mixtures displayed synergistic interactions at effect levels ϕ > 0.5 against C. albicans. The dosis of the most synergistic (Lϕ = 0.62) mixture was 65.96 µg/ml (ZpE = 28%; LnE = 72%) containing 8 and 36% of flavonoids and lignans respectively. On the other hand, one over four IC50ZpE/IC50LnE mixtures displays synergistic interactions at ϕ > 0.5 against C. glabrata. The dosis of the most synergistic (Lϕ = 0.67) mixture was 168.23 µg/ml (ZpE = 27%; LnE = 73%) with 9.7 and 31.6% of flavonoids and lignans respectively.
  
  
  CONCLUSIONS
  Studies with the statistical-based MixLow method, allowed for the finding of the most ZpE-LnE synergistic mixtures, giving support to a proper joint use of both antifungal herbs in traditional medicine.
authors.affiliation: Área Farmacognosia, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina.
authors.forename: Estefanía
authors.initials: E
authors.lastname: Butassi
authors.affiliation: Área Farmacognosia, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina.
authors.forename: Laura A
authors.initials: LA
authors.lastname: Svetaz
authors.affiliation: Área Estadística, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina.
authors.forename: Juan J
authors.initials: JJ
authors.lastname: Ivancovich
authors.affiliation: Instituto de Biotecnología, Facultad de Ingeniería, Universidad Nacional de San Juan, Av. Libertador General San Martín 1109 (O), 5400 San Juan, Argentina.
authors.forename: Gabriela E
authors.initials: GE
authors.lastname: Feresin
authors.affiliation: Instituto de Biotecnología, Facultad de Ingeniería, Universidad Nacional de San Juan, Av. Libertador General San Martín 1109 (O), 5400 San Juan, Argentina.
authors.forename: Alejandro
authors.initials: A
authors.lastname: Tapia
authors.affiliation: Área Farmacognosia, Facultad de Ciencias Bioquímicas y Farmacéuticas, Universidad Nacional de Rosario, Suipacha 531, 2000 Rosario, Argentina. Electronic address: szaabgil@citynet.net.ar.
authors.forename: Susana A
authors.initials: SA
authors.lastname: Zacchino
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D002176:Candida albicans / Q000187:drug effects*; D041221:Candida glabrata / Q000187:drug effects*; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D007887:Fabaceae / Q000737:chemistry*; D032442:Larrea / Q000737:chemistry*; D008826:Microbial Sensitivity Tests; D015394:Molecular Structure; D035261:Plant Components, Aerial / Q000737:chemistry; D010936:Plant Extracts / Q000494:pharmacology*; D053719:Tandem Mass Spectrometry
chemical_list: D000935:Antifungal Agents; D010936:Plant Extracts
keywords: Antifungal; Bi-herbal combinations; Larrea nitida; MixLow method; Synergism; Zuccagnia punctata