Bioassay guided purification of the antimicrobial fraction of a Brazilian propolis from Bahia state.
BACKGROUND
Brazilian propolis type 6 (Atlantic forest, Bahia) is distinct from the other types of propolis especially due to absence of flavonoids and presence of other non-polar, long chain compounds, but presenting good in vitro and in vivo antimicrobial activity. Several authors have suggested that fatty acids found in this propolis might be responsible for its antimicrobial activity; however, so far no evidence concerning this finding has been reported in the literature. The goals of this study were to evaluate the antibacterial activity of the main pure fatty acids in the ethanolic extract and fractions and elucidate the chemical nature of the bioactive compounds isolated from Brazilian propolis type 6.


METHODS
Brazilian propolis type 6 ethanolic extract (EEP), hexane fraction (H-Fr), major fatty acids, and isolated sub-fractions were analyzed using high performance liquid chromatography (HPLC), high resolution gas chromatography with flame ionization detection (HRGC-FID), and gas chromatography-mass spectrometry (GC-MS). Three sub-fractions of H-Fr were obtained through preparative HPLC. Antimicrobial activity of EEP, H-Fr, sub-fractions, and fatty acids were tested against Staphyloccus aureus ATCC 25923 and Streptococcus mutans Ingbritt 1600 using minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC).


RESULTS
EEP and H-Fr inhibited the growth of the microorganisms tested; nevertheless, no antimicrobial activity was found for the major fatty acids. The three sub-fractions (1, 2, and 3) were isolated from H-Fr by preparative HPLC and only sub-fraction 1 showed antimicrobial activity.


CONCLUSIONS
a) The major fatty acids tested were not responsible for the antimicrobial activity of propolis type 6; b) Sub-fraction 1, belonging to the benzophenone class, was responsible for the antimicrobial activity observed in the present study. The identification of the bioactive compound will improve the development of more efficient uses of this natural product.
Castro
Myrella Lessio
ML
Department of Physiological Sciences, Piracicaba Dental School, University of Campinas; Avenida Limeira, Piracicaba, SP 13414-903, Brazil.
Vilela
Walquíria Ribeiro
WR
Zauli
Rogéria Cristina
RC
Ikegaki
Masaharu
M
Rehder
Vera Lúcia Garcia
VL
Foglio
Mary Ann
MA
de Alencar
Severino Matias
SM
Rosalen
Pedro Luiz
PL
D000818:Animals; D000900:Anti-Bacterial Agents / Q000302:isolation & purification / Q000494:pharmacology*; D001516:Bees; D001577:Benzophenones / Q000494:pharmacology*; D001938:Brazil; D002849:Chromatography, Gas; D002851:Chromatography, High Pressure Liquid; D005227:Fatty Acids / Q000302:isolation & purification / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D011429:Propolis / Q000737:chemistry / Q000494:pharmacology*; D013211:Staphylococcus aureus / Q000187:drug effects*; D013295:Streptococcus mutans / Q000187:drug effects*
D000900:Anti-Bacterial Agents; D001577:Benzophenones; D005227:Fatty Acids; C047723:benzophenone; D011429:Propolis