Brazilian red propolis: phytochemical screening, antioxidant activity and effect against cancer cells

Abstract

Background: The implementation of new public healthcare models that stimulate the use of natural products from traditional medicine, as a so-called integrated medicine, refers to an approach that use best of both conventional medicine and traditional medicine. Propolis is a widely used natural product by different ancient cultures and known to exhibit biological activities beneficial for health. The large number of studies conducted with propolis had shown that its chemical composition differs as a function of the climate, plant diversity and bee species and plays an important role on its therapeutic properties. The aim of this study was to analyse the phytochemical profile of the ethanolic extract of red propolis (EEP) and its fractionation, antioxidant action of EEP and its fractions hexane, cloroform and ethyl acetate and cytotoxic activity of EEP on human tumour cell lines SF-295 (glioblastoma), OVCAR-8 (ovary) and HCT-116 (colon).

Methods: EEP was obtained by maceration with absolute ethanol, then it was concentrated in rotaevaporator up to complete evaporation of the solvent. The crude extract was fractionated with hexane, ethyl acetate, chloroform and methanol and they were subjected to phytochemical screening and total phenolic compounds. Antioxidant activity of EEP and fractions was done by means of the 2,2-diphenyl-1-picryhydrazyl (DPPH) method. Biomarkers of red propolis were identified by LC-Orbitrap-FTMS. To assess cytotoxic activity of the extract, cells were exposed to EEP over 72 h. Cell viability was assessed by means of MTT assay. The percentage of cell growth inhibition (IC50) was analysed by means of non-linear regression, and the absorbance values of the various investigated concentrations were subjected to one-factor analysis of variance (ANOVA) followed by Tukey's or Tamhane's tests (α = 0.05).

Results: The results obtained using phytochemical screening and LC-Orbitrap-FTMS indicated the presence of phlobaphene tannins, catechins, chalcones, aurones, flavonones, flavonols, xanthones, pentacyclic triterpenoids and guttiferones in Brazilian red propolis. EEP and its hexane, chloroform and ethyl acetate fractions obtained by liquid-liquid partitioning exhibited satisfactory antioxidant percentages. EEP (IC50 < 34.27 μg/mL) exhibited high levels of cytotoxicity on all human tumour cell lines tested when compared to negative control.

Conclusions: C-Orbitrap-FTMS was useful to establish the chemical profile of the red propolis. Brazilian red propolis has antioxidant properties and decreases substantially the percentage of cell survival of human tumour cells; thus, it has potential to serve as an anticancer drug.

Fig. 1

Cell viability 72 h after exposure to various EEP concentrations. Cell survival is expressed as a percentage relative to the negative control (DMSO). EEP at a concentration of 50 μg/mL, exhibited cytotoxic activity against HCT-116 and OVCAR-8 cells, without a significant difference compared with the positive control (0.5 μM doxorubicin). Compared with the other two cell lines, SF-295 cells exposed to EEP at a concentration of 50 μg/mL exhibited a significantly higher survival rate compared with those exposed to doxorubicin. Each bar graph represents the mean, and the error bars represent ± SD of three independent experiments performed in triplicate

Fig. 2

Concentration of phenolic compounds in mg of gallic acid equivalents/g of extract detected in EEP and fractions diluted in hexane, chloroform and ethyl acetate. The highest values corresponded to the chloroform fraction

Fig. 3

Chromatogram of EEP obtained by means of LC-Orbitrap-FTMS