Chemical composition, immunostimulatory, cytotoxic and antiparasitic activities of the essential oil from Brazilian red propolis

Most studies of Brazilian red propolis have explored the composition and biological properties of its ethanolic extracts. In this work, we chemically extracted and characterized the essential oil of Brazilian red propolis (EOP) and assessed its adjuvant, antiparasitic and cytotoxic activities. The chemical composition of EOP was analyzed using gas chromatography with mass spectrometry (GC-MS). EOP was tested for in vitro activity against Trichomonas vaginalis (ATCC 30236 isolate); trophozoites were treated with different concentrations of EOP (ranging from 25 to 500 mu g/mL) in order to establish the MIC and IC50 values. A cytotoxicity assay was performed in CHO-K1 cells submitted to different EOP concentrations. BALB/c mice were used to test the adjuvant effect of EOP. The animals were divided in 3 groups and inoculated as follows: 0.4 ng/kg BW EOP (G1); 50 mu g of rCP40 protein (G2); or a combination of 0.4 ng/kg BW EOP and 50 mu g of rCP40 (G3). Total IgG, IgG1 and IgG2a levels were assessed by ELISA. The major constituent compounds of EOP were methyl eugenol (13.1%), (E)-beta-farnesene (2.50%), and d-amorphene (2.3%). Exposure to EOP inhibited the growth of T. vaginalis, with an IC50 value of 100 mu g/mL of EOP. An EOP concentration of 500 mu g/mL was able to kill 100% of the T. vaginalis trophozoites. The EOP kinetic growth curve showed a 36% decrease in trophozoite growth after a 12 h exposure to 500 mu g/mL of EOP, while complete parasite death was induced at 24 h. With regard to CHO-K1 cells, the CC50 was 266 mu g/mL, and 92% cytotoxicity was observed after exposure to 500 mu g/mL of EOP. Otherwise, a concentration of 200 mu g/mL of EOP was able to reduce parasite proliferation by 70% and was not cytotoxic to CHO-K1 cells. As an adjuvant, a synergistic effect was observed when EOP was combined with the rCP40 protein (G3) in comparison to the administration of each component alone (G1 and G2), resulting in higher concentrations of IgG, IgG1 and IgG2a. EOP is constituted by biologically active components with promising antiparasitic and immunostimulatory activities and can be investigated for the formulation of new vaccines or trichomonacidal drugs.