Genotoxic, phytotoxic and protective effects of Portuguese propolis
Propolis is a mixture produced by bees (Apis mellifera L.) from various plant sources with a diverse
chemical composition including many bioactive phenolic compounds and terpenes characteristic of plant secondary
metabolism. The bees use propolis mainly as a sealant for cracks in the beehive, but it has been used in human folk
medicine for several millennia. A renewed interest in its study emerged in recent years due to the increasing popularity
of natural products in foods, beverages and medicines.
Propolis extracts have been associated with varied biological activities, like antioxidant, antimicrobial and
anti-inflammatory among others, and indicated for the treatment of several pathologies such as cancer or
neurodegenerative diseases. Portuguese propolis, usually regarded as a by-product without any value in apiaries,
remains insufficiently studied, hence the need for a better characterization of its properties.
In this study Portuguese propolis samples from different apiaries and different collection dates were
selected and ethanolic and n-hexane extracts were prepared and tested using a yeast (Saccharomyces cerevisiae) and in
vitro flax (Linum usitatissimum) cultures as models. Viability assays with yeast cells and plant growth analysis were
performed to assess extracts toxicity. The yeast model was used to investigate mechanisms of cytotoxicity by the
mitochondrial membrane potential-targeting fluorochrome rhodamine 123 approach and to evaluate genotoxicity by the
comet assay. The in vitro plant model was used to evaluate extracts effects at the multicellular developmental level as
well as in the photosynthetic function by pulse amplitude modulated fluorometry. Both photoinhibitiory and
photoprotective potential against oxidative damage of the extracts were investigated using plants grown under high light
intensities. Extracts antioxidant properties were also studied in yeast by flow cytometry using the redox-sensitive
fluorochrome dichlorofluorescein diacetate.
Our results suggest that ethanolic extracts from Pereiro and Povoa do Varzim apiaries are among the most
toxic for both yeast and plants. They decrease yeast viability and mitochondrial membrane potential and dramatically
affect early plant development inhibiting particularly root growth, photosynthetic efficiency and increasing nonphotochemical
quenching in a dose-dependent manner. Significant genotoxicity was found only in ethanolic extracts
from Pereiro collected in 2010, also one of the most toxic in both models tested. However, some ethanolic extracts were
also able to revert oxidative-induced damage. They reduced intracellular oxidation induced by hydrogen peroxide in
yeast, and greatly recovered the total chlorophyll content reduced by high-light-induced photooxidative stress. Here, also
n-hexane extracts were effective.
Globally our results are in line with the antioxidant properties revealed by propolis worldwide but also
underline the strong toxicity in different cellular models suggesting that eukaryotic universal…