Most of the antioxidant compounds present in propolis affect the viability of a number of undesirable microorganisms, but their role on probiotic microorganisms has not yet been properly elucidated. This research aims to assess the effect of propolis extracts on pathogenic microorganisms, as well as on potential probiotic lactobacilli. It was observed that the maceration in water does not extract the bioactive compounds present in these samples unlike the ethanolic extracts that showed antimicrobial activity against Staphylococcus aureus, including MRSA strain and yeasts. Inhibition of Candida albicans and Saccharomyces boulardii by propolis extracts was comparable to amphotericin B. C. albicans was not inhibited in the absence of total phenolics. Regarding lactobacilli, the agar diffusion method did not correlate well with the minimum inhibitory concentration (MIC) assay. Moreover, MIC values of propolis extracts for Lacticaseibacillus and Limosilactobacillus were higher than for pathogen microorganisms, making propolis attractive to be used as a preservative food additive, including fermented foods.