Geographic Area of Collection Determines the Chemical Composition and Antimicrobial Potential of Three Extracts of Chilean Propolis.

The biological properties of chilean propolis have been described and include antibacterial, antifungal and antibiofilm activities. Propolis has a strong antimicrobial potential. Clinical experiences with synthetic antibiotics indicated the need to discover new sources of bioactive compounds associated with ethnopharmacological knowledge or natural sources such as propolis. The microscopic analysis of pollen grains from plants allows us to determine the botanical origin of the propolis samples. In Angol, sample pollen grains were obtained from fodder plants (Sorghum bicolor; Lotus sp.) and trees, such as Acacia sp., Pinus radiata, Eucalyptus sp. and Salix babylonica. Propolis from the Maule region contains pollen grains from endemic plants such as Quillaja saponaria. Finally, the sample obtained from Melipilla presented a wider variety of pollen extracted from vegetable species.Colorimetric assays performed to quantify the total polyphenols present in Chilean propolis samples established that PCP2 (Angol sample) showed high amounts of phenolics compounds, with significant statistical differences in comparison with the other samples. The main compounds identified were pinocembrin, quercetin and caffeic acid phenethyl ester (CAPE). The Angol sample showed a high content of polyphenols.Studies that determine the influence of geographical and floral variables on the chemical composition of propolis are a valuable source of information for the study of its biological properties.

Evaluation of Alternative Methods to Assess the Biological Properties of Propolis on Metabolic Activity and Biofilm Formation in Streptococcus mutans.

Several biological activities have been reported for the Chilean propolis, among their antimicrobial and antibiofilm properties, due to its high polyphenol content. In this study, we evaluate alternative methods to assess the effect of Chilean propolis on biofilm formation and metabolic activity of Streptococcus mutans (S. mutans), a major cariogenic agent in oral cavity. Biofilm formation was studied by using crystal violet and by confocal microscopy. The metabolic activity of biofilm was evaluated by MTT and by flow cytometry analysis. The results show that propolis reduces biofilm formation and biofilm metabolic activity in S. mutans. When the variability of the methods to measure biofilm formation was compared, the coefficient of variation (CV) fluctuated between 12.8 and 23.1% when using crystal violet methodology. On the other hand, the CV ranged between 2.2 and 3.3% with confocal microscopy analysis. The CV for biofilm's metabolic activity measured by MTT methodology ranged between 5.0 and 11.6%, in comparison with 1.9 to 3.2% when flow cytometry analysis was used. Besides, it is possible to conclude that the methods based on colored compounds presented lower precision to study the effect of propolis on biofilm properties. Therefore, we recommend the use of flow cytometry and confocal microscopy in S. mutans biofilm analysis.

Antimicrobial and Antibiofilm Activity against Streptococcus mutans of Individual and Mixtures of the Main Polyphenolic Compounds Found in Chilean Propolis.

Dental caries is multifactorial disease and an important health problem worldwide. Streptococcus mutans is considered as a major cariogenic agent in oral cavity. This bacteria can synthetize soluble and insoluble glucans from sucrose by glucosyltransferases enzymes and generate stable biofilm on the tooth surface. Biological properties of Chilean propolis have been described and it includes antimicrobial, antifungal, and antibiofilm activities. The main goal of this study was to quantify the concentrations of main flavonoids presents in Chilean propolis and compare some biological properties such as antimicrobial and antibiofilm activity of individual compounds and the mixture of this compounds, against S. mutans cultures. Chilean propolis was studied and some polyphenols present in this extract were quantified by HPLC-DAD using commercial standards of apigenin, quercetin, pinocembrin, and caffeic acid phenethyl ester (CAPE). MIC for antimicrobial activity was determined by serial dilution method and biofilm thickness on S. mutans was quantified by confocal microscopy. Pinocembrin, apigenin, quercetin, and caffeic acid phenethyl ester (CAPE) are the most abundant compounds in Chilean propolis. These polyphenols have strong antimicrobial and antibiofilm potential at low concentrations. However, pinocembrin and apigenin have a greater contribution to this action. The effect of polyphenols on S. mutans is produced by a combination of mechanisms to decrease bacterial growth and affect biofilm proliferation due to changes in their architecture.

Polyphenol-Rich Extract from Propolis Reduces the Expression and Activity of Streptococcus mutans Glucosyltransferases at Subinhibitory Concentrations.

Tooth decay is an infectious disease, whose main causative agent identified is Streptococcus mutans (S. mutans). Diverse treatments have been used to eradicate this microorganism, including propolis. To date, it has been shown that polyphenols from Chilean propolis inhibit S. mutans growth and biofilm formation. However, the molecular mechanisms underlying this process are unclear. In the present study, we assessed the effect of Chilean propolis on the expression and activity of the glycosyltransferases enzymes and their related genes. Polyphenol-rich extract from propolis inhibited gene expression of glycosyltransferases (GtfB, GtfC, and GtfD) and their related regulatory genes, for example, VicK, VicR, and CcpA. Moreover, the treatment inhibited glucosyltransferases activity measured by the formation of sucrose-derived glucans. Additionally, an inhibitory effect was observed in the expression of SpaP involved in sucrose-independent virulence of S. mutans. In summary, our results suggest that Chilean propolis has a dose-dependent effect on the inhibition of genes involved in S. mutans virulence and adherence through the inhibition of glucosyltransferases, showing an anticariogenic potential of polyphenols from propolis beyond S. mutans growth inhibition.

Antibiofilm Activity of Chilean Propolis on Streptococcus mutans Is Influenced by the Year of Collection.

The chemical composition of propolis varies according to factors that could have an influence on its biological properties. Polyphenols from propolis have demonstrated an inhibitory effect on Streptococcus mutans growth. However, it is not known if different years of propolis collection may affect its activity. We aimed to elucidate if the year of collection of propolis influences its activity on Streptococcus mutans. Polyphenol-rich extracts were prepared from propolis collected in three different years, characterized by LC-MS and quantified the content of total polyphenols and flavonoids groups. Finally, was evaluated the antibacterial effect on Streptococcus mutans and the biofilm formation. Qualitative differences were observed in total polyphenols, flavones, and flavonols and the chemical composition between the extracts, affecting the strength of inhibition of biofilm formation but not the antimicrobial assays. In conclusion, chemical composition of propolis depends on the year of collection and influences the strength of the inhibition of biofilm formation.

Chemical and botanical characterization of Chilean propolis and biological activity on cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus.

Propolis is a non-toxic natural substance with multiple pharmacological properties including anti-cancer, antioxidant, fungicidal, antibacterial, antiviral, and anti-inflammatory among others. The aim of this study was to determine the chemical and botanical characterization of Chilean propolis samples and to evaluate their biological activity against the cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus. Twenty propolis samples were obtained from beekeeping producers from the central and southern regions of Chile. The botanical profile was determined by palynological analysis. Total phenolic contents were determined using colorimetric assays. Reverse phase HPLC and HPLC-MS were used to determine the chemical composition. The minimum inhibitory concentration (MIC) was determined on S. mutans and S. sobrinus. All propolis samples were dominated by structures from native plant species. The characterization by HPLC/MS, evidenced the presence of quercetin, myricetin, kaempferol, rutine, pinocembrin, coumaric acid, caffeic acid and caffeic acid phenethyl ester, that have already been described in these propolis with conventional HPLC. Although all propolis samples inhibited the mutans streptococci growth, it was observed a wide spectrum of action (MIC 0.90 to 8.22 µg mL(-1)). Given that results it becomes increasingly evident the need of standardization procedures, where we combine both the determination of botanical and the chemical characterization of the extracts. Research conducted to date, describes a promising effectiveness of propolis in the prevention of caries and other diseases of the oral cavity, making it necessary to develop studies to identify and understand the therapeutic targets or mechanisms of molecular action of the various compounds present on them.

Chemical and botanical characterization of Chilean propolis and biological activity on cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus

Propolis is a non-toxic natural substance with multiple pharmacological properties including anticancer, antioxidant, fungicidal, antibacterial, antiviral, and anti-inflammatory among others. The aim of this study was to determine the chemical and botanical characterization of Chilean propolis samples and to evaluate their biological activity against the cariogenic bacteria Streptococcus mutans and Streptococcus sobrinus. Twenty propolis samples were obtained from beekeeping producers from the central and southern regions of Chile. The botanical profile was determined by palynological analysis. Total phenolic contents were determined using colorimetric assays. Reverse phase HPLC and HPLC-MS were used to determine the chemical composition. The minimum inhibitory concentration (MIC) was determined on S. mutans and S. sobrinus. All propolis samples were dominated by structures from native plant species. The characterization by HPLC/MS, evidenced the presence of quercetin, myricetin, kaempferol, rutine, pinocembrin, coumaric acid, caffeic acid and caffeic acid phenethyl ester, that have already been described in these propolis with conventional HPLC. Although all propolis samples inhibited the mutans streptococci growth, it was observed a wide spectrum of action (MIC 0.90 to 8.22 µgmL-1). Given that results it becomes increasingly evident the need of standardization procedures, where we combine both the determination of botanical and the chemical characterization of the extracts. Research conducted to date, describes a promising effectiveness of propolis in the prevention of caries and other diseases of the oral cavity, making it necessary to develop studies to identify and understand the therapeutic targets or mechanisms of molecular action of the various compounds present on them.