Prenylated (iso)flavonoids as antimicrobial agents : production, activity and mode of action

Abstract

Prenylated flavonoids and isoflavonoids (i.e. (iso)flavonoids) are typically produced in the legume (Fabaceae) family as part of plant's defence mechanism. Prenylation, i.e. the substitution with a C5-isoprenoid moiety, is acknowledged for conferring the antimicrobial properties of prenylated (iso)flavonoids. The first aim of this thesis was to enhance the amounts and diversity of prenylated (iso)flavonoids in planta using the well-studied legume, soybean. Second, the antimicrobial properties of prenylated (iso)flavonoids against the Gram-positive pathogen, methicillin-resistant Staphylococcus aureus (MRSA) and the food spoilage yeast, Zygosaccharomyces parabailii were explored. Lastly, insights into the mode of action of antimicrobial prenylated (iso)flavonoids were sought through in silico and in vitro means.Sensitization of soybean seedlings with reactive oxygen species (ROS), generated through H2O2 and FeSO4, prior to microbial elicitation enhanced the production of prenylated isoflavonoids (glyceollins), compared to the traditional fungal elicitation without priming. Application of the most stable ROS, H2O2, together with AgNO3, prior to microbial elicitation enhanced the production of prenylated isoflavones and a coumestan with promising antibacterial activity, in addition to glyceollins. A systematic study on 106 prenylated (iso)flavonoids tested against MRSA showed that 75% of the diprenylated (iso)flavonoids and 40% of the mono-prenylated ones were active with minimum inhibitory concentrations (MIC) ≤ 25 μg/mL. Formal and partial charge, hydrophobic volume and uneven distribution of hydrophobic moieties were correlated to antimicrobial activity. The different molecular properties suggested potentially distinct interactions of the different prenylated (iso)flavonoid subclasses with the microbial membrane. Two mono-prenylated isoflavonoids from different subclasses were found to be very active towards the food spoiler and weak organic acid-resistant Z. parabailii (MIC ≤ 12.5 μg/mL, pH 4.0). Killing of Z. parabailii was evidenced within 15 min of exposure to these compounds, accompanied with different deformations of the yeast membrane. Prenylated (iso)flavonoids can be powerful antimicrobials to be used as therapeutics or food preservatives, when their toxicity profile against human cells has been verified. The quantitative models developed provide useful information on the design of new antimicrobial prenylated (iso)flavonoids. Table of contents Chapter 1General Introduction Chapter 2 Enhanced biosynthesis of the natural antimicrobial glyceollins in soybean seedlings by priming and elicitation Chapter 3 Induction of promising antibacterial prenylated isoflavonoids from different subclasses by sequential elicitation of soybean Chapter 4 Prenylated (iso)flavonoids as agents against methicillin-resistant Staphylococcus aureus -Insights into the molecular properties underlying antibacterial activity Chapter 5 Prenylated (iso)flavonoids as antifungal agents against the food spoiler Zygosaccharomyces parabailii