Abstract
It has been hypothesized that the therapeutic effects of artepillin C, a natural compound derived from Brazilian green propolis, are likely related to its partition in the lipid bilayer component of biological membranes. To test this hypothesis, we investigated the effects of the major compound of green propolis, artepillin C, on model membranes (small and giant unilamelar vesicles) composed of ternary lipid mixtures containing cholesterol, which display liquid-ordered (lo) and liquid-disordered (ld) phase coexistence. Specifically, we explored potential changes in relevant membrane parameters upon addition of artepillin C presenting both neutral and deprotonated states by means of small angle X-ray scattering (SAXS), differential scanning calorimetry (DSC), and confocal and multiphoton excitation fluorescence microscopy. Thermotropic analysis obtained from DSC experiments indicated a loss in the lipid cooperativity of lo phase at equilibrium conditions, while at similar conditions spontaneous formation of unilamellar vesicles from SAXS experiments showed that deprotonated artepillin C preferentially located at the surface of the membrane. Time-resolved experiments using fluorescence microscopy showed that at doses above 100 µM, artepillin C in its neutral state interacted with both liquid-ordered and liquid-disordered phases, inducing curvature stress and promoting dehydration at the membrane interface.
Highlights
Depending on the lateral organization of the lipid bilayer and the nature of the lipid’s polar head group, bioactive compounds can interact with biological membranes and display a range of therapeutic effects by altering their structural and dynamical properties, leading to cell death [1,2,3,4]
In order to evaluate the effect of this compound on lipid membranes showing higher compositional complexity, we examined the interaction of artepillin C on membrane systems composed of mixtures of DPPC, 1,2-dioleoyl-sn-glycero3-phosphocholine (DOPC), and cholesterol suspended in ultra-pure Milli-Q water, which displays coexistence of liquid-ordered and liquid-disordered phases [25,26,27]
The broad thermal transition measured for the lipid suspension in the absence of artepillin C in this temperature range, results mainly from the presence of cholesterol embedded in DPPC enriched domains (Lo domains), resulting in a substantial loss of cooperativity observed for pure DPPC [38]
Summary
Depending on the lateral organization of the lipid bilayer and the nature of the lipid’s polar head group, bioactive compounds can interact with biological membranes and display a range of therapeutic effects by altering their structural and dynamical properties, leading to cell death [1,2,3,4]. Natural compounds found in Brazilian biomes have been a target of several studies due to the variety of species spread across the country and their large spectrum of biological activities Due to their particular chemical structures, these compounds are able to interact directly with lipid bilayers without the presence of specific membrane receptors. Among all the propolis types, Brazilian green propolis, collected by the bee species Apis mellifera, has been extensively investigated worldwide, due to the presence of artepillin C (3,5-diprenyl-4-hydroxycinnamic acid), a phenolic acid derivative compound, which presents antioxidant, anti-inflammatory, and antitumor properties [10,11,12] This molecule has two prenylated groups bound to a phenyl group, which enhances its hydrophobicity (see Supplementary Figure S1)
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