Nanodisks protect amphotericin B from ultraviolet light and oxidation-induced damage.

BACKGROUND: Macrolide polyene antibiotics possess potent broad-spectrum antifungal properties. Use of these agents in the field or in controlled environments is impeded by their poor water solubility and susceptibility to oxidation- and/or light-induced degradation. While typically used for human disease therapy, there is potential to expand the utility of polyene macrolide antibiotics, such as amphotericin B, for control of fungal disease infestation in agricultural settings. Thus, the susceptibility of this antibiotic to exposure-induced activity loss was evaluated. RESULTS: Incubation of the prototype polyene amphotericin B (AMB) with phospholipid vesicles and apolipoprotein A-I results in the formation of nanoscale complexes, termed nanodisks (NDs), capable of solubilizing significant quantities of AMB. To evaluate whether AMB incorporation into NDs conferred protection against light- or oxidation-induced damage, yeast growth inhibition assays were conducted. Compared with AMB solubilized in detergent micelles, AMB incorporated into NDs was protected from damage caused by exposure to UV light as well as by KMnO(4)-induced oxidation. Furthermore, AMB-NDs inhibited growth of the turfgrass fungus Marasmius oreades Fr. CONCLUSION: Results suggest that this water-soluble formulation of a natural, biodegradable, antifungal agent represents a potential cost-effective, non-toxic and environmentally friendly substitute for chemical agents currently employed to control a range of fungal infestations.

Nanodisks derived from amphotericin B lipid complex.

The goal of this study was to determine the effect of apolipoproteins on Amphotericin B lipid complex (ABLC). We report that incubation of ABLC with recombinant human apolipoprotein A-I (apoA-I) induces solubilization of ABLC by transforming the micron sized phospholipid/AMB assemblies into discrete nanoscale disk-shaped complexes termed nanodisks (ND). ApoA-I induced changes in ABLC solubility and morphology were monitored by spectroscopy and electron microscopy. AMB efficacy was evaluated in yeast and pathogenic fungi growth inhibition assays and the effect of AMB formulation on cell toxicity was assessed in cultured Hep3B cells. AMB associated with ND were more efficiently nebulized than AMB associated with ABLC. Thus, transformation of ABLC into ND preserves the potent biological activity of AMB as well as the reduced toxicity of the ABLC formulation. ABLC derived AMB-ND offer advantages over conventional ABLC in terms of stability, storage, nebulization efficiency and provides an intrinsic "handle" for tissue specific targeting via genetic engineering of its protein component.

Peptide stabilized amphotericin B nanodisks.

Nanometer scale apolipoprotein A-I stabilized phospholipid disk complexes (nanodisks; ND) have been formulated with the polyene antibiotic amphotericin B (AMB). The present studies were designed to evaluate if a peptide can substitute for the function of the apolipoprotein component of ND with respect to particle formation and stability. An 18-residue synthetic amphipathic alpha-helical peptide, termed 4F (Ac-D-W-F-K-A-F-Y-D-K-V-A-E-K-F-K-E-A-F-NH(2)), solubilized vesicles comprised of egg phosphatidylcholine (egg PC), dipentadecanoyl PC or dimyristoylphosphatidylcholine (DMPC) at rates greater than or equal to solubilization rates observed with human apolipoprotein A-I (apoA-I; 243 amino acids). Characterization studies revealed that interaction with DMPC induced a near doubling of 4F tryptophan fluorescence emission quantum yield (excitation 280 nm) and a approximately 7 nm blue shift in emission wavelength maximum. Inclusion of AMB in the vesicle substrate resulted in formation of 4F AMB-ND. Spectra of AMB containing particles revealed the antibiotic is a highly effective quencher of 4F tryptophan fluorescence emission, giving rise to a Ksv=7.7 x 10(4). Negative stain electron microscopy revealed that AMB-ND prepared with 4F possessed a disk shaped morphology similar to ND prepared without AMB or prepared with apoA-I. In yeast and pathogenic fungi growth inhibition assays, 4F AMB-ND was as effective as apoA-I AMB-ND. The data indicate that AMB-ND generated using an amphipathic peptide in lieu of apoA-I form a discrete population of particles that possess potent biological activity. Given their intrinsic versatility, peptides may be preferred for scale up and clinical application of AMB-ND.