Spectroscopic studies of amphotericin B solubilized in nanoscale bilayer membranes

• Published in: Biochimica et biophysica acta Vol. 1758; no. 1; pp. 38 - 44
• Main Authors: Hargreaves, Peter L., Nguyen, Thanh-Son, Ryan, Robert O.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 2006
• Subjects: Amphotericin B; Amphotericin B - chemistry; Amphotericin B - pharmacology; Antifungal Agents - chemistry; Antifungal Agents - pharmacology; Apolipoprotein; Cholesterol - metabolism; Cholesterol - pharmacology; Dose-Response Relationship, Drug; Drug Carriers - chemistry; Ergosterol - metabolism; Ergosterol - pharmacology; Kinetics; Lipid Bilayers - chemistry; Nanodisk; Nanostructures - chemistry; Phospholipid; Saccharomyces cerevisiae - drug effects; Spectrophotometry, Ultraviolet - methods; Spectroscopy; Temperature; DMPG; PBS; CD; apo; Spectroscopy; DMSO; Phospholipid; Nanodisk; PAGE; Apolipoprotein; Amphotericin B; AMB; ND; DMPC
• ISSN: 0005-2736; 0006-3002; 1879-2642; 1878-2434
• DOI: 10.1016/j.bbamem.2006.01.001

Nanodisks (ND) are discrete nanometer scale phospholipid bilayers whose perimeter is circumscribed by amphipathic apolipoproteins. The membranous environment of ND serves as a matrix for solubilizing the polyene antibiotic amphotericin B (AMB). The spectral properties of AMB in ND are dependent upon AMB concentration. Whereas AMB-ND prepared at a concentration of 2.5 mg AMB per 10 mg phospholipid are consistent with AMB self association in the ND membrane environment, AMB-ND prepared at 0.25 or 0.025 mg AMB per 10 mg phospholipid give rise to spectra reminiscent of AMB in organic solvent. Incubation of ND prepared at a phospholipid/AMB ratio of 400:1 (w/w) at 37 °C for 1 h induced a shift in absorbance and near UV circular dichroism spectra consistent with antibiotic self-association. The kinetics of this spectral transition were investigated as a function of incubation temperature. While no change in A 388 nm occurred in incubations at 20 °C, a time-dependent decrease in A 388 nm was observed at 25, 30 and 37 °C. Inclusion of ergosterol in the ND membrane attenuated temperature-induced AMB spectral changes. In Saccharomyces cerevisiae growth inhibition assays, ND containing self associated AMB were somewhat less effective than ND possessing a greater proportion of monomeric AMB. On the other hand, inclusion of ergosterol or cholesterol in the ND particle did not alter the growth inhibition properties of AMB-ND. The miniature membrane environment of ND provides a novel milieu for solubilization and characterization of lipophilic biomolecules.