Study on the aggregation states of Amphotericin B with Raman spectroscopy and UV-vis spectroscopy

Abstract

Amphotericin B (AmB), the polyene macrolide antifungal agent, formulated as a micellar system was reported as a cause of nephrotoxicity. The aggregation of AmB plays an important role on its nephrotoxicity. It is necessary to understand the aggregation behavior of AmB, because it can help in developing new AmB formulations. Therefore, the aim of this work was to evaluate the molecular aggregation states of AmB at different molar ratios of AmB to sodium deoxycholate by Raman spectroscopy and UV-vis spectroscopy. At low concentrations ranging from 1 x 10-5 M to 1 x 10-7 M, UV-vis spectroscopy could detect the molecular states of AmB as soluble oligomer, and a monomer at maximum wavelength around 325 nm and 408 nm, respectively. However, UV-vis spectroscopy could not be used to study the molecular states of AmB at higher concentrations. On the other hand, Raman spectroscopy could detect remarkable intense peaks relating to molecular structure of AmB at higher concentrations. Heat treatment at 70°C for 20 min caused the slight blue-shifts in UV peak around 325 nm, indicating the superaggregates formation. In the Raman spectra, the heat treatment led to the slight shift of the peak around 1556 cm-1 to lower wavenumber. This result indicated the change in intermolecular bonding between chromophore parts of AmB molecules which was related to its aggregation states. In conclusion, both Raman spectroscopy and UVvis spectroscopy could be used to study the molecular states of AmB in micellar system depending on its concentration.