Gamma-irradiation of liposomes composed of saturated phospholipids. Effect of bilayer composition, size, concentration and absorbed dose on chemical degradation and physical destabilization of liposomes

Abstract

Liposomes composed of dipalmitoylphosphatidylcholine (DPPC), dipalmitoylphosphatidylglycerol (DPPG), or mixtures of these two phospholipids were exposed to gamma-irradiation in an air environment. Disappearance of the mother compounds was monitored by HPLC analysis. Plotting of the logarithmic values of residual DPPC or DPPG concentration versus irradiation dose resulted in straight lines. The slopes of these lines (overall degradation constants) depended on the type of phospholipids, concentration of the liposomes and the size of the liposomes. Under the chosen conditions, addition of DPPG in DPPC-liposomes did not affect the degradation rate constant of DPPC and visa versa. The presence of phosphate buffer (pH 7.4), pH or presence of sodium chloride did not affect the irradiation damage either. Minor changes were found upon analysis of total fatty acids by GLC and upon measurement of water soluble phosphate compounds. These changes were less pronounced than the changes monitored by HPLC of phospholipids, because the HPLC analysis monitored the overall degradation of the liposomal phospholipids. Thin-layer chromatography/fast atom bombardment mass spectrometry (TLC/FAB-MS) analysis of irradiated and non-irradiated DPPC or DPPG provided information on the structure of several degradation products. Degradation routes which include these degradation products are proposed. Gamma-irradiation neither affected the size of the liposomes nor the bilayer rigidity as determined by dynamic light scattering and fluorescence anisotropy of the probe 1,6-diphenyl-1,3,5-hexatriene (DPH), respectively. However, upon gamma-irradiation, changes in the melting characteristics of the liposomes were found by differential scanning calorimetry (DSC) measurements. The pre-transition melting enthalpy of the liposomal bilayer decreased or disappeared and the main-transition broadened. The changes found in DSC scans correlated qualitatively well with the changes recorded after HPLC analysis of phospholipids.