Background: Amphotericin B (AmB) is a drug of choice in the therapy of systemic
fungal infection because of its board-spectrum antifungal activity. However, its
conventional formulation has many side effects such as acute and chronic nephrotoxicity.
Liposomes have been developed to reduce the drug’s toxicity. However, they had to meet
strict stability criteria. In general, liposomes can be freeze-dried to inhibit liposomes infusion,
phospholipids degradation during storage. Liposomal size usually increases after
freeze-drying because of being influenced by many factors in freezing, lyophilizing and
rehydration processes. Therefore, cryoprotectants are used to stabilize liposomal vesicles
during freeze-drying process.
Objective: In the present study, we developed AmB liposomal suspension and lyophilized
liposomes loaded with AmB, evaluated the effect of different cryoprotectants on the characterization
of freeze-dried AmB liposomes.
Methods: In this study, AmB liposomes were prepared from hydrogenated soy phosphatidylcholine,
distearoylphosphatidylglycerol and cholesterol by thin lipid film hydration
method using different hydrate mediums likely: Glucose solution, citrate buffer,
phosphate buffer. High-pressure homogenization and extrusion methods were used to
reducing vesicles size. Dynamic light scattering was used to characterize liposomal size,
and size distribution. HPLC method was used to assay drug and determine entrapment
efficiency. Liposomal suspension was lyophilized with different cryoprotectants: Sucrose,
mannitol, lactose, trehalose and glycerol. Differential scanning calorimetry was used to
study lyophilized cake.
Results: We found that liposomal suspension with hydration medium10 mM citrate buffer
pH 5.5 had a small average size (<100nm) and narrow distribution (PDI <0.2). Sucrose
and trehalose stabilized vesicles size during freezing process, and lyophilized liposomes
with sucrose and trehalose had an elegant appearance, yellow, compact cake. DSC study
showed that sucrose and trehalose in lyophilized cake were amorphous. The cake was
rehydrated within 10 seconds to form liposomal suspension, in which vesicles size was
less than 140 nm.
Conclusion: We have developed successfully AmB liposomal suspension and lyophilized
liposomes loaded with AmB. Sucrose and trehalose can be used as cryoprotectants in the
freeze-drying and reconstitution process.