Preparation and Characterization of Freeze-dried Liposomes Loaded with Amphotericin B

Abstract

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. </P><P> 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. </P><P> 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. </P><P> 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. </P><P> 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.

Preparation, Characterization, and In Vivo Pharmacokinetic Study of the Supercritical Fluid-Processed Liposomal Amphotericin B

“…It has been reported that the liposomes containing drug molecules can be lyophilized and reconstituted with significant drug retention, which is measured as the percent of the encapsulated drug and without a significant change in the mean vesicle size. Cryoprotectants are needed during lyophilization, and sugars, such as sucrose, lactose, and trehalose, are used to protect the liposomes during the freezing stage of the lyophilization cycle [39,40]. Lyophilization with 9% lactose as a cryoprotectant provided physical stability to the liposomes as well as high protection against liposomal drug leakage.…”

A Comprehensive Review on Novel Liposomal Methodologies, Commercial Formulations, Clinical Trials and Patents

“…This method involves dissolving of lipid and drug in tert-butyl alcohol at 45 0 C, and lyo-protectant dissolves in water at 45 0 C. Both of the obtained solutions are mixed to form third identical monophasic solution, followed by filtration, and freeze drying of solution is performed to produce proliposomes. Freeze drying involves 2 steps: in first, the sample is frozen at − 40ºC and then dried at room temperature which upon hydration eventually forms liposomes with mean dimension of 100-300 nm [76]. Advantages…”