Generic placeholder image

Current Drug Therapy


ISSN (Print): 1574-8855
ISSN (Online): 2212-3903

Research Article

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

Author(s): Tran Thi Hai Yen, Le Nho Dan, Le Hoang Duc, Bui Thanh Tung and Pham Thi Minh Hue*

Volume 14, Issue 1, 2019

Page: [65 - 73] Pages: 9

DOI: 10.2174/1574885514666181217130259

Price: $65


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.

Keywords: Liposomes, amphotericin B, freeze-drying, sucrose, trehalose, cryoprotectants.

Graphical Abstract
Adler-Moore JP, Proffitt RT. Amphotericin B lipid preparations: what are the differences? Clin Microbiol Infect 2008; 14: 25-36.
Torchilin VP. Recent advances with liposomes as pharmaceutical carriers. Nat Rev Drug Discov 2005; 4: 145-60.
Levacheva I, Samsonova O, Tazina E, et al. Optimized thermosensitive liposomes for selective doxorubicin delivery: Formulation development, quality analysis and bioactivity proof. Colloids Surf B Biointerfaces 2014; 121: 248-56.
Yen TTH, Meerovich IG, Meerovich GA, et al. Comparative studies in vivo of free and liposomal forms of photosensitizer on a base of hydrophilic derivative of chlorin e6. J Drug Deliv Sci Technol 2012; 22: 291-4.
Slingerland M, Guchelaar HJ, Rosing H, et al. Bioequivalence of liposome-entrapped paclitaxel easy-to-use (lep-etu) formulation and paclitaxel in polyethoxylated castor oil: A randomized, two-period crossover study in patients with advanced cancer. Clin Ther 2013; 35: 1946-54.
Proffitt Richard T, Adler-More J, Chiang S-M. Amphotericin B liposome preparation. US5965156, US, 1999.
Robert Abra; Francis C. Szoka. Stabilized liposome amphotericn B. US4766046, 1988.
Moribe K, Maruyama K, Iwatsuru M. Encapsulation characteristics of nystatin in liposomes: effects of cholesterol and polyethylene glycol derivatives. Int J Pharm 1999; 188: 193-202.
Ueda S, Miyamoto S, Kaida K, et al. Safety and efficacy of treatment with liposomal amphotericin B in elderly patients at least 65 years old with hematological diseases. J Infect Chemother 2016; 22: 287-91.
Hamill RJ. Amphotericin B formulations: A comparative review of efficacy and toxicity. Drugs 2013; 73: 919-34.
Stark B, Pabst G, Prassl R. Long-term stability of sterically stabilized liposomes by freezing and freeze-drying: Effects of cryoprotectants on structure. Eur J Pharm Sci 2010; 41: 546-55.
Torchilin VPC, Torchilin VPC, Torchilin VPC. Liposomes : A practical approach. Oxford University Press 2003.
Hua ZZ, Li BG, Liu ZJ, et al. Freeze-Drying of liposomes with cryoprotectants and its effect on retention rate of encapsulated ftorafur and vitamin A. Dry Technol 2003; 21: 1491-505.
Van Winden ECA, Crommelin DJA. Long term stability of freeze-dried, lyoprotected doxorubicin liposomes. Eur J Pharm Biopharm 1997; 43(3): 295-307.
Bangham AD, Standish MM, Watkins JC. Diffusion of univalent ions across the lamellae of swollen phospholipids. J Mol Biol 1965; 13: 238-52.
Hamilton-Miller JM. The effect of pH and of temperature on the stability and bioactivity of nystatin and amphotericin B. J Pharm Pharmacol 1973; 25: 401-7.
Crowe JH, Crowe LM, Carpenter JF, et al. Interactions of sugars with membranes. Biochim Biophys Acta 1988; 947: 367-84.
Crowe JH, Crowe LM, Carpenter JF, et al. Stabilization of dry phospholipid bilayers and proteins by sugars. Biochem J 1987; 242: 1-10.
Crowe JH, Crowe LM, Jackson SA. Preservation of structural and functional activity in lyophilized sarcoplasmic reticulum. Arch Biochem Biophys 1983; 220: 477-84.
Crowe LM, Crowe JH, Rudolph A, et al. Preservation of freeze-dried liposomes by trehalose. Arch Biochem Biophys 1985; 242: 240-7.
Crowe JH, Carpenter JF, Crowe LM. The role of vitrification in anhydrobioisis. Annu Rev Physiol 1998; 60: 73-103.
Chen C, Han D, Cai C, et al. An overview of liposome lyophilization and its future potential. J Control Release 2010; 142: 299-311.

Rights & Permissions Print Export Cite as
© 2023 Bentham Science Publishers | Privacy Policy