Pharmaceutical NanotechnologyA pilot study of freeze drying of poly(epsilon-caprolactone) nanocapsules stabilized by poly(vinyl alcohol): Formulation and process optimization
Introduction
Submicronic colloidal vectors have gained a considerable interest in the last few years because of their ability to ensure a specific drug targeting by both the oral route (Ponchel and Irache, 1998) and the parenteral route (Marty et al., 1978, Soppimath et al., 2001).
Such particulate systems have been widely investigated for gene delivery to cells and tissues (Panyam and Labhasetwar, 2003) as in the delivery of anti-sense oligonucleotides (Lambert et al., 2001) and also in cancer therapy and diagnosis (Brigger et al., 2002).
Among these vectors, liposomes and nanoparticles have special advantages with regard to the modulation of an active ingredient distribution within the human body (Soppimath et al., 2001).
Nanoparticles can be classified into nanospheres and nanocapsules. Nanocapsules are vesicular systems in which the drug is confined to an oily or aqueous cavity surrounded by a unique polymeric membrane while nanospheres are matrix systems in which the drug is physically and uniformly dispersed (Soppimath et al., 2001).
The major obstacle that limits the use of such vectors is their instability in an aqueous medium (Chacon et al., 1999). Aggregation and particle fusion are frequently noticed after a long period of storage of such systems (Auvillain et al., 1989). Furthermore, drug leakage out of the nanocapsules and non-enzymatic polymer hydrolysis can happen. Thus, the stabilization of colloidal vectors is deeply explored in order to reach a shelf-life of several years.
Freeze drying, also termed lyophilization, is an industrial process of drying by freezing and sublimation of ice; it is used to convert solutions of labile materials into solids of sufficient stability for distribution and storage (Franks, 1998). This technique was considered as a good method to conserve the integrity of colloidal vectors. In literature, there are many papers that investigate in detail the stabilization by freeze drying of liposomes (Crowe et al., 1986, Crowe et al., 1994, Anchordoguy et al., 1987) and nanospheres (De Chasteigner et al., 1996, Schwarz and Mehnert, 1997, Chacon et al., 1999, Saez et al., 2000;) but few researchers, to the best of our knowledge, studied the lyophilization of nanocapsules which have a very thin and fragile envelope that may not withstand the mechanical stress of freezing (Auvillain et al., 1989, De Chasteigner et al., 1995, Schaffazick et al., 2003, Choi et al., 2002).
Auvillain et al. (1989) found that the freeze drying of polycaprolactone nanocapsules is possible but 30% trehalose as cryoprotectant was necessary to preserve the integrity of nanocapsules. In such a case, rapid freezing has been favourable and the conservation of encapsulated oil in liquid state during the freezing process improved the success of lyophilization.
De Chasteigner et al. (1995) have reported the lyophilization of polylactide nanocapsules with 10% glucose or sucrose but the produced lyophilizates showed a two-fold increase in their size after redispersion in water. The authors explained this result by a clustering of nanocapsules.
Schaffazick et al. (2003) reported the lyophilization of polycaprolactone and eudragit nanocapsules upon the addition of colloidal silicon dioxide, however such an addition makes intravenous administration of the nanocapsules impossible.
Finally, Choi et al. (2002) freeze dried nanocapsules of polycaprolactone with pluronic F68 as a stabilizer without a cryoprotectant. These authors found that the freeze process can break the nanocapsules and promote leakage of their contents. They concluded that the nanocapsules may not have been broken by water crystallization in the external phase but by the solidification of the oil (miglyol) in the internal phase.
The aim of this study is to investigate the factors which can influence the nanocapsules stability during the different steps of lyophilization. Different parameters have been tested throughout the freeze–thawing study including PVA concentration, polymer concentration, cooling rate, cryoprotectant concentration, nature of encapsulated oil and nanocapsules purification. Nanocapsules have been freeze dried both before and after purification. Freeze dried purified nanocapsules were characterized by particle size measurement, collapse temperature and determination, scanning electron microscope observation, environmental scanning electron microscope imaging and residual humidity quantification. Finally, the effect of annealing on the nanocapsules stability and the sublimation rate has been well explored. A successful nanocapsules lyophilization requires a good formulation and optimal conditions of freezing and desiccation. Such process must produce an acceptable non-collapsed cake, which can rehydrate immediately with the conservation of nanocapsules properties.
Section snippets
Materials
Poly(epsilon-caprolactone) (PCL) (Mw: 14,000 g/mol) and hydroxypropyl beta cyclodextrin (HPβCD) were obtained from Sigma–Aldrich (France). Poly(vinyl alcohol) (PVA) Moviol 4-88 (88% hydrolyzed, Mw: 31,000 g/mol) was purchased from Clariant (France). Miglyol 810 and miglyol 829 were supplied from Condea chemie (Germany). Ethyl acetate was obtained from Carlo Erba (France). d(+)Sucrose from prolabo (France). d(+)Anhydrous glucose, d(+)trehalose and d-mannitol from Flucka biochemika (Switzerland).
Influence of cooling rate
Nanocapsules were prepared in this study by the emulsification diffusion method which is capable of preparing nanocapsules in a simple, efficient and reproducible manner. This method was preferred to the other techniques of nanocapsules preparation such as emulsification evaporation of solvent and nanoprecipitation. Emulsification evaporation technique requires working with toxic solvents, whereas nanoprecipitation method produces low yields and poor entrapment efficacy (Quintanar-Guerrero et
Discussion
PCL NC prepared by emulsion–diffusion method produces NC with a mean size of 294–401 nm in a reproducible and an efficient way. In this present study, PVA was used as stabilizer for preparing PCL NC. This polymer is one of the most frequently used stabilizers to produce stable nanoparticles, since it enhances the production of stable particles with a small size and narrow size distribution (Zambaux et al., 1998, Sahoo et al., 2002). Many papers have mentioned that a fraction of PVA used in the
Conclusion
The results of this study demonstrate that polycaprolactone nanocapsules stabilized by PVA and prepared using the emulsion–diffusion method can be freeze dried without a cryoprotectant when the concentration of PVA is sufficient to prevent the NC from aggregation during freezing. However, for purified NC, the addition of cryoprotectant at a concentration of 5% seems necessary. The type of cryoprotectants had practically negligible effects on the size and the rehydration of freeze dried
Acknowledgement
The authors are grateful to Annie Rivoire from “Le Centre Technologique des Microstructures de l’Université Lyon 1” for her technical assistance with the SEM and ESEM imaging.
References (36)
- et al.
Modes of interaction of cryoprotectants with membrane phospholipids during freezing
Cryobiology
(1987) - et al.
Dynamic properties of poly (dl-lactide) and polyvinyl alcohol monolayers at the air/water and dichloromethane/water interfaces
J. Colloid Interface Sci.
(1995) - et al.
Nanoparticles in cancer therapy and diagnosis
Adv. Drug Deliv. Rev.
(2002) - et al.
Stability and freeze drying of cyclosporine loaded poly(d,l lactide-glycolide) carriers
Eur. J. Pharm. Sci.
(1999) - et al.
Prevention of fusion and leakage in freeze-dried liposomes by carbohydrates
Biochim. Biophys. Acta
(1986) - et al.
Is vitrification sufficient to preserve liposomes during freeze-drying?
Cryobiology
(1994) Freeze-drying of bioproducts: putting principles into practice
Eur. J. Pharm. Biopharm.
(1998)- et al.
Nanoparticulate systems for the delivery of antisense oligonucleotides
Adv. Drug Deliv. Rev.
(2001) - et al.
Influence of the degree of hydrolyzation and polymerisation of pol(vinylalcohol) on the preparation and properties of poly(dl-lactide-co-glycolide) nanoparticles
Int. J. Pharm.
(1997) - et al.
preparation of poly(dl-lactide-co-glycolide) nanoparticles by modified spontaneous emulsification solvent diffusion method
Int. J. Pharm.
(1999)
Lyophilization of protein formulations in vials: investigation of the relationship between resistance to vapor flow during primary drying and small-scale product collapse
J. Pharm. Sci.
Biodegradable nanoparticles for drug and gene delivery to cells and tissue
Adv. Drug Deliv. Rev.
The collapse temperature in freeze drying: dependance on measurement methodology and rate of water removal from the glassy state
Int. J. Pharm.
Specific and non-specific bioadhesive particulate systems for oral delivery to the gastrointestinal tract
Adv. Drug Deliv. Rev.
Influence of stabilizing agents and preparative variables on the formation of poly (d,l-lactic acid) nanoparticles by an emulsification–diffusion technique
Int. J. Pharm.
Freeze-drying of polycaprolactone and poly(d,l-lactic-glycolic) nanoparticles induce minor particle size changes affecting the oral pharmacokinetics of loaded drugs
Eur. J. Pharm. Biopharm.
Residual polyvinyl alcohol associated with poly (d,l-lactide-co-glycolide) nanoparticles affects their physical properties and cellular uptake
J. Control. Release
Freeze-drying polymeric colloidal suspensions: nanocapsules, nanospheres and nanodispersion. A comparative study
Eur. J. Pharm. Biopharm.
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