Physico-chemical characterisation of cationic DOTAP liposomes as drug delivery system for a hydrophilic decapeptide before and after freeze-drying (original) (raw)
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Freeze-Dried Liposome Formulation for Small Molecules, Nucleic Acid, and Protein Delivery
Systematic Reviews in Pharmacy, 2020
Liposomes are one of the means in the drug delivery system that is quite promising because of its ability to carry active ingredients, both hydrophilic and lipophilic. In addition, the structure of lipid bilayers that resemble cell bilayer membranes is useful in optimizing liposomes' ability through increased internalization of active ingredients into cells. However, liposomes themselves are physically and chemically vulnerable systems, so increasing liposome stability is one of the primary considerations in liposome formulations. Lyophilization or freeze-drying can increase the shelf-life of liposomes, but the process itself is very extreme and prone to triggering damage to vesicle structures. Lyoprotectans from sugars and polyols are usually employed to improve the formulations' stability during the drying process. Physical characterization of the liposome end product is done by looking at the system's morphological appearance, crystallinity profile, thermal analysis, ...
Trehalose preserves DDA/TDB liposomes and their adjuvant effect during freeze-drying
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2007
Disaccharides are well-known reagents to protect biostructures like proteins and phospholipid-based liposomes during freezing and drying. We have investigated the ability of the two disaccharides trehalose and sucrose to stabilize a novel, non-phospholipid-based liposomal adjuvant composed of the cationic dimethyldioctadecylammonium (DDA) and trehalose 6,6′-dibehenate (TDB) upon freeze-drying. The liposomes were freeze-dried using a human dose concentration containing 2.5 mg/ml DDA and 0.5 mg/ml TDB with varying concentrations of the two sugars. The influence on particle size upon rehydration was investigated using photon correlation spectroscopy (PCS) and the gel to fluid phase transition was examined by differential scanning calorimetry (DSC). Data revealed that concentrations above 211 mM trehalose protected and preserved DDA/ TDB during freeze-drying, and the liposomes were readily rehydrated. Sucrose was less efficient as a stabilizer and had to be used in concentrations above 396 mM in order to obtain the same effect. Immunization of mice with the tuberculosis vaccine candidate Ag85B-ESAT-6 in combination with the trehalose stabilized adjuvant showed that freeze-dried DDA/TDB liposomes retained their ability to stimulate both a strong cell-mediated immune response and an antibody response. These findings show that trehalose at isotonic concentrations protects cationic DDA/ TDB-liposomes during freeze-drying. Since this is not the case for liposomes based on DDA solely, we suggest that the protection is facilitated via direct interaction with the headgroup of TDB and a kosmotropic effect, whereas direct interaction with DDA plays a minor role.
A Comprehensive Production Method of Self-cryoprotected Nano-Liposome Powders
International journal of pharmaceutics, 2015
This study provided a convenient approach for large scale production of hydrogenated soya phosphatidylcholine nano-liposome powders using beclometasone dipropionate as model drug and sucrose as proliposome carrier. Fluid-bed coating was employed to manufacture proliposomes by coating sucrose with the phospholipid (5%, 10%, 15% and 20% weight gains), followed by hydration, size reduction using high pressure homogenization, and freeze-drying to yield stable nano-vesicles. High pressure homogenization was compared with probe-sonication in terms of liposome size, zeta potential and drug entrapment. Furthermore, the effect of freeze-drying on vesicle properties generated using both size reduction methods was evaluated. Results have shown that high-pressure homogenization followed by freeze-drying and rehydration tended to yield liposomes smaller than the corresponding vesicles downsized via probe-sonication, and all size measurements were in the range of 72.64-152.50nm, indicating that f...
Journal of Controlled Release, 1994
Liposomes were prepared from natural (EPC) and hydrogenated (HEPC) egg phosphatidylcholine, with and without cholesterol (CHOL), from sucrose fatty acid ester (SPS7; sucrose-palmitate/stearate) with CHOL and dicetylphosphate (DCP) or from EPC and HEPC with the mono-, di-and tri-ester of SPS7. The cryoprotective activity of sucrose or membrane-bound sucrose fatty esters was assessed. Vesicles were frozen and thawed, or freeze-dried and reconstituted, and retention of the encapsulated marker 5,6-carboxyfluorescein (CF) was monitored. CF retention decreased with decreasing freezing temperature, while increasing concentrations of sucrose provided increasing cryoprotection during freezing and thawing. SPS7 vesicles were fully protected by 0.6 M sucrose, whereas equimolar mixtures of EPC and HEPC with SPS7 required 1 M sucrose for complete protection. EPC/CHOL liposomes retained maximally 85% and HEPC/CHOL liposomes 95% marker at the highest sucrose concentration. Lyophilized liposomes without sucrose or in mixture with the SPS mono-or diester retained < 10% CF. Lyophilization of EPC and HEPC liposomes in the presence of 0.4 M sucrose resulted in 75% retention of originally encapsulated marker. Differential scanning calorimetry showed a significant reduction of the transition temperature (T c) of lyophilized HEPC liposomes in the presence of sucrose and the SPS monoester. Infrared spectroscopy indicated sucrose and the SPS monoester forming strong hydrogen bonds with phosphate head groups which supports the water replacement or 'pseudohydration' hypothesis.
Background: The present study aims to design formulation of liposomes that are well-preserved during freeze-drying. The combination of Hydroxy Propyl Methyl Cellulose (HPMC) as dispersion matrix and lyoprotectants; maltodextrin or mannitol, was employed to prevent aggregation and/or recrystallization. The obtained dry products were investigated in terms of their physical characteristics. Methods: Liposomes were prepared using thin film method and hydrated with the lyoprotectant solution. The formed liposomes were mixed with HPMC gel and freeze-dried. The obtained solid products were characterized using Differential Scanning Calorimetry (DSC), X-Ray Diffraction (XRD), and Scanning Electron Microscopy (SEM). Results: The DSC thermograms of formulations with maltodextrin were relatively homogenous, yet exhibiting meta-stable properties. In contrast, the formulations using mannitol showed phase separation. These results were confirmed by XRD data, in which formulations with maltodextrin showed no intensive peaks, indicating amorphous solid while the formulations with mannitol exhibited more intensive peaks, indicating the presence of crystalline solids. The SEM images of both maltodextrin and mannitol-containing formulations showed porous matrix with spherical liposomes trapped in the matrices. The SEM images also correspond to the DSC and XRD data, wherecrystalline solid existed in the mannitol-containing formula. Conclusion: The developed liposomes formulation using combination of HPMC matrix and maltodextrin showed potential in preserving liposomes structure, contrary to those of using mannitol.
Lyophilization of Liposomal Formulations: Still Necessary, Still Challenging
Pharmaceutics
Nowadays, the freeze-drying of liposome dispersions is still necessary to provide a solid dosage form intended for different routes of administration (i.e., parenteral, oral, nasal and/or pulmonary). However, after decades of studies the optimization of process conditions remains still challenging since the freezing and the dehydration destabilize the vesicle organization with the concomitant drug leakage. Starting from the thermal properties of phospholipids, this work reviews the main formulation and process parameters which can guarantee a product with suitable characteristics and increase the efficiency of the manufacturing process. In particular, an overview of the cryo- and/or lyo-protective mechanisms of several excipients and the possible use of co-solvent mixtures is provided. Attention is also focused on the imaging methods recently proposed to characterize the appearance of freeze-dried products and liposome dispersions upon reconstitution. The combination of such data wo...
International Journal.of Pharm Tech Research, 2017
The present study describes the effect of saccharides and hydroxypropyl methylcellulose (HPMC) matrix on phase behavior of dehydrated cationic dimethyl-dioctadecylammonium (DDA)-based liposomes. Saccharides such as sucrose, lactose and mannitol,have been reportedpreserve the lipid membranes during drying, whilst HPMC matrix is widely used in solid dispersion to prevent aggregation and/or recrystallization.The study revealed that addition of sucrose and HPMCin the formulation demonstrated a miscible mixture that might construct a stable dried liposomal formulation. DTA data showed that sucrose (5%w/v) and HPMC added to DDA liposomal formulation were relatively more miscible with the mixtures; whereas lactose and mannitol at the same concentration of 5% showed phase separation from the mixtures in the dehydrated state. Furthermore, XRD and SEM analysis exhibited supporting evidences in which formulation using sucrose and lactose showedrelatively less crystalline-forming properties compared to formulation using mannitol. Recrystallization that cause phase separation might trigger leakage and further affect the efficacy of the entrapped drug/antigen. From these data, it might be concluded that a dried-liposomal formulation can be prepared in the presence of sucrose (lyoprotectant) that is dispersed in HPMC matrix. The protective mechanism of sucrose (5%w/v) and HPMC matrix is proposed through inhibition of the recrystallization which causes phase separation; indicated by DTA, SEM and XRDdata. The present study revealed prospective advantages of using sucrose and HPMC in development of dried – DDA liposomal formulations.
International Journal of Pharmaceutics - INT J PHARM, 2008
The purpose of this study was to determine optimal lipid concentration range for lyophilization of sterically stabilized phospholipid nanomicelles (SSM) and the freeze drying feasibility of selfassociated therapeutic peptide-SSM assemblies. SSM at 5 to 20 mM 1,2-distearoyl-sn-glycero-3phosphoethanolamine-N-methoxy-poly(ethylene glycol 2000 (DSPE-PEG 2000) were analyzed for particle size and viscosity before and after freeze drying which showed no significant changes (p>0.05). However, a steep increase in viscosity was seen for SSM above 15 mM phospholipid implying micelle-micelle interaction. Greater shrinkage of lyophilized cakes was observed below 10 mM phospholipid while they were more fibrous above 15 mM. Therefore, 10-15 mM DSPE-PEG 2000 was chosen as the optimal phospholipid concentration for lyophilized SSM. When vasoactive intestinal peptide (VIP), glucagon-like peptide 1 (GLP-1) or gastric inhibitory peptide (GIP) (each, 67 μM) was added to SSM (10 mM), formulations showed no significant change in particle size, peptide fluorescence and peptide α-helicity before and after lyophilization. In conclusion, we found that peptide drug-SSM interactions are conserved during lyophilization.
Freeze-drying of ATP entrapped in cationic, low lipid liposomes
Cryobiology, 2010
Concerning the instability of ATP liposomes formulated to easily diffuse through the liver (size 100nm),thisworktargetsthekeyparametersthatinfluencethefreeze−dryingofapreparationthatcombinescholesterol,DOTAPandphosphatidylcholine(eithernaturalsoybeanoregg(SPCorEPC)orhydrogenated(HSPC)).Afterfreeze−dryingblankliposomes,sizeincreasedsignificantlywheninitiallipidconcentrationwasloweredfrom20to5mM(p=0.0018).Withlowlipidconcentrationpreparation(5mM),SPClimitedsizeincrease(SI)moreefficientlycomparedtoEPCorHSPC.WithSPCandEPC,sucroseshowedbettersizeresultscomparedtotrehalose(Lyoprotectant/Lipidratio(w/w)avoidinganySI:100 nm), this work targets the key parameters that influence the freeze-drying of a preparation that combines cholesterol, DOTAP and phosphatidylcholine (either natural soybean or egg (SPC or EPC) or hydrogenated (HSPC)). After freeze-drying blank liposomes, size increased significantly when initial lipid concentration was lowered from 20 to 5 mM (p = 0.0018). With low lipid concentration preparation (5 mM), SPC limited size increase (SI) more efficiently compared to EPC or HSPC. With SPC and EPC, sucrose showed better size results compared to trehalose (Lyoprotectant/Lipid ratio (w/w) avoiding any SI: 100nm),thisworktargetsthekeyparametersthatinfluencethefreeze−dryingofapreparationthatcombinescholesterol,DOTAPandphosphatidylcholine(eithernaturalsoybeanoregg(SPCorEPC)orhydrogenated(HSPC)).Afterfreeze−dryingblankliposomes,sizeincreasedsignificantlywheninitiallipidconcentrationwasloweredfrom20to5mM(p=0.0018).Withlowlipidconcentrationpreparation(5mM),SPClimitedsizeincrease(SI)moreefficientlycomparedtoEPCorHSPC.WithSPCandEPC,sucroseshowedbettersizeresultscomparedtotrehalose(Lyoprotectant/Lipidratio(w/w)avoidinganySI:5 and 10(forSPC),10 (for SPC), 10(forSPC),10 and $15 (for EPC), for sucrose and trehalose, respectively), but the opposite was evidenced with HSPC liposomes where a Trehalose/Lipid ratio of 25 barely prevented SI. In addition, slow versus quick cooling rate led to limiting SI for HSPC liposomes (p = 0.0035). With sucrose or trehalose at both Lyoprotectant/Lipid ratios ensuring size stabilisation (10:1 and 15:1, respectively), ATP leakage ranged between 38.8 ± 7.9% and 58.2 ± 1.4%. In conclusion, this study emphasizes that using strict size maintenance as the primary objective does not result in drug complete retention inside the liposome core.