Effect of lyophilization on liposomal encapsulation of salmon calcitonin.
OBJECTIVE
The intent of this work was to assess the impact of lyophilization on the encapsulation of salmon calcitonin (sCT) into liposomes.


METHODS
Four different liposomal formulations were investigated, i.e. DPPC:Chol:DSPE-PEG(2000) (75:20:5 and 65:30:5) and DPPC:Chol (80:20 and 66.7:33.3). Lipid films were prepared and hydrated with loading buffer containing sCT and different concentrations of the cryoprotectant, trehalose dihydrate. The liposomes were lyophilized, reconstituted and extruded to obtain small unilamellar vesicles. Non-encapsulated sCT was separated by gel filtration. Non-lyophilized formulations and liposomes lyophilized without the cryoprotectant were used as controls. Liposomes were analyzed for particle size, polydispersity index, zeta-potential and encapsulation efficiency. ³¹P-NMR (phosphorous nuclear magnetic resonance spectroscopy) was performed on selected formulations.


RESULTS
Post-lyophilization, no significant change in particle sizes and zeta-potentials were noted, regardless of the presence or absence of the cryoprotectant. Encapsulation efficiencies, however, increased following lyophilization, in both PEGylated (lyophilization control batch) and non-PEGylated liposomes (cryoprotectant batches only). ³¹P-NMR revealed the presence of two distinct vesicle populations--liposomes and micelles--in PEGylated formulation. The presence of micelles might be responsible for the observed encapsulation enhancement of sCT in the PEGylated formulation.


CONCLUSIONS
Lyophilization resulted in an increase in encapsulation efficiency of sCT in PEGylated liposomes, even in the absence of a cryoprotectant, due to presence of micellar vesicles.
School of Pharmacy and Pharmaceutical Sciences, Trinity College Dublin , Dublin 2 , Ireland.
Janani
J
Swaminathan
Carsten
C
Ehrhardt
D000818:Animals; D050071:Bone Density Conservation Agents / Q000008:administration & dosage / Q000737:chemistry*; D002116:Calcitonin / Q000008:administration & dosage / Q000737:chemistry*; D003451:Cryoprotective Agents / Q000737:chemistry; D004337:Drug Carriers / Q000008:administration & dosage / Q000737:chemistry*; D004339:Drug Compounding; D004355:Drug Stability; D029941:Fish Proteins / Q000008:administration & dosage / Q000737:chemistry*; D005612:Freeze Drying; D008081:Liposomes; D010316:Particle Size; D055550:Protein Stability; D012474:Salmon; D014199:Trehalose / Q000737:chemistry
D050071:Bone Density Conservation Agents; D003451:Cryoprotective Agents; D004337:Drug Carriers; D029941:Fish Proteins; D008081:Liposomes; D002116:Calcitonin; D014199:Trehalose
31P-NMR; Biopharmaceuticals; PEGylated liposomes; freeze drying