Biomedical Engineering Reference
In-Depth Information
Preparation
Liposomes were prepared by film cast method as reported elsewhere; phosphati-
dylcholine, cholesterol, stearylamine, and drug (7:3:1.5:1.0 molar ratio) were
dissolved in minimum quantity of chloroform and methanol (2:1) in a round-
bottomed flask, and a thin film was casted by evaporating the solvent under
reduced pressure using rotatory flash. Finally, traces of solvent were removed
under vacuum. The dried lipid film was hydrated with phosphate buffer saline
(PBS) (pH 7.4, 0.1 mL/mg lipid) [
5
]. To 5 ml suspension of liposomes, 5 mg
chondroitin sulfate was dissolved in PBS (pH 7.4). To it 10 mg of EDC per ml was
added. The mixture was incubated at room temperature (25 ± 2 C) for 3 h, and
coupling was confirmed by IR spectroscopy [
6
].
Characterization
Surface Morphology
The morphological examination of liposomes was performed using a transmission
electron microscope (TEM) following negative staining with sodium phospho-
tungstate solution (0.2 %, w/v), at various magnifications.
Zeta Potential Measurements
Suitably diluted aqueous suspension of liposomes at concentration of
1.0 ± 0.3 mg/mL was taken for zeta potential measurements using Malvern
Zetasizer (DTS Ver, 4.10, Malvern Instruments).
Entrapment Efficiency
Entrapment efficiency was determined by separating the unentrapped drug by
minicolumn centrifugation and disrupting the liposomes with 0.1 % Triton X-100.
Drug
content
was
determined
spectrophotometrically
using
UV-Vis
spectrophotometer.
In Vitro Drug Release
Liposomal suspension was held in dialysis membrane (mol. wt. cut off 3,500) and
hung in a beaker with PBS (pH 7.4); the system was shaken with a magnetic
stirrer. Samples were withdrawn periodically and analyzed for drug content.
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