chitosan and plga microspheres as drug delivery ... - UniCA Eprints

6. RFP Loaded PLGA Microspheres Prepared by Solvent Evaporation Methodmethods were chosen. In the first PLGA and RFP were dissolved in the organic phase and inthe second the drug was dissolved in the aqueous phase. Briefly twenty milligram of PLGAwas dissolved in 1 ml of dichloromethane (DCM). This was dispersed in 4 ml of an aqueousphase of 4% PVA. The resultant emulsion was homogenised for 10 min with an Ultraturax®homogeniser at 800 rpm. Subsequent evaporation of the DCM was carried out withmechanical stirring over night at room temperature. Microparticles were collected bycentrifugation and washed by dispersion in water with subsequent centrifugation, this stepwas repeated three times. Microspheres were than freeze dried.6.1.3. Characterization of RFP-Loaded PLGA Microspheres6.1.3.1. Particle Sizing and MorphologyThe microspheres were analysed for their size and polydispersity index on Zetasizer Nano ZS,Malvern instruments, based on photon correlation spectroscopy, at a scattering angle of 90°and temperature of 25°. Each measurement was the results of 12 run.Measurements were carried out both for fresh and freeze-dried samples. Before counting, thesamples were diluted with a 0.05% (w/v) tween 80 water solution in order to preventprecipitation during the measurements. Results were the means of triplicate experiments.6.1.3.2. Surface Charge (Zeta-Potential)The surface charge of the microspheres was determined with Zetasizer Nano ZS, Malverninstruments. The measurements were carried out in an aqueous solution of KCl 0.1N.Immediately before the determinations microspheres were diluted with KCl solution. Themeasured values were corrected to a standard reference at temperature of 20°. Results are themeans of triplicate experiments.6.1.3.3. Particle MorphologyIn preparation for scanning electron microscopy (SEM) several drops of the microspheresuspension were placed on an aluminum stub having previously been coated with adhesive.The samples were evaporated at room temperature until completely dried, leaving only a thinlayer of particles on the stub. All samples were sputter coated with gold-palladium (Polaron5200,VG Microtech,West Sussex, UK) for 90 seconds (2.2 kV; 20 mA; 150–200A°) under anargon atmosphere. The SEM (Model 6300, JEOL, Peabody, NY) was operated using anacceleration voltage of 10 kV.87