Chitosan microparticles for delivery of proteins were prepared by spray-drying technique. The effects of formulation (molecular weight and concentration of chitosan) and process variables (inlet drying air temperature and spray rate) on size and morphology of microparticles were characterized. Size of microparticles was mainly controlled by formulation variables, while particle morphology was influenced by both formulation and process variables investigated in this study. Bovine serum albumin (BSA), as a model protein, was loaded into microparticles at different levels. BSA-loaded chitosan microparticles were characterized in terms of physicochemical properties and integrity of encapsulated protein, which was studied by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and circular dichroism. Size of microparticles ranged between 3.760–8.681 µm, of which BSA-loaded microparticles were larger in size than their corresponding blank microparticles. All microparticles showed dented or distorted surface, especially when BSA was incorporated, with positive surface charge exposed. Burst release of protein was observed. The effect was more pronounced as BSA loading level was increased. Integrity of entrapped protein could be retained when BSA was incorporated at high loading level. In conclusion, chitosan microparticles for delivery of protein could be efficiently prepared by spray-drying technique. The encapsulated protein was capable of retaining its integrity after the preparation process.

Diclofenac sodium (DS) controlled release solid dispersions were prepared by spray drying using ethylcellulose (EC), methacrylic acid copolymer (Eudragit), chitosan, hydroxypropyl methylcellulose (HPMC), and carbomer as single carriers and EC-chitosan as combined carriers. Among solid dispersions of 3:1 drugsingle carrier, the system containing chitosan exhibited the slowest dissolution followed by the systems containing Eudragit, EC, HPMC, and carbomer, respectively. Combined carriers of EC-chitosan exhibited more dissolution retarding effect than single carrier of EC or chitosan. An Hadamard matrix H[8] was employed to estimate the main effects of four parameters: spray feeding volume and contents of absolute ethanol, EC, and chitosan. Optimization strategy using multiple linear regression and a feasibility computer program was utilized to obtain the optimum quantities of the four parameters that would result in a required DS controlled release solid dispersion. The validation of the optimum DS solid dispersion was confirmed by statistical analysis. The optimized 10: (2.5+0.02) DS:(EC+chitosan) controlled release solid dispersion exhibited a dissolution profile that was well fitted to Higuchi model.

Squalene-based oil-in-water (O/W) emulsions have been used as effective and safe adjuvants in approved influenza vaccines. However, there are concerns regarding the safety and side effects of increasing risk of narcolepsy. In present study, novel O/W microemulsions (MEs) containing wheat germ oil, D-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS) and Cremophor EL (CreEL) or Solutol HS15 were formulated with/without a cationic surfactant, cetyltrimethylammonium bromide (CTAB) and then sterilized by autoclaving. Their physical properties and biological efficacies were evaluated. The results demonstrated that autoclaving reduced the droplet size to ∼20 nm with narrow size distributions resulting in monodisperse systems with good stability up to 3 years. Hemolytic activity, viscosity, pH, and osmolality were appropriate for parenteral use. Bovine serum albumin (BSA), a model antigen, after mixing with MEs retained the protein integrity, assessed by SDS-PAGE and CD spectroscopy. Greater percentages of 28SC cell viability were observed from CreEL-based MEs. Uptake of FITC-BSA-MEs increased with the increasing concentration of CTAB confirmed by CLSM images. Furthermore, cationic CreEL-based MEs could induce Th1 cytokine synthesis with an increase in TNF-α and IL-12 levels and a decrease in IL-10 level. In vivo immunization study in mice of adjuvants admixed with influenza virus solution revealed that nonionic and selected cationic CreEL-MEs enhanced immune responses as measured by influenza-specific serum antibody titers and hemagglutination inhibition titers. Particularly, cationic CreEL-based ME showed better humoral and cellular immunity with higher IgG2a titer than nonionic CreEL-based ME and antigen alone. No differences in immune responses were observed between mice immunized with selected cationic CreEL-based ME and marketed adjuvant. In addition, the selected ME induced antigen-sparing while retained immune stimulating effects compared to antigen alone. No inflammatory change in muscle fiber structure was observed. Accordingly, the developed cationic CreEL-based ME had potential as novel adjuvant for parenteral influenza vaccine.