ABSTRACT
PURPOSE: Spray-dried chitosan microparticles for cellular delivery of antigen to dendritic cells (DC) and macrophages (MÏ) were investigated. METHODS: Chitosan microparticles were prepared by spray drying. For comparison, poly(lactic-co-glycolic acid) (PLGA) and poly(α-butyl cyanoacrylate) (BCA) micro-/nanoparticles were generated. Bovine serum albumin (BSA) was used as a model antigen. The particles were characterized in terms of size, morphology, surface charge, surface composition, protein content, entrapment efficiency, in vitro release, and protein integrity. Additionally, they were subject to cell viability and cellular uptake study with DC and MÏ. RESULTS: Size of chitosan, PLGA, and BCA micro-/nanoparticles ranged between 3.11-7.18, 0.94-6.26, and 0.30-6.34 µm, respectively. Particle morphology and in vitro protein release varied, depending on polymer type, particle composition and preparation process parameters. Chitosan microparticles were cationic, while PLGA microparticles were neutral. BCA micro-/nanoparticles were either anionic or cationic, according to polymerization pH. Protein content and entrapment efficiency of chitosan and PLGA microparticles were relatively consistent. Only integrity and conformational structure of protein encapsulated in chitosan microparticles were completely retained. Chitosan and PLGA microparticles were non-toxic to DC and MÏ, but the former were internalized more efficiently. CONCLUSIONS: Spray-dried chitosan microparticles delivered the antigen efficiently to DC and MÏ.
Subject(s)
Antigens/administration & dosage , Antigens/chemistry , Chitosan/chemistry , Dendritic Cells/metabolism , Macrophages/metabolism , Proteins/administration & dosage , Proteins/chemistry , Animals , Cell Survival/drug effects , Cells, Cultured , Drug Delivery Systems/methods , Hydrogen-Ion Concentration , Lactic Acid/chemistry , Mice , Microspheres , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Particle Size , Polyglycolic Acid/chemistry , Polylactic Acid-Polyglycolic Acid Copolymer , Polymers/administration & dosage , Polymers/chemistry , Serum Albumin, Bovine/administration & dosage , Serum Albumin, Bovine/chemistryABSTRACT
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 microm, 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.
Subject(s)
Capsules/chemistry , Chitosan/chemistry , Proteins/chemistry , Animals , Cattle , Chemistry, Physical/methods , Circular Dichroism , Drug Compounding , Electrophoresis, Polyacrylamide Gel , Molecular Weight , Particle Size , Serum Albumin, Bovine/chemistry , Temperature , Time FactorsABSTRACT
Anionic pH-sensitive membrane-disruptive polymers have evolved as a new class of bioactive excipients for the cytosolic delivery of therapeutic macromolecules. A large variety of anionic copolymers and analogues of poly(acrylic acid) (PA) was investigated and compared to a cationic PA copolymer. The pH-responsive membrane-disruptive properties were characterized by employing three in vitro models, such as pH dependent shift of pyrene fluorescence, liposome leakage and lysis of red blood cells. The pH-dependent increase of polarity and membrane disruption in the different model systems was in good agreement for all tested PA polymers. The efficacy of polymer-induced membrane disruption was concentration-dependent and significantly affected by the composition of the membrane. The sensitivity of relatively complex membranes of mammalian cells can be ranked between plain diphosphatidylcholine (DPPC) liposomal membranes and the more rigid cholesterol-containing DPPC membranes. Among the various studied PA polymers, medium and low molecular poly(ethacrylic acid) (PEA) and poly(propacrylic acid) (PPA) were identified as displaying significant pH-dependent disruptive activity. Relative to the disruptive cationic PA polymer (PDMAEM) the ranking is PEA < PPA < PDMAEM. The fine tuning of the pH-responsive hydrophilic-hydrophobic balance is likely to be responsible for the superior effect of PEA and PPA compared to other anionic PA polymers. This thorough investigation of a large variety of different anionic PA polymers and the comparison with an efficient, although rather toxic cationic PA polymer provides a good assessment for further therapeutic applications.
