Protein encapsulation into nanostructured CaCO3 microparticles templated by PEO-b-polyphosphoester using a supercritical CO2 process

The use of calcium carbonate microparticles is becoming more and more attractive in biomedical applications as proteins carriers for their controlled delivery in the body. We investigate the protein encapsulation by in situ precipitation of CaCO3 particles prepared by a process based on supercritical CO2 and using a new type of degradable well-defined double hydrophilic block copolymer composed of poly(ethylene oxide) and polyphosphoester block with an affinity for calcium like poly(phosphotriester)s bearing pendent carboxylic acids on each repeating monomer unit which evidenced an efficient structure for templating the formation of CaCO3 leading to unprecedented small-sized particles. Lysozyme was chosen as a model for therapeutic protein for its availability and ease of detection. It was found that by this green process, loading into the CaCO3 microparticles with a diameter about 2 μm can be obtained as determined by scanning electron microscopy. A protein loading up to 6.5% active lysozyme was measured by a specific bioassay (Micrococcus lysodeikticus). By encapsulating fluorescent-labelled lysozyme (lysozyme-FITC), the confocal microscopy images confirmed its encapsulation and suggested a core–shell distribution of lysozyme into CaCO3.