European Journal of Pharmaceutical Sciences 25 (2005) 427–437 Microencapsulated chitosan nanoparticles for lung protein delivery Ana Grenha, Bego˜ na Seijo, Carmen Remu ˜ n´ an-L´ opez ∗ University of Santiago de Compostela, Pharmacy and Pharmaceutical Technology, 15782 Santiago de Compostela, Spain Received 15 November 2004; received in revised form 17 March 2005; accepted 9 April 2005 Available online 11 May 2005 Abstract It has already been demonstrated that spray drying is a very valuable technique for producing dry powders adequate for pulmonary delivery of drugs. We have developed chitosan/tripolyphosphate nanoparticles that promote peptide absorption across mucosal surfaces. The aim of this work was to microencapsulate protein-loaded chitosan nanoparticles using typical aerosol excipients, such as mannitol and lactose, producing microspheres as carriers of protein-loaded nanoparticles to the lung. The results showed that the obtained microspheres are mostly spherical and possess appropriate aerodynamic properties for pulmonary delivery (aerodynamic diameters between 2 and 3 m, apparent density lower than 0.45 g/cm 3 ). Moreover, microspheres morphology was strongly affected by the content of chitosan nanoparticles. These nanoparticles show a good protein loading capacity (65–80%), providing the release of 75–80% insulin within 15 min, and can be easily recovered from microspheres after contact with an aqueous medium with no significant changes in their size and zeta potential values. Therefore, this work demonstrated that protein-loaded nanoparticles could be successfully incorporated into microspheres with adequate characteristics to reach the deep lung, which after contact with its aqueous environment are expected to be able to release the nanoparticles, and thus, the therapeutic macromolecule. © 2005 Elsevier B.V. All rights reserved. Keywords: Chitosan nanoparticles; Dry powders; Ionic gelation; Microspheres; Pulmonary protein delivery; Spray drying 1. Introduction Over the last few years, absorption of therapeutic macro- molecules administered by pulmonary route has received great attention. The large alveolar surface area suitable for drug absorption, low thickness epithelial barrier, extensive vascularization and relatively low proteolytic activity com- pared to other administration routes, together with the ab- sence of the first-pass effect, make the pulmonary delivery of peptides and proteins an outstanding target (Patton and Platz, 1992; Clark, 2002; Courrier et al., 2002). The prerequisite for reliable and specific lung protein delivery is the design of adequate carrier systems. Microspheres have recently been proposed for pulmonary administration, once they can be designed to achieve appro- ∗ Corresponding author. Tel.: +34 981563100x15045; fax: +34 981547148. E-mail address: ffcarelo@usc.es (C. Remu˜ n´ an-L´ opez). priate morphological and aerodynamic characteristics for that purpose. The success of the inhaled particles depends mostly on their size and density, and hence, aerodynamic diameter (Taylor and Kellaway, 2001). The respirable fraction of these powders, generally the fraction of particles with an aerody- namic diameter ranging from 1 to 5 m, should be as high as possible to guarantee a maximum deposition in the deep lung (Bosquillon et al., 2001a). Independently of the method used to produce the aerosol, before reaching the deep lung, inhaled particles must over- come certain obstacles and lung defence mechanisms, essen- tially the effect of the airways structure and the mucus layer, which protects the epithelium in the tracheobronchial region. Particles targeted to the deep lung should be small enough to pass through the mouth, throat and conducting airways and reach the deep lung, but not so small that they fail to deposit and are breathed out again. Therefore, they should have an aerodynamic diameter between 1 and 5 m. Even so, a certain number of particles will be transported away 0928-0987/$ – see front matter © 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.ejps.2005.04.009