Biopharmaceutical characterisation of insulin and recombinant human growth hormone loaded lipid submicron particles produced by supercritical gas micro-atomisation

Abstract

Homogeneous dispersions of insulin and recombinant human growth hormone (rh-GH) in tristearin/phosphatidylcholine/PEG mixtures (1.3:1.3:0.25:0.15 w/w ratio) were processed by supercritical carbon dioxide gas micro-atomisation to produce protein-loaded lipid particles. The process yielded spherical particles, with a 197 ± 94 nm mean diameter, and the insulin and rh-GH recovery in the final product was 57 ± 8% and 48 ± 5%, respectively. In vitro, the proteins were slowly released for about 70–80 h according to a diffusive mechanism. In vivo, the insulin and glucose profiles in plasma obtained by subcutaneous administration of a dose of particles containing 2 μg insulin to diabetic mice overlapped that obtained with 2 μg of insulin in solution. Administration of a dose of particles containing 5 μg insulin resulted in faster and longer glycaemia reduction. Oral administration of 20 and 50 μg insulin equivalent particles produced a significant hypoglycaemic effect. The glucose levels decreased since 2 h after administration, reaching about 50% and 70% glucose reduction in 1–2 h with the lower and higher dose, respectively. As compared to subcutaneous administration, the relative pharmacological bioavailability obtained with 20 and 50 μg equivalent insulin particles was 7.7% and 6.7%, respectively. Daily subcutaneous administration of 40 μg of rh-GH-loaded particles to hypophysectomised rats induced similar body weight increase as 40 μg rh-GH in solution. The daily oral administration of 400 μg rh-GH equivalent particles elicited a slight body weight increase, which corresponded to a relative pharmacological bioavailability of 3.4% compared to subcutaneous administration.