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Supercritical Fluids Processing of Recombinant Human Growth Hormone
Uppsala university, Department of Pharmacy.
2005 (English)In: Drug Development and Industrial Pharmacy, ISSN 0363-9045, E-ISSN 1520-5762, Vol. 31, no 2, p. 135-49Article in journal (Refereed) Published
Abstract [en]

The aim of the study was to investigate the feasibility of precipitating recombinant human growth hormone (hGH) from aqueous solutions using conventional and modified techniques of solution-enhanced dispersion (SEDS) by supercritical fluids. The study investigated the effect on hGH stability of adding isopropanol either as a cosolvent with the original aqueous protein solution (conventional process) or to the supercritical carbon dioxide before mixing with the aqueous protein solution (modified process). The influence of the addition of sucrose (with or without isopropanol) on the precipitation behavior and stability of the protein was also studied. Experiments were performed under various processing conditions (pressure 100-200 bars and temperature 40-50°C), and with various flow rates and solution compositions (CO2/isopropanol and protein solution). Bioanalytical characterization of the resulting powders involved spectrophotometry, sodium dodecyl sulfate-polycrylamide gel electrophoresis, reverse-phase high performance liquid chromatography (RP-HPLC), and size exclusion chromatography. Solid-state characterization was performed using differential scanning calorimetry, X-ray powder diffraction, scanning electron microscopy, and Karl Fischer techniques. Results showed that with both conventional and modified methods, under optimum processing conditions, the presence of sucrose in the solution decreased the destabilizing effects of the solvent and/or process on the structure of hGH. More hGH was dissolved from the precipitated powders containing sucrose than from those containing only isopropanol. Reverse-phase HPLC indicated that about 94% of the hGH was recovered in its native form. The proportion of dimers and oligomers was reduced in the presence of sucrose; about 92% of the soluble protein was present in monomer form under optimal conditions. The remaining undissolved protein was in monomeric form. The precipitated powders were amorphous, containing particulate aggregates in the size range 1-6 µm with 5-10% residual moisture content. In conclusion, hGH was successfully precipitated from aqueous solution using SEDS technology. The presence of sucrose in the protein solution promoted the precipitation of hGH and reduced aggregation and improved dissolution

Place, publisher, year, edition, pages
2005. Vol. 31, no 2, p. 135-49
Identifiers
URN: urn:nbn:se:ltu:diva-7615DOI: 10.1081/DDC-200047368Local ID: 60250bc0-e8ca-11db-b9a9-000ea68e967bOAI: oai:DiVA.org:ltu-7615DiVA: diva2:980505
Note
Upprättat; 2005; 20070412 (andbra)Available from: 2016-09-29 Created: 2016-09-29 Last updated: 2017-11-24Bibliographically approved

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