High-throughput system for determining dissolution kinetics of inclusion bodies.
Efficient solubilization is a crucial step during inclusion body processing and dissolving conditions were usually empirically established. Here we describe a new methodology for rapid screening of solubilization conditions and evaluation of dissolution kinetics in microtiter plates. Increase of protein in solution over time was directly related to decrease of turbidity measured by absorbance at 600 nm. Dissolution kinetics of inclusion bodies were described by a first-order reaction kinetics, which was used for drug dissolution modeling. Reaction constants were in the range of 0.01-0.03 s(-1) for buffer conditions providing sufficient solubilization power. This method is not limited to the screening of optimal buffer conditions for solubilization and can be applied for studying other parameters involved in the solubility of IBs, such as pI of the protein, influence of fermentation conditions, influence of initial protein concentration, and more.
Dürauer
Astrid
A
Austrian Center of Biopharmaceutical Technology, Department of Biotechnology, University of Natural Resources and Applied Life Sciences Vienna, Austria. astrid.duerauer@boku.ac.at
Mayer
Sabrina
S
Sprinzl
Wolfgang
W
Jungbauer
Alois
A
Hahn
Rainer
R
D002021:Buffers; D004926:Escherichia coli / Q000378:metabolism; D005612:Freeze Drying; D002479:Inclusion Bodies / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D007700:Kinetics; D008956:Models, Chemical; D009391:Nephelometry and Turbidimetry; D011506:Proteins; D012995:Solubility
D002021:Buffers; D011506:Proteins