Endotoxin Reduction in Protein Solutions Using Octyl β-D-1-thioglucopyranoside Wash on Chromatography Media

Dhanesh Gadre, Marcia Carlson, Ashley Mullan, Ivy Kabundi, Nichole Pedowitz, Sadiye Amcaoglu Rieder, Natalie White, Kathy Yu, Ellen O'Connor

J Chromatogr A. 2018 Nov 9;1575:49-58.

PMID: 30262393

Abstract:

Endotoxins are complex molecules and one of the most challenging impurities requiring separation in biopharmaceutical protein purification processes. Usually these contaminants are cleared during the downstream process, but if endotoxin interacts with the target protein it becomes difficult to remove. In the present study we identified a detergent, octyl-β-D-1-thioglucopyranoside (OTG), that disrupted endotoxin-protein interactions. The integration of this detergent into washes on several chromatography media was demonstrated to provide a separation tool for decreasing endotoxin from target proteins. This study also examined the mechanism of OTG endotoxin-protein disruption through phase modification incubation and chromatographic studies. The non-ionic OTG wash was shown to break both hydrophobic and electrostatic interactions between the endotoxin and protein. This mechanism contrasts with the breaking of hydrophobic interactions by washing with known endotoxin decreasing Triton X-100 detergent. The difference in mechanisms likely results in the ability of OTG to decrease endotoxin to levels less than those resulting from a detergent wash such as Triton X-100. Finally, we show the impact of the OTG endotoxin removal tool on the biopharmaceutical industry. While maintaining monomer purity and activity levels, endotoxin removal from a fusion protein allowed for decreased background levels in a T cell functional assay. The lowered baseline of T cell responses allowed for more effective detection of molecular interaction with the cells. The detergent wash can be used to both decrease the overall level of endotoxin in a purified protein solution and to enable more effective screening of lead candidate molecules.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP85618219	Octyl β-D-1-thioglucopyranoside		85618-21-9	Price