LPS extraction process

Patent metadata

Field	Value
Publication number	US-9499639-B2
Application number	US-201514592894-A
Country	US
Kind code	B2
Filing date	Jan 8, 2015
Priority date	May 20, 2010
Publication date	Nov 22, 2016
Grant date	Nov 22, 2016

Abstract

The present invention provides a method of lipopolysaccharide (LPS) extraction from gram negative bacterial cells comprising the step: extracting LPS from the cell culture in a composition (LPS extraction composition) comprising water, an alcohol and a further organic solvent.

First claim

The invention claimed is:

1. A method of lipopolysaccharide (LPS) extraction from gram negative bacterial cells comprising the step, extracting LPS from the gram negative bacterial cells in an LPS extraction composition comprising water, an alcohol and a further organic solvent, wherein the LPS extraction composition is single-phase, and wherein the amount of water in the LPS extraction composition is between about 0.8 to 1.2% (v/v).

2. The method of claim 1 wherein the extraction of LPS is performed at a temperature of between about 35° C. and about 65° C.

3. The method of claim 1 wherein LPS extraction is performed at a pH of between 7.8 and 9.

4. The method of claim 1 wherein the LPS extraction is performed in about 0.5 to about 20 hours.

5. The method of claim 1 further comprising the steps:
i. Washing cells with a solution of ethanol or methanol prior to the step of extraction; and optionally
ii. Washing cells a second time with ethanol or methanol prior to the step of extraction.

6. The method of claim 1 further comprising the step: evaporating the water, alcohol and further organic solvent from the LPS solution, thereby yielding a dry LPS residue.

7. The method of claim 1 further comprising the step: subjecting the LPS to sequential acid hydrolysis and base hydrolysis, to form 3D-MPL.

8. The method of claim 1 wherein the amount of water is about 1% (v/v).

9. The method of claim 1 wherein the alcohol is selected from the list: methanol, ethanol, isopropanol or butanol.

10. The method of claim 1 wherein the percentage of alcohol in the LPS extraction composition is between about 5% and about 40% (v/v).

11. The method of claim 1 wherein the LPS extraction composition comprises chloroform, methanol and water.

12. The method of claim 1 wherein the LPS extraction composition comprises an alkane, ethanol and water.

Assignees

• Glaxosmithkline Biologicals Sa

Inventors

• Charles Philippe
• Geldhof Geoffroy
• Mancuso Vincent

Classifications

• C08B37/0003 Primary

  General processes for their isolation or fractionation, e.g. purification or extraction from biomass · CPC title

• C12P19/04

  Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds · CPC title

• C07H1/00

  Processes for the preparation of sugar derivatives · CPC title

• C09K3/00

  Materials not provided for elsewhere · CPC title