Method for removal of impurities from bacterial capsular polysaccharide based preparations
Inventors
DHERE, Rajeev Mhalasakant • Pisal, Sambhaji Shankar • ANNAMRAJU, Dattatreya Sarma
Assignees
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Publication Number
US-12180527-B2
Publication Date
2024-12-31
Expiration Date
Abstract
The present invention relates to an improved process for purification of bacterial capsular polysaccharides, more specifically capsular polysaccharides of gram negative bacteria. The process comprises of concentration and diafiltration of harvest, treatment with anionic detergent and strong alkali followed by centrifugation, diafiltration and cationic detergent based precipitation of bacterial polysaccharides. The process results in significant reduction of endotoxin, protein and nucleic acid impurities thereby providing higher recovery of capsular polysaccharide with the desired O-acetyl levels. Said process is scalable, non-enzymatic, and employs fewer purification steps.
Core Innovation
The invention provides an improved, scalable, non-enzymatic purification process for Neisseria meningitidis capsular polysaccharides selected from serotype C, serotype W, and serotype Y. The process is directed to reducing endotoxin, protein impurity, and nucleic acid impurity while maintaining desired O-acetylation levels.
In the disclosed process, an anionic detergent is added to a crude polysaccharide solution, followed by adding alkali and neutralizing the pH. The resulting solution is subjected to alcohol-based precipitation with centrifugation and retention of the supernatant, and the anionic detergent present in the supernatant is removed.
The polysaccharide is then concentrated and diafiltrated, supporting a purified polysaccharide recovery of about 60–80%. The summary indicates that anionic detergent removal can include potassium salt precipitation, followed by cationic detergent precipitation, and the process is described as maintaining O-acetylation while meeting impurity-related constraints and regulatory/Halal compliance themes.
Claims Coverage
The independent claim covers a method for isolating polysaccharide from Neisseria meningitidis selected from serotype C, serotype W, and serotype Y, with purification steps including detergent/alkali/neutralization, alcohol-based precipitation, anionic detergent removal, and subsequent concentration/diafiltration, where the obtained purified polysaccharide has a recovery of 60–80%. Only one independent claim is provided in the partial content.
Detergent–alkali–pH neutralization and alcohol precipitation workflow
A method for isolation of polysaccharide from Neisseria meningitidis selected from serotype C, serotype W, and serotype Y comprising adding an anionic detergent to a crude polysaccharide solution; adding alkali to the solution; neutralizing the pH of the solution; subjecting the solution to alcohol-based precipitation, followed by centrifugation and retention of supernatant.
Anionic detergent removal after supernatant precipitation
Removing the anionic detergent present in the supernatant.
Concentration and diafiltration after detergent removal
Concentrating and diafiltrating the polysaccharide after removal of the anionic detergent.
Recovery requirement for purified polysaccharide
Wherein the obtained purified polysaccharide from Neisseria meningitidis selected from serotype C, serotype W, and serotype Y has a recovery of 60–80%.
Across the provided material, the independent claim is centered on a purification workflow that combines anionic detergent addition, alkali addition and pH neutralization, alcohol-based precipitation with centrifugation and supernatant retention, subsequent removal of the anionic detergent from the retained supernatant, and final concentration/diafiltration, with the purified polysaccharide recovery constrained to 60–80%.
Stated Advantages
Reduces endotoxin, protein, and nucleic acid impurities while maintaining desired O-acetyl levels.
Scalable, non-enzymatic purification process.
Reduced chemical/consumable use and regulatory/Halal and operational benefits.
Documented Applications
Purified polysaccharide used in conjugation and immunogenic composition embodiments.
Production contexts requiring control of O-acetylation and impurity limits for Neisseria meningitidis serotypes C, W, and Y.
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