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Hib polysaccharide purifying technology

A polysaccharide and process technology, applied in the field of Hib polysaccharide purification process, can solve the problems of many steps, difficult to remove nucleic acid, large difference in stability between batches, etc., and achieve the effects of reducing protein impurities, easy enrichment and harvesting, and saving usage.

Inactive Publication Date: 2012-08-15
CHENGDU OLYMVAX BIOPHARM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to overcome the deficiencies in the prior art, to provide a Hib polysaccharide purification process, which solves the problem that nucleic acid is not easy to remove in the existing Hib preparation process, the purification process is cumbersome, there are many steps, the one-time qualified rate is low, and the batch-to-batch There is a large difference in stability, a large amount of chemicals are used and a lot of waste (reagents, drugs) is generated, and the recovery rate of refining is low, etc.

Method used

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Effect test

Embodiment 1

[0031] Hib polysaccharide purification process, which includes the following eight steps: collecting polysaccharide complex precipitates, centrifuging to collect rough sugar precipitates, washing rough sugar precipitates, drying raw sugar, dissolving raw sugar, phenol extraction, phenol removal, and refining refined sugar:

[0032] S1: Collect the polysaccharide complex precipitate:

[0033] S11: Centrifuge the sterilized culture solution, remove bacteria by centrifugation, and collect the supernatant;

[0034] S12: Add 10% cetyltrimethylammonium bromide solution to the collected supernatant until the final concentration is 2‰, stir and mix evenly, let stand at 5°C for 5 hours, and then collect by centrifugation Precipitation of polysaccharide complexes;

[0035] S2: Centrifuge to collect crude sugar precipitate:

[0036] S21: Use an appropriate amount of 0.7M NaCl to depolymerize the polysaccharide complex, then add 95% ethanol to a final concentration of 25% (v / v), let sta...

Embodiment 2

[0053] Hib polysaccharide purification process, which includes the following eight steps: collecting polysaccharide complex precipitates, centrifuging to collect rough sugar precipitates, washing rough sugar precipitates, drying raw sugar, dissolving raw sugar, phenol extraction, phenol removal, and refining refined sugar:

[0054] S1: Collect the polysaccharide complex precipitate:

[0055] S11: Centrifuge the sterilized culture solution, remove bacteria by centrifugation, and collect the supernatant;

[0056] S12: Add 5% cetyltrimethylammonium bromide solution to the collected supernatant until the final concentration is 1‰, stir and mix evenly, let stand at 8°C for 6 hours, and then collect by centrifugation Precipitation of polysaccharide complexes;

[0057] S2: Centrifuge to collect crude sugar precipitate:

[0058] S21: Use an appropriate amount of 0.4M NaCl to depolymerize the polysaccharide complex, then add 95% ethanol to a final concentration of 30% (v / v), let stan...

Embodiment 3

[0069] Hib polysaccharide purification process, which includes the following eight steps: collecting polysaccharide complex precipitates, centrifuging to collect rough sugar precipitates, washing rough sugar precipitates, drying raw sugar, dissolving raw sugar, phenol extraction, phenol removal, and refining refined sugar:

[0070] S1: Collect the polysaccharide complex precipitate:

[0071] S11: Centrifuge the sterilized culture solution, remove bacteria by centrifugation, and collect the supernatant;

[0072] S12: Add 15% cetyltrimethylammonium bromide solution to the collected supernatant until the final concentration is 3‰, stir and mix evenly, let stand at 2°C for 4 hours, and then collect by centrifugation Precipitation of polysaccharide complexes;

[0073] S2: Centrifuge to collect crude sugar precipitate:

[0074] S21: Use an appropriate amount of 1.0M NaCl to depolymerize the polysaccharide complex, then add 95% ethanol to a final concentration of 20% (v / v), let sta...

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Abstract

The invention discloses a Hib polysaccharide purifying technology which comprises eight steps of: collecting polysaccharide composite precipitate, centrifugally collecting the raw sugar precipitate, washing the raw sugar precipitate, drying the raw sugar, dissolving the raw sugar, performing phenol extraction, removing phenol and refining to obtain refined sugar. The invention provides a Hib polysaccharide purifying technology; each index of the Hib polysaccharide manually purified by the technology in the continuous production batch can meet the pharmacopeia standard at the intermediate level; the impurity residue amount is far lower than the standard limit value, and thus the product quality is improved; the operation steps and reagent usage amount are reduced, and the production of waste is correspondingly reduced, thereby being conducive to environmental protection; the purifying conditions are relatively mild, and the product is treated in a tight environment so as to meet the requirements of the existing GMP (good manufacturing practice) on the production process and protect the product from pollution; and the refining recovery rate of Hib polysaccharide can be improved by 20-30% over the traditional technology.

Description

technical field [0001] The invention relates to a Hib polysaccharide purification process. Background technique [0002] Haemophilus influenzae type b Hib (Hamephilus influenzae type b, referred to as Hib) polysaccharide vaccine has produced a good immune effect on older children since it was launched in the United States in 1985, but it is not effective in inducing infants under 18 months of age. Bactericidal antibodies can not induce immune memory, and the actual immune efficacy is very low, which hinders the popularization and use of Hib polysaccharide vaccine. The reason for this phenomenon is that polysaccharides are T cell-independent antigens. In infants under the age of 2 with immature immune system functions, polysaccharide antigens cannot stimulate the body to produce effective antibodies. High-risk groups cannot play an effective protective role. In order to change the T-cell-independent polysaccharide, the polysaccharide can be covalently coupled to a protein c...

Claims

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Application Information

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IPC IPC(8): C08B37/00
Inventor 李洪光罗力心陈庚朱冲
Owner CHENGDU OLYMVAX BIOPHARM
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