Method for preparing type b Haemophilus influenzae and polysaccharide conjugate vaccine

A technology of Haemophilus influenzae and conjugated vaccines, which is applied in the field of preparation of Haemophilus influenzae type b polysaccharide conjugated vaccines, which can solve the problems affecting the stability of the production process, the stability of finished vaccines between batches, the quality consistency of finished vaccines, the combination of high molecular weight polysaccharides and proteins Low product yield, uneven molecular size distribution, etc., to achieve the effect of saving raw materials, reducing production costs, and reducing differences between batches

Inactive Publication Date: 2013-08-21
CHENGDU OLYMVAX BIOPHARM
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] However, because there are a large amount of small molecule polysaccharides with a molecular weight lower than 700kd in the refined Hib capsular polysaccharide prepared, the molecular weight of this part of the polysaccharide is small after being combined with the carrier protein, and it is separated after gel filtration chromatography, resulting in a high concentration during the binding process. The yield of molecular polysaccharide-protein conjugates is very low, and the yield of Hib capsular polysaccharide polysaccharide in this p

Method used

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  • Method for preparing type b Haemophilus influenzae and polysaccharide conjugate vaccine
  • Method for preparing type b Haemophilus influenzae and polysaccharide conjugate vaccine
  • Method for preparing type b Haemophilus influenzae and polysaccharide conjugate vaccine

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Embodiment 1

[0019] A method for preparing Haemophilus influenzae type b polysaccharide conjugate vaccine, comprising the following steps:

[0020] S1. Gel filtration chromatography purification of polysaccharides. Dilute the Hib capsular polysaccharides with water for injection to a solution with a concentration of 5 mg / ml, and filter them with a filter membrane with a pore size of 0.45 μm. The filtrate is purified by Sepharose4FF gel chromatography column chromatography. , load the sample on Sepharose4FF gel chromatography column with 10% column volume, elute with 0.15M sodium chloride as the eluent, collect the chromatographic eluate with KD≤0.3, and then use 300KD ultrafiltration to collect the eluate Concentrate by membrane ultrafiltration to a polysaccharide concentration of 6 mg / ml to obtain the macromolecular Hib capsular polysaccharide;

[0021] According to this method, macromolecular Hib capsular polysaccharides were prepared in total, and the preparation results were as follows...

Embodiment 2

[0029] A method for preparing Haemophilus influenzae type b polysaccharide conjugate vaccine, comprising the following steps:

[0030] S1. Gel filtration chromatography purification of polysaccharides. Dilute the Hib capsular polysaccharides with water for injection to a solution with a concentration of 3 mg / ml, and filter them with a filter membrane with a pore size of 0.45 μm. The filtrate is purified by Sepharose4FF gel chromatography column chromatography. , load the sample on Sepharose4FF gel chromatography column with 8% column volume, elute with 0.15M sodium chloride as the eluent, collect the chromatographic eluate with KD≤0.3, and then use 300KD ultrafiltration to collect the eluate Concentrate by membrane ultrafiltration to a polysaccharide concentration of 4 mg / ml to obtain the macromolecular Hib capsular polysaccharide;

[0031] According to this method, macromolecular Hib capsular polysaccharides were prepared in total, and the preparation results were as follows:...

Embodiment 3

[0039] A method for preparing Haemophilus influenzae type b polysaccharide conjugate vaccine, comprising the following steps:

[0040] S1. Gel filtration chromatography purification of polysaccharides. Dilute the Hib capsular polysaccharides with water for injection to a solution with a concentration of 6 mg / ml, and filter them with a filter membrane with a pore size of 0.45 μm. The filtrate is purified by Sepharose4FF gel chromatography column chromatography. , load the sample on Sepharose4FF gel chromatography column with 5% column volume, elute with 0.15M sodium chloride as the eluent, collect the chromatographic eluate with KD≤0.3, and then use 300KD ultrafiltration to collect the eluate Concentrate by membrane ultrafiltration to a polysaccharide concentration of 5 mg / ml to obtain a macromolecular Hib capsular polysaccharide;

[0041] According to this method, macromolecular Hib capsular polysaccharides were prepared in total, and the preparation results were as follows: ...

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Abstract

The invention discloses a method for preparing a type b Haemophilus influenzae and polysaccharide conjugate vaccine. The method comprises the following steps: S1, performing gel filtration chromatography purification of polysaccharide to obtain macromolecular Hib capsular polysaccharide; and S2, preparation of a polysaccharide-TT conjugate, which comprises the following sub-steps: S21, adding CNBr into a macromolecular Hib capsular polysaccharide liquor to activate for 15-25 minutes, then, adding adipic dihydrazide to derivate for 10-20 minutes to remove residual CNBr so as to obtain a polysaccharide-AH derivative, and S22, adding TT liquor to the polysaccharide-AH derivative, uniformly mixing and placing in an ice-bath, and adding EDAC powder, and separating, purifying, degerming and filtering the reactant to obtain the finished product. The method disclosed by the invention has the beneficial effects that the yield of qualified conjugates is improved, the raw material consumption in the conjugation process is reduced, and the stability of the production process and the consistency of quality of finished products are improved.

Description

technical field [0001] The invention relates to the technical field of vaccine preparation methods, in particular to a method for preparing Haemophilus influenzae type b polysaccharide-conjugated vaccine. Background technique [0002] Haemophilus influenzae (Haemophilus Influenza type b, Hib) is still the main pathogen causing human invasive diseases so far, most of which are caused by Haemophilus influenzae type b (Hib). Hib meningitis ranks first among bacterial meningitis, and has the characteristics of high morbidity, high mortality and high disability rate; another characteristic is that the age of onset is young, the infection rate of infants is high, and the age of onset is mainly concentrated under the age of 5 , especially under the age of 2. It is estimated that it causes at least 3 million cases of severe disease each year, of which about 386,000 people die, and has become a major global public health problem. In developing countries, Hib pneumonia plays an impo...

Claims

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Application Information

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IPC IPC(8): A61K39/385A61K39/102A61P31/04C08B37/00
Inventor 李洪光伍长华关晓峰吴强吴畏
Owner CHENGDU OLYMVAX BIOPHARM
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