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Process for refining group C/Y/W135 meningococcal polysaccharides

A meningococcal and polysaccharide technology, which is applied to antibacterial drugs, medical preparations containing active ingredients, and antibody medical ingredients, etc. No phenol residues, the effect of reducing endotoxin content

Active Publication Date: 2014-03-19
罗益(无锡)生物制药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] In order to overcome the disadvantages of using cold phenol to extract protein in crude polysaccharides in the prior art, the residual cold phenol is harmful to the human body, and other laboratory methods cannot be mass-produced, the present invention provides a new chromatography technology, the protein content is far Far lower than the range specified in the Pharmacopoeia, the residual toxicity is lower than that of the cold phenol method, and suitable for mass production

Method used

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  • Process for refining group C/Y/W135 meningococcal polysaccharides
  • Process for refining group C/Y/W135 meningococcal polysaccharides
  • Process for refining group C/Y/W135 meningococcal polysaccharides

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Purification of group C meningococcal polysaccharide

[0029] The raw material is crude group C meningococcal polysaccharide, the solution is 10mM Tris-HCl, pH9.0; the equilibration buffer is 20mM Tris-HCl, pH8.0, which contains 1% DOC; the eluent is 20mM Tris-HCl , which contains 1M NaCl, pH8.0.

[0030] 1. Dissolve the crude group C meningococcal polysaccharide with a dissolving solution to 5 mg / ml;

[0031] 2. Use a SepHarose-G25 (26 / 10 desalting) column to replace the dissolved polysaccharide into the equilibrium buffer, collect the first peak of the external water as the polysaccharide peak, and the final concentration is about 2.5mg / ml;

[0032] 3. Put the collected polysaccharide solution on DEAE-4FF and Capto TM Adhere tandem column, the loading volume is 20~50mg / ml gel;

[0033] 4. Using linear elution method from DEAE-4FF and Capto TM The adsorbed substance was eluted on the adhere series column, the eluent used was 20mM Tris-HCl, pH8.0, and th...

Embodiment 2

[0037] Example 2 Purification of group C meningococcal polysaccharide

[0038] The method as described in Example 1, the difference is that the aforementioned step 2 is changed to: use a 100KD ultrafiltration membrane, and replace the dissolved polysaccharide into the equilibrium buffer on the ultrafilter, with a final concentration of about 2.5 mg / ml. Compared with Example 1, this method can also achieve the effect of replacing the polysaccharide into the equilibrium buffer, and at the same time can omit the operation of collecting the elution peak, so it is more suitable for industrial production.

Embodiment 3

[0039] Example 3 Refinement of Group Y meningococcal polysaccharide

[0040] The raw material is crude group Y meningococcal polysaccharide. The solution is 10mM Tris-HCl, pH9.0; the equilibration buffer is 10mM Tris-HCl, pH8.0; the eluent is 20mM Tris-HCl, 1 mol / lNaCl, pH8.0.

[0041] 1. Dissolve the crude group Y meningococcal polysaccharide with a dissolving solution to 5 mg / ml;

[0042] 2. Use SepHarose-G25 (26 / 10desalting) column to replace the polysaccharide into the equilibrium buffer, collect the first peak of the external water as the polysaccharide peak, and the final concentration is 2.5mg / ml;

[0043] 3. Put the collected polysaccharide solution on Capto TM Adhere anion exchange column, the loading amount is 20~50mg / ml gel;

[0044] 4. Elute the polysaccharide by combining gradient elution and linear elution, specifically, gradient elution with 35% B eluent for 2 column volumes, and then linear elution with 35%-100% B eluent 5 column volumes. B eluent is 20mM...

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Abstract

The invention discloses a process for refining group C / Y / W135 meningococcal polysaccharides. The process includes dissolving one of a coarse group C meningococcal polysaccharide, a coarse group Y meningococcal polysaccharide and a coarse group W135 meningococcal polysaccharide, displacing the polysaccharide into an equilibration buffer solution, feeding a collected polysaccharide to an anion exchange column, using an eluant to elute absorption substances from the anion exchange column, piecewise collecting absorption peaks with wavelength smaller than 206 according to different elution peaks and using a desalting method to remove micromolecule materials. According to the process for refining the group C / Y / W135 meningococcal polysaccharides, protein contents can be effectively controlled, so that the polysaccharide protein content is far lower than the polysaccharide protein content in a cold phenol method, the recovery rate can reach around 80%, and the process is applicable to mass production with 100g of coarse polysaccharide yield per batch.

Description

technical field [0001] The invention relates to the field of pharmaceutical products containing antigens, in particular to the refining process of C / Y / W135 group meningococcal polysaccharides. Background technique [0002] Meningococcal meningitis is an acute suppurative meningitis caused by Neisseria meningitidis. Its main clinical manifestations are sudden high fever, severe headache, frequent vomiting, skin and mucous membrane petechiae, ecchymosis, and meningeal irritation. In severe cases, septic shock and brain parenchymal damage may occur, which can often be life-threatening. Some patients have outbreaks and can quickly die. A+C+Y+W135 quadrivalent vaccine can play a positive role in the prevention of the disease. After vaccination, the body can produce a humoral immune response to prevent epidemic cerebrospinal meningitis caused by meningococci of groups A, C, Y, and W135. [0003] Bacterial capsular polysaccharide is an active ingredient in the preparation of men...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C08B37/00A61K39/095A61P31/04
Inventor 周家富陈宣洪周富昌任晓莉顾玉林
Owner 罗益(无锡)生物制药有限公司
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