CN105037579A - Preparation process of crude polysaccharide of A-group Neisseria meningitidis bacterium capsule - Google Patents
Preparation process of crude polysaccharide of A-group Neisseria meningitidis bacterium capsule Download PDFInfo
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- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
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Abstract
The invention discloses a preparation process of crude polysaccharide of an A-group Neisseria meningitidis bacterium capsule. The preparation process of the crude polysaccharide of the A-group Neisseria meningitidis bacterium capsule comprises the following steps that (1) cultivated Neisseria meningitidis is sterilized by utilizing formaldehyde, and supernate is centrifugally collected from a sterilized culture solution; (2) CTAB is added into the supernate till the final concentration of the CTAB is 1.0 g/L, then standing is performed overnight after full stirring, and polysaccharide is centrifugally collected; (3) the precipitated polysaccharide is stirred by using a calcium chloride solution for 3 hours, and the supernate is centrifugally collected; (4) ethyl alcohol is added into the supernate till the final concentration by volume of the ethyl alcohol is up to 20%-25%, standing is performed at the temperature of 2 DEG C to 8 DEG C overnight, the supernate is centrifugally collected, then ethyl alcohol is added into the supernate till the final concentration by volume of the ethyl alcohol is up to 75%-80%, full shaking is performed, standing is performed for more than 18 hours, and a precipitate is centrifugally collected; (5) absolute ethyl alcohol and acetone are adopted to perform washing for three times respectively, and then drying is performed to obtain the crude polysaccharide. The preparation process of the crude polysaccharide of the A-group Neisseria meningitidis bacterium capsule has the advantages of being high in extraction efficiency and the like.
Description
Technical field
The present invention relates to a kind of preparation technology of Crude polysaccharides, that be specifically related to is the preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides.
Background technology
Meningococcal meningitis (being called for short epidemic meningitis, lower same) is a kind of Acute respiratory infectious disease caused by the scorching coccus (Neisseriameningitidis) of neisseria meningitis.Over more than 100 year, always popular all over the world or be dispersed in generation, septicemia, meningitis can be caused after infection pathogen.Susceptible population is mainly children, the highest with fulminant type case fatality rate, can reach 40% ~ 60%.Each continent, world today sickness rate is 1,/10 ten thousand ~ 10,/10 ten thousand, and total case fatality rate, 5% ~ 15%, has neural system sequela up to the meningitis patient of 20%, comprises intellectual impairment and deafness etc.Carry out serological classification according to capsular polysaccharide type and can divide 13 serotypes, wherein A group, B group, C group account for 90% of Epidemic bacterial flora.A group meningitis cocci is more pandemic principal causative serogroups, particularly at so-called Africa " the popular band of epidemic meningitis ", just there will be once comparatively be very popular every 7-14.
China in 1938,1949, once to there are 5 national epidemic meningitis in nineteen fifty-nine, 1967 and 1977 popular; Wherein popular serious with spring in 1967, sickness rate is up to 4,03/,100,000, and case fatality rate is 5.49%.The case in China's past more than 90% is that A group germ is caused a disease, and present B group or C group germ also cause epidemic meningitis to break out sometimes.2003 start, and the sickness rate of C group's epidemic meningitis obviously rises.Current A group and C group account for more than 50% of all serogroupss altogether, and C group still has the trend increased further.Therefore, the emphasis of current China prevention epidemic meningitis work prevents A group and C group to be master.
The most effective means of current prevention meningococcal meningitis is vaccination.Research shows, the capsular polysaccharide of A group C group meningitis cocci has good immunogenicity, extracts capsular polysaccharide and directly can be prepared into vaccine, and the crowd after injecting immune can adaptive immune protection.China included vaccine of epidemic menigitis in National immunization Program from 2007, in the whole country to the universal inoculation of school-age children.The epidemic meningitis polysaccharide vaccine including National Immunization Program at present in comprises A meningococcal polysaccharide vaccine, A group C meningococcal polysaccharide vaccine.
At present, the production technique of A group C meningococcal polysaccharide vaccine is all extract Crude polysaccharides from meningococcal capsular, Crude polysaccharides obtains refined sugar after purifying, and refined sugar makes polysaccharide-ADH derivative through overactivation with derivative, and then effectively makes A group C meningococcal polysaccharide vaccine.
Prior art extracts in the production process of Crude polysaccharides from meningococcal capsular, and the not high cost that causes of Crude polysaccharides extraction efficiency increases.
Summary of the invention
The object of the invention is to solve the not high problem causing cost to increase of efficiency extracting Crude polysaccharides in prior art in pod membrane, a kind of preparation technology of the A group meningitis cocci bacterial capsule Crude polysaccharides solved the problem is provided.
For solving above-mentioned shortcoming, technical scheme of the present invention is as follows:
The preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides, comprises the following steps:
(1) meningococcus turned out is utilized formaldehyde sterilization, the medium centrifugal of sterilization is collected supernatant liquor;
(2) till when the final concentration adding CTAB to CTAB in supernatant liquor is 1.0g/L, hold over night after then fully stirring, collected by centrifugation polysaccharide;
(3) the polysaccharide calcium chloride solution of precipitation is stirred 3h, collected by centrifugation supernatant liquor;
(4) in supernatant liquor, ethanol is added until the volume final concentration of ethanol reaches 20 ~ 25%, 2 ~ 8 DEG C of hold over night, collected by centrifugation supernatant liquor; In supernatant liquor, add ethanol again until the volume final concentration of ethanol reaches 75 ~ 78%, shake well, then leave standstill more than 18 hours, centrifugal collecting precipitation;
(5) dry after respectively washing three times with dehydrated alcohol and acetone, be rough polysaccharide after drying.
Further, in described step (2), the final concentration of CTAB is 1.0g/L.
Further, in described step (3), churning time is 3h, and calcium chloride solution concentration is 0.5mol/L.
Preferably, in described step (4), first time adds the volume final concentration of ethanol is 25%, and the volume final concentration that second time adds ethanol is 75%.
Preferably, the drying in described step (5) adopts freeze-drying method.
In the present invention, CTAB is cetyl trimethylammonium bromide.
The present invention compared with prior art, has the following advantages and beneficial effect:
Method of the present invention optimizes ratio when each chemical substance adds reaction and concentration, effectively improves in pod membrane the efficiency extracting Crude polysaccharides, and then effectively reduces cost and drop into.
Embodiment
Below in conjunction with embodiment, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.
Embodiment 1
The preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides, comprises the following steps:
(1) meningococcal for 1L nutrient solution is utilized formaldehyde sterilization, the medium centrifugal of sterilization is collected supernatant liquor;
(2) till when the final concentration adding CTAB to CTAB in supernatant liquor is 1.0g/L, hold over night after then fully stirring, collected by centrifugation polysaccharide;
(3) the polysaccharide calcium chloride solution of precipitation being stirred 3h makes polysaccharide and CTAB dissociate, and calcium chloride solution concentration is 0.5mol/L, collected by centrifugation supernatant liquor;
(4) in supernatant liquor, ethanol is added until the volume final concentration of ethanol reaches 25%, 2 ~ 8 DEG C of hold over night, collected by centrifugation supernatant liquor; In supernatant liquor, add ethanol again until the volume final concentration of ethanol reaches 75%, shake well makes polysaccharide precipitation, then leaves standstill more than 18 hours, centrifugal collecting precipitation;
(5) dry after respectively washing three times with dehydrated alcohol and acetone, be rough polysaccharide after drying; The quality detecting the rough polysaccharide that the present embodiment extracts is 146mg.
Embodiment 2
The difference of the present embodiment and embodiment 1 is only, ratio when each chemical substance adds reaction in the present embodiment is different with concentration, specifically arranges as follows:
In step (2), the final concentration of CTAB is 1.0g/L, hold over night after then fully stirring, collected by centrifugation polysaccharide; In step (4), first time adds the volume final concentration of ethanol is 20%, and the volume final concentration that second time adds ethanol is 78%; The quality detecting the rough polysaccharide that the present embodiment extracts is 152mg.
Embodiment 3
The difference of the present embodiment and embodiment 1 is only, ratio when each chemical substance adds reaction in the present embodiment is different with concentration, specifically arranges as follows:
In step (2), the final concentration of CTAB is 1.5g/L; In step (3), the concentration of calcium chloride solution is 1mol/L; In step (4), first time adds the volume final concentration of ethanol is 25%, and the volume final concentration that second time adds ethanol is 80%, detects that the quality of the rough polysaccharide that the present embodiment extracts is 115mg.
Above-described embodiment is only the preferred embodiments of the present invention, not limiting the scope of the invention, as long as adopt principle of design of the present invention, and the change carried out non-creativeness work on this basis and make, all should belong within protection scope of the present invention.
Claims (5)
- The preparation technology of 1.A group meningitis cocci bacterial capsule Crude polysaccharides, is characterized in that, comprise the following steps:(1) meningococcus turned out is utilized formaldehyde sterilization, the medium centrifugal of sterilization is collected supernatant liquor;(2) till when the final concentration adding CTAB to CTAB in supernatant liquor is 1.0g/L, hold over night after then fully stirring, collected by centrifugation polysaccharide;(3) the polysaccharide calcium chloride solution of precipitation is stirred 3h, collected by centrifugation supernatant liquor;(4) in supernatant liquor, ethanol is added until the volume final concentration of ethanol reaches 20 ~ 25%, 2 ~ 8 DEG C of hold over night, collected by centrifugation supernatant liquor; In supernatant liquor, add ethanol again until the volume final concentration of ethanol reaches 75 ~ 78%, shake well, then leave standstill more than 18 hours, centrifugal collecting precipitation;(5) dry after respectively washing three times with dehydrated alcohol and acetone, be rough polysaccharide after drying.
- 2. the preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides according to claim 1, is characterized in that, in described step (2), the final concentration of CTAB is 1.0g/L.
- 3. the preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides according to claim 1, is characterized in that, in described step (3), churning time is 3h, and calcium chloride solution concentration is 0.5mol/L.
- 4. the preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides according to claim 1, is characterized in that, in described step (4), first time adds the volume final concentration of ethanol is 25%, and the volume final concentration that second time adds ethanol is 75%.
- 5. the preparation technology of A group meningitis cocci bacterial capsule Crude polysaccharides according to claim 1, is characterized in that, the drying in described step (5) adopts freeze-drying method.
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| Application Number | Priority Date | Filing Date | Title |
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| CN201510543283.9A CN105037579A (en) | 2015-08-31 | 2015-08-31 | Preparation process of crude polysaccharide of A-group Neisseria meningitidis bacterium capsule |
| PCT/CN2016/078085 WO2017036139A1 (en) | 2015-08-31 | 2016-03-31 | Preparation process for a-group neisseria meningitidis bacterium capsule crude polysaccharide |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017036139A1 (en) * | 2015-08-31 | 2017-03-09 | 成都欧林生物科技股份有限公司 | Preparation process for a-group neisseria meningitidis bacterium capsule crude polysaccharide |
| CN107488237A (en) * | 2017-08-08 | 2017-12-19 | 江苏大学 | A kind of easy low endotoxin Acinetobacter bauamnnii capsular polysaccharide extracting method |
| CN112521520A (en) * | 2020-12-04 | 2021-03-19 | 苏州微超生物科技有限公司 | Preparation method of meningococcal capsular polysaccharide |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1518456A (en) * | 2001-06-20 | 2004-08-04 | Capsular polysaccharide solubilisation and combination vaccines |
Family Cites Families (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4235994A (en) * | 1978-06-26 | 1980-11-25 | Merck & Co., Inc. | High molecular weight meningococcal group C vaccine and method for preparation thereof |
| CN100540675C (en) * | 2006-12-06 | 2009-09-16 | 云南沃森生物技术有限公司 | A kind of Sepharose of use 4FF gel carries out the method for purifying to bacterial polysaccharides |
| CN100462102C (en) * | 2007-02-07 | 2009-02-18 | 重庆智仁生物技术有限公司 | Mucosal meningococcal multivalence combined vaccines |
| WO2011148382A1 (en) * | 2010-05-28 | 2011-12-01 | Biological E Limited | An improved process for the purification of capsular polysaccharides of haemophilus influenza - b, neisseria meningitis such as serotypes a, c, y and w-135, and other similar related capsular polysaccharides produced from both gram negative and gram positive microorganisms using aluminium phosphate with alcohol. |
| CN104689309A (en) * | 2015-03-27 | 2015-06-10 | 成都欧林生物科技股份有限公司 | Separated and purified acellular pertussis-diphtheria-tetanus, b-type haemophilus influenzae and A-group and C-group meningococcus combined vaccine and preparation method thereof |
| CN105031634A (en) * | 2015-08-31 | 2015-11-11 | 成都欧林生物科技股份有限公司 | A-group C-group Neisseria meningitidis polysaccharide conjugate vaccine activating process |
| CN105037579A (en) * | 2015-08-31 | 2015-11-11 | 成都欧林生物科技股份有限公司 | Preparation process of crude polysaccharide of A-group Neisseria meningitidis bacterium capsule |
-
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1518456A (en) * | 2001-06-20 | 2004-08-04 | Capsular polysaccharide solubilisation and combination vaccines |
Non-Patent Citations (2)
| Title |
|---|
| 王晓杰等: "《免疫技术》", 31 August 2010, 化学工业出版社 * |
| 陈作江等: "A群脑膜炎球菌荚膜多糖纯化工艺的优化", 《微生物学免疫学进展》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2017036139A1 (en) * | 2015-08-31 | 2017-03-09 | 成都欧林生物科技股份有限公司 | Preparation process for a-group neisseria meningitidis bacterium capsule crude polysaccharide |
| CN107488237A (en) * | 2017-08-08 | 2017-12-19 | 江苏大学 | A kind of easy low endotoxin Acinetobacter bauamnnii capsular polysaccharide extracting method |
| CN112521520A (en) * | 2020-12-04 | 2021-03-19 | 苏州微超生物科技有限公司 | Preparation method of meningococcal capsular polysaccharide |
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