CN111303228A - Method for synthesizing cobamamide - Google Patents
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- 238000000034 method Methods 0.000 title claims abstract description 30
- 235000006279 cobamamide Nutrition 0.000 title claims abstract description 27
- 239000011789 cobamamide Substances 0.000 title claims abstract description 27
- 229960005452 cobamamide Drugs 0.000 title claims abstract description 27
- ZIHHMGTYZOSFRC-UWWAPWIJSA-M cobamamide Chemical compound C1(/[C@](C)(CCC(=O)NC[C@H](C)OP(O)(=O)OC2[C@H]([C@H](O[C@@H]2CO)N2C3=CC(C)=C(C)C=C3N=C2)O)[C@@H](CC(N)=O)[C@]2(N1[Co+]C[C@@H]1[C@H]([C@@H](O)[C@@H](O1)N1C3=NC=NC(N)=C3N=C1)O)[H])=C(C)\C([C@H](C/1(C)C)CCC(N)=O)=N\C\1=C/C([C@H]([C@@]\1(CC(N)=O)C)CCC(N)=O)=N/C/1=C(C)\C1=N[C@]2(C)[C@@](C)(CC(N)=O)[C@@H]1CCC(N)=O ZIHHMGTYZOSFRC-UWWAPWIJSA-M 0.000 title claims abstract description 27
- 230000002194 synthesizing effect Effects 0.000 title claims abstract description 13
- RMRCNWBMXRMIRW-BYFNXCQMSA-M cyanocobalamin Chemical compound N#C[Co+]N([C@]1([H])[C@H](CC(N)=O)[C@]\2(CCC(=O)NC[C@H](C)OP(O)(=O)OC3[C@H]([C@H](O[C@@H]3CO)N3C4=CC(C)=C(C)C=C4N=C3)O)C)C/2=C(C)\C([C@H](C/2(C)C)CCC(N)=O)=N\C\2=C\C([C@H]([C@@]/2(CC(N)=O)C)CCC(N)=O)=N\C\2=C(C)/C2=N[C@]1(C)[C@@](C)(CC(N)=O)[C@@H]2CCC(N)=O RMRCNWBMXRMIRW-BYFNXCQMSA-M 0.000 claims abstract description 18
- 235000000639 cyanocobalamin Nutrition 0.000 claims abstract description 15
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- ZKHQWZAMYRWXGA-KQYNXXCUSA-J ATP(4-) Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](COP([O-])(=O)OP([O-])(=O)OP([O-])([O-])=O)[C@@H](O)[C@H]1O ZKHQWZAMYRWXGA-KQYNXXCUSA-J 0.000 claims description 3
- ZKHQWZAMYRWXGA-UHFFFAOYSA-N Adenosine triphosphate Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(COP(O)(=O)OP(O)(=O)OP(O)(O)=O)C(O)C1O ZKHQWZAMYRWXGA-UHFFFAOYSA-N 0.000 claims description 3
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Images
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H23/00—Compounds containing boron, silicon, or a metal, e.g. chelates, vitamin B12
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
- C07H1/06—Separation; Purification
Abstract
The invention discloses a method for synthesizing cobamamide, belonging to the technical field of production of vitamin B12 series products. The invention adopts a chemical synthesis method based on cyanocobalamine, sodium borohydride and adenine nucleoside triphosphate to prepare the adenosyl cobalamine, and then the adenosyl cobalamine is further refined and purified to obtain a finished product of the adenosyl cobalamine. The process can greatly improve the yield of the cobamamide from about 50 percent to over 99 percent.
Description
Technical Field
The invention relates to the technical field of production of vitamin B12 series products, in particular to a method for synthesizing cobamamide.
Background
The cobamamide is one of vitamin B12 series products, has a chemical name of 5, 6-dimethylbenzimidazolyl-5' -deoxyadenyl cobamamide and a molecular formula of C72H100CoN18O 17P; the molecular weight is 1579. Cobamamide is a dark red crystalline or amorphous powder, has strong hygroscopicity, and is easily decomposed by light. It is slightly soluble in water, almost insoluble in ethanol, and insoluble in acetone, diethyl ether, and chloroform. Cobamamide is mainly used for megaloblastic anemia, nutritional anemia and gestational anemia, and is also used for nervous diseases such as polyneuritis, radiculitis, trigeminal neuralgia, sciatica, nerve palsy, nutritional nervous diseases and leukopenia caused by radioactive rays and medicines.
At present, the production of cobamamide in China always adopts a fermentation liquor extraction method, which comprises the following steps:
a. hydrolyzing and filtering the aerobic vitamin B12 fermentation liquor under the condition of keeping out of the sun to obtain filtrate;
b. adsorbing and desorbing the filtrate by weak acid cation exchange resin to obtain a first-hydrolyzed solution;
c. adding a flocculating agent into the primary hydrolysate, and filtering to obtain a purified solution;
d. adsorbing, spreading and desorbing the purified solution by macroporous resin to obtain secondary decomposition solution;
e. decolorizing the second-hydrolyzed solution with strong-base anion exchange resin to obtain third-hydrolyzed solution;
f. adsorbing the third-stage hydrolyzed solution by chromatography resin, and spreading to obtain a fourth-stage hydrolyzed solution;
g. concentrating the four-decomposition liquid by macroporous resin to obtain a crystallization stock solution;
h. and under the stirring state, adding acetone into the crystallization stock solution for crystallization, suction filtration and drying to obtain the cobamamide finished product.
However, the yield of the process is low, about 50%, and the process has room for improvement.
Disclosure of Invention
In view of the above, the invention provides a cobamamide synthesis method, which has a simple process and a yield of over 99%.
In order to achieve the purpose, the invention adopts the technical scheme that:
a method for synthesizing cobamamide comprises the following steps:
(1) dissolving cyanocobalamine in water to prepare a solution;
(2) pouring sodium borohydride into absolute ethyl alcohol to prepare a solution;
(3) adding the cyanocobalamine solution prepared in the step (1) and the sodium borohydride solution prepared in the step (2) into a reaction tank, enabling the weight ratio of the cyanocobalamine to the sodium borohydride in the solution to be 2.2-2.6: 1, slowly adding adenosine triphosphate into the reaction tank, controlling the pH of the feed liquid in the reaction tank to be 3.0-3.5, reacting for more than 90 minutes, and centrifugally filtering;
(4) adsorbing the filtrate obtained in the step (3) through a macroporous resin column at the flow rate of 160-300L/h, introducing a spreading agent into the macroporous resin column to remove impurities, introducing an analytic agent into the macroporous resin column, and collecting the eluted concentrated material into a crystallization stock solution tank;
(5) adding the concentrated material in the crystallization raw liquid tank into a crystallization tank, and adding crystalline acetone into the crystallization tank; stirring the feed liquid in the crystallizing tank, controlling the feeding speed of the crystallized acetone to be 600-1000L/h, stopping feeding the crystallized acetone when the density of the feed liquid reaches 0.820-0.840 g/ml, continuing stirring for a period of time, stopping stirring and standing;
(6) carrying out vacuum filtration on the mixed solution in the crystallization tank, and separating the mother solution from the crystals;
(7) washing the crystal with three acetone water solutions with the density being more than, equal to or less than that of the mother solution in sequence, and then pumping to dry; then, washing the crystal with crystalline acetone, finally removing the washing liquid through vacuum filtration, and obtaining wet crystal after pumping;
(8) and (4) sieving the wet crystals by using a 80-mesh sieve, and drying undersize products to obtain dried cobamamide finished products.
Further, the concentration of the cyanocobalamin solution in the step (1) is more than 15 g/L.
Further, the concentration of the sodium borohydride solution in the step (2) is 42-52 g/L.
Further, the resolving agent in the step (4) is an acetone aqueous solution, and the concentration of the acetone aqueous solution is 0.940 +/-0.005 g/mL.
Further, in the step (5), the concentrated material and the crystallized acetone are filtered by a filter before being added into the crystallizing tank.
Further, in the step (5), the stirring is continued for 3 hours, and the standing time is more than 2 hours.
Further, in the step (7), the dosage of the three acetone aqueous solutions is controlled to be 100-150L, each acetone aqueous solution is washed twice, and crystals are fully stirred in the washing process.
Further, in the step (7), the vacuum filtration time is 6-8 hours, and the vacuum degree is kept to be more than 0.02MPa in the filtration process.
Further, the drying in the step (8) is specifically performed by:
and (3) putting the undersize into a double-cone dryer, starting the double-cone dryer, starting a hot water pump when the vacuum degree reaches 0.09-0.1 MPa, controlling the temperature at 55-70 ℃ and the drying time at 4-8 h, and cooling the material for 60-80 min after drying.
The beneficial effect who adopts above-mentioned technical scheme to obtain lies in:
1. the invention adopts a chemical synthesis method based on cyanocobalamine, sodium borohydride and ATP (adenosine triphosphate) to prepare the adenosyl cobalamine, and then the adenosyl cobalamine is further refined and purified to obtain a finished product of the adenosyl cobalamine. The process is completely different from the prior art and has outstanding substantive characteristics.
2. The yield of the method can reach more than 99 percent, is far higher than that of the prior art, and has obvious technical progress.
3. The method is simple and easy to implement, is suitable for large-scale production, and has higher economic benefit.
Drawings
FIG. 1 is a process flow diagram of a cobamamide synthesis method according to an embodiment of the present invention.
Detailed Description
The present invention will be described in further detail with reference to the accompanying drawings.
A method for synthesizing cobamamide comprises the following steps:
(1) dissolving cyanocobalamine in water to prepare a solution;
(2) pouring sodium borohydride into absolute ethyl alcohol to prepare a solution;
(3) adding the cyanocobalamine solution prepared in the step (1) and the sodium borohydride solution prepared in the step (2) into a reaction tank, enabling the weight ratio of the cyanocobalamine to the sodium borohydride in the solution to be 2.2-2.6: 1, slowly adding adenosine triphosphate into the reaction tank, controlling the pH of the feed liquid in the reaction tank to be 3.0-3.5, reacting for more than 90 minutes, and centrifugally filtering;
(4) adsorbing the filtrate obtained in the step (3) through a macroporous resin column at the flow rate of 160-300L/h, introducing a spreading agent into the macroporous resin column to remove impurities, introducing an analytic agent into the macroporous resin column, and collecting the eluted concentrated material into a crystallization stock solution tank;
(5) adding the concentrated material in the crystallization raw liquid tank into a crystallization tank, and adding crystalline acetone into the crystallization tank; stirring the feed liquid in the crystallizing tank, controlling the feeding speed of the crystallized acetone to be 600-1000L/h, stopping feeding the crystallized acetone when the density of the feed liquid reaches 0.820-0.840 g/ml, continuing stirring for a period of time, stopping stirring and standing;
(6) carrying out vacuum filtration on the mixed solution in the crystallization tank, and separating the mother solution from the crystals;
(7) washing the crystal with three acetone water solutions with the density being more than, equal to or less than that of the mother solution in sequence, and then pumping to dry; then, washing the crystal with crystalline acetone, finally removing the washing liquid through vacuum filtration, and obtaining wet crystal after pumping;
(8) and (4) sieving the wet crystals by using a 80-mesh sieve, and drying undersize products to obtain dried cobamamide finished products.
Further, the concentration of the cyanocobalamin solution in the step (1) is more than 15 g/L.
Further, the concentration of the sodium borohydride solution in the step (2) is 42-52 g/L.
Further, the resolving agent in the step (4) is an acetone aqueous solution, and the concentration of the acetone aqueous solution is 0.940 +/-0.005 g/mL.
Further, in the step (5), the concentrated material and the crystallized acetone are filtered by a filter before being added into the crystallizing tank.
Further, in the step (5), the stirring is continued for 3 hours, and the standing time is more than 2 hours.
Further, in the step (7), the dosage of the three acetone aqueous solutions is controlled to be 100-150L, each acetone aqueous solution is washed twice, and crystals are fully stirred in the washing process.
Further, in the step (7), the vacuum filtration time is 6-8 hours, and the vacuum degree is kept to be more than 0.02MPa in the filtration process.
Further, the drying in the step (8) is specifically performed by:
and (3) putting the undersize into a double-cone dryer, starting the double-cone dryer, starting a hot water pump when the vacuum degree reaches 0.09-0.1 MPa, controlling the temperature at 55-70 ℃ and the drying time at 4-8 h, and cooling the material for 60-80 min after drying.
In the above examples, the specific values of the condition ranges have no substantial influence on the product yield of the process, and the end values and the middle values of the condition ranges can be used, which are not described herein again.
FIG. 1 shows a more specific process flow, which includes the following steps:
1. dissolving cyanocobalamine in water, and obtaining solution unit of about 15000 mu g/ml after conversion;
2. sodium borohydride is poured into about 50L of absolute ethyl alcohol;
3. slowly adding ATP (adenosine triphosphate) and controlling the pH value of the feed liquid to be 3.0-3.5;
4. reacting for more than 90 minutes, and centrifuging and filtering;
5. adsorbing the feed liquid on a macroporous resin column at the flow rate of 160-300L/h;
6. introducing 0.990 + -0.005 g/ml of spreading agent into the macroporous resin column to remove impurities, then introducing 0.940 + -0.005 g/ml of acetone aqueous solution, and collecting the eluted concentrated material into a crystallization raw liquid tank;
7. and (3) crystallization: pressing the crystallization stock solution into a crystallization tank through a filter, pressing the crystallization acetone into the crystallization tank through the filter, starting stirring, controlling the feeding speed of the crystallization acetone to be 600-1000L/h, stopping adding the acetone when the density of the feed liquid reaches 0.820-0.840 g/ml, continuing stirring for 3 hours, and stopping stirring and standing for more than 2 hours;
and (3) suction filtration: the mother liquor and crystals were separated by vacuum through the filter and the crystals were trapped in the filter bag of the filter.
8. And (3) washing crystals: washing the crystal with three acetone aqueous solutions with the density of 0.01g/ml or more and less than that of the mother liquor respectively, and finally washing with crystalline acetone, wherein the dosage of each detergent is controlled to be 100-150L, and washing is carried out twice; the crystals should be thoroughly stirred during the washing process and then drained. And (4) carrying out vacuum filtration to pump out residual washing liquor for 6-8 hours. Observing the vacuum degree at any time during the period of pumping out the residual washing liquid, keeping the vacuum degree to be more than or equal to 0.02MPa, and obtaining wet crystals after pumping out.
9. Sieving: the wet crystals are sieved (80 mesh).
10. And (3) drying: starting the double-cone dryer, starting a hot water pump when the vacuum degree reaches 0.09-0.1 MPa, controlling the temperature at 55-70 ℃ and the drying time at 4-8 h, cooling the material for 60-80 min after the drying and mixing process is finished, and taking out and weighing crystals.
11. Packaging: and packaging the product into a product with a fixed specification by using a double-layer plastic bag, verifying the product subjected to plastic packaging by using a metal detector, filling the product into an aluminum bottle after the product is verified to be qualified, pasting a label after a cover is pressed by a capping machine, and finishing packaging.
The method is simple and easy to implement, is suitable for large-scale production, and has high economic benefit. The yield of the method can reach more than 99 percent, is far higher than about 50 percent of the prior art, and has remarkable technical progress.
Claims (9)
1. A method for synthesizing cobamamide is characterized by comprising the following steps:
(1) dissolving cyanocobalamine in water to prepare a solution;
(2) pouring sodium borohydride into absolute ethyl alcohol to prepare a solution;
(3) adding the cyanocobalamine solution prepared in the step (1) and the sodium borohydride solution prepared in the step (2) into a reaction tank, enabling the weight ratio of the cyanocobalamine to the sodium borohydride in the solution to be 2.2-2.6: 1, slowly adding adenosine triphosphate into the reaction tank, controlling the pH of the feed liquid in the reaction tank to be 3.0-3.5, reacting for more than 90 minutes, and centrifugally filtering;
(4) adsorbing the filtrate obtained in the step (3) through a macroporous resin column at the flow rate of 160-300L/h, introducing a spreading agent into the macroporous resin column to remove impurities, introducing an analytic agent into the macroporous resin column, and collecting the eluted concentrated material into a crystallization stock solution tank;
(5) adding the concentrated material in the crystallization raw liquid tank into a crystallization tank, and adding crystalline acetone into the crystallization tank; stirring the feed liquid in the crystallizing tank, controlling the feeding speed of the crystallized acetone to be 600-1000L/h, stopping feeding the crystallized acetone when the density of the feed liquid reaches 0.820-0.840 g/ml, continuing stirring for a period of time, stopping stirring and standing;
(6) carrying out vacuum filtration on the mixed solution in the crystallization tank, and separating the mother solution from the crystals;
(7) washing the crystal with three acetone water solutions with the density being more than, equal to or less than that of the mother solution in sequence, and then pumping to dry; then, washing the crystal with crystalline acetone, finally removing the washing liquid through vacuum filtration, and obtaining wet crystal after pumping;
(8) and (4) sieving the wet crystals by using a 80-mesh sieve, and drying undersize products to obtain dried cobamamide finished products.
2. The method for synthesizing cobamamide according to claim 1, wherein the concentration of the solution of cyanocobalamin in the step (1) is more than 15 g/L.
3. The method for synthesizing cobamamide according to claim 1, wherein the concentration of the sodium borohydride solution in the step (2) is 42-52 g/L.
4. The method for synthesizing cobamamide according to claim 1, wherein the resolving agent in step (4) is acetone aqueous solution with concentration of 0.940 ± 0.005 g/mL.
5. The cobamamide synthesis process according to claim 1, wherein in said step (5), the concentrate and the crystallized acetone are filtered by a filter before being fed into the crystallization tank.
6. The method for synthesizing cobamamide according to claim 1, wherein in the step (5), the stirring is continued for 3 hours and the standing time is more than 2 hours.
7. The method for synthesizing cobamamide according to claim 1, wherein in the step (7), the dosage of all three acetone aqueous solutions is controlled to be 100-150L, each acetone aqueous solution is washed twice, and the crystals are fully turned over in the washing process.
8. The method for synthesizing cobamamide according to claim 1, wherein in the step (7), the time of vacuum filtration is 6-8 hours, and the vacuum degree is kept more than 0.02MPa in the process of filtration.
9. The cobamamide synthesis method according to claim 1, wherein the drying in step (8) is carried out in a specific manner:
and (3) putting the undersize into a double-cone dryer, starting the double-cone dryer, starting a hot water pump when the vacuum degree reaches 0.09-0.1 MPa, controlling the temperature at 55-70 ℃ and the drying time at 4-8 h, and cooling the material for 60-80 min after drying.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102391340A (en) * | 2011-10-31 | 2012-03-28 | 河北玉星生物工程有限公司 | Preparation method of mecobalamin |
| CN107698642A (en) * | 2017-10-09 | 2018-02-16 | 广州普星药业有限公司 | A kind of method for preparing Mecobalamin |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102391340A (en) * | 2011-10-31 | 2012-03-28 | 河北玉星生物工程有限公司 | Preparation method of mecobalamin |
| CN107698642A (en) * | 2017-10-09 | 2018-02-16 | 广州普星药业有限公司 | A kind of method for preparing Mecobalamin |
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| 权爽等: "甲钴胺的环境友好生产工艺过程研究", 《分析仪器》 * |
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