CN110898014A - Preparation method of polyuridylic acid - Google Patents
Preparation method of polyuridylic acid Download PDFInfo
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- CN110898014A CN110898014A CN201911250698.1A CN201911250698A CN110898014A CN 110898014 A CN110898014 A CN 110898014A CN 201911250698 A CN201911250698 A CN 201911250698A CN 110898014 A CN110898014 A CN 110898014A
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- polyglandular
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- 239000002253 acid Substances 0.000 title claims abstract description 47
- 238000002360 preparation method Methods 0.000 title abstract description 19
- 239000003381 stabilizer Substances 0.000 claims abstract description 37
- 238000002156 mixing Methods 0.000 claims abstract description 33
- 239000012295 chemical reaction liquid Substances 0.000 claims abstract description 28
- 108091034057 RNA (poly(A)) Proteins 0.000 claims abstract description 23
- 239000003960 organic solvent Substances 0.000 claims abstract description 23
- 238000001035 drying Methods 0.000 claims abstract description 22
- 239000002244 precipitate Substances 0.000 claims abstract description 21
- DJJCXFVJDGTHFX-UHFFFAOYSA-N Uridinemonophosphate Natural products OC1C(O)C(COP(O)(O)=O)OC1N1C(=O)NC(=O)C=C1 DJJCXFVJDGTHFX-UHFFFAOYSA-N 0.000 claims abstract description 20
- FOGRQMPFHUHIGU-UHFFFAOYSA-N Uridylic acid Natural products OC1C(OP(O)(O)=O)C(CO)OC1N1C(=O)NC(=O)C=C1 FOGRQMPFHUHIGU-UHFFFAOYSA-N 0.000 claims abstract description 20
- DJJCXFVJDGTHFX-XVFCMESISA-N uridine 5'-monophosphate Chemical compound O[C@@H]1[C@H](O)[C@@H](COP(O)(O)=O)O[C@H]1N1C(=O)NC(=O)C=C1 DJJCXFVJDGTHFX-XVFCMESISA-N 0.000 claims abstract description 20
- 239000000843 powder Substances 0.000 claims abstract description 19
- 210000002700 urine Anatomy 0.000 claims abstract description 15
- 238000001816 cooling Methods 0.000 claims abstract description 11
- 239000008055 phosphate buffer solution Substances 0.000 claims abstract description 11
- 238000000034 method Methods 0.000 claims abstract description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 22
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 20
- 239000000203 mixture Substances 0.000 claims description 19
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 claims description 15
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 14
- 108010039918 Polylysine Proteins 0.000 claims description 14
- BDERNNFJNOPAEC-UHFFFAOYSA-N propan-1-ol Chemical compound CCCO BDERNNFJNOPAEC-UHFFFAOYSA-N 0.000 claims description 14
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 12
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 claims description 10
- 239000001110 calcium chloride Substances 0.000 claims description 10
- 229910001628 calcium chloride Inorganic materials 0.000 claims description 10
- 239000004202 carbamide Substances 0.000 claims description 10
- RQFQJYYMBWVMQG-IXDPLRRUSA-N chitotriose Chemical compound O[C@@H]1[C@@H](N)[C@H](O)O[C@H](CO)[C@H]1O[C@H]1[C@H](N)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)[C@@H](CO)O1 RQFQJYYMBWVMQG-IXDPLRRUSA-N 0.000 claims description 9
- 230000001376 precipitating effect Effects 0.000 claims description 9
- 239000004227 calcium gluconate Substances 0.000 claims description 8
- 229960004494 calcium gluconate Drugs 0.000 claims description 8
- 235000013927 calcium gluconate Nutrition 0.000 claims description 8
- NEEHYRZPVYRGPP-UHFFFAOYSA-L calcium;2,3,4,5,6-pentahydroxyhexanoate Chemical compound [Ca+2].OCC(O)C(O)C(O)C(O)C([O-])=O.OCC(O)C(O)C(O)C(O)C([O-])=O NEEHYRZPVYRGPP-UHFFFAOYSA-L 0.000 claims description 8
- 229930024421 Adenine Natural products 0.000 claims description 7
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 claims description 7
- 229960000643 adenine Drugs 0.000 claims description 7
- 229930027917 kanamycin Natural products 0.000 claims description 7
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 claims description 7
- 229960000318 kanamycin Drugs 0.000 claims description 7
- 229930182823 kanamycin A Natural products 0.000 claims description 7
- 230000000694 effects Effects 0.000 abstract description 12
- 239000000243 solution Substances 0.000 abstract description 7
- 238000001556 precipitation Methods 0.000 abstract description 4
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- 230000001766 physiological effect Effects 0.000 abstract description 3
- 230000008569 process Effects 0.000 abstract description 3
- 231100000331 toxic Toxicity 0.000 abstract description 3
- 230000002588 toxic effect Effects 0.000 abstract description 3
- 229940115272 polyinosinic:polycytidylic acid Drugs 0.000 abstract description 2
- 150000001875 compounds Chemical class 0.000 description 6
- 239000003814 drug Substances 0.000 description 4
- 230000002708 enhancing effect Effects 0.000 description 4
- 230000036737 immune function Effects 0.000 description 4
- 230000036039 immunity Effects 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 3
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- 150000001768 cations Chemical class 0.000 description 3
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- 102000014150 Interferons Human genes 0.000 description 2
- 108010050904 Interferons Proteins 0.000 description 2
- 241001465754 Metazoa Species 0.000 description 2
- 208000004880 Polyuria Diseases 0.000 description 2
- ISAKRJDGNUQOIC-UHFFFAOYSA-N Uracil Chemical compound O=C1C=CNC(=O)N1 ISAKRJDGNUQOIC-UHFFFAOYSA-N 0.000 description 2
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 description 2
- 230000015556 catabolic process Effects 0.000 description 2
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- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 229940079322 interferon Drugs 0.000 description 2
- 239000002799 interferon inducing agent Substances 0.000 description 2
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- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 229920002477 rna polymer Polymers 0.000 description 2
- 201000010153 skin papilloma Diseases 0.000 description 2
- AUHDWARTFSKSAC-HEIFUQTGSA-N (2S,3R,4S,5R)-3,4-dihydroxy-5-(hydroxymethyl)-2-(6-oxo-1H-purin-9-yl)oxolane-2-carboxylic acid Chemical compound [C@]1([C@H](O)[C@H](O)[C@@H](CO)O1)(N1C=NC=2C(O)=NC=NC12)C(=O)O AUHDWARTFSKSAC-HEIFUQTGSA-N 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 206010019799 Hepatitis viral Diseases 0.000 description 1
- XQFRJNBWHJMXHO-RRKCRQDMSA-N IDUR Chemical compound C1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C(I)=C1 XQFRJNBWHJMXHO-RRKCRQDMSA-N 0.000 description 1
- 241000934136 Verruca Species 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 208000000260 Warts Diseases 0.000 description 1
- 238000009098 adjuvant therapy Methods 0.000 description 1
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- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 210000002421 cell wall Anatomy 0.000 description 1
- 230000001684 chronic effect Effects 0.000 description 1
- 201000004196 common wart Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 210000003494 hepatocyte Anatomy 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000008799 immune stress Effects 0.000 description 1
- 239000000568 immunological adjuvant Substances 0.000 description 1
- 230000000091 immunopotentiator Effects 0.000 description 1
- 229940028843 inosinic acid Drugs 0.000 description 1
- 239000004245 inosinic acid Substances 0.000 description 1
- 206010023332 keratitis Diseases 0.000 description 1
- 210000005229 liver cell Anatomy 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 125000003588 lysine group Chemical group [H]N([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 230000000149 penetrating effect Effects 0.000 description 1
- 210000001539 phagocyte Anatomy 0.000 description 1
- 229920000656 polylysine Polymers 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 229940035893 uracil Drugs 0.000 description 1
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 description 1
- 229940045145 uridine Drugs 0.000 description 1
- 229960005486 vaccine Drugs 0.000 description 1
- 201000001862 viral hepatitis Diseases 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
- A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
- A61K9/146—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with organic macromolecular compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/7088—Compounds having three or more nucleosides or nucleotides
- A61K31/713—Double-stranded nucleic acids or oligonucleotides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K9/00—Medicinal preparations characterised by special physical form
- A61K9/14—Particulate form, e.g. powders, Processes for size reducing of pure drugs or the resulting products, Pure drug nanoparticles
- A61K9/141—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers
- A61K9/145—Intimate drug-carrier mixtures characterised by the carrier, e.g. ordered mixtures, adsorbates, solid solutions, eutectica, co-dried, co-solubilised, co-kneaded, co-milled, co-ground products, co-precipitates, co-evaporates, co-extrudates, co-melts; Drug nanoparticles with adsorbed surface modifiers with organic compounds
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
- A61P37/04—Immunostimulants
Abstract
The invention discloses a preparation method of poly (adenine-urea) nucleotide, which comprises the following steps: A. respectively dissolving poly (A) and poly (uridylic acid) with phosphate buffer solution, mixing, adding stabilizer, mixing, and keeping temperature at 40-80 deg.C; B. naturally cooling the reaction liquid obtained in the step A, mixing the reaction liquid with an organic solvent, standing for precipitation, and drying the precipitate to obtain the double-stranded polyuridylic acid; the invention provides a preparation method of double-chain polyglandular urine dry powder, the prepared polyinosinic-polycytidylic acid dry powder has complete double-spiral structural characteristics and physiological activity, the quality and the stability are much better than those of polyglandular uridylic acid in a solution state, and the polyglandular urine which is exposed in a living body has stronger enzymolysis resistance, so that the curative effect is enhanced. And the double-chain polyglandular urine solution is purified in the process of preparing the dry powder, so that a plurality of small molecular substances and impurities are removed, the toxic and side effects are reduced, and the quality of the double-chain polyglandular urine acid is improved.
Description
Technical Field
The invention relates to the technical field of polyuridylic acid preparation, in particular to a preparation method of polyuridylic acid.
Background
Polyuridylic acid, alternative name: polyadenylic acid uridine, double-chain polyglandular urine and PolyA: U (polyadenogenic-polymeric acid) are high-efficiency interferon inducers. Polyadenylic acid is double-stranded ribonucleic acid formed by pairing artificially synthesized Poly A (polyadenosine acid, Poly A) and Poly uracil (Polyuridinic acid, Poly U). The product is an artificially synthesized interferon inducer, and has the similar action of enhancing the immune function of lymphocytes to that of poly-inosinic acid. Can efficiently induce the action of interferon, inhibit the replication of infected viruses, strengthen the activity of phagocytes, improve the immune function of organisms, promote the non-specific immune function and certain specific immune functions of human bodies, and achieve the effects of resisting hepatocyte necrosis, resisting tumors and the like. Can be used for adjuvant treatment of viral infectious diseases and tumor, and is clinically suitable for chronic viral hepatitis, herpes, verruca plana, verruca vulgaris, viral keratitis, etc.
The double-stranded polyinosinic-polycytidylic acid is artificially synthesized double-stranded ribonucleic acid, can induce to generate interferon, and has broad-spectrum antiviral effect; in addition, the medicine also has the functions of regulating the immunity of the organism, enhancing the nonspecific immunity function and certain specific immunity function of the human body, achieving the effects of resisting the necrosis of liver cells, resisting tumors and the like. In addition, the double-chain polyuria is also an animal immunopotentiator, can be used as an auxiliary medicine for treating diseases, enhances the immunity of animal bodies and improves the curative effect of the medicine. Meanwhile, the double-chain polyuria is also an immunologic adjuvant, and can be used while being immunized by the vaccine, so that the immunologic effect can be obviously improved, and the immune stress and other viral diseases can be prevented.
Epsilon-polylysine (. epsilon. -PL) is a homo-type monomer polymer containing 25 to 30 lysine residues and was discovered by Japanese scholars in 1977. The epsilon-polylysine has good bactericidal ability and thermal stability. The action mechanism of epsilon-polylysine is mainly shown in acting on cell walls and cell membrane systems; genetic material or genetic microparticulate structure, and enzymes or functional proteins.
Because epsilon-polylysine is rich in cations, has strong electrostatic force with substances with anions and has good penetrating power to biological membranes, the polylysine can be used as a carrier of certain medicines, and thus is widely applied to the aspects of medical treatment and pharmacy.
Disclosure of Invention
The present invention is directed to a method for preparing poly (adenine dinucleotide), which solves the above problems of the background art.
In order to achieve the purpose, the invention provides the following technical scheme: a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 40-80 deg.C for 0.5-5 hr;
B. and (C) naturally cooling the reaction liquid in the step (A), mixing the reaction liquid with an organic solvent with the volume of 0.5-20 times, standing and precipitating for 0.5-72 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
Preferably, the stabilizing agent in the step A is one or a mixture of more of epsilon-polylysine, kanamycin, chitosan oligosaccharide, calcium chloride and calcium gluconate.
Preferably, the mass ratio of the poly A to the poly A is 1: 0.5-1.5, and the mass of the stabilizer is 0.2-2 times of the sum of the mass of polyinosinic acid and polycytidylic acid.
Preferably, the organic solvent in step B is one or more of ethanol, methanol, acetone, diethyl ether, isopropanol and n-propanol.
Preferably, in the step B, the precipitate is dried in a drying oven at the temperature of 20-60 ℃ for 15-25 min.
Compared with the prior art, the invention has the beneficial effects that:
(1) the invention provides a preparation method of double-stranded polyuridylic acid dry powder, which is simple in preparation process, polyadenylic acid and polyuridylic acid are polymerized by adopting a base complementary pairing principle and adding a stabilizer, and then the polyuridylic acid dry powder prepared by precipitation with an organic solvent has complete double-spiral structural characteristics and physiological activity.
(2) The double-chain poly (adenine dinucleotide) is introduced with stabilizers such as epsilon-polylysine, kanamycin, chitosan oligosaccharide, calcium chloride, calcium gluconate and the like to form a compound with good hydrophilicity; the mutual compound form of the double-stranded polyuridylic acid and the stabilizing agent above the cation ensures that the compound has stronger biological enzyme degradation resistance effect than the single naked double-stranded polyuridylic acid, thereby enhancing the curative effect.
(3) The double-stranded polyuridylic acid solution is purified in the process of preparing the dry powder from the double-stranded polyuridylic acid solution, so that a plurality of small molecular substances and impurities are removed, the toxic and side effects are reduced, and the quality of the double-stranded polyuridylic acid is improved.
Detailed Description
The technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
The first embodiment is as follows:
the invention provides the following technical scheme: a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 40 deg.C for 0.5 hr;
B. and (C) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with 0.5-time volume of organic solvent, standing for precipitation for 0.5 hour, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer used in step a was a mixture of epsilon-polylysine and calcium gluconate.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 0.5, the mass of the stabilizer is 0.2 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol and methanol.
In this example, the precipitate in step B was dried in a drying oven at 20 ℃ for 25 min.
Example two:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 80 deg.C for 5 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume 20 times that of the reaction liquid, standing and precipitating for 72 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer in step A is a mixture of kanamycin, chitosan oligosaccharide and calcium chloride.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1.5, the mass of the stabilizer is 2 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol and acetone.
In this example, the precipitate in step B was dried in a drying oven at 60 ℃ for 15 min.
Example three:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 45 deg.C for 1 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume of 1 time, standing and precipitating for 7 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer used in step A was a mixture of epsilon-polylysine and kanamycin.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1, the mass of the stabilizer is 0.5 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol, isopropanol, and n-propanol.
In this example, the precipitate in step B was dried in a drying oven at 30 deg.C for 17 min.
Example four:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 75 deg.C for 0.8 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume being 18 times that of the reaction liquid, standing and precipitating for 60 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer used in step a was a mixture of epsilon-polylysine, calcium chloride, and calcium gluconate.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 0.9, the mass of the stabilizer is 0.8 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol, diethyl ether and n-propanol.
In this example, the precipitate in step B was dried in a drying oven at 55 deg.C for 22 min.
Example five:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 65 deg.C for 4 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume 17 times that of the reaction liquid, standing and precipitating for 60 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer in step a is a mixture of chitosan oligosaccharide and calcium chloride.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1.2, the mass of the stabilizer is 1.7 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this example, the organic solvent in step B is a mixture of ethyl ether, isopropyl alcohol, and n-propyl alcohol.
In this example, the precipitate in step B was dried in a drying oven at 40 ℃ for 17 min.
Example six:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 68 deg.C for 3.2 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume 14 times that of the reaction liquid, standing and precipitating for 20 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer used in step a was a mixture of epsilon-polylysine, chitosan oligosaccharide, and calcium chloride.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1.4, the mass of the stabilizer is 1.4 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of acetone, diethyl ether and isopropanol.
In this example, the precipitate in step B was dried in a drying oven at 35 ℃ for 16 min.
Example seven:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 62 deg.C for 3.2 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume 13 times that of the reaction liquid, standing and precipitating for 33 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this embodiment, the stabilizer in step a is a mixture of chitosan oligosaccharide, calcium chloride, and calcium gluconate.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1.2, the mass of the stabilizer is 1.3 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol, diethyl ether, isopropanol, and n-propanol.
In this example, the precipitate in step B was dried in a drying oven at 48 ℃ for 17 min.
Example eight:
a preparation method of poly (adenine-urea) comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 60 deg.C for 3 hr;
B. and (3) naturally cooling the reaction liquid obtained in the step (A), mixing the reaction liquid with an organic solvent with the volume 10 times that of the reaction liquid, standing and precipitating for 35 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
In this example, the stabilizer in step a was a mixture of epsilon-polylysine, kanamycin, chitosan oligosaccharide, calcium chloride, and calcium gluconate.
In this example, the mass ratio of poly (A) to poly (uridylic acid) is 1: 1, the mass of the stabilizer is 1.1 times of the sum of the masses of polyinosinic acid and polycytidylic acid.
In this embodiment, the organic solvent in step B is a mixture of ethanol, methanol, acetone, diethyl ether, isopropanol, and n-propanol.
In this example, the precipitate in step B was dried in a drying oven at 40 ℃ for 20 min.
In conclusion, the invention provides a preparation method of double-stranded polyuridylic acid dry powder, the preparation process is simple, polyadenylic acid and polyuridylic acid are polymerized by adding a stabilizer by adopting a base complementary pairing principle, and then the polyuridylic acid dry powder prepared by precipitation by using an organic solvent has complete double-helix structural characteristics and physiological activity; the double-chain poly (adenine dinucleotide) is introduced with stabilizers such as epsilon-polylysine, kanamycin, chitosan oligosaccharide, calcium chloride, calcium gluconate and the like to form a compound with good hydrophilicity; the mutual compound form of the double-chain polyuridylic acid and the stabilizing agent above the cation ensures that the compound has stronger biological enzyme degradation resistance effect than the single naked double-chain polyuridylic acid, thereby enhancing the curative effect; the double-stranded polyuridylic acid solution is purified in the process of preparing the dry powder from the double-stranded polyuridylic acid solution, so that a plurality of small molecular substances and impurities are removed, the toxic and side effects are reduced, and the quality of the double-stranded polyuridylic acid is improved.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (5)
1. A method for preparing poly (adenine-urea) acid, which is characterized in that: the method comprises the following steps:
A. respectively dissolving poly (A) and poly (uridylic acid) with appropriate amount of phosphate buffer solution with pH of 6.0-8.0, mixing, adding stabilizer, mixing, and keeping temperature at 40-80 deg.C for 0.5-5 hr;
B. and (C) naturally cooling the reaction liquid in the step (A), mixing the reaction liquid with an organic solvent with the volume of 0.5-20 times, standing and precipitating for 0.5-72 hours, and drying the precipitate to obtain the double-chain polyglandular urine dry powder.
2. The method according to claim 1, wherein the step of preparing poly (adenine) comprises: and the stabilizer in the step A is one or a mixture of more of epsilon-polylysine, kanamycin, chitosan oligosaccharide, calcium chloride and calcium gluconate.
3. The method according to claim 1, wherein the step of preparing poly (adenine) comprises: the mass ratio of the poly A to the poly A is 1: 0.5-1.5, and the mass of the stabilizer is 0.2-2 times of the sum of the mass of polyinosinic acid and polycytidylic acid.
4. The method according to claim 1, wherein the step of preparing poly (adenine) comprises: and the organic solvent in the step B is one or a mixture of more of ethanol, methanol, acetone, diethyl ether, isopropanol and n-propanol.
5. The method according to claim 1, wherein the step of preparing poly (adenine) comprises: and B, drying the precipitate in a drying oven at the temperature of 20-60 ℃ for 15-25 min.
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