CN1054132C - Process for separating thymidine from mixture - Google Patents
Process for separating thymidine from mixture Download PDFInfo
- Publication number
- CN1054132C CN1054132C CN98110601A CN98110601A CN1054132C CN 1054132 C CN1054132 C CN 1054132C CN 98110601 A CN98110601 A CN 98110601A CN 98110601 A CN98110601 A CN 98110601A CN 1054132 C CN1054132 C CN 1054132C
- Authority
- CN
- China
- Prior art keywords
- thymidine
- exchange resin
- macropore
- post
- impurity
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Images
Landscapes
- Saccharide Compounds (AREA)
Abstract
The present invention belongs to the technical field of biologic chemical engineering, which relates to a method for separating thymidine from a mixture. In the method, a thymidine solution obtained by chemical synthesis and containing impurities flows through a weak acid cation exchange resin of a macroporous phenolic aldehyde series and an anion exchange resin of a macroporous weak alkali acrylic acid series in sequence, and a proper amount of borate is added into the solution to improve the purity of a thymidine product; a thymidine solution obtained in an enzymatic method can add a strong acidic cation exchange resin to well remove the impurities in the solution. The method has the advantages of simple process and high product yield, the purity can reach more than 99 %, and the requirements of industrial production can be satisfied.
Description
The invention belongs to technical field of biochemical industry, relate to a kind of from mixture the method for separating thymidine.
Thymidine, (code name dT) are a kind of important biochemical reagents, have very in the research of biological chemical field with in producing and use widely.In recent years, because AIDS spreads in the whole world, human life in serious threat, therefore the development research of AIDS resisting cytotoxic drug has obtained people's attention, thymidine is preparation AIDS resisting medicine---the important source material of Zidovodine (AZT), no matter as biochemical reagents still as the raw material of preparation Zidovodine (AZT), its content must be more than 98%, and the content of 5-methyluridine wherein can not surpass 0.3%.The preparation of thymidine at present mainly contains chemosynthesis and two kinds of methods of DNA enzymolysis, contain a large amount of impurity in the resulting thymidine crude: by containing oily matter in the resulting thymidine crude of chemical synthesis, pigment, thymidine derivatives such as 5-methyluridine, then contain Desoxyadenosine (dA) in the resulting thymidine crude of enzymolysis process, Deoxyribose cytidine (dC), deoxidation crow glycosides (dG), Hypoxanthine deoxyriboside impurity such as (dI), how from the mixture of thymidine, to isolate highly purified thymidine, the problem that then becomes many scientific and technical personnel and paid close attention to.
At present, the separation method commonly used of chemical synthesis gained thymidine crude is such: evaporation earlier removes desolvates, and adds methyl alcohol or dissolve with ethanol then, drips ethyl acetate, reflux, crystallisation by cooling, thus the thymidine product obtained, and this method is because periodic crystallisation, yield is low, shade deviation, the content height of 5-methyluridine, and crystalline mother solution can not reclaim; Document (E. Cha Jiafu, J.N. Da Dafusheng: " nucleic acid " the 1st volume, 19,279 pages) a kind of method of utilizing strong base anion exchange chromatography separating thymidine in the borate system disclosed, but this method can not be removed impurity and pigment, is unwell to suitability for industrialized production.Therefore, must research and develop a kind of high yield as early as possible, be suitable for the method for the separating thymidine of suitability for industrialized production, to satisfy industrial needs.
The objective of the invention is to disclose a kind of macropore weakly acidic cation-exchange resin and macroporous weakly basic anion exchange resin absorption and adding borate of adopting to remove the impurity in the thymidine solution, thereby obtain the method for high purity thymidine, in the hope of overcoming the existing in prior technology shortcoming, satisfy industrial needs, the content of gained thymidine of the present invention can reach more than 98%, and can be suitable for suitability for industrialized production.
Since the source difference of thymidine mixture, thereby the content of thymidine is also different in the mixture, the kind of impurity is also different, and according to different thymidine mixture and product requirements, the contriver proposes the design of following removal impurity according to a large amount of tests:
(1) for adopting chemical method synthetic thymidine mixture, be most of pigment and small amount of impurities in the weakly acidic cationic exchanger resin absorption mixing solutions with the macropore phenolic aldehyde earlier, and then with remaining a spot of pigment and a large amount of impurity in the macroreticular weakly base ion exchange resin absorption mixing solutions, as thymidine derivatives such as 5-methyluridines, add simultaneously a spot of borate in solution or during crystallization, make itself and a small amount of 5-methyluridine that residues in the solution form complex compound, increase its solubleness, the crystallization method by routine can obtain highly purified thymidine product;
(2) for the thymidine mixture that adopts the DNA enzymolysis process to obtain, then earlier remove most of dG with storng-acid cation exchange resin, dC, dI, impurity such as dA, and then obtain highly purified thymidine with the method for (1).
(3) for the thymidine mixture that only contains a small amount of 5-methyluridine, can directly add the small amount of boron hydrochlorate, reduce the content of 5-methyluridine in the product, the crystallization method by routine obtains highly purified thymidine product then.
It is to contain thymidine 50% in the raw material synthetic thymidine crude that the present invention also is achieved in that by the 5-methyluridine, 5-methyluridine 10% and other impurity, the thymidine crude solid that will contain impurity is dissolved in the deionized water, its aqueous solution is dark-brown, and oily matter and suspended substance arranged, it is the weakly acidic cationic exchanger resin post that this aqueous solution is flow through the macropore phenolic aldehyde with the flow velocity of 40～60ml/h, remove most oily matter, suspended substance, pigment and part thymidine derivative, Zeo-karb commonly used is commercially available HD-1,122,125 etc., its consumption is dT: resin=1: 6 (W: V); And then flow through macropore weak base acrylic acid type anion exchange resin post with the flow velocity of 40～60ml/h, remove remaining pigment and thymidine derivative, macropore weak base acrylic acid type anion exchange resin commonly used is commercially available D301, D309, D396, D351,709,710A, 710B, Amberlite IRA-63, IRA-93, IRA-94, IRA-943, Amberlyst-A-21, XE-1003, Diaon WA-20, WA-21, Lewatit MP-60, Duolite A-305 etc., and its consumption is dT: resin=1: 12 (W: V).Above-mentioned weakly acidic cationic exchanger resin post and weak basic anion exchange resin post are connected mutually, fuse.Series connection by effusive behind two posts be the colourless or faint yellow thymidine aqueous solution, the impurity of the overwhelming majority has been adsorbed to be removed, this aqueous solution adopts conventional concentrated, crystallization, drying means, can obtain content and be higher than 98% thymidine xln.In order to remove the 5-methyluridine in the solution effectively, can in the aqueous solution of thymidine crude or when crystallization, add the small amount of boron hydrochlorate, as sodium tetraborate etc., the purity of thymidine xln is reached more than 99%.The borate add-on is: the 5-methyluridine: borate=1: 0.1～1, the best are 1: 0.3～0.5 (mass ratio).Because weakly acidic cationic exchanger resin and weak basic anion exchange resin do not adsorb thymidine, so the rate of recovery of thymidine almost can reach 100%, and crystalline mother solution can be reclaimed by aforesaid method once more.
For the thymidine crude that is obtained by the DNA enzymolysis process, because of wherein containing dG, dC, dI, impurity such as dA, thus can make the aqueous solution of this thymidine crude pass through storng-acid cation exchange resin earlier, to remove dG wherein earlier, dC, dI, impurity such as dA, and then obtain highly purified thymidine xln with aforesaid method, the rate of recovery also can reach more than 95%, and storng-acid cation exchange resin commonly used is commercially available 732,001 * 7,001 * 6,001 * 8, Amberlite IR-12 etc.
For the thymidine that only contains a small amount of 5-methyluridine, then can directly add the small amount of boron hydrochlorate, as sodium tetraborate etc., the purity of thymidine xln is reached more than 99%.The borate add-on is: the 5-methyluridine: borate=1: 0 1～1, the best are 1: 0 3～0.5 (mass ratio).
Further illustrate content of the present invention below in conjunction with accompanying drawing.
Fig. 1 is the process flow diagram of the thymidine crude of separating the purification chemical synthesis and obtaining.
Wherein:
1---------macropore phenolic aldehyde is the weakly acidic cationic exchanger resin post
2---------macropore weak base acrylic acid type anion exchange resin posts
3---------concentration kettle
4---------crystallizers
5---------strainers
It is weakly acidic cationic exchanger resin post 1, macropore weak base acrylic acid type anion exchange resin post 2 that thymidine crude soluble in water or crystalline mother solution are flow through the macropore phenolic aldehyde according to the order of sequence, its flow rate control is about 40～60ml/h, the thymidine solution that flows out anion-exchange resin column enters concentration kettle 3 and concentrates, crystallization in crystallizer 4 can obtain highly purified thymidine crystal by strainer 5; Owing to concentrate, crystallization, be filtered into prior art, repeat no more herein.In order to obtain highly purified thymidine crystal, can be in solution or add the small amount of boron hydrochlorate during crystallization, general add-on is: the 5-methyluridine: borate=1: 0.1～1, the best are 1: 0.3～0.5.
Fig. 2 is the process flow diagram of the thymidine crude of separating the enzyme purification solution and obtaining.
------strong acid cation exchange resin column wherein: 6
With thymidine crude soluble in water or crystalline mother solution flow through strong acid cation exchange resin column 6 according to the order of sequence, the macropore phenolic aldehyde is weakly acidic cationic exchanger resin post 1, macropore weak base acrylic acid type anion exchange resin post 2, its flow rate control is about 40～60ml/h, the thymidine solution that flows out anion-exchange resin column enters concentration kettle and concentrates 3, crystallization in crystallizer 4 can obtain highly purified thymidine crystal by strainer 5.
Further illustrate content of the present invention below in conjunction with embodiment; because storng-acid cation exchange resin 7, macropore phenolic aldehyde are the of a great variety of weakly acidic cationic exchanger resin 1 and macropore weak base acrylic acid type anion exchange resin 2; relevant engineering technical personnel can be according to design disclosed in this invention and technical scheme; lift one and return the realization the present invention of three ground, so embodiment itself does not limit protection scope of the present invention.
The thymidine crude that 15 grams is contained 50% thymidine, 10%5-methyluridine and other impurity is dissolved in the 120ml deionized water, this thymidine solution is flow through HD-1 cation exchange resin column 1 and D315 anion-exchange resin column 2 with the flow velocity of 40ml/h, the consumption of HD-1 is 45ml, the diameter of post is 12mm, the length of post is 600mm, the consumption of D315 is 90ml, the diameter of post is 20mm, the length of post is 600mm, last sample finishes, add the 200ml deionized water again, the thymidine solution of staying in the post is discharged.Beginning effusive is the water of 100ml, follow effusive for the thymidine solution of 300ml, the rate of recovery is 95%, concentrate the back and add 0.4 gram sodium tetraborate, can get thymidine 4.5 grams after crystallization, filtration and the drying, content 99.2%, crystalline mother solution can reclaim thymidine 1.8 grams as stated above once more, and the secondary total yield is 84%.
Take by weighing 54.7 kilograms of crystalline mother solutions, wherein thymidine content is 19%, and 5-methyluridine content is 7%, adds 1 kilogram of Sodium Tetraborate in this solution, is diluted to 200L with deionized water.Flow velocity with 60L/h flow through HD-1 cation exchange resin column and D315 anion-exchange resin column then, the consumption of HD-1 is 90L, the diameter of post is 400mm, the length of post is 2000mm, the consumption of D315 is 120L, the diameter of post is 400mm, the length of post is 2000mm, and then adds deionized water, and the thymidine solution of staying in the post is discharged, collect the solution of discharging in the post, the thymidine rate of recovery is 99%, concentrates the back and adds 1 kilogram of sodium tetraborate, crystallization, can get 5 kilograms of thymidines after the drying, content 99.6%, crystalline mother solution can reclaim as stated above once more, and the secondary total yield is 70%.
Take by weighing thymidine solution 160ml by the preparation of DNA enzymolysis process, wherein contain 2 gram thymidine and dA, dC, dG, impurity such as dI, this solution is flow through 732 storng-acid cation exchange resin 30ml earlier, and then flow through HD-1 cation exchange resin column and D315 anion-exchange resin column according to the order of sequence with the flow velocity of 60L/h, the consumption of HD-1 is 30ml, the diameter of post is 12mm, the length of post is 600mm, the consumption of D315 is 120ml, the diameter of post is 12mm, the length of post is 60mm, and then adds deionized water, and the thymidine solution of staying in the post is discharged, collect the solution of discharging in the post, through concentrating, crystallization, can get thymidine 1.4 grams after the drying, content 98.2%, yield is 70%.
Adopt the raw material identical with embodiment 1, processing parameter and exchange column adopt 122 Zeo-karbs and D301 anionite-exchange resin, can get thymidine 6.1 and restrain content 99.3%, yield 81%.
Adopt the raw material identical with embodiment 1, processing parameter and exchange column adopt 125 Zeo-karbs and D709 anionite-exchange resin, can get thymidine 6.0 and restrain content 99.7%, yield 80%.
Adopt the raw material identical with embodiment 1, processing parameter and exchange column adopt HD-1 Zeo-karb and Amberlite IRA-63 anionite-exchange resin, can get thymidine 6.2 and restrain content 99.5%, yield 83%.
Embodiment 7
Adopt raw material, processing parameter and the exchange column identical, do not add borate during crystallization, can get thymidine 6.1 and restrain content 98.6%, yield 82.0% with embodiment 1.
Embodiment 8
Adopt raw material, processing parameter and the exchange column identical, do not add borate during crystallization, and in material solution, add sodium tetraborate 0.15 gram, can get thymidine 6.2 to restrain content 99.0%, yield 82.5% with embodiment 1.
Embodiment 9
Adopt raw material, processing parameter and the exchange column identical, add sodium tetraborate 1.5 grams during crystallization, can get thymidine 6.3 and restrain content 99.2%, yield 82.6% with embodiment 1.
Claims (5)
1. the method for a separating thymidine from mixture, constituted by the removal of impurity, concentrated (2), crystallization (3), (4) process of filtering, it is characterized in that the removal of impurity is performed such: the thymidine aqueous solution that will contain impurity flows through two placed in-line ion exchange resin columns according to the order of sequence, and said ion exchange resin column is weakly acidic cationic exchanger resin post (1) and macropore weak base acrylic acid type anion exchange resin post (2) for the macropore phenolic aldehyde.
2. according to the described method of claim 1, it is characterized in that: the thymidine aqueous solution that contains impurity at first flows through a strong acid cation exchange resin column (6), and then to flow through the macropore phenolic aldehyde be weakly acidic cationic exchanger resin post (1) and macropore weak base acrylic acid type anion exchange resin post (2).
3. according to the described method of claim 2, it is characterized in that: storng-acid cation exchange resin is 732 Zeo-karbs.
4. according to claim 1 or 2 described methods, it is characterized in that: said macropore phenolic aldehyde is that weakly acidic cationic exchanger resin is a kind of among the HD-1,122,125; Said macropore weak base acrylic acid type anion exchange resin is a kind of among D301, D309, D396, D315,709,710A, 710B, AmberliteIRA-63, IRA-93, IRA-94, IRA-943, Amberlyst-A-21, XE-1003, Diaon WA-20, WA-21, Lewatit MP-60, the Duolite A-305.
5. according to the described method of claim 1, add borate when it is characterized in that crystallization or in the aqueous solution of thymidine, said borate is a sodium tetraborate, makes sodium tetraborate and 5-methyluridine form complex compound, and add-on is: the 5-methyluridine: borate=1: 0.1～1 (mass ratio).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN98110601A CN1054132C (en) | 1998-01-05 | 1998-01-05 | Process for separating thymidine from mixture |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN98110601A CN1054132C (en) | 1998-01-05 | 1998-01-05 | Process for separating thymidine from mixture |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1190656A CN1190656A (en) | 1998-08-19 |
| CN1054132C true CN1054132C (en) | 2000-07-05 |
Family
ID=5220634
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN98110601A Expired - Fee Related CN1054132C (en) | 1998-01-05 | 1998-01-05 | Process for separating thymidine from mixture |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1054132C (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103435670B (en) * | 2013-08-26 | 2015-08-26 | 江苏诚意药业有限公司 | A kind of method reclaiming inosine from inosine mother liquor |
| CN105859809B (en) * | 2016-04-09 | 2018-12-04 | 南昌大学 | A method of extracting beta-thymidine from fermentation liquid |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1107847A (en) * | 1993-11-15 | 1995-09-06 | 布里斯托尔-迈尔斯斯奎布公司 | Process for preparing AZT and derivatives thereof |
| CN1107856A (en) * | 1993-11-15 | 1995-09-06 | 布里斯托尔-迈尔斯斯奎布公司 | Process for large-scale preparation of 2',3'-didehydro-2',3'-dideoxynucleosides |
-
1998
- 1998-01-05 CN CN98110601A patent/CN1054132C/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1107847A (en) * | 1993-11-15 | 1995-09-06 | 布里斯托尔-迈尔斯斯奎布公司 | Process for preparing AZT and derivatives thereof |
| CN1107856A (en) * | 1993-11-15 | 1995-09-06 | 布里斯托尔-迈尔斯斯奎布公司 | Process for large-scale preparation of 2',3'-didehydro-2',3'-dideoxynucleosides |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1190656A (en) | 1998-08-19 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN100338017C (en) | Method for preparing high-purity chlorogenic acid from honeysuckle crude extract | |
| CN1257182C (en) | Method for preparing enoxolone | |
| CN102220387B (en) | Method for extracting and purifying resveratrol and emodin from fresh giant knotweed | |
| CN102093236B (en) | Ornithine aspartate compound and new preparation method thereof | |
| CN1054132C (en) | Process for separating thymidine from mixture | |
| CN101759557A (en) | Preparation method of shikimic acid | |
| EP4215538A1 (en) | Method for purifying sucralose | |
| CN101096380B (en) | Method for purifying citicoline from biotransformation or multienzyme reaction liquid | |
| CN1081191C (en) | Method for separating D-ribose from fermentation liquor | |
| CN101066989A (en) | Process of separating and purifying glutathione from fermented liquid in a four-area simulated moving bed | |
| CN1067677C (en) | L-glutamine separating and purifying process | |
| CN1177859C (en) | Process for separating nucleotide from ribonucleic acid enzymolysis liquid using cationic exchanging resin | |
| CN100344608C (en) | Production of high-purity capsaicin crystal by ion exchange resin method | |
| FI115293B (en) | Separation of nucleosides using solid, cross-linked xylose isomerase | |
| CN1169817C (en) | Prepn of high-purity acephate powder (2) | |
| US20040082776A1 (en) | Process for recovery of uridine from molasses | |
| CN109796333B (en) | Method for extracting and purifying pravastatin sodium from fermentation liquor | |
| CN1286259A (en) | Process for separating nucleotide from enzymolyzed ribonucleic acid liquid | |
| CN1206009A (en) | Extraction of anhydro tetraodotoxin | |
| TWI670278B (en) | High purity low endotoxin carbohydrate (hple) compositions, and methods of isolation thereof | |
| SK69795A3 (en) | Method of producing alpha, alpha-trehaloze | |
| CN111067965B (en) | Preparation method of bacopa monnieri saponin | |
| CN103193837A (en) | Etimicin sulfate preparation method | |
| CN113621010A (en) | Method for separating and extracting cytidine from fermentation liquor | |
| WO2003014135A1 (en) | Method for purification of acarbose |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C06 | Publication | ||
| PB01 | Publication | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| C19 | Lapse of patent right due to non-payment of the annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |