CA1315464C - Polymers containing thiol groups, a process for their preparation, and their use - Google Patents
Polymers containing thiol groups, a process for their preparation, and their useInfo
- Publication number
- CA1315464C CA1315464C CA000573912A CA573912A CA1315464C CA 1315464 C CA1315464 C CA 1315464C CA 000573912 A CA000573912 A CA 000573912A CA 573912 A CA573912 A CA 573912A CA 1315464 C CA1315464 C CA 1315464C
- Authority
- CA
- Canada
- Prior art keywords
- polymer
- groups
- water
- hydroxyl groups
- polymer containing
- 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
Classifications
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08F—MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS
- C08F8/00—Chemical modification by after-treatment
- C08F8/34—Introducing sulfur atoms or sulfur-containing groups
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08F—MACROMOLECULAR COMPOUNDS OBTAINED BY REACTIONS ONLY INVOLVING CARBON-TO-CARBON UNSATURATED BONDS
- C08F8/00—Chemical modification by after-treatment
- C08F8/08—Epoxidation
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N11/00—Carrier-bound or immobilised enzymes; Carrier-bound or immobilised microbial cells; Preparation thereof
- C12N11/02—Enzymes or microbial cells immobilised on or in an organic carrier
- C12N11/08—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a synthetic polymer
- C12N11/082—Enzymes or microbial cells immobilised on or in an organic carrier the carrier being a synthetic polymer obtained by reactions only involving carbon-to-carbon unsaturated bonds
- C12N11/087—Acrylic polymers
Abstract
Abstract of the Disclosure Polymers containing thiol groups are described which can be obtained by introducing oxirane groups into polymers containing hydroxyl groups and treating the product with a salt of hydrogen sulfide.
Description
BEHRING~ERKE AKTIEN~ESELLSC~AFT 87/~ OZ9 - Ma ~4Z
Dr. Ha/Sd.
Polymers containing thiol groups, a process fGr their preparation, and the_r use The invention relates to polymers containing thiol groups, a process for their preparation, and their use.
Polymeric carrier ~aterials which are derivatized by mercapto groups are of great interest since, on the one hand, substances which usually interact with sulfhydryl groups are purified using them and, on the other hand, substance~ can be irreversibly i~mobilized on sulfhydryl groups. Such substances can be, for example, enzymes, antibodiesO proenzymes or antigens. Hitherto, deriva-tives of agarose, in particular, have been employed as the carrier matrix. During the preparation of these poLymeric carriers, the chemical introduction of the sulfhydryl functions deserves particular attention.
Polymer derivatizations using homocystein thiolactone, glutathione coupling reactions with prior syanogen bro-mide activation, and activation using epichlorohydrin and subsequent reaction with th;osulfate along ~ith a reduction step, and reaction of an oxirane group with dithiothreitol are known and thus correspond to the prior art (R. Axén et al., Acta Chemica Scan. ~ 29, 471-474, 1975).
Ho~ever, these thiol-containing polymers and their pre-paration processes are afflicted ~ith disadvantages and are thus capable of improvement. The reasons for this are, on the one hand, in the matrix, which, due to the carbohydrate structure, is mechanically sensitive and can aLso be at~acked by microbes, and, on the other hand, are in the process for introduction of the sulfhydryL group, which is either expensive or inconvenient, and in some cases also includes the production of unstable bonds.
~P
1 ~3 154 64 The object of the inven~ion ~as thus to develop a process which allo~s polymers containing thiol groups to be pre-par~d simply and economicalLy. In addition, the condi-tion must be fulfilled that ~he chemical bonds produced are distinguished by high stability.
The object of the invention was also to prepare thiol-containing polymers which, as 3 consequence of the matrix empLoyed for derivatization, are distinguished by superior mechanical and chemical stabilityn Surprisingly, it has now been found that polymers which were derivatized using oxirane groups reacted with water-soluble salts of hydrogen sulfide to form immobilized sulfhydryl functions.
The invention relates to a polymer containing thiol groups Mhich is obtainable by introducing oxirane groups into a water-insol~ble polymer containing'hydroxyl groups and treating the polymer containing oxirane groups with a ~ater-soluble salt of hydrogen sulfide, preferably an alkali metal salt or alkaline earth metal salt.
Suitable polymers containing hydroxyl groups are carbo-hydrate-based polymers, for example agarose gels or dextran gels, or polymers based on methacrylamide, N-methylene-bis-methacrylamide, glycidyl methacry(ate/
polyethylene glycol derivatives and pentaerythritol dimethacrylate.
Polymers which, as a consequence of their chemical nature, are kno~n as copolymers and cons~ructed from acrylic acid der;vatives or methacrylic acid derivatives ancl are provided with oxirane groups are suitable in a preferred manner for the preparation of polymers containing thiol groups. The monomer components of such copolymers are, for example~ me~hacrylam;de, N-methyLene-bis-methacryl-amide, glycidyl methacrylates, glycidyl meehcrylate/polyethylene glycol derivatives or pentaerythritol methacrylic acid derivatives. In particular, resins which are commercially available under the names RFractogel or RSepabeads are suitable.
S The hydroxyl groups of these water-insoluble resi~s are reacted by known methods, for example in accordance with EP-A-0,203,463, ~ith 1-chloro-2,3-epoxypropane (epi-chLorohydrin) or with 1,4-bis(2,3-epoxypropoxy)butane, and oxirane groups are thus introduced into the polymers.
The epoxidization is carried out, for example, by sus-pending 1 kg of water-moist polymer in 1,5D0 ~l of water and 650 ml of 2 N sodium hydroxide solu~ion and reacting the suspension with 200 ml of ep;chlorohydrin at eleva-ted temperature, for example 40-50C. After 1-4 hours, the polymer is washed ~ith water and sucked dry.
The oxirane-containing polymer is then taken up in water or an aqueous buffer solution, 500 ml to 3 l, preferably about 1 l, usually being added per kg of resin. 9uffers ~hich can be employed are salt solut;ons, for example sodium phosphate buffer, sodium c;trate buffer or HEPES
buffer, having a pH of 4 to p~ 13. 1 liter of a 0.25 ~olar sodium hydrogen carbonate solut;on is preferably added per kg of poly~er~
10-200 g of a ~ater-soluble sulfide or hydrogen sulfide, preferably an alkali metal sulfide, are then added.
Particularly preferably, 70 80 9 of sodium sulfide x 9H20 ar~ added, and the batch is stirred at a tempera-ture of 0-70GC, preferably 35-1l5C. The reaction time chosen is 2 to 4R hours, preferably 12-14 hours. The polymer ;s then washed ~ith water and/or O.S ml/100 ml strength acet;c acid.
The invention also relates to a process for the prepara-tion of a ~olymer con~aining thiol groups, wherein a ~ater-insoluble polymer containing hydroxyl groups is reacted for one to four hours at 40-50C w;th l-chloro-2,3-epoxypropane (epichlorohydrin) or 1,4-bist2,3-epoxypro-po~y)butane, whereby oxirane grsups are introduced into the polymer, the polymer is washed with ~ater, taken up in aqueous solution and treated for 2 to 48 hours at a tPmperature of 0-70C with a water-soluble sulfide, and the polymer is washed with water or dilute acid.
A pre~erred polymer according to the invention is on~
~hich can be obtained by introducing oxirane groups, by reaction ~ith epichlorohydrin, into a copoLymer prepared from pentaerythritol, methacrylic acid derivatives and polyethylene glycol and which is crosslinked with divinylbenzene and treating ~ith sodium suLf;de.
A further preferred polymer according to the invention is one which can be obtained by introducing oxirane groups ;nto a polymer based on methacrylic acid deriva-tives (RSepabeads) by reaction with butanediol digly-cidyl ether, and treating with sodium sulfide.
The polymeric carriers which are provided~with thiol functions by the above process are very particularly preferably suitable, due to their mechanical and chemical stability, for immobilization of enzymes, proenzymes, antibodies or antigens and for chromatography of sub-stances which interact w;th mercapto groups.
The thiol-containing polymers prepared ;n this way are furthermore suitable, a~ter prior act;vation using a di-sulf;de, for reaction ~ith thiol-containing substances, for example proteins, ~ith disulfide exchange. Such pro-3D teins ca~ thereby be concentrated or eluted from thepolymer and thus purified. ParticuLarly suitable di-sulfides for activation of polymeric mercapto groups are, for example, 2,2'-dipyridyL disulfide, bis-(2-pyridyl-N-oxide) disuLfide and 5,5'-dithiobis-(2-nitrobenzoic acid)~
A polymer according to the invention can preferably be used for purification of alpha1-antitrypsin.
The examples below illustrate the invention.
Exa~ple 1 _ _ Preparation of a thiol-containing polymer based on RFractogel 1,000 ml of RFractogel H~-65 (F), a copolymer prepared fro~ pentaerythritol, methacrylic acid derivatives and poLyethylene glycol (manufacturer: Toyo Soda~, were treated with 350 ml of ~ater and 275 ml of 5 N sodium ~ hydroxide solution, and the ~ixture was warmed to 45C.
; 200 ml of epichlorohydrin ~ere added, and the mixture was stirred at this temperature for 2 hours. The mixture was fil~ered and washed w;th ~ater until the pH had dropped to 7. The suction-dried resin was taken up in 1,400 ml of 0.2 molar sodium hydrogen carbonate solution and treated with 72 9 of NazS x 9H20. The batch was stirred a~ 40C for 15 hours, filtered under suction and washed thoroughly with water and subsecluently with 0.5 ml/100 ml strength acetic acid.
xample 2 Preparation of a thiol-containing polymer based on RSepabeads (manufacturer: Mitsubishi) 1,000 ml of RSepabeads HG05 were suspended in 1,000 ml of 0.6 N NaOH containing 2 9 of Na8H4. 1,000 ml of butanediol diglycidyl ether were subsequently added, and the suspension was st;rred at room temperature for 8 hours~ The epoxy-activated RSepabeads WG05 material was subsequently washed with 15 L of water on a glass frit.
The reaction ~ith Na2S x 9H20 takes place as described in Example l.
1 3 1 546~
Example 3 Activation of thiol-containing RFractogel using 2,2'-dithiob;s(pyridine N-oxide) 500 ml of thiol-containing RFractogel H~-65, which was prepared as described in Example 1, ~as equilibrated with a buffer A containing 50 mmol/l of tris/HCl and 150 mmol/l of NaCl, pH 7.5. The resin material ~as subsequently incubated 4 ti~es for 320 minutes at room temperature ~ith 1 l of buffer A ~ 1.5 mmol/l of 2,Z'-dithiobis-(pyridine N-oxide) in each case and then again washed with buffer Ao The resin material activated in this ~ay can now be used~ for example, for disulfide exchange reactions.
Example 4 Activation of thiol-containing RSepabeads HG05 using di-(2-pyridyl) disulfide 500 ml of thiol-containing RSepabeads HG05, which were prepared as described in Example 2, were activated using 1.5 mmol/L of di-(2-pyridyl) disulfide under the condi-: 20 tions of Example 3.
Example S
Purification of alpha1-antitrypsin 500 ml of human plasma ~ere diluted ~ith 1,000 ml of water containing 30 mmol/l of mercaptoethanol, and the mixture was incubated at room temperature for 1 hour and subsequently subjected to fractional ammonium sulfate precipitation between 50% saturation (~/v) and 75g saturation ~/v?. The 50%-75X (w/v) residue was dis-solved in a solution containing 50 mmol/l of Tris, 200 mmol/l of NaCl and 1 mmol/l of EDTA, pH 8.0, and 1~1546~
chromatographed over a 500 ~l RACA 202 column. The fractions containing alpha1-antitrypsin were purified and transferred to a th;ol-RFractogel column (100 ml) activated using 2,2'-dithiobis(pyridine N-oxide). The S column was washed with the same buffer, and alpha1-antitrypsin was eluted by means of a solution containing 10 mmol/l of dithiothreitol~ The alphal-antitrypsin thus obtained was substantially freed from impurities.
Dr. Ha/Sd.
Polymers containing thiol groups, a process fGr their preparation, and the_r use The invention relates to polymers containing thiol groups, a process for their preparation, and their use.
Polymeric carrier ~aterials which are derivatized by mercapto groups are of great interest since, on the one hand, substances which usually interact with sulfhydryl groups are purified using them and, on the other hand, substance~ can be irreversibly i~mobilized on sulfhydryl groups. Such substances can be, for example, enzymes, antibodiesO proenzymes or antigens. Hitherto, deriva-tives of agarose, in particular, have been employed as the carrier matrix. During the preparation of these poLymeric carriers, the chemical introduction of the sulfhydryl functions deserves particular attention.
Polymer derivatizations using homocystein thiolactone, glutathione coupling reactions with prior syanogen bro-mide activation, and activation using epichlorohydrin and subsequent reaction with th;osulfate along ~ith a reduction step, and reaction of an oxirane group with dithiothreitol are known and thus correspond to the prior art (R. Axén et al., Acta Chemica Scan. ~ 29, 471-474, 1975).
Ho~ever, these thiol-containing polymers and their pre-paration processes are afflicted ~ith disadvantages and are thus capable of improvement. The reasons for this are, on the one hand, in the matrix, which, due to the carbohydrate structure, is mechanically sensitive and can aLso be at~acked by microbes, and, on the other hand, are in the process for introduction of the sulfhydryL group, which is either expensive or inconvenient, and in some cases also includes the production of unstable bonds.
~P
1 ~3 154 64 The object of the inven~ion ~as thus to develop a process which allo~s polymers containing thiol groups to be pre-par~d simply and economicalLy. In addition, the condi-tion must be fulfilled that ~he chemical bonds produced are distinguished by high stability.
The object of the invention was also to prepare thiol-containing polymers which, as 3 consequence of the matrix empLoyed for derivatization, are distinguished by superior mechanical and chemical stabilityn Surprisingly, it has now been found that polymers which were derivatized using oxirane groups reacted with water-soluble salts of hydrogen sulfide to form immobilized sulfhydryl functions.
The invention relates to a polymer containing thiol groups Mhich is obtainable by introducing oxirane groups into a water-insol~ble polymer containing'hydroxyl groups and treating the polymer containing oxirane groups with a ~ater-soluble salt of hydrogen sulfide, preferably an alkali metal salt or alkaline earth metal salt.
Suitable polymers containing hydroxyl groups are carbo-hydrate-based polymers, for example agarose gels or dextran gels, or polymers based on methacrylamide, N-methylene-bis-methacrylamide, glycidyl methacry(ate/
polyethylene glycol derivatives and pentaerythritol dimethacrylate.
Polymers which, as a consequence of their chemical nature, are kno~n as copolymers and cons~ructed from acrylic acid der;vatives or methacrylic acid derivatives ancl are provided with oxirane groups are suitable in a preferred manner for the preparation of polymers containing thiol groups. The monomer components of such copolymers are, for example~ me~hacrylam;de, N-methyLene-bis-methacryl-amide, glycidyl methacrylates, glycidyl meehcrylate/polyethylene glycol derivatives or pentaerythritol methacrylic acid derivatives. In particular, resins which are commercially available under the names RFractogel or RSepabeads are suitable.
S The hydroxyl groups of these water-insoluble resi~s are reacted by known methods, for example in accordance with EP-A-0,203,463, ~ith 1-chloro-2,3-epoxypropane (epi-chLorohydrin) or with 1,4-bis(2,3-epoxypropoxy)butane, and oxirane groups are thus introduced into the polymers.
The epoxidization is carried out, for example, by sus-pending 1 kg of water-moist polymer in 1,5D0 ~l of water and 650 ml of 2 N sodium hydroxide solu~ion and reacting the suspension with 200 ml of ep;chlorohydrin at eleva-ted temperature, for example 40-50C. After 1-4 hours, the polymer is washed ~ith water and sucked dry.
The oxirane-containing polymer is then taken up in water or an aqueous buffer solution, 500 ml to 3 l, preferably about 1 l, usually being added per kg of resin. 9uffers ~hich can be employed are salt solut;ons, for example sodium phosphate buffer, sodium c;trate buffer or HEPES
buffer, having a pH of 4 to p~ 13. 1 liter of a 0.25 ~olar sodium hydrogen carbonate solut;on is preferably added per kg of poly~er~
10-200 g of a ~ater-soluble sulfide or hydrogen sulfide, preferably an alkali metal sulfide, are then added.
Particularly preferably, 70 80 9 of sodium sulfide x 9H20 ar~ added, and the batch is stirred at a tempera-ture of 0-70GC, preferably 35-1l5C. The reaction time chosen is 2 to 4R hours, preferably 12-14 hours. The polymer ;s then washed ~ith water and/or O.S ml/100 ml strength acet;c acid.
The invention also relates to a process for the prepara-tion of a ~olymer con~aining thiol groups, wherein a ~ater-insoluble polymer containing hydroxyl groups is reacted for one to four hours at 40-50C w;th l-chloro-2,3-epoxypropane (epichlorohydrin) or 1,4-bist2,3-epoxypro-po~y)butane, whereby oxirane grsups are introduced into the polymer, the polymer is washed with ~ater, taken up in aqueous solution and treated for 2 to 48 hours at a tPmperature of 0-70C with a water-soluble sulfide, and the polymer is washed with water or dilute acid.
A pre~erred polymer according to the invention is on~
~hich can be obtained by introducing oxirane groups, by reaction ~ith epichlorohydrin, into a copoLymer prepared from pentaerythritol, methacrylic acid derivatives and polyethylene glycol and which is crosslinked with divinylbenzene and treating ~ith sodium suLf;de.
A further preferred polymer according to the invention is one which can be obtained by introducing oxirane groups ;nto a polymer based on methacrylic acid deriva-tives (RSepabeads) by reaction with butanediol digly-cidyl ether, and treating with sodium sulfide.
The polymeric carriers which are provided~with thiol functions by the above process are very particularly preferably suitable, due to their mechanical and chemical stability, for immobilization of enzymes, proenzymes, antibodies or antigens and for chromatography of sub-stances which interact w;th mercapto groups.
The thiol-containing polymers prepared ;n this way are furthermore suitable, a~ter prior act;vation using a di-sulf;de, for reaction ~ith thiol-containing substances, for example proteins, ~ith disulfide exchange. Such pro-3D teins ca~ thereby be concentrated or eluted from thepolymer and thus purified. ParticuLarly suitable di-sulfides for activation of polymeric mercapto groups are, for example, 2,2'-dipyridyL disulfide, bis-(2-pyridyl-N-oxide) disuLfide and 5,5'-dithiobis-(2-nitrobenzoic acid)~
A polymer according to the invention can preferably be used for purification of alpha1-antitrypsin.
The examples below illustrate the invention.
Exa~ple 1 _ _ Preparation of a thiol-containing polymer based on RFractogel 1,000 ml of RFractogel H~-65 (F), a copolymer prepared fro~ pentaerythritol, methacrylic acid derivatives and poLyethylene glycol (manufacturer: Toyo Soda~, were treated with 350 ml of ~ater and 275 ml of 5 N sodium ~ hydroxide solution, and the ~ixture was warmed to 45C.
; 200 ml of epichlorohydrin ~ere added, and the mixture was stirred at this temperature for 2 hours. The mixture was fil~ered and washed w;th ~ater until the pH had dropped to 7. The suction-dried resin was taken up in 1,400 ml of 0.2 molar sodium hydrogen carbonate solution and treated with 72 9 of NazS x 9H20. The batch was stirred a~ 40C for 15 hours, filtered under suction and washed thoroughly with water and subsecluently with 0.5 ml/100 ml strength acetic acid.
xample 2 Preparation of a thiol-containing polymer based on RSepabeads (manufacturer: Mitsubishi) 1,000 ml of RSepabeads HG05 were suspended in 1,000 ml of 0.6 N NaOH containing 2 9 of Na8H4. 1,000 ml of butanediol diglycidyl ether were subsequently added, and the suspension was st;rred at room temperature for 8 hours~ The epoxy-activated RSepabeads WG05 material was subsequently washed with 15 L of water on a glass frit.
The reaction ~ith Na2S x 9H20 takes place as described in Example l.
1 3 1 546~
Example 3 Activation of thiol-containing RFractogel using 2,2'-dithiob;s(pyridine N-oxide) 500 ml of thiol-containing RFractogel H~-65, which was prepared as described in Example 1, ~as equilibrated with a buffer A containing 50 mmol/l of tris/HCl and 150 mmol/l of NaCl, pH 7.5. The resin material ~as subsequently incubated 4 ti~es for 320 minutes at room temperature ~ith 1 l of buffer A ~ 1.5 mmol/l of 2,Z'-dithiobis-(pyridine N-oxide) in each case and then again washed with buffer Ao The resin material activated in this ~ay can now be used~ for example, for disulfide exchange reactions.
Example 4 Activation of thiol-containing RSepabeads HG05 using di-(2-pyridyl) disulfide 500 ml of thiol-containing RSepabeads HG05, which were prepared as described in Example 2, were activated using 1.5 mmol/L of di-(2-pyridyl) disulfide under the condi-: 20 tions of Example 3.
Example S
Purification of alpha1-antitrypsin 500 ml of human plasma ~ere diluted ~ith 1,000 ml of water containing 30 mmol/l of mercaptoethanol, and the mixture was incubated at room temperature for 1 hour and subsequently subjected to fractional ammonium sulfate precipitation between 50% saturation (~/v) and 75g saturation ~/v?. The 50%-75X (w/v) residue was dis-solved in a solution containing 50 mmol/l of Tris, 200 mmol/l of NaCl and 1 mmol/l of EDTA, pH 8.0, and 1~1546~
chromatographed over a 500 ~l RACA 202 column. The fractions containing alpha1-antitrypsin were purified and transferred to a th;ol-RFractogel column (100 ml) activated using 2,2'-dithiobis(pyridine N-oxide). The S column was washed with the same buffer, and alpha1-antitrypsin was eluted by means of a solution containing 10 mmol/l of dithiothreitol~ The alphal-antitrypsin thus obtained was substantially freed from impurities.
Claims (11)
1. A polymer containing thiol groups which is obtainable by introducing oxirane groups into a water-insoluble polymer containing hydroxyl groups and treating the polymer containing oxirane groups with a water-soluble salt of hydrogen sulfide.
2. A Polymer as claimed in claim 1 wherein the polymer containing hydroxyl groups is a carbohydrate-based polymer or a polymer based on methacrylamide N-methylene-bis-methacrylamide glycidyl methacrylate/
polyethylene glycol derivatives or pentaerythritol dimethacrylate.
polyethylene glycol derivatives or pentaerythritol dimethacrylate.
3. A polymer as claimed in claim 1 wherein the polymer containing hydroxyl groups is an agarose gel.
4. A polymer as claimed in claim 1 wherein the salt of hydrogen sulfide is an alkali metal salt or an alkaline earth metal salt.
5. A polymer as claimed in claim 1 wherein the polymer containing oxirane groups contains acrylic acid derivatives or methacrylic acid derivatives.
6. A polymer as claimed in claim 1 wherein the water-insoluble polymer containing hydroxyl groups is a copolymer prepared from pentaerythritol methacrylic acid derivatives and polyethylene glycol and which is crosslinked with divinylbenzene whereon the oxirane groups are introduced by reacting this copolymer with epichlorohydrin, and Wherein it is treated with sodium sulfide.
7. A polymer as claimed in claim 1 wherein the water-insoluble polymer containing hydroxyl groups is a polymer based on methacrylic acid derivatives (RSepabeads), wherein the oxirane groups are intro-duced by reacting this polymer with butanediol di-glycidyl ether, and wherein it is treated with sodium sulfide.
8. A process for the preparation of a polymer containing thiol groups, wherein a water-insoluble polymer con-taining hydroxyl groups is reacted for one to four hours at 40-50°C with 1-chloro-2,3-epoxypropane (epichlorohydrin) or 1,4-bis(2,3-epoxypropoxy)butane, whereby oxirane groups are introduced into the poly-mer, the polymer is washed with water, taken up in aqueous solution and treated for 2 to 48 hours at a temperature of 3-70°C with a water-soluble sulfide, and the polymer is washed with water or dilute acid.
9. The use of a polymer as claimed in claim 1 for immobilization of enzymes, proenzymes, antibodies or antigens and for chromatography of substances which interact with mercapto groups.
10. The use of a polymer as claimed in claim 1, after activation using a disulfide, for binding substances containing thiol groups.
11. The use of a polymer as claimed in claim 1 for purification of alpha1-antitrypsin.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| DE19873726454 DE3726454A1 (en) | 1987-08-08 | 1987-08-08 | POLYMERS CONTAINING THIOL GROUPS, METHOD FOR THE PRODUCTION THEREOF AND THEIR USE |
| DEP3726454.0 | 1987-08-08 |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CA1315464C true CA1315464C (en) | 1993-03-30 |
Family
ID=6333374
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000573912A Expired - Fee Related CA1315464C (en) | 1987-08-08 | 1988-08-05 | Polymers containing thiol groups, a process for their preparation, and their use |
Country Status (12)
| Country | Link |
|---|---|
| EP (1) | EP0305749B1 (en) |
| JP (1) | JP2703937B2 (en) |
| KR (1) | KR0131766B1 (en) |
| AT (1) | ATE67509T1 (en) |
| AU (1) | AU616898B2 (en) |
| CA (1) | CA1315464C (en) |
| DE (2) | DE3726454A1 (en) |
| DK (1) | DK439988A (en) |
| ES (1) | ES2025743T3 (en) |
| FI (1) | FI90779C (en) |
| GR (1) | GR3003361T3 (en) |
| PT (1) | PT88191B (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004092223A1 (en) * | 2003-04-17 | 2004-10-28 | Ultraceuticals R & D Pty Limited | Cross-linked polysaccharide composition |
| CN103709280A (en) * | 2009-09-30 | 2014-04-09 | 蒂奥迈里克斯研究与指导有限公司 | Mucoadhesive polymers having vitamin B partialstructures |
Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB9007384D0 (en) * | 1990-04-02 | 1990-05-30 | Duncan Ruth | Coupling between polymers and other organic molecular entities utilising thiol-specific reactive groups |
| DE4106984A1 (en) * | 1991-03-05 | 1992-09-10 | Bio Rad Lab Gmbh | METHOD FOR SEPARATING SPECIES CONTAINING PRIMARY AMINO GROUPS FROM SOLUTIONS |
| US5710211A (en) * | 1995-08-01 | 1998-01-20 | Kuraray Co., Ltd. | Process for producing vinyl alcohol polymer |
| CA2369869C (en) * | 1999-04-02 | 2008-06-10 | Ajinomoto Co., Inc. | Process for producing subunit peptide originating in polymer protein |
| DE10004565A1 (en) * | 2000-02-02 | 2001-08-09 | Merck Patent Gmbh | Alkaline-stable coating on substrates |
| CN102532372B (en) * | 2010-12-17 | 2015-06-10 | 通用电气公司 | Polymer and preparation and using method thereof |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE1495319A1 (en) * | 1963-08-28 | 1969-08-28 | Huels Chemische Werke Ag | Process for the production of thermosetting resins |
| JPS4867382A (en) * | 1971-12-06 | 1973-09-14 | ||
| US3900451A (en) * | 1973-08-24 | 1975-08-19 | Dow Chemical Co | Method of making sulfhydryl-containing polymers |
| BE821435A (en) * | 1974-10-24 | 1975-02-17 | PROCESS FOR PREPARING COPOLYMERS. | |
| DE2732301C3 (en) * | 1977-07-16 | 1980-12-11 | Roehm Gmbh, 6100 Darmstadt | Process for the production of stabilized carrier-bound proteins |
| US4565854A (en) * | 1983-04-07 | 1986-01-21 | Kuraray Co., Ltd. | Polymer having thiol end group |
-
1987
- 1987-08-08 DE DE19873726454 patent/DE3726454A1/en not_active Withdrawn
-
1988
- 1988-08-02 AT AT88112531T patent/ATE67509T1/en not_active IP Right Cessation
- 1988-08-02 ES ES198888112531T patent/ES2025743T3/en not_active Expired - Lifetime
- 1988-08-02 EP EP88112531A patent/EP0305749B1/en not_active Expired - Lifetime
- 1988-08-02 DE DE8888112531T patent/DE3864958D1/en not_active Expired - Fee Related
- 1988-08-04 FI FI883648A patent/FI90779C/en not_active IP Right Cessation
- 1988-08-04 PT PT88191A patent/PT88191B/en not_active IP Right Cessation
- 1988-08-05 DK DK439988A patent/DK439988A/en not_active Application Discontinuation
- 1988-08-05 JP JP63194740A patent/JP2703937B2/en not_active Expired - Fee Related
- 1988-08-05 CA CA000573912A patent/CA1315464C/en not_active Expired - Fee Related
- 1988-08-05 AU AU20431/88A patent/AU616898B2/en not_active Ceased
- 1988-08-08 KR KR1019880010085A patent/KR0131766B1/en not_active IP Right Cessation
-
1991
- 1991-12-17 GR GR91401549T patent/GR3003361T3/en unknown
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2004092223A1 (en) * | 2003-04-17 | 2004-10-28 | Ultraceuticals R & D Pty Limited | Cross-linked polysaccharide composition |
| CN103709280A (en) * | 2009-09-30 | 2014-04-09 | 蒂奥迈里克斯研究与指导有限公司 | Mucoadhesive polymers having vitamin B partialstructures |
| CN103709280B (en) * | 2009-09-30 | 2016-03-23 | 蒂奥迈里克斯研究与指导有限公司 | Mucoadhesive polymers containing vitamins B substructure |
Also Published As
| Publication number | Publication date |
|---|---|
| EP0305749A1 (en) | 1989-03-08 |
| FI90779C (en) | 1994-03-25 |
| ATE67509T1 (en) | 1991-10-15 |
| JPS6466213A (en) | 1989-03-13 |
| DE3864958D1 (en) | 1991-10-24 |
| DK439988D0 (en) | 1988-08-05 |
| KR890003842A (en) | 1989-04-18 |
| FI883648A (en) | 1989-02-09 |
| DK439988A (en) | 1989-02-09 |
| JP2703937B2 (en) | 1998-01-26 |
| EP0305749B1 (en) | 1991-09-18 |
| FI90779B (en) | 1993-12-15 |
| DE3726454A1 (en) | 1989-02-16 |
| PT88191A (en) | 1989-06-30 |
| ES2025743T3 (en) | 1992-04-01 |
| FI883648A0 (en) | 1988-08-04 |
| KR0131766B1 (en) | 1998-04-13 |
| PT88191B (en) | 1995-03-01 |
| GR3003361T3 (en) | 1993-02-17 |
| AU616898B2 (en) | 1991-11-14 |
| AU2043188A (en) | 1989-02-09 |
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| MKLA | Lapsed |