CH636628A5 - Process for the preparation of biologically active compounds, and the use thereof - Google Patents

Description

The invention relates to a process for the production of new carboxyls, which are often contained in these natural products.

Compounds of biologically active substances and water groups, because of their non-specific affinities, are perceived as disturbingly insoluble high-molecular compounds, their use. In addition, these substances have a low thermal and chemical stability relative to carrying out immune reactions in vitro and agents.

contain these connections. Many of these disadvantages could be overcome by using

In recent years, a new technique within lyvinylene glycol, a synthetic polymer, as a template of the biochemical working methods has been more and more eliminated. Polyvinylene glycol, also polyhydric

sets, whose primary characteristic is the affinity of 15 oximethylene, is produced by acidic or basic hydrolysis of carrier-bound, biologically active substances to selectively from polyvinylene carbonate. To insert each carbon atom to be carried out reactions. carries a hydroxyl group and a hydrogen atom. By-

Through a specific complex formation of the C-C bond bound to the carrier, the carrier gives the carrier a special substance with a second substance, which is present in a mixture, with high stability.

the substance in question can be removed from the mixture and 20 It has now been found that, instead of polyvinylene glycol, if desired, vinylene glycol copolymers are then advantageously isolated as a carrier matrix by desorption

will. can turn. By polymerizing suitable como

Carrier-bound enzymes have the advantage that substance monomers in polyvinylene carbonate can do the physical and

Changes in preparative, continuous processes due to chemical properties of the polyvinyl

feed and to get the reaction products enzyme-free. 25 glycols can be varied widely.

In biochemical, enzymatic analysis, the “hydrophilicity” or “swelling” of the poly-bound water-insoluble enzymes as multi-use vinylene glycol is available by polymerizing in hydrophobic mobile reagents. nomers, such as ethylene, vinyl chloride or styrene in the polyviny- due to the property of the enzymes, affinities cannot be changed to lencarbonate. Due to the copolymerization of only against the substrates, but also against the 30 vinyl acetate, to show a vinylene glycol-specific inhibitors in the saponified end product, the extraction group has been replaced by a vinyl alcohol group, which increases the enzyme inhibitors with the help of affinity chromatography to swellability in an aqueous medium Consequence. proven particularly favorable on carrier-bound enzymes. It is possible to use comonomers such as e.g. On the other hand, the binding of inhibitors to water-ethylene glycol divinyl ether and divinylbenzene allows crosslinking soluble matrices to prepare the corresponding preparations and thus improve the mechanical properties of the enzymes. of the wearer. By polymerizing electro-as so-called immunoadsorbents, antigens or pure functions, e.g. Oxirane groups (epoxy groups) from Vi antibodies bound to water-insoluble matrices and erlaunylglycidyl ether comonomers, a direct implementation of the subsequent isolation of the corresponding antibody matrix with nucleophilic functions (-NH2, -OH etc.) is possible or. Antigens. 40 lent. Carboxyl groups can be introduced by acrylic acid ester-Comono-As biologically active substances according to the mere. After copolymerization, the previous statements in vivo and in vitro are effective and the ester groups are saponified to carboxyl groups. The introduction of natural and artificially produced substances understood in the form of -NH2 or -COOH groups allows the broadest sense of activity as enzymes, activators, inhibitors, antigens with the aid of carbodiimides, isoxazolium salts, glutaral or antibodies, vitamins and hormones are called 45 dehyd etc. copolymers-vinylene glycol / vinyl alcohol. For these biologically active substances, since they represent a higher specific surface area and thus a higher binding principle of action of the water-insoluble systems, there is capacity for e.g. Proteins were introduced to the surface of the poly-the term effectors. merpulvers compared to vinylene glycol homopolymer.

Most of the carrier-bound Ef objects of the invention described so far are accordingly methods for fectors are much more stable than the corresponding biolo production of biologically active compounds from a gisch active substances in solution. preferably water-insoluble copolymer of polyvinylene. Advantageous glycols and a substance chemically, while maintaining their biologically compatible substances, are used as carrier materials, so-called matrices, which, in addition to being insoluble, see the activity of a biologically active substance. In the aqueous systems, the lowest possible non-specific adene compounds are the carrier matrix have a polyvinylene glycol sorption. For this purpose, hydrophobic, hydrophilic and 55 copolymer containing preferably up to 45%, preferably ionic interactions between the matrix and the react 5-20% of a comonomer. The biologically active substitution partners of the effector are largely prevented. Zen are substances such as Enzymes, activators, inhibitors, anti-mit substances whose binding to the effector is undesirable or antibodies, other plasma proteins, blood groups, e.g. Those that are not specific reactants of the substance, phythaemagglutinins, antibiotics, vitamins or effector should be excluded from binding. 60 hormones, peptides or amino acids or synthetically produced

The matrices used so far as carriers for biological effectors.

Active substances can be subdivided into those that bind the polymeric carrier and the biologically active substance to the effectors through physical adsorption. For this purpose, polystyrene, activated carbon and glass beads and (spacers) are covalently linked to one another either directly or via a side chain .

those linked to the effector via a covalent linkage. Binding the latter (spacer) is advantageous if the molecular weights

Vinyl polymers as homopolymers and copolymers, e.g. Polyacrylics of their affine partners are very different, or if, in the case of acids, polyacrylic acid amides and amino, carboxy or sulfo, the use in a biospecific process is very highly

5

636 628

molecular proteins or proteins made up of several subunits. Spacers are side chains of a certain length anchored to the polymer matrix, which are obtained by polymerizing in, by attachment or incorporation (binding of bi- or polyfunctional compounds - for example a tripeptide - to the matrix) or by incorporation of functional groups and gradual extension on them can.

The invention relates to methods for the covalent connection of the biologically active substances to the carrier.

A number of generally known reactions are available for this.

Ri "Ri

H? C = C "" H, C = C; '

3rd

They initially relate to the processes for producing the polyvinylene glycol copolymer. They are e.g. characterized in that vinylene carbonate with comonomers of the formulas:

H, C = C.

R,

h2c = c:

H, C = C '

R]

COOR,

"D-CO-CH,

\

Cl

10th

H2C = C.

Ri

(0CH2-CH2) n-0-CH = CH2

-CH = CH2

Ri / Ri h2c = c ^ ch2 h2c = c /

\ (CH2) "\ (CH2) n-CH-CH2

II \ /

oc ch2 °

Ri ^

h2c = c ^ h2c = c;

0-CH-CH2 CH-CH2

V 1 1

u o o

\ /

c

II

o in which Rj is hydrogen, methyl, ethyl, R3 is methyl, ethyl, by polymerizing monomers with epoxy-pyl and n are an integer between 1 and 4, polymeric groups, e.g. with epoxyalkyl vinyl ethers in the Polyvinylencar-siert. bonat and due to the very large differences in verse

The polymerization of the polyvinylene carbonate takes place between cyclic carbonate and epoxy trains with up to 45% comonomers, particularly preferably with 5 4oxy groups, a polyvinylene glycol matrix of up to 20% is easily obtained. with reactive epoxy groups with nucleophilic functions

The production of the biologically active compound can be implemented. The direct binding of a bio, for example, is possible in that an electrophilic group of logically active substance and / or the introduction (extension — neither into the biologically active substance or into the polyvinylation) of a side chain (spacer) is possible.

The glycol copolymer is introduced and via this the bio-45 Another advantage is the use of gisch active substance with the polyvinylene glycol copolymers also the possibility of direct introduction is implemented. (Polymerization) of electrophilic functions, the one

Allow the introduction of the electrophilic groups in the carrier implementation by known methods.

In general, matrix is carried out either by copolymerization. Comonomers are obtained by copolymerizing vinylene carbonate with ethyl or acrylate of vinylene carbonate with electrophilic groups and subsequent saponification of the polymers, or by reacting the support matrix with ak-a matrix with -OH and -COOH functions. The activating, low molecular weight carboxyl groups carrying electrophilic groups can be activated with the aid of carbodiimides. and then an amide bond with amino groups of

Effector. Carboxyl groups also allow the formation. Generally, the chemical coupling of a reaction bond of amide bonds with the help of Woodward's own to the support matrix is simple if, on the one hand, isoxazolium salts are added.

Cleophile and on the other electrophilic functions exist- Another method for coupling the biologically active ones. Amino-sulfhydryl and hydroxyl groups, which with the aid of N-ethoxycarbonyl-2-ethoxy 1,2-dihydroquinoline, are usually used either in the matrix or in the reactant 60, as reactive nucleophilic functions before compounds on carboxyl-containing polymers (EEDQ) executed.

are already included. Electrophilic functions must first be carried out. The carboxyl groups can be carried out by known methods. For this purpose, carboxal groups can be esterified in acid haloesters, then the ester can be converted into the hydrazide and genide, acid azides, acid anhydrides, imidazolides, and finally the resulting azide can be activated with the ami or directly with carbodiimides. Isocyanate, isothiocyanate, 65 trjx will also be linked as a reactive group of the effector protein with the polyvinylene glycol Ma electrophilic groups.

Diazonium groups or cyclic imidocarbonate esters possess the comonomers polymerized into the matrix. The introduction of reactive groups by subsequent reactions of freely available amino groups using copolymerization is particularly favorable. pen, so with vinyl sulfonate containing arylamino groups

636 628 6

derivatives or sulfuric acid esters of ß-hydroxyethyl sulfones and then egg- with correspondingly reactive groups

the introduction of arylamino groups is possible, whereby the nes effector can be connected, e.g. Arylamino groups then diazo by known methods

K

C —C

OH OH

H H

C - C - C-

K

n

OH H 0 H

CH.

HO-CH 1

CH.

d.

NH-C CH2) 6-NH-CH2-CH2-S02-

W //

• ivî.

An extension of a spa can also be carried out in this way.

Another way of binding proteins via the amino groups is with the help of glutardialdehyde.

It is simple to link the effector to the matrix after activating the hydroxyl or amino groups of the polyvinylene glycol copolymer by means of cyanogen halides, advantageously with cyanogen bromide and subsequently reacting the biologically active effectors containing amino groups via these activated groups.

A further possibility for linking the effector to a polyvinylene glycol or polyvinylene glycol copolyme risat matrix is to acylate the hydroxyl groups with bromoacetyl bromide, followed by alkylating the amino group of the effector.

Similar reaction processes can be achieved, for example, by reacting the hydroxyl groups of the support with reactive triazines, some of the reactive groups of the triazine reacting with the polyvinylene glycol compound and some with amino groups of the effector.

Diazotizable aromatic amines, which on the one hand can connect to the hydroxyl groups of the carrier via a further reactive group, on the other hand, allow coupling with enabled, activated amino acids, for example the tyrosine or histidine residues of the protein detector.

Arylamino group-containing vinylsulfone derivatives and sulfuric acid half-esters of β-hydroxyethylsulfones can be reacted with the hydroxyl groups of the support. They enable the binding of the effector after the above-described diazotization reaction.

Particularly stable ether bonds are formed when the hydroxyl groups of the polyvinylene glycol copoly merisate are reacted with non-ion-forming epoxides which contain at least 2 reactive groups, such as the epihalohydrins or the polyepoxides, for example epichlorohydrin or bisepoxides.

Another type of modification is the copolymerization of comonomers into the polyvinylene carbonate chain (such as 35 e.g. of vinyl pyrrolidone) with subsequent partial reaction of the cyclocarbonate rings of the polyvinylene carbonate with an amine, e.g. with hexamethylenediamine, to a polyvinylene carbonate copolymer substituted by a urethane bond with a spacer or effector. The remaining Zyklocar-40 bonate groups are then saponified to hydroxyl groups:

Polyvinylene carbonate

H

.N

iX

OH OH

n-1

CH-CH -

I I

OH 0 I

C = 0 I

NH

i

R

-CH-

0 0

\ /

C II

0

- GH-

I.

OH n-1

-CH -.

I.

0

1

C = 0 I

NH I

R

Saponification lead and thereby establish the covalent connection between the two; so the well-known implementation with complex-form the metal compounds, e.g. Titanium compounds. All of these methods are known to the person skilled in the art.

When binding low-molecular biologically active compounds, it is often advantageous to activate the effector rather than the carrier.

fi0 In principle, all methods can be used that are known in macromolecular chemistry for the modification of synthetic or natural macromolecular compounds.

The polyvinylene glycol copolymers stand out alongside

In addition to the methods exemplified here for producing the covalent connection between the carrier matrix 65 of chemical and thermal stability by means of advantageous and the protein effector or other effectors, there are processing properties and thus lead to still further methods which lead to a reaction of the hydroxyl groups of superiority over the hitherto found as optimal or certain groups of the copolymer with an effective carrier matrix based on natural carbohydrates

636 628

They are e.g. Can be produced in the form of fibers, threads, foils or spherical particles, so that the most suitable shape can be selected depending on the application of the effector to be bound to it. These copolymers are preferably used in the form of finely divided powders with a high specific surface area.

The size of the surface accessible for the binding of the effectors or the spacers, which can be controlled in terms of production technology, shows a further advantage of the polyvinylene glycol copolymers over the known carrier materials. 10th

The new biologically active compounds are suitable for most application processes which have hitherto become known for other effectors bound to hydrophilic, water-insoluble carriers. Accordingly, enzymes can be made water-insoluble. The insoluble enzymes are increasingly used to determine substrates in automatic analyzers and as so-called enzyme electrodes. Because of the increased stability, a number of carrier-bound enzymes are suitable for carrying out technical enzymatic reactions. 20th

Carrier-bound, biologically active substances have found widespread use in affinity chromatography because of their property as specific adsorbents. With the help of carrier-bound natural or synthetic enzyme inhibitors, the high purification of enzymes is possible, while the enzymes have been found to be extremely suitable as effectors, in particular for obtaining natural enzyme inhibitors from crude extracts. Carrier-bound water-insoluble antigens can be used to isolate the associated antibodies, which are thus free of other serum components and free of other antigens. Affinity chromatography can also be used to isolate and quantitate antibodies that cannot be precipitated and those that cannot be precipitated in the serum due to their low concentration. 3S

Examples

Production of vinylene carbonate copolymers

The vinylene carbonate used in the preparation of the copolymers (CP) was refluxed for one hour over sodium borohydride (100 parts by weight of vinylene carbonate to 2 parts by weight of NaBH4) at a pressure of 33 mm, then at 75733 mm over a 50 cm long , distilled silver-coated column filled with Ra-schig glass rings and used as soon as possible for the copolymerization. 45 Likewise, the comonomers used had previously been purified by distillation.

1. Copolymer-vinylene glycol / vinyl alcohol from saponified copolymer-vinylene carbonate / vinyl acetate 50

a) 0.05 part by weight of azobisisobutyronitrile is dissolved in 9 parts by weight of vinylene carbonate and 1 part by weight of vinyl acetate under nitrogen. The monomer mixture is filled under nitrogen into a flattened (4 mm thick, 60 mm wide, 150 mm long) aluminum tube with a screw cap (total filling volume approx. 35 ml), sealed under N2 and suspended in a 50 ° warm water bath for 48 hours. Hard, brittle plastic plates are obtained which are pre-crushed in a hammer mill. The granules are then heated to 120 ° at 1-2 mm pressure for 5 hours in order to separate off the non-polymerized residual monomer.

The demonomerized granules are further ground in a mill to a grain size of <0.1 mm, introduced into 500 parts by weight of 0.5 N sodium hydroxide solution, mixed at 20 ° with a fast-running stirrer for one hour, the carbonate 65 and Acetyl groups are saponified. The water-insoluble CP-vinylene glycol / vinyl alcohol is suctioned off, washed well with water from inorganic salts and freeze-dried.

Yield: 4.7 parts by weight of CP vinylene glycol / vinyl alcohol with a specific surface area measured according to BET of 34m2g_1.

b) As described under 1. a), except that 3 parts by weight of vinyl acetate are used for 7 parts by weight of vinylene carbonate.

Yield: 4.1 parts by weight of CP vinylene glycol / vinyl alcohol with a specific surface area of the freeze-dried polymer powder of 62.5 m2g_1.

c) Comparative experiment as described under 1. a), except that 10 parts by weight of vinylene carbonate and no comonomer are used.

Yield: 4.7 parts by weight of homopolymer-vinylene glycol with a specific surface area of the freeze-dried polymer powder of 20.5 m2g_1.

2. Copolymer vinylene glycol / vinyl glycidyl ether

A mixture of 7 parts by weight of vinylene carbonate, 3 parts by weight of vinyl glycidyl ether and 0.1 part by weight of azobisisobutyronitrile is heated as described under 1.) a) in flat aluminum tubes to 50 ° C. under N2 for 3 days. The plastic plates obtained are comminuted as described under 1.) a), demonomerized, ground to a grain size <0.1 mm, suspended in 500 parts by weight of ice-cold, 0.5 N NaOH with a high-speed stirrer for 30 minutes, then the saponified polymer centrifuged off immediately, slurried in ice water, neutralized to pH 7 with ice-cold with 2N H2S04, suction filtered, washed with ice water and freeze-dried.

Yield: 3.9 parts by weight of CP-vinylene glycol / vinylglycidyl ether containing 1.3 milliequivalents of oxirane groups / g (determined according to: "Internship in Macromolecular Organic Chemistry", p. 221, von Braun, D., Cherdron, H., Kern, W., Hü-thig Verlag, Heidelberg 1966).

3. Amino-substituted copolymer support material

3.9 parts by weight of CP vinylene glycol / vinyl glycidyl ether (from Example 2) are suspended in 100 parts by weight of H20 using an Ultra-Turrax stirrer and 10 ml of 30% ammonia are added. With a small magnetic stirrer, the mixture is then stirred for a further 10 hours at 50 °, finally suctioned off and washed well with water and freeze-dried.

Yield: 3.5 parts by weight of copolymer support material containing 0.9 m equivalent amino groups per gram, in the form of 3-amino-2-hydroxypropyl groups (formed from the reaction of ammonia with glycidyl groups).

4. CP vinylene glycol / acrylic acid

9 parts by weight of vinylene carbonate, 1 part by weight of ethyl acrylate and 0.05 part by weight of azobisisobutyronitrile are polymerized under N2 as in Example 1. a), comminuted, demonomerized and ground to a particle size <0.1 mm, saponified, suction filtered, washed out and freeze-dried.

Yield: 5.0 parts by weight of hydrophilic polymer powder, containing 1.1 milliequivalents of carboxyl groups per gram of carrier with a BET specific surface area of 27.2 mV1.

5. (O-aminohexyl group-substituted poly-vinylene glycol

5 parts by weight of CP-vinylene glycol / acrylic acid from Example 4 are suspended with the Ultra-Turrax in 100 ml of H20, cooled to 5 ° with 2.5 g of N-cyclohexyl-N '- (N-methylmorpholino) ethyl) carbodiimide p-toluene sulfonate was added, the mixture was stirred at pH 5 and 5 ° for 30 minutes, filtered off and washed quickly with ice water. The activated carrier is then immediately suspended in a solution of 5 g of hexamethylenediamine in 100 ml of water, which has been brought to pH 7.5 with 2N HCl and cooled to 5 °, and for a further 24 hours under these conditions with a

636 628

Magnetic stirrer stirred, suctioned off, washed well with water and freeze-dried.

Yield: 4.5 parts by weight of co-aminohexyl group-substituted poly-vinylene glycol with 0.7 milliequivalents of amino groups per gram of carrier.

6. Conversion of CP-vinylene glycol / vinyl alcohol from Example 1. a) with a BET specific surface area of 34 m2g ~ '

a) with epichlorohydrin

50 g of a CP-vinylene glycol / vinyl alcohol, prepared according to Example 1. a) are suspended in 1 liter of 2N NaOH, mixed with 250 ml of epichlorohydrin and stirred at 55-60 ° for 2 hours. The pH of the suspension drops to 10-11 after a short time. This pH is maintained for a further 1 hour by adding NaOH. After a reaction time of 2 hours, the solid is filtered off with suction, washed with water, acetone and finally with water.

b) with hexamethylenediamine

50 g of hexamethylenediamine are dissolved in 1.5 l of water and mixed with HCl to pH 10. That becomes the solution

6. a) activated CP-vinylene glycol / vinyl alcohol and stirred at 50-55 ° for 6 hours. The product is then filtered off and washed with water free of hexamethylene diamine.

c) with l-aminobenzene-4-ß-hydroxyethylsulfonic acid ester

The product obtained in Example 6. b) is stirred with 50 g of 1-aminobenzene-4-β-hydroxyethylsulfonic acid at 55 ° and pH 10 for one hour. The solid is then filtered off, washed with water, acetone and again with water.

d) Diazotization 10 g of the product obtained in Example 6. c) are washed on a suction filter with 200 ml of 0.1 N HCl and then suspended in 300 ml of 0.5 N HCl. The suspension is mixed with 0.1 N NaNO 2 solution at 0-4 ° with stirring until a small excess of nitrite is found in the diazotization with KJ starch paper. After 10 minutes, the mixture is filtered through a suction filter and the residue is washed with ice water and then with 0.15 M sodium phosphate buffer pH 7.5 at 0-4 °.

e) Covalent binding with protein

0.8 g of albumin is dissolved in 350 ml of pH 7.5 phosphate buffer, cooled to 4 ° and the product prepared under 6.d) is added. The suspension is stirred for 20 hours at 4 °, then filtered off and the solid with 1 M NaCl and phosphate-buffered saline (PBS) (aqueous 0.9% NaCl solution containing Vis Mol Na2HP04-KH2P04 buffer of pH 7.2) washed.

Filtrate and washing systems are examined for albumin using the radial immunodiffusion method. 60 mg of albumin are bound to 1 g of the carrier thus produced.

7. Conversion of CP-vinylene glycol / vinyl alcohol from Example 1. b) with a specific surface area according to BET of 62.5 rr? G ~ 1

Following analogous reactions as described under 6. a) to e), 80 mg of albumin can be bound quantitatively to 1 g of activated carrier.

8. Implementation of homopolymer vinylene glycol from Example 1. c) with a BET specific surface area of 20.5 m? G '

Following analogous reactions as described under 6. a) to e), 45 mg of albumin can be bound quantitatively to 1 g of activated carrier.

9. m-aminohexyl group-substituted vinylene glycol / vinyl pyrrolidone copolymer

In 9 parts by weight Vinylene carbonate and 1 part by weight Vinylpyrroli-don are 0.05 parts by weight Abzobisisobutyrolnitril dissolved under nitrogen and polymerized and worked up analogously to Example 1. a).

After demonomerization, the granules are dissolved in a 12% by weight dimethylformanide solution, and this solution is sprayed into a methanol precipitation bath with a pressure of 15 atm. A fine-fiber precipitate of vinylene carbonate / vinyl pyrrolidone copolymer is obtained, which is filtered off with suction, washed with methanol and in 200 parts by weight. Methanol is resuspended. 6 parts by weight Hexamethy-lo-lendiamine, dissolved in 50 parts by weight Methanol, added, the suspension stirred at room temperature for 2 days, suction filtered and washed out with methanol. The washed filter residue is now in a solution of 10 parts by weight. Sodium methylate in 300 parts by weight 96% methanol suspended for 4 days at room temperature sus-15, washed out with methanol and then very intensively with water and freeze-dried.

Yield: 5.0 parts by weight a vinylene glycol / vinyl pyrrolidone copolymer substituted by urethane bonds with to-aminohexyl groups containing 1.6 m equivalent of NH2-20 groups / g of support.

10. Binding of IgG to co-aminohexyl group-substituted copolymer from Example 9 using succinic anhydride and water-soluble carbodiimide 25 5 parts by weight of Cu-aminohexyl-substituted carrier prepared according to Example 9 are mixed at 10 ° and pH 6 with 2.5 parts by weight. Succinic anhydride, which in 200 parts by weight Water are suspended, succinoylated for 4 hours. The pH is adjusted with 2N NaOH. After washing the solid 30 with water, the product is washed with 1.25 parts by weight of N-cyclohexyl-N '(N-methylmorpholino) ethyl) -carbodiimide-p-toluenesulfonate at pH 5 and 5 ° 30 min. stirred, filtered off and quickly washed with ice water.

0.5 part by weight IgG are in 150 parts by weight Phosphate buffer 35 pH 7.5 dissolved and stirred with the activated carrier at 4 ° for 24 hours. After filtration, the product is washed with 1 M sodium chloride and with PBS.

75 mg of IgG are covalently bound to 1 g of the carrier.

40 11. Copolymer vinylene glycol / diethylene glycol divinyl ether

In 9 parts by weight Vinylene carbonate and 1 part by weight Diethylene glycol kivivinyl ether 0.1 parts by weight Azobisisobutyrolnitrile dissolved under nitrogen and as described under 1. a) in flat aluminum tubes under N2 heated to 50 ° for 3 days and polymerized. After working up and saponification as described under 1. a), 3.5 parts by weight are Obtained CP-Vinylenglykol / Diethylengly-koldivinyläther.

12. Copolymer - vinylene glycol / N-acryloylaminoacetaldehyde-50 dimethylacetal a) in 9 parts by weight Vinylene carbonate and 1 part by weight N-Acryloyl-aminoacetaldehyde dimethyl acetal, 0.05 part by weight of azobisisobutyrene nitrile are dissolved under nitrogen. The monomer mixture is polymerized as described under 1. a), worked up.

55 prepared and saponified to 4.1 parts by weight N-Acryloylaminoacetalde hyddimethylacetal / vinylene glycol copolymer.

10 parts by weight N-Acryloylaminoacetaldehyde dimethyl acetal / vinylene glycol copolymers were in 100 parts by weight. 1N HCl stirred for 4-5 hours. The activated carrier was washed with water 60 and phosphate buffer pH 7.5.

b) 0.5 g of albumin was dissolved in 200 ml of PBS and stirred with the product prepared under a) at 4 ° for 14 hours. After filtration, the carrier-bound protein was washed with 1 M sodium chloride and with PBS. 1 g of the carrier thus produced binds

65 40 mg albumin.

C.

Claims (5)

636 628 1. A process for the preparation of a compound from a copolymer of vinylene glycol and a biologically active substance chemically bonded thereto, characterized in that a copolymer of vinylene carbonate is reacted with a biologically active substance and then the cyclocarbonate groups still present are converted into hydroxyl groups. 2nd Ri / Ri H, C = C ^ H2c = c ^ O-CH-CH2 CH-CH2 \ / il O O \ / c II O O \ / o in which R! Hydrogen, methyl, ethyl, R3 methyl, ethyl, Pro 9. Agent containing a compound from a copolyme-pyl and n is an integer between 1 and 4, veins of the vinylene glycol and one chemically bound to it, turns. biologically active substance as an active component. 636 628 4 10. A method of a compound made from a copolymer of nyl-substituted polystyrene, furthermore the cellulose and its waste of the vinylene glycol and a chemically bound, organic and finally natural and synthetic polylogically active substance for carrying out immunoreactive peptides and proteins. Because of the balanced changes in vitro. Use has an effect between the matrix and the effector 2. A process for the preparation of a compound from a copolymer of vinylene glycol and a biologically active substance chemically bound thereto, characterized in that the cyclocarbonate groups of a copolymer of vinylene carbonate are converted into hydroxyl groups and electrophilic groups in the copolymer with a biologically active substance implements. 15 2nd PATENT CLAIMS 3rd 636 628 Rj Rj in which Rx is hydrogen, methyl, ethyl, R3 methyl, ethyl, pro- H2C = C ^ H2C = C / pyl and n represent a number between 1 and 4. ^ O-CH-CHs ^ CH-CH2 7- Method according to claim 3, characterized in \ / | | that a copolymer containing at least 55% mono- O O O 5 mer units of vinylene glycol and at most 45% mono- \ / mer units, at least one connection of the formulas C Rj R, R] II H2C = C ^ H2C = C ^ H2C = C ^ o xcoor3 o-co-ch, ci 10 y R1 R, Ri H-, C = C HiC = C H2C = C ^ \ \, (0CH2-CH2) n ~ 0-CH = CH2 <_J> V _ / - ch = CH2 Ri H, C = Q = r / R, CH-> OC- H, C = C ^ (CH2) n I. ch2 \ (CH2) n-CH-CH2 O Ri / R H-.C = QT H2C = C ' o-ch-ch2 xch-ch2 V 1 1 u O O \ / c II O in which Rx is hydrogen, methyl, ethyl, R3 is methyl, ethyl, protein units of vinylene glycol and at most 45% mono-pyl and n are an integer between 1 and 4, number units, at least one compound of the formulas is used. R, JRi 8. The method according to claim 4, characterized in that r c-c H2C = C H2C = C that a copolymer containing at least 55% mono- 40 2 ^ COOR 2 ^ 0-CO-CH3 ^ Cl Ri Ri / R> H, C = C ^ H, C = C / H2C = C. / 12V_ = L. x (0CHi-CH2) n-0-CH = CH2 <^> - ch = CH> R. ^ H, C = C / CH2 H2C = C ^ \ (CH2) n (CH2) n-CH-CH2 OC CH 3. A process for the preparation of a compound from a copolymer of vinylene glycol and a biologically active substance chemically bonded thereto, characterized in that the cyclocarbonate groups of a copolymer of vinylene carbonate are converted into hydroxyl groups and the copolymer first with an electrophilic group-containing compound and then with a biologically active substance. 4. A process for the preparation of a compound from a copolymer of vinylene glycol and a biologically active substance chemically bonded thereto, characterized in that the cyclocarbonate groups of a copolymer of vinylene carbonate are converted into hydroxyl groups and the copolymer is reacted with a biologically active substance carrying electrophilic groups. 5. The method according to claim 1, characterized in that a copolymer containing at least 55% monomer units of vinylene glycol and at most 45% monomer units, at least one compound of the formulas H, C = C. \ h2c = c: COOR, -Ri "D-CO-CH, H2C = C Ri Cl h2c = c: Ri (0CH2-CH2) n-0-CH = CH2 Ri Rt H2c = qf ch2 H2C = C ^ No \ (CH2) n (CH2) n-CH-CH2 II \ / OC CH2 O ßl / R1 h2c = c ^ H2c = ef o-ch-ch2 ^ ch-ch2 V 1 I u 00 \ / c II o in which Rx is hydrogen, methyl, ethyl, R3 is methyl, ethyl, propyl and n is an integer between 1 and 4. 6. The method according to claim 2, characterized in that a copolymer containing at least 55% mono- 5 mer units of vinylene glycol and at most 45% monomer units, at least one compound of the formulas JLi / Rl h2c = <h2c = <h2c = c ^ COOR3 O-CO-CH3 C1 H2C = C ^ (0CH2-CH2) n-0-CH = CH2 CH2 R, Ri H2C = C / CH2 H2C = Cf \ (CH2) n \ (ch2) "- ch-ch2 II \ / oc ch2 0 5 of carbohydrates, especially cellulose, dextran, the starch, the agar or its descendants, found the most widespread as a matrix in aqueous systems, if