Polymerizable composition

A polymerizable composition comprises (a) an ethylenically unsaturated diluent monomer comprising an ethylenically unsaturated fluorine-containing monomer; (b) an ethylenically unsaturated monomer containing a reactive ester group capable of coupling with an amino group-containing compound by the formation of an amide link; and, (c) a polymerization initiator. A polymer produced from the composition is capable of immobilizing an amino group-containing compound e.g. a protein. Such polymers are suitable for use in a variety of biomedical applications.

The invention relates to a polymerisable composition and to a polymer 
produced therefrom. Polymers which are biocompatible and which may be 
employed in a variety of biomedical applications may be produced from the 
compositions of the invention. 
More particularly, polymers are provided which are capable of immobilizing 
compounds containing amino groups. Such compounds include proteins and 
amino acids. Specific applications of the polymers of the invention 
include affinity chromatography wherein an amino group-containing ligand 
is attached to the polymer and peptide synthesis. 
For example, the polymers of the invention could be used for the separation 
of a component of a body fluid e.g. blood using a bioaffinity separation 
procedure. This could be achieved by bringing the body fluid into contact 
with the polymer having an appropriate protein ligand attached to its 
surface. 
Preferred polymer compositions of the invention are those from which 
hydrogels may be produced. A hydrogel is a polymeric material that imbibes 
a significant proportion of water within a three dimensional network 
without causing dissolution of the polymer. 
Die Makromolekulare Chemie 177, 683-689 (1976) describes the synthesis of 
monomers containing a reactive ester group capable of coupling with an 
amine by the formation of an amide link. More particularly, it suggests 
that copolymers of succinimido esters of 
.omega.-methacryloylaminocarboxylic acid and methacrylamide may be used as 
carriers for enzymes and drugs. 
U.S. Pat. No. 4,330,440 describes an activated polymer matrix for use in 
affinity chromatography. A macroporous polymer having surface hydroxyl 
groups e.g. hydroxyethyl methacrylate is treated with a carbonylating 
agent to provide active groups which are capable of immobilising compounds 
containing amino groups. 
U.S. Pat. No. 4,433,111 describes polymeric materials suitable for 
biomedical applications, particularly for making contact lenses. The 
materials have enhanced surface properties which improve their protein 
repellency. Examples of other biomedical applications which are mentioned 
in the specification include surgical implants and prosthetic devices e.g. 
blood vessels, artificial urethers, heart valves and artificial breast 
tissue. The polymeric materials are also said to be useful for contact 
with body fluids outside the body e.g. in manufacturing membranes for 
kidney dialysis and heart/lung machines, swabs, nappy liners and wound 
dressings. 
The hydrogel-forming polymeric material of U.S. Pat. No. 4,433,111 
comprises units derived from (1) an olefinically unsaturated carboxylic 
acid amide, (2) an N-vinyl lactam, (3) an olefinically unsaturated 
carboxylic acid ester, (4) an olefinically unsaturated carboxylic acid and 
(5) a hydrophobic monomer comprising (a) a fluorine-containing 
polymerisable monomer having a fluoroaliphatic side chain and (b) a 
non-fluorine-containing polymerisable hydrophobic vinyl monomer. The 
various units are present in specified amounts and the copolymer is 
cross-linked with a cross-linking agent. The disclosure demonstrates the 
ability of the fluorine-containing monomer to affect the surface energy of 
the polymer and increase its protein repellency. 
Unlike the polymer compositions of the present invention, the hydrogels 
according to U.S. Pat. No. 4,433,111 are specifically designed to be 
unreactive i.e. they do not contain reactive groups for the purpose of 
reacting with other compounds. While the polymer compositions of U.S. Pat. 
No. 4,330,440 do contain such reactive groups, the compositions and their 
preparation have a number of disadvantages. In this respect, the 
compositions require the provision of a macroporous polymer followed by 
separate steps to activate the polymer. Further, no action is taken to 
minimise non-specific adsorption to the polymer i.e. the adsorption of 
compounds other than those intended to react with the active groups. 
Similarly, the polymer compositions of Die Makromolekulare Chemie 177, 
683-689 (1976) are reactive but make no provision for minimising 
non-specific adsorption. 
The present invention aims to overcome disadvantages associated with prior 
art compositions by providing a polymerisable composition from which a 
desired activated polymer may be rapidly prepared. The method of 
preparation offers a high degree of control over the composition of the 
polymer and the monomers are chosen such that non-specific adsorption is 
reduced. 
The invention provides a polymerisable composition comprising 
an ethylenically unsaturated diluent monomer comprising an ethylenically 
unsaturated fluorine-containing monomer; 
an ethylenically unsaturated monomer containing a reactive ester group 
capable of coupling with an amino group-containing compound by the 
formation of an amide link; and, 
a polymerisation initiator. 
The invention also provides a method of making a polymer having reactive 
ester groups which method comprises forming the polymerisable composition 
of the invention and subjecting the composition to conditions which 
generate free radicals from the polymerisation initiator. 
Preferably, the diluent monomer is present in an amount from 65 to 99 mole 
percent and the monomer containing the reactive ester group is present in 
an amount from 1 to 35 mole percent, said percentages being based on the 
total monomer present. 
The diluent monomer is chosen to provide the composition with desired 
physical properties. It is preferred that it comprises 
non-fluorine-containing monomer in addition to the fluorine-containing 
monomer. Any non-fluorine-containing monomer is preferably hydrophilic to 
minimise the non-specific adsorption of proteins to the polymer. 
Preferably, the diluent monomer or monomers are chosen to ensure that the 
polymerisable composition is coatable and film-forming either with or 
without the aid of a solvent. In a particularly preferred embodiment, the 
combination of monomers in the polymerisable composition form a solution 
without requiring a non-polymerisable solvent. An advantage of such a 
totally polymerisable composition is that it overcomes the problem of 
leaching out of small molecules, for example molecules associated with the 
initiation of polymerisation, which occurs with polymer membranes prepared 
by other methods. The concentration of the diluent monomer can be varied 
to adjust the level of reactive groups in the polymer to the desired 
range. 
Preferred non-fluorine-containing diluent monomers are selected from esters 
of ethylenically unsaturated carboxylic acids (e.g. substituted or 
unsubstituted alkyl esters of acrylic or methacrylic acid), amides of 
ethylenically unsaturated carboxylic acids (e.g. N-alkyl substituted or 
unsubstituted amides of acrylic or methacrylic acid), N-vinyl substituted 
amides of carboxylic acids or N-vinyl substituted nitrogen-containing 
heterocyclic monomers. Examples of suitable diluent monomers include 
acrylamide, methacrylamide, N-substituted acrylamide and methacrylamide 
e.g. N-alkyl acrylamide and N,N-dialkyl acrylamide, alkyl acrylates and 
alkyl methacrylates wherein the alkyl groups are optionally substituted, 
N-vinyl-2-pyrrolidone and N-methyl-N-vinylacetamide. 
For the formation of hydrogels, the diluent monomer is preferably a 
hydroxyalkyl acrylate, hydroxyalkyl methacrylate, glycidyl acrylate, 
glycidyl methacrylate, hydroxyalkylacrylamide or 
hydroxyalkylmethacrylamide monomer in which the alkyl group preferably 
contains from 1 to 6 carbon atoms. 
Preferably, the fluorine-containing diluent monomer is a fluoroalkyl ester 
or amide of an ethylenically unsaturated carboxylic acid. 
Examples of preferred ethylenically unsaturated fluorine-containing 
monomers include fluoroalkyl acrylates, fluoroalkyl methacrylates, 
fluoroalkylacrylamides and fluoroalkyl methacrylamides. The fluoroalkyl 
group may be partially or fully fluorinated and preferably contains from 1 
to 6 carbon atoms. Particularly preferred fluoroalkyl groups terminate in 
a trifluoromethyl group and include trifluoroethyl. 
All or part of the diluent monomer may be a fluorine-containing monomer. 
Preferably, the fluorine-containing monomer is present in an amount from 5 
to 40 mole percent and the non-fluorine-containing monomer is present in 
an amount from 25 to 94 mole percent based on the total monomer present in 
the composition. 
The monomer containing a reactive ester group capable of coupling with an 
amino group-containing compound, hereinafter also referred to as the 
reactive ester monomer, may be derived from an ester or amide of an 
ethylenically unsaturated carboxylic acid e.g. an acrylate, methacrylate, 
acrylamide or methacrylamide monomer. 
Preferred reactive ester groups are represented by the formula --COOX 
wherein X represents an electron-withdrawing group. Functional groups are 
classified as electron-withdrawing groups relative to hydrogen, e.g. 
--NO.sub.2 and --I groups draw electrons to themselves more than a 
hydrogen atom occupying the same position in the molecule, J. March, 
Advanced Organic Chemistry, 2nd edition, McGraw Hill, p 20,246. Specific 
examples of X groups include N-succinimido, benzylidene aniline, 
pentafluorophenyl, 4-nitrophenyl, 4-cyanophenyl, 4-alkylsulphonylphenyl, 
acyl, 4-acylphenyl, 4-dialkylaminocarbonylphenyl, 4-alkoxycarbonylphenyl 
and 4-alkoxysulphonylphenyl. 
Preferably, a chain of from 4 to 15 atoms separates the reactive ester 
group from the ethylenically unsaturated portion of the monomer which 
undergoes polymerisation. Such a chain may comprise an alkylene chain. The 
purpose of the chain is to ensure that the reactive ester group is spaced 
away from the polymer backbone after polymerisation. 
The reactive ester group reacts directly with the amino group-containing 
compound. Preferably, such reaction will take place under physiological 
reaction conditions. 
Preferred polymerisable compositions may comprise from 5 to 25 mole percent 
reactive ester monomer and from 75 to 95 mole percent diluent monomer. 
The polymerisation initiator is a compound or a combination of compounds 
which is capable of generating the free radicals required for 
polymerisation to occur. A wide variety of polymerisation initiators are 
known including thermal and photoinitiators. Such initiators include 
carbonyl compounds, organic sulphur compounds, peroxides, redox systems, 
azo and diazo compounds and halogen compounds. 
The composition of the invention preferably comprises a photopolymerisation 
initiator. A particularly preferred photopolymerisation initiator is a 
combination of an aromatic carbonyl compound and an amine compound. 
Advantages associated with the use of such an initiator system are that 
polymerisation proceeds rapidly and can be carried out at room 
temperature. 
Particularly preferred aromatic carbonyl compounds include ketocoumarin 
compounds. Specific examples of preferred aromatic carbonyl compounds 
include 2,2'-dimethoxy-2-phenylacetophenone, 
3,3'-carbonyl-bis-(5,7-di-n-propoxycoumarin), 
3,3'-carbonyl-bis-(7-diethylaminocoumarin) and 
7-diethylamino-3-thenoylcoumarin. 
A preferred example of an amine coinitiator compound is N-phenylglycine. 
In addition to the components described above, the polymer composition of 
the invention may comprise a crosslinking agent. Many suitable 
crosslinking agents are known and include alkylene glycol diacrylates and 
dimethacrylates e.g. ethylene glycol dimethacrylate, and other 
polyfunctional compounds such as N,N'-methylene-bis-acrylamide and 
divinylbenzene. 
The monomers used in the invention may be readily prepared and some are 
commercially available. 
The fluorine-containing monomers and the monomers containing a reactive 
ester group used in the invention may be prepared by appropriate 
modifications of established literature techniques e.g. H. -G Batz, J. 
Koldehoff; Makromol. Chem. 177, 683 (1976) and W de Winter, A. Marien; 
Makromol. Chem., Rapid Commun. 5, 593 (1984). 
In order to produce the reactive ester-containing monomer, the basic 
monomer e.g. acrylamide may be converted into a carboxy terminated 
derivative e.g. acrylamidocaproic acid which in turn may be esterified to 
provide a terminal reactive ester group e.g. a succinimido ester. A 
representative preparative method is given in Die Makromolekulare Chemie 
177, 683-689 (1976). 
The polymerisable composition of the invention may be prepared by mixing 
the individual components using a solvent if required. By the appropriate 
choice of monomers, no solvent is necessary. For example, all the monomers 
may be liquids or the diluent monomer can act as a solvent for the other 
monomers present. 
By way of example, the polymerisable composition of the invention may be 
prepared by dissolving the fluorine-containing monomer, the reactive ester 
monomer and, optionally, a cross-linking agent in a solvent monomer. 
Subsequently, the polymerisation initiator e.g. a combination of 
ketocoumarin and amine compounds dissolved in solvent monomer, may be 
added to and mixed with the polymer composition. 
A reactive ester-containing polymer is produced as a result of polymerising 
the polymerisable composition of the invention under conditions which 
generate free radicals from the polymerisation initiator e.g. using heat 
and/or radiation when required. 
For example, using a thermal initiator the polymerisable composition may be 
heated to a temperature from 50.degree. to 80.degree. C. and 
polymerisation allowed to proceed for from 0.5 to 30 hours. Using a 
photoinitiator, polymerisation may be carried out at ambient temperature 
for from 0.5 to 4 hours. 
The invention includes xerogels and hydrogels derived from the 
polymerisable composition of the invention. 
The polymers of the invention may be used in a variety of forms. 
The polymerisable composition may be formed into a shaped polymeric article 
by introducing the composition into a mould of the desired configuration 
before polymerisation is effected. 
For example, a xerogel membrane may be prepared by injecting the polymer 
composition into a polymerisation cell formed by two glass plates which 
are clamped together and separated by a gasket. Preferably, the surfaces 
of the mould in contact with the polymerisable composition are treated 
with a mould release agent. Examples of suitable mould release agents 
include silicones and fluorocarbon compounds. Polymerisation e.g. by 
exposure to UV light, results in the formation of a xerogel membrane. 
The shaped article may be immersed in water or an aqueous medium until 
equilibrium is reached. The water content of the hydrogel so produced will 
depend on the nature of the copolymer and its structure. 
Alternatively, the polymerisable composition may be coated as a layer on a 
support. 
An amino group containing-compound may be coupled to the polymer by 
contacting the polymer with the compound. The compound may be a ligand 
capable of interacting selectively with another compound whereby the 
polymer may be used for affinity chromatography. Examples of amino 
group-containing ligands include proteins.

The invention is further illustrated by way of example as follows. (The 
molar ratio of monomer components is given in parenthesis after each 
polymer). 
EXAMPLE 1 
Synthesis of 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloy 
laminocaproic acid, succinimido ester) (7:3:1) 
The following was placed in a round-bottomed flask, fitted with a reflux 
condenser, stirrer and nitrogen inlet: 
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acrylamide 4.97 g 
2,2,2-trifluoroethylmethacrylamide 
5.01 g 
methacrylamidocaproic acid, 
2.96 g 
N-hydroxysuccinimido ester 
azobisisobutyronitrile 0.06 g 
dimethylformamide 30 ml 
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The reaction mixture was stirred for 5 hours at 60.degree. C. under a 
nitrogen blanket. At the end of this period, the viscous solution was 
diluted with dimethylformamide (30 ml) and, after standing overnight, the 
polymer was precipitated into diethyl ether. The polymer was washed with 
acetone. 
Yield: 10.3 g. 
Analysis: Theory C 49.15, H 6.11, F 13.21, N 12.98, O 18.55%. Found C 
47.52, H 6.53, F 12.84, N 11.92, O 21.19%. 
Using the synthesis procedure described above, the following polymers were 
prepared: 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)-methacrylamide-N-methacryloylam 
inocaproic acid, succinimido ester) (10:1:1, 8:2:1, 6:4:1, 5:5:2) 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)-methacrylamide-co-N-methacryloy 
l-beta-alanine, succinimido ester) (8:2:1) 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloyl 
aminocaproic acid, p-nitrophenyl ester (16:4:1) 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)-methacrylamide-co-N-methacryloy 
lglyclglycine, succinimido ester) (8:1:2, 8:2:1) 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloyl 
-omega-aminoundecanoic acid, succinimido ester) 8:2:1) 
poly(2-hydroxyethyl methacrylate-co-2,2,2-trifluoroethyl 
methacrylate-co-N-methacryloylaminocaproic acid, succinimido ester) 
(18:1:1) 
poly(2-hydroxypropylmethacrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamid 
e-co-N-methacryloylaminocaproic acid, pentafluorophenyl ester) (8:2:1) 
poly(2-hydroxypropyl methacrylate-co-2,2,2-trifluoroethyl 
methacrylate-co-N-methacryloylaminocaproic acid, succinimido ester) 
(8:1:2, 8:2:1) 
poly(2-hydroxypropyl methacrylate-co-2,2,2, -trifluoroethyl 
methacrylate-co-N-methacryloylglycylglycine, p-nitrophenyl ester) (8:2:1) 
poly(2-hydroxypropyl methacrylate-co-2,2,2-trifluoroethyl 
methacrylate-co-N-methacryloylglycylglycine, succinimido ester) (8:2:1) 
poly(N-methyl-N-vinylacetamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co 
-N-methacryloylaminocaproic acid, succinimido ester) (8:2:1) 
poly(N,N-dimethylacrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co-N- 
methacryloyl-beta-alanine, succinimido ester) (8:2:1) 
poly(N,N-dimethylacrylamide-N-(2,2,2-trifluoroethyl)methacrylamide-co-N-met 
hacryloylaminocaproic acid, succinimido ester) (8:2:1) 
poly(2-hydroxypropylmethacrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamid 
e-co-N-methacryloylaminocaproic acid, p-nitrophenyl 
ester-co-N-methacryloyl-omega-aminoundecanoic acid, p-nitrophenyl ester) 
(16:4:1:1, 8:2:1:1) 
poly(N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloylaminocaproic 
acid, succinimido ester) (5:1) 
poly(N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloylaminocaproic 
acid, p-nitrophenyl ester) (10:1) 
A coating solution was prepared by dissolving 
poly(acrylamide-co-N-(2,2,2-trifluoroethyl)methacrylamide-co-N-methacryloy 
laminocaproic acid, succinimido ester) (7:3:1) (10% w/w) in 
dimethylformamide. The coating solution also contained glutaraldehyde (10% 
w/w based on the polymer) as a crosslinking agent. 
The solution was coated on a polyester (ESTAR) sheet using a gravure roller 
at a coating speed of 1 to 2 m/min to provide a wet laydown of 2.5 mls per 
250 cm.sup.2. 
A sample of the dried, crosslinked coated product was treated with a 
solution of albumin (an amino group-containing protein). Infra-red 
spectral analysis of the treated and untreated coating confirmed that the 
protein had coupled to the polymer at the active ester sites in the 
polymer as a result of amide formation. 
EXAMPLE 2 
Preparation of poly(2-hydroxypropyl methacrylate-co-epsilon 
methacrylamidocaproic acid succinimido ester 
(MCS)-2,2,2-trifluoroethylmethacrylamide (TFEMA) 
MCS (13.5 mmoles, 4.0 g), TFEMA (12.0 mmoles, 2.0 g) and the bifunctional 
crosslinking agent, ethylene glycol dimethacrylate (EGDMA) (1.68 mmoles, 
0.34 g) were dissolved in 2-hydroxypropyl methacrylate (103.6 mmoles, 14.0 
mls), immersing the mixture in an ultrasonic bath to hasten dissolution. 
7.9 mls of the following initiator stock solution was added: 
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3,3'-carbonyl-bis-(5,7-di-N- 
(0.55 mmoles) 0.30 g 
propoxycoumarin) 
N-phenylglycine (NPG) 
(4.63 mmoles) 0.70 g 
2-hydroxypropyl methacrylate 
50 ml 
______________________________________ 
Mixing was effected through brief re-immersion in the ultrasonic bath, and 
three identical polymerisation cells were completely filled with the 
resultant solution. 
The photopolymerisation cells were constructed from two glass plates, 
separated by a poly(tetrafluoroethylene) gasket. Prior to positioning of 
the gasket, the internal glass faces of the cell were covered with a mould 
release agent. The appropriate volumes of monomer, attendant 
photoinitiator and cross-linking agent were injected into the cell, held 
together with spring release clips, with a glass syringe and needle 
pre-positioned within the cell. 
The cells were placed on the plate glass diffuser of an exposure frame, 
where they were exposed to an array of four 125 watt medium pressure 
vapour UV lamps for a period of 1.5 hours. 
After exposure, photopolymerised xerogels were removed from the cell by 
release of the clips and separation of the glass plates. Surface 
characterisation of the xerogels was performed by electron spectroscopy. 
The xerogel membranes were transparent indicating that the homogeneity of 
the polymers was good. 
The polymer membranes produced in this manner were readily hydrated to form 
hydrogels.