Method to produce polyacrylamide hydrogel

FIELD: biotechnologies.

SUBSTANCE: method is realised by polymerisation of an aqueous solution containing 7-15 wt % of acrylamide and 0.5-1.5 wt % of N,N'-methylenebisacrylamide, in presence of polymerisation initiator, besides, the polymerisation initiator is a mixture of 4,4'-azobis(4-cyanopentane acid) and ammonium salt of 4-8-dithiobenzoate of 4-cyanopentane acid with their content in the aqueous solution as 0.03-0.07 wt % and 0.07-0.35 wt %, accordingly, and polymerisation is performed at temperature of 70-80°C and pH 3.0-4.0.

EFFECT: increased structural homogeneity of a hydrogel.

2 tbl, 5 ex

The invention relates to the field of polymer chemistry, biochemistry and medicine, namely to a process for the preparation of polyacrylamide hydrogel, which is due to the high porosity is used as the separating medium in liquid chromatography, as a carrier of immobilized biologically active substances, as well as for the manufacture of implants soft tissue.

A method of obtaining polyacrylamide hydrogel by γ-irradiation dose of 0.5 to 5.0 Mrad powdered polyacrylamide followed by the addition of water and mechanical homogenization of the mixture [RF patent 2114867, C08F 120/56, 1998].

The disadvantage of this method is the heterogeneous structure of the obtained hydrogel, due to the presence of pores of different sizes. All the pores of the hydrogel accessible to molecules of insulin with a molecular mass (MM) 6000 and molecules of serum albumin with MM 67000, 70-80% of the pores accessible to molecules of alcohol dehydrogenase with MM is 141,000 and 40-50% of the pores accessible to molecules of fibrinogen with MM 340000.

The closest in technical essence and the achieved results is the method of obtaining a polyacrylamide hydrogel by polymerization of an aqueous solution containing 7-15 wt.% acrylamide and 0.5 to 1.5 wt.% N,N'-methylenebisacrylamide, in the presence of initiator, polymerizati [research Methods in immunology, Ed. by I. Lefkovits and of B. Pernis World Moscow, s-107].

As an initiator of polymerization using a redox system: ammonium persulfate and N,N,N',N'-tetramethylethylenediamine, and polymerization was performed at room temperature.

The disadvantage of this method is the structural heterogeneity of the obtained hydrogel, due to the wide range of pore sizes. Depending on the amount of acrylamide and N,N'-methylenebisacrylamide obtained hydrogels swollen in the state contain 80-95% water, all the pores of hydrogels available for molecules of insulin, 90-95% of the pores accessible to molecules of ovomucoid with MM 31000, 80-90% of the pores accessible to molecules of serum albumin, 50-60% of the pores accessible to molecules of alcohol dehydrogenase and 30-40% of the pores accessible to molecules of fibrinogen.

The objective of the invention is to increase the structural homogeneity of the hydrogel.

The technical result that is achievable with the use of the invention is to increase the structural homogeneity of the hydrogel.

The technical result is achieved in that in the method of obtaining a polyacrylamide hydrogel by polymerization of an aqueous solution containing 7-15 wt.% acrylamide and 0.5 to 1.5 wt.% N,N'-methylenebisacrylamide, in the presence of a polymerization initiator as a polymerization initiator, a mixture of 4,4'-azobis(4-cyanopentanoate acid) and ammonium salts of 4-S-dithio is enzoate 4-cyanopentanoate acid in their content in an aqueous solution of 0.03-0.07 wt.% and 0.07 and 0.35 wt.% accordingly, and the polymerization is carried out at a temperature of 70-80°C and pH 3.0 to 4.0.

Example 1.

In to 89.5 ml of water at pH of 3.6 dissolve 10 g of acrylamide (AA), 0.5 g of N,N'-methylenebisacrylamide (BIS), 0,065 g of 4,4'-azobis(4-cyanopentanoate acid) (CTC) and 0.07 g of ammonium salt of 4-S-dithiobenzoate 4-cyanopentanoate acid (DTB). The vacuum solution with 10^-3mm Hg and incubated for 20 hours at 78°C. the resulting gel is crushed by pushing through a sieve with a pore diameter of 1 mm and washed with 10-fold excess of distilled water. The water content of the hydrogel assessed by weighing the swollen hydrogel in water and lyophilized hydrogel. To assess content in the hydrogel pores of different size to 2 ml of gel add 4 ml of the protein solution and the mixture was incubated for 48 hours at 4°C. the Concentration of the original protein solution and the protein solution after contact with the hydrogel was measured spectrophotometrically at 280 nm using pre-built calibration dependence. Given the volume ratio of hydrogel and solution of protein, counting the number of pores available for each protein, taking as 100% the number of pores available for water. The properties of the hydrogel are shown in table 2.

Examples 2-4.

The process is conducted according to example 1, using different numbers of components. The compositions of the reaction mixture and the polymerization conditions are shown in table 1.

Example 5 (the end of the roll, the prototype).

At room temperature and stirring in 89 ml of distilled water was dissolved 10 g of acrylamide and 0.5 g of N,N-methylenebisacrylamide. After complete dissolution the solution was added 0.08 g of ammonium persulfate. The solution is vacuum at 10-12 mm Hg, to it add 0,08 ml of N,N,N',N'-tetramethylethylenediamine and incubated at room temperature (18-21°C) for 5 hours. The obtained gel is then removed from the vessel, crushed by pushing through a sieve with a pore diameter of 1 mm and washed with 10-fold excess of distilled water.

It is seen that using as an initiator of polymerization of a mixture of 4,4'-azobis(4-cyanopentanoate acid) and ammonium salts of 4-S-dithiobenzoate 4-cyanopentanoate acid in the process of copolymerization of acrylamide and N,N'-methylenebisacrylamide results in a more homogeneous porous hydrogels, in which very little of the content of pores of large size. So, if the hydrogel obtained by the method prototype, all the pores have dimensions that provide access for water molecules and protein molecules with 6000 MM, of which 38 % is available for protein molecules with MM 360000, hydrogels, the scientists on the proposed method, the number of pores available for protein molecules with MM 360000, reduced 3.8-7.6 times.

The limiting values of the components of the initiator due to the fact that at concentrations below these values, the polymerization proceeds at a very slow speed, and at concentrations above these values, the hydrogel is formed. Only at pH values of 3.0 to 4.0 is possible to obtain a homogeneous solution all used compounds. At temperatures above 80°C the polymerization is explosive in nature, and at temperatures below 70°C it does not leak.

A method of obtaining a polyacrylamide hydrogel by polymerization of an aqueous solution containing 7-15 wt.% acrylamide and 0.5 to 1.5 wt.% N,N'-methylenebisacrylamide, in the presence of a polymerization initiator, characterized in that as the initiator of polymerization, a mixture of 4,4'-azobis(4-cyanopentanoate acid) and ammonium salts of 4-S-dithiobenzoate 4-cyanopentanoate acid in their content in an aqueous solution of 0.03-0.07 wt.% and 0.07 and 0.35 wt.%, respectively, and the polymerization is carried out at a temperature of 70-80°C and pH 3.0 to 4.0.