CS252916B1 - Substrate for growing living tissues and organisms - Google Patents

Abstract

Synthetic substrate for growing living tissues and organisms (agar substitute), consisting of multiblock copolymers of acrylonitrile with acrylamide and/or acrylic acid or its salts, containing at least 80%, preferably over 95% water in the state of equilibrium swelling at 20 00, and optionally with nutrients for the organisms or their parts to be grown. The powdered substrate is dissolved in water under heat at atmospheric pressure, and the solution solidifies into a flexible gel upon cooling.

Description

The invention relates to a substrate for growing living tissues and organisms.

By living tissue is meant parts of living plants or animals or humans, ie individual cells or more or less organized sets of cells. Plants are understood to be all organisms from single cell algae and bacteria to fungi and fungi to higher telomic plants, including parts such as leaves, roots, rhizomes, cuttings, eyes and tissue pieces to individual cells. Cultivation is understood to mean all operations aimed at the preservation, testing and multiplication of those organisms, including cloning and genetic engineering, and similar biotechnological procedures. The substrates are macromolecules at temperatures up to 45 ° C in water-swellable substances, neither harmful to the cultivated organisms, nor eaten by them, nor decomposed by them, but forming a suitable medium containing nutrient solutions.

So far, mostly marine agar has been used for these purposes. They have all the desired properties in a pure state and can be reversibly dissolved in water and thus cast, by cooling, solidifying into a gel which, with a suitable water content, has sufficient cohesion but does not impose excessive resistance to the growing parts of the organisms. Its disadvantage, however, is the relatively limited amount occurring in nature and the relatively difficult cleaning, which implies the current high cost (in 1985 about 40 U.S. doi. Or 100 DM per kg).

Agar - the original Malaysian name agar-agar - is a polysaccharide whose long macromolecules are not chemically cross-linked but only physically. It is physiologically harmless and has therefore been used in the food industry.

It has now been found that multiblock acrylonitrile-acrylamide-containing multiblock copolymers containing similar

252 916 there are some acrylic acid units, respectively. their salts with physiologically acceptable cations, in particular sodium, if strongly swellable, above 80 preferably above 95% of the water content at equilibrium swelling at 20 ° C. Since these hydrogels are generally prepared by controlled hydrolysis of the poles (acrylonitrile) with sulfuric or nitric acid to a concentration such that the poles (acrylonitrile) dissolve in them at least warmly, this means that they are copolymers with a relatively high degree of hydrolysis.

These synthetic hydrogels are easy to prepare in very pure form in any amount and are relatively inexpensive, substantially cheaper than agar. Their thermoreversible solubility in water has not previously been observed or mentioned in the literature. So far, thermoreversible gels of multibiocrylic copolymers of acrylonitrile with acrylamide and / or acrylic acid have been prepared using only a solvent mixture of poly (acrylonitrile) (e.g. dimethylsulfoxide) with a precipitate (e.g. glycerol), see e.g.

AO 178 638 and 179 179 · There is also an indication in the patent literature about the solubility of polyacrylonitrile in water alone at temperatures above 180 ° C under autoclave pressure, but the accuracy of this data for at least polymers in the usual range of molecular weights has not been verified. Multiblock copolymers of acrylonitrile, containing distinct, X-ray diffraction domains (acrylonitrile), behave in the same way as the poles (acrylonitrile), and therefore the solubility of some of them in hot water at atmospheric pressure is surprising. It was necessary to assume that as the poles (acrylonitrile) they would dissolve themselves only in highly polar solvents such as dimethyl sulfoxide, dimethylformamide, strongly concentrated nitric or sulfuric acid and concentrated aqueous solutions of some salts such as zinc chloride, sodium or calcium rhodanide or lithium bromide, but not in water alone at atmospheric pressure. Therefore, in the available literature, these multiblock copolymers have generally been referred to as water-insoluble, water-swellable gels.

The substrates of the present invention can be readily prepared in pure form also in the form of dry powder, in swellable cold water and in hot water well soluble in pourable, by cooling the solidifying solution similar to agar. The powder form is very suitable for transport, storage and convenient fast application. Against agar, the synthetic gel of the type described has the advantage of selecting the appropriate ones

The 252 916 production conditions can be adapted to the specific requirements of the consumer, such as the swellability, dissolution temperature and the stiffness of the cooled gel, as well as the content of acidic, cation-binding groups. The physical properties of the resulting gel largely depend on the concentration of the copolymer in the aqueous hot solution.

Example 1 wt. parts of acrylonitrile were dissolved in 80 parts of 65% colorless nitric acid, 0.3 parts of urea dissolved in a minimum amount of water and 0.2 parts of a 10% aqueous ammonium persulfate solution were added. The well-stirred solution was left under nitrogen for 12 days at room temperature and then extruded with pressurized nitrogen through a 6 mm diameter orifice into cold water. The coagulated copolymer was washed first with tap water and finally with deionized water until neutral. The heavily swollen mass was freed from excess water by dripping on a sieve and melted by heating. The solution was partially poured directly into petri dishes where it solidified into a gel directly usable as a substrate, partially converted in a spray dryer into a powder readily soluble in hot water.

Example 2 wt. 150 parts by weight of an average molecular weight of 150,000 (finely divided acrylonitrile) powder is stirred at temperatures below 5 ° C in 80% sulfuric acid to a homogeneous suspension, then the temperature is raised to 30 ° C with stirring for 3 hours. It is then poured in a thin stream into an excess of cold water. After washing with tap water until the acidic reaction disappears, it is washed again with deionized water, the swollen gel is separated and dried. After grinding, a white powder is obtained which can be redissolved in water with heat. By pouring the solution and cooling to ambient temperature, a transparent, flexible gel is obtained. If necessary, conventional nutrient solutions may be used instead of pure water to prepare the solution, depending on the purpose for which the substrate is to be used.

In the examples, nitric or sulfuric acid can be replaced in whole or in part by 9900% orthophosphoric acid252 91B

Or any mixture of said mineral acids may be used.

If the copolymer is to contain a large number of acrylic acid groups, partial alkaline hydrolysis with poly (acrylonitrile) in concentrated sodium rodanide solutions according to U.S. Pat.

AO No 179 567.

SUBJECT OF THE INVENTION

Claims (1)

SUBJECT OF THE INVENTION A substrate for the cultivation of living tissues and organisms, characterized in that it consists of a multiblock copolymer of acrylonitrile with acrylamide and / or acrylic acid or its salts, with a swellability of above 80%, preferably above 95% of water at equilibrium swelling at 20 ° C heat in water at atmospheric pressure and solidifying by cooling the obtained solution to a flexible gel, optionally containing the usual nutrients.