GB2083825A

GB2083825A - Biologically-active composites

Info

Publication number: GB2083825A
Application number: GB8126580A
Authority: GB
Original assignee: UK Atomic Energy Authority
Current assignee: UK Atomic Energy Authority
Priority date: 1980-09-11
Filing date: 1981-09-02
Publication date: 1982-03-31

Abstract

A method is disclosed for the preparation of a composite containing an active material which comprises applying an active material composition of selected rheological properties to a support material and then providing a cover for the active material composition in the form of a permeable membrane. The active material may be, for example, a biologically active species or a material capable of selectively retaining chemical species from a fluid.

Description

SPECIFICATION Improvements in or relating to composites The present invention relates to composites.

According to one aspect of the present invention there is provided a method for the preparation of a composite containing an active material which comprises applying an active material composition having selected rheological properties to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

According to one embodiment of the present invention there is provided a method for the preparation of a biologically active composite which comprises applying an active material composition having selected rheological properties and containing a biologically active species to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

According to another embodiment of the present invention there is provided a method for the preparation of a composite for the selective retention of chemical species from a fluid containing said chemical species which comprises applying an active material composition having selected rheological properties and containing an active material capable of selectively retaining the chemical species (or precursor for such an active material to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

Preferably the rheological properties of the active material composition are selected to faciiitate application thereof to support material and retention thereof on the support material prior to and during providing the permeable membrane. It is preferred that the choice of support material and the rheological properties (e.g. viscosity) of the active material composition is such that substantially none of the composition applied to the support material leaves the support material during providing the membrane.

It will be appreciated that the permeable membrane is provided to retain the active material composition on the support material.

The permeable membrane may be formed in situ, for example, around the active material composition on the support material or a preformed permeable membrane can be provided around the active material composition on the support material.

By "permeable membrane" it is meant in this specification that the membrane is capable of permitting the passage therethrough of species which are to interact (e.g. by sorption or biochemical reaction) with the active material around which the permeable membrane is provided.

Thus, in accordance with one embodiment of the invention the permeable membrane should be permeable to chemical species to be selectively retained by an active material whilst in accordance with another embodiment the permeable membrane should be permeable to species which are to participate in biochemical reactions with a biologically active species.

A permeable membrane can be formed in situ around an active material composition on a support material, for example, by applying a film of a membrane precursor dope and subsequently setting the dope to give a permeable membrane.

Thus, for example, known membrane forming techniques can be utilised wherein a membrane forming polymer dope comprising a polymer in a solvent therefor is applied as a film and the film is subsequently contacted with water to remove the solvent to precipitate a permeable polymer membrane.

In one particular embodiment the active material composition is applied to a support material and the support material and active material composition are immersed in a membrane precursor dope and then immersed in water.

In the case of forming a permeable membrane around a biologically active species it is desirable to keep the contact between the biologically active species and any polymer solvent to a minimum.

As an alternative to forming a permeable membrane in situ, a preformed permeable membrane can be fixed around the active material composition on the support material.

Thus, for example, where an active material composition and support material in accordance with the present invention is in sheet form, preformed membrane sheets can be glued around the sheet thereby to enclose it.

The preformed membrane sheets (e.g. made from cellophane, visking tube or cast cellulose acetate) can be glued together with cyanoacrylate glue.

With membrane materials which tend to change shape on becoming wet it is preferred to pre-soak preformed membrane and remove superficial water before gluing.

In circumstances where excessive gaseous products are expected to be evolved a pressure release valve can be inrorporated (e.g. at a corner of a sheet). By way of example, gaseous products will be evolved in the use of a composite to effect fermentation since CO2 will be produced.

In accordance with an embodiment of the invention the active material can be chosen, for example to have sorptive properties such that it can interact with chemical species thereby selectively to retain the chemical species. Sorptive properties include absorption and adsorption (which includes physisorption and chemisorption).

The active material can be chosen such that the interaction between it and the chemical species is predominantly chemical (e.g. ion exchange) or predominantly physical (e.g. gel permeation and gel filtration).

The active material can be chosen from a wide range of organic and inorganic materials which have useful active properties in being able selectively to retain chemical species from a fluid.

A large number of organic and inorganic materials have useful active properties in being able selectively to retain chemical species from a fluid. However many suffer from undesirable physical characteristics which can give rise to difficulties in use particularly use on a large, industrial scale. Thus, some sorbent organic materials (e.g. ion exchangers) are highly compressible and can change size at various stages in use. Some sorbent inorganic materials are finely divided powders which can be difficult to use in industrial applications, for example, due to problems in quantitative removal of suspended material from a batch stirred tank reactor or due to poor flow rates when used in a pack bed or column reactor.

The present invention may thus be utilised to make inorganic and organic materials having undesirable physical properties (as discussed above) easier to handle and use by supporting them and providing them with a permeable membrane.

Materials which may be used as the active material in accordance with the present invention include many minerals, clays and inorganic powders, (e.g. alumino silicates (such as those available under the trade names Alusil and Fullers Earth), zinc oxide, titania and hydroxylapatite) and organic materials (e.g.

ion exchangers based on a carbohydrate backbone such as DEAE-cellulose).

Alternatively, the active material may be a biologically active species.

The term "biologically active species" as used in this specification embraces, infer alia, whole cells, viable and non-viable sub-cellular particles which are biologically active and proteinaceous substances (e.g. enzymes, soluble enzyme complexes, non-soluble enzyme complexes, proteins, glyco-proteins and nucleic acids). Furthermore it is to be understood that the term "biologically active species" embraces, inter alia, those substances capable of participating in biologically specific interactions, such substances including, for example, substances of biological origin and those which act on living organisms.Substances of synthetic origin which can participate in reactions involving specific biological interactions analogous to those which can occur with naturally occuring substances are also embraced within the term "biologically active species The term "active material" as used herein includes a precursor therefor.

Thus, for example where the composite is to contain biologically active species the active material in the active material composition may not be biologically active per se but can be such that it can be activated to make it biologically active after the membrane has been provided. An example of an inactive precursor for a biologically active species is mercuripapain which can be immobilized in accordance with the present invention and subsequently "activated", to make it biologically active, in a manner known in the art by the use of EDTA. Where the active material composition has biologically active species the composite when formed will contain immobilized biologically active species. Further examples of inactive precursors are spores of microorganisms.

Preferably the active material composition of selected rheological properties is prepared by mixing the active material and a viscous substance in controlled proportions to give the required rheological properties.

The active material composition can be formed by dissolving or suspending the active material in a viscous substance.

Where the active material composition containing an active material is to be applied by spreading (e.g. with the aid of a paint brush, glass rod or plate spreader) onto a support material the composition is controlled to have a viscosity which whilst permitting even application is also sufficiently high to enable the applied composition containing the active material to remain on the support material during the provision of the membrane.

Similarly where the composition containing the active material is introduced into the pores of a support material prior to provision of the membrane, the rheological properties are selected to permit entry of the composition into the pores.

The viscous substance should be such that there is no adverse interaction with either the active material or the support material.

It has been found that active material compositions with a viscosity of approximately 1 000cp are convenient for spreading onto horizontal or sloping surfaces. However the rheological properties of an active material composition can be chosen to suit a particular circumstance. Thus, compositions having other viscosities (e.g. between a 100 and 1 0,000cp) can be useful in carrying out the method of the present invention.

The following aqueous solutions and hydrocolloid suspensions are examples of viscous substances suitable for use with an active material to form an active material composition in accordance with the present invention where the said composition is to be applied to a support material by spreading: Xanthan gum (1%) Sodium alginate (21%) Carboxymethyl cellulose-sodium salt (3%) Cellulose ethers (2%) (e.g. "Cellacol" pro duced by British Celanese, Derby) Polyvinyl alcohol (10%) Agarose (21%) Cold water soluble starch (sold under the name LAP) (5%) Cellulose paste (sold under the name Poly cel) (5%) Carrageenam (3%) The above concentrations may need minor adjustments depending upon the nature of the active material.Other non-toxic viscous substances may be used, for example, acrylic acid, polymers and co-polymers, pectins and other non-toxic polymers.

The viscous substance is chosen such that it is capable of permitting the passage therethrough of species which are to interact (e.g.

by sorption or biochemical reaction) with the active material.

Where a support material is used to provide dimensional stability to the composite it can be any suitable two- or three-dimensionally stable material capable of retaining the active material composition and the membrane.

Examples of "two-dimensionally'' stable materials are nylon mesh, steel mesh and woven materials such as loosely woven cloth (e.g. that available under the trade name "Jcloth" available from Johnson and Johnson Ltd.). In the cases of meshes and loosely woven cloth the active material composition can be enmeshed with the support in the sense that some of the composition will have passed through holes of the mesh or cloth weave with the result that, after provision of the membrane, there is composition on both sides of the mesh or cloth so that the support material is encased by and enmeshed with, active material composition. The support material thereby acts to "reinforce" the active material composition.In the case of a mesh or cloth the thickness of the active material composition is preferably not greater than that which can be protected by the mesh or cloth (e.g. typically not more than 3mm away from the face of the mesh or cloth. Where the active material composition is to be retained predominantly on the surface of a support material it is preferred that the surface of the support material is either rough, irregular, porous or otherwise capable of allowing the composition to "key" to it.

Examples of three-dimensionally stable support materials are porous materials such as porous particles made by a method as disclosed in British Patent No. 1421 531 (UK AEA).

Other examples of support materials are thin section sponges, roughened metal plates and ceramic surfaces.

It will be appreciated that in the case of a porous support material (e.g. porous particles) with inter-connected internal porosity, it can be arranged if desired, that the majority of the active material composition is present within the pores of the porous support material rather than on the surface thereof.

Thus it is to be understood that in this context "retained on the support material" is used in this specification to embrace also "retained in the support material".

Where the active material composition is to be retained on the support material predominately by being in the pores of the support material the composition containing active material may be applied to the support material in a volume sufficient just to fill the pore volume.

In accordance with an embodiment of the present invention the active material composition is applied to the support material by spreading the active material composition thereon. The spreading may be effected, for example, with the aid of a brush.

For example a sheet of composite containing an active material may be prepared by pouring an active material composition onto a horizontally stretched cloth or mesh, spreading the composition substantially evenly over the cloth or mesh and thereafter forming a membrane around the composition.

The cloth or mesh carrying the active material composition may be treated to form a membrane by dipping in a membrane precurosor dope and then dipping into water.

A sheet of composite prepared in accordance with the present invention may be used in its flat form for example to effect biochemical reactions or to effect selective retention.

Alternatively, by way of example, a sheet of composite prepared in accordance with the present invention can be wound co-axially to give a cylindrical module for use in effecting biochemical processes or selective retention.

It will be appreciated that a sheet of composite when wound co-axially gives a "swiss roll" configuration.

It is preferred that an inert spacer is provided between adjacent membrane surfaces to provide adequate lumen.

It has been found that with certain sheet support materials and certain active material compositions the strength of a coating of composition on the support material may be enhanced by forming bands of composition on the support material thereby leaving areas uncoated between the bands and around the edges.

It has also been found that in certain circumstances the integrity of the membrane may be increased by sandwiching sheets of support material, coated with active material composition, between two dry sheets of mesh.

(Where the support material is a mesh, the two sheets of mesh used to form the sandwich are preferably of larger mesh).

In the case of co-axiaX wound configuration this may be achieved by rolling a sheet of composite and a sheet of inert material together to give a substantially cylindrical "swiss roll" configuration in which radially adjacent membrane surfaces are separated by the inert material. When using this configuration it is preferred that the support material is thin (e.g. < 5mm) to give as high a membrane area as possible.

A composite may be prepared in filamentatious form in accordance with the present invention and tied together aso as to pack coaxially in a column.

According to a further aspect of the present invention there is provided a method for effecting a biochemical reaction which comprises contacting a fluid containing species for participation in a biochemical reaction with a composite wherein the composite comprises an active material composition of selected rheological properties on a support material and a cover for the active material composition, the cover being in the form of a permeable membrane.

According to yet a further aspect of the present invention there is provided a method for effecting a biochemical reaction which comprises contacting a fluid containing species for participation in a biochemical reaction with a composite prepared by a method in accordance with the present invention.

According to a further aspect of the present invention there is provided a method for the selective retention of chemical species from a fluid substance containing said chemical species which comprises contacting a fluid containing chemical species to be selectively retained with a composite wherein the composite comprises an active material composition of selected rheological properties on a support material and a cover for the active material composition, said cover being in the form of a permeable membrane.

According to yet a further aspect of the present invention that is provided a method for the selective retention of chemical species from a fluid substance containing said chemical species which comprises contacting a fluid containing chemical species to be selectively retained with a composite prepared by a method in accordance with the present invention.

Examples of chemical species which can be selectively retained using a composite material in accordance with the present invention are macromolecules (i.e. molecules of high molecular weight such as protein molecules). Particular examples of macromolecules are serum albumin, y-globulin, haemoglobin, lyzozyme, ribonuclease, phosphoglycerate kinase, lactate dehydrogenase, cytochrome c, urease, oralchumin, myoglobin, thymus DNA and yeast RNA.

The present invention provides, in a further aspect a composite containing an active material comprising an active material composition of selected rheological properties on a support material and a cover for the active material composition, the cover being in the form of a permeable membrane.

The invention also provides a composite prepared by a method in accordance with the present invention.

The invention will now be further described, by way of example only as follows: Example 1 Yeast (Sigma YSC, 1 g) was dispersed in 21% sodium alginate (Sigma, 1 Oml) to form an active material composition paste and a portion of the paste was spread onto nylon mesh (400,u, 7 x 3cm) support material. The resulting coated mesh was dropped into a membrane forming dope comprising a 10% w/v solution of cellulose acetate (Hopkin and Williams) in acetone and subsequently transferred, using forceps, to a beaker of cold water to form a membrane film around the coated mesh. The resulting composite was washed in water.

Example 2 Yeast (Sigma YSC, 1 g) was dispersed in 1% xanthan gum (Sigma, 10ml) to form an active material composition paste and a portion was spread in two narrow bands onto nylon mesh (400y, 7 X 3cm) support material. This resulting coated mesh was then sandwiched between two similar pieces of mesh and the ends held with bulldog clips.

The mesh sandwich was dipped first into a membrane forming dope comprising a 10% w/v solution of cellulose diacetate in dioxan/foramide (1 3:5) and then into water to form the membrane around the mesh sandwich.

The resulting composite was washed with water.

Example 3 Yeast (Sigma YSC, 19) was dispersed in 1% xanthan gum (Sigma, 1 Oml) to form an active material composition paste and a portion of this paste was spread onto crimped Fecralloy (Reg. Trade Mark) steel sheet (14 X 4cm) support material. The coated sheet thus produced was placed on a 15 X 7cm piece of visking tube membrane which had been pre-soaked in water. The edges of the membrane were wiped, coated with cyanoacrylate glue and then folded to seal the edge of the membrane.

Pressure was maintained on the joint for 5 minutes to allow setting, the integrity of the seal tested and the composite then washed in water.

Example 4 Yeast cells (Sigma YSC, 0.139 in 0.5ml) were suspended in 4% xanthan gum (Sigma 2.59) and basic nutrient media (7my) to form an active material composition paste and the paste was spread onto a 1 2 X 1 2cm square of J-cloth support material. This was then laid in a length of pre-soaked visking tube membrane and the ends of the tube glued with cyanoacrylate tube. The resulting composite was washed in water.

Example 5 The composite prepared in Example 1 was washed with 5% glucose solution and subsequently incubated for 1 > hours at 37' with 5% glucose solution (1 5ml) during which fermentation occurred with a loss of 1.5mg/ml of the glucose.

Example 6 The composite prepared in Example 2 was washed with a 5% glucose solution and subsequently incubated at room temperature with 5% glucose solution (1 5ml) for 1 6 hours during which fermentation occured with a loss of 4.5mg/ml glucose.

Example 7 The composite prepared in Example 3 was washed with 5% glucose solution and subsequently incubated at 37 for 2 hours with 5% glucose solution (1 5ml). Fermentation occurs resulting in a loss of 1 mg/ml of the glucose.

Example 8 The composite prepared in Example 4 was washed in 5% glucose solution and afterwards incubated at 35 with a 5% glucose solution (30ml). After 24 hours incubation there was a loss of 20.7 mg/ml glucose and a synthesis of 6.5 mg/ml ethanol.

Claims

1. A method for the preparation of a composite containing an active material which comprises applying an active material composition having selected rheological properties to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

2. A method as claimed in Claim 1 for the preparation of a biologically active composite which comprises applying an active material composition having selected rheological properties and containing a biologically active species to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

3. A method as claimed in Claim 1 for the preparation of a composite for the selective retention of chemical species from a fluid containing said chemical species which comprises applying an active material composition having selected rheological properties and containing an active material capable of selectively retaining the chemical species (or precursor for such an active material) to a support material and providing a cover for the active material composition on the support material, the cover being in the form of a permeable membrane.

4. A method as claimed in any one of Claims 1 to 3 wherein the rheological properties of the active material composition are selected to facilitate application thereof to the support material and retention thereof on the support material prior to and during providing the permeable membrane.

5. A method as claimed in any one of Claims 1 to 4 wherein the permeable membrane is formed in situ around the active material composition on the support material.

6. A method as claimed in Claim 5 wherein the permeable membrane is formed in situ by applying a film of membrane precursor dope and subsequently setting the dope to give a permeable membrane.

7. A method as claimed in Claim 6 wherein a membrane forming polymer dope comprising a polymer in a solvent therefor is applied as a film and the film is subsequently contacted with water to remove the solvent to precipitate a permeable polymer membrane.

8. A method as claimed in Claim 6 or Claim 7 wherein the active material composition is applied to a support material and the support material and active material composition are immersed in a membrane precursor dope and then immersed in water.

9. A method as claimed in any one of Claims 1 to 4 wherein a preformed permeable membrane is provided around the active material composition on the support material.

1 0. A method as claimed in Claim 9 wherein the active material composition and the support material is in sheet form and preformed membrane sheets are glued around the sheet to enclose it.

11. A method as claimed in Claim 9 or 10, wherein the preformed membrane is made from Cellophane (registered Trade Mark), visking tube or cast cellulose acetate.

1 2. A method as claimed in any one of Claims 1 and 3 to 11 wherein the active material has sorptive properties.

1 3. A method as laimed in any one of Claims 1 and 3 to 11 wherein the active material has ion exchange properties, gel permeation properties or gel filtration properties.

1 4. A method as claimed in any one of Claims 1 and 3 to 11 wherein the active material is an alimino silicate, zinc oxide, titania, hydroxylapatite or an ion exchanger based on a carbohydrate back-bone.

1 5. A method as claimed in any one of Claims 1, 2 and 4 to 11 wherein the active material is a biologically active species comprising whole cells, biologically active viable sub-cellular particles, biologically active non viable sub-cellular particles, an enzyme, a soluble enzyme complex, a non-soluble enzyme complex, a protein, a glyco-protein or a nucleic acid.

16. A method as claimed in any one of the preceding Claims wherein the active material composition having selected rheological properties is prepared by mixing the active material and a viscous substance in controlled proportions to give the required rheological properties.

17. A method as claimed in Claim 1 6 wherein the viscous substance is an aqueous solution or a hydro-colloid suspension.

18. A method as claimed in Claim 1 7 wherein the aqueous solution or hydro-colloid suspension is 1% xanthan gum, 22% sodium alginate, 3% carboxymethyl cellulose-sodium salt, 2% cellulose ether, 10% polyvinyl alcohol, 22% agarose, 5% cold water soluble starch, 5% cellulose paste or 3% carra geenam.

19. A method as claimed in any one of the preceding Claims wherein the support material is nylon mesh, steel mesh, loosely woven cloth, porous particles, thin section sponge, roughened metal plate or a ceramic surface.

20. A method as claimed in any one of the preceding Claims wherein the active material composition is poured onto a horizontally stretched cloth or mesh, the composition is spread substantially evenly over the cloth or mesh and thereafter a membrane is formed around the composition.

21. A method as claimed in any one of the preceding Claims wherein a sheet of composite is prepared and subsequently wound into a swiss roll configuration.

22. A method for the preparation of a composite material substantially as hereinbefore described with reference to any one of Examples 1 to 4.

23. A method for effecting a biochemical reaction which comprises contacting a fluid containing species for participation in biochemical reaction with a composite prepared by a method as claimed in any one of the preceding Claims.

24. A method for the selective retention of chemical species from a fluid substance containing said chemical species which comprises contacting a fluid containing chemical species to be selectively retained with a composite prepared by a method as claimed in any one of the preceding Claims.

25. A method for effecting a biochemical reaction which comprises contacting a fluid containing species for participation in a biochemical reaction with a composite wherein the composite comprises an active material composition of selected rheological properties on a support material and a cover for the active material composition, the cover being in the form of a permeable membrane.

26. A method for the selective retention of chemical species from a fluid substance containing said chemical species which comprises contacting a fluid containing chemical species to be selectively retained with a composite wherein the composite comprises an active material composition of selected rheological properties on a support material and a cover for the active material composition, said cover being in the form of a permeable membrane.

27. A composite containing an active material comprising an active material composition of selected rheological properties on a support material and a cover for the active material composition, the cover being in the form of a permeable membrane.

28. A composite prepared by a method as claimed in any one of Claims 1 to 22.