Enzyme stabilisation

• US 5,240,843 A
• Filed: 06/27/1991
• Issued: 08/31/1993
• Est. Priority Date: 11/11/1988
• Status: Expired due to Term

• Current Assignee: University of Leeds
• Original Assignee: Cranfield Biotechnology, Ltd.
• Inventors: Gibson, Timothy D., Woodward, John R.
• Primary Examiner(s): Lilling, Herbert J.
• Application Number:
  US07/721,424
• Time in Patent Office: 796 Days
• Field of Search: 435/188, 435/189, 435/25
• US Class Current: 435/188
• CPC Class Codes: C12N 9/96 Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates

Abstract

A method of stabilising enzymes or other proteins against denaturation or drying comprises mixing the protein with a cyclic polyol and a cationic polyelectrolyte.

Claims

1. A method for preparing a stabilised dried enzyme comprising the steps of:

mixing an aqueous solution of an enzyme with a soluble polymeric cationic polyelectrolyte and a polyol comprising mono-, di-, trisaccharides, or reduced derivative thereof to form a mixture, and

removing water from the mixture so as to obtain the stabilised dried enzyme.

2. A method as claimed in claim 1, wherein said polyelectrolyte comprises a quaternary ammonium functionalised polysaccharide.

3. A method as claimed in claim 2, wherein said polyelectrolyte comprises diethylaminoethyl-dextran or chitosan.

4. A method as claimed in claim 1, wherein the polyelectrolyte comprises polyethyleneimine.

5. A method as claimed in claim 1, wherein the polyol is a di or trisaccharide.

6. A method as claimed in claim 5 wherein the polyol comprises lactitol, lactose, maltose, sucrose or cellobiose.

7. A method as claimed in claim 1, wherein water is removed at a temperature ranging between 4°

and 50°

C.

8. A method as claimed in claim 7, wherein the temperature ranges between 25°

and 35°

C.

9. A method as claimed in claim 1, wherein the polyol comprises lactitol, sorbitol, inositol, lactose, maltose, sucrose, or cellobiose.

10. A method as claimed in claim 1, wherein the enzyme is present in the aqueous solution at a concentration ranging from 0.5 to 200 mg/mL.

11. A method as claimed in claim 10, wherein the enzyme is present in the aqueous solution at a concentration of 1 mg/mL.

12. A method as claimed in claim 1, wherein the enzyme is present in the aqueous solution at a concentration ranging from 0.5 to 1000 units/mL.

13. A method as claimed in claim 1, wherein the amount of the polyol ranges from 1 to 20%.

14. A method as claimed in claim 13, wherein the amount of the polyol ranges from 2 to 10%.

15. A method as claimed in claim 14, wherein the amount of said polyol ranges from 5 to 10%.

16. A method as claimed in claim 1, wherein the amount of said polyelectrolyte ranges from 0.1 to 10%.

17. A method as claimed in claim 16, wherein the amount of said polyelectrolyte ranges from 0.5 to 2%.

18. A dried product containing an oxidase, a soluble polymeric cation polyelectrolyte and a polyol comprising mono-, di-, trisaccharides, or reduced derivative thereof.

19. A dried product as claimed in claim 18 wherein said polyelectrolyte is a quaternary ammonium functionalised polymeric polysaccharide.

20. A dried product as claimed in claim 18, wherein said polyelectrolyte is polyethyleneimine.

21. A dried product as claimed in claim 18, wherein said polyol is a di or trisaccharide.

22. A dried products as claimed in claim 18, wherein the amount of said polyol ranges from 1 to 20%.

23. A dried product as claimed in claim 22, wherein the amount of said polyol ranges from 2 to 10%.

24. A dried product as claimed in claim 23, wherein the amount iof said polyol ranges from 5 to 10%.

25. A dried product as claimed in claim 18, wherein the amount of said polyelectrolyte ranges from 0.1 to 10%.

26. A dried product as claimed in claim 25, wherein the amount of said polyelectrolyte ranges from 0.5 to 10%.

27. A dried product as claimed in claim 18, wherein the polyol comprises lactitol, sorbitol, inositol, lactose, maltose, sucrose, or cellobiose.