Process for the preparation of purified vancomycin class antibiotics

• US 4,667,024 A
• Filed: 07/13/1983
• Issued: 05/19/1987
• Est. Priority Date: 07/13/1983
• Status: Expired due to Fees

• Current Assignee: Pfizer Inc.
• Original Assignee: Smithkline Beecham Corporation (Smithkline Beecham PLC)
• Inventors: Snader, Kenneth M., Sitrin, Robert D., Wasserman, Gail F.
• Primary Examiner(s): Brown, Johnnie R.
• Application Number:
  US06/513,511
• Time in Patent Office: 1,406 Days
• Field of Search: 536/16.8, 536/16.9, 536/128, 536/18.5, 536/127, 536/22
• US Class Current: 536/16.9
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54353 with ligand attached to the carrier via a chemical coupling agent coatings G01N33/54393
• CPC Class Codes: G01N 33/9446 Antibacterials

Abstract

A novel process for the isolation and purification of vancomycin class antibiotics which utilized affinity chromatography by the formation of a sorption complex between the antibiotic and an immobilizing ligand selected from -D-alanyl-D-alanine or -X-D-alanyl-D-alanine, wherein X is an amino acid radical and the novel affinity chromatography sorbent employed therein are disclosed.

Claims

1. An affinity chromatography process for the preparation of a purified vancomycin class antibiotic which comprises:

(1) contacting an impure solution containing the antibiotic with a solid carrier matrix to which immobilizing ligands selected from -D-alanyl-D-alanine or -X-D-alanyl-D-alanine, wherein X is an amino acid radical, has been attached;

(2) thereby, selectively forming an affinity sorption complex between the antibiotic and the immobilizing ligand;

(3) washing the impurities from the matrix and complex attached thereto; and

(4) dissociating the complex between the antibiotic and the immobilizing ligand to recover said purified antibiotic.

2. A process according to claim 1 wherein the antibiotic is selected from the group consisting of vancomycin, ristocetin, actinoidin, avoparcin, actaplanin, teichomycin A₂, LL-AM-374, A 477, OA 7653, A35512B, AAD 216 complex and the individual factors of said antibiotics.

3. A process according to claim 2 wherein the antibiotic is vancomycin.

4. A process according to claim 2 wherein the antibiotic is ristocetin.

5. A process according to claim 2 wherein the antibiotic is AAD 216 complex.

6. A process according to claim 2 wherein the antibiotic is avoparcin.

7. A process according to claim 2 wherein the antibiotic is teichomycin A₂.

8. A process according to claim 2 wherein the antibiotic is A35512B.

9. A process according to claim 1 wherein the dissociation of the sorption complex is accomplished by employing a basic buffer solution and a polar organic solvent.

10. A process according to claim 9 wherein the basic buffer solution is selected from the group consisting of 0.4 M sodium bicarbonate at pH 9.5, 0.5M triethylammonium carbonate at pH 9, 0.25M sodium bicarbonate at pH 9 and 0.25M sodium bicarbonate at pH 9.5.

11. A process according to claim 9 wherein the polar organic solvent is acetonitrile.

12. A process according to claim 1 wherein tne impure solution is a fermentation broth.

13. A process according to claim 1 wherein the immobilizing ligand is -D-alanyl-D-alanine.

14. A process according to claim 1 wherein the immobilizing ligand is selected from the group consisting of α

-N-acetyl-L-lysyl-D-alanyl-D-alanine, ε

-N-acetyl-L-lysyl-D-alanyl-D-alanine, glycyl-D-alanyl-D-alanine, L-alanyl-D-alanyl-D-alanine, L-ornithyl-D-alanyl-D-alanine, L-tryrosyl-D-alanyl-D-alanine and D-alanyl-D-alanyl-D-alanine.

15. A process according to claim 14 wherein the immobilizing ligand is α

-N-acetyl-L-lysyl-D-alanyl-D-alanine.

16. A process according to claim 1 wherein the solid matrix support is selected from the group consisting of agarose, dextran, cellulose, polystyrene, polyacrylamide, silica, controlled pore glass, nylon, acrylic copolymers and polyesters.

17. A process according to claim 16 wherein the solid matrix support is selected from the group consisting of agarose, Sepharose and acrylic copolymers.

18. A process according to claim 17 wherein the solid matrix support contains spacer arms.

19. A process according to claim 18 wherein the solid matrix support is Affi-Gel®

10.