# US 10,308,923 B2 · Medium composition for preparing botulinum toxin

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- Number: US 10,308,923 B2
- Document ID: US10308923B2
- Status: Granted
- Kind: B2
- Application number: 15/521,852
- Filed: 2016-04-28
- Published: 2017-09-14
- Granted: 2019-06-04
- Earliest claimed: 2015-04-28
- Assignees: Daewoong Co., Ltd.
- Applicants: Daewoong Co., Ltd.
- Inventors: Kyoung-Yun Kim; Hye-Young Sul; Kyoung-Min Min
- Web page: https://patentgrep.com/patent/US10308923B2

## CPC

- C12N 9/52: derived from bacteria {or Archaea}
- C07K 14/33: from Clostridium (G)
- C12N 1/20: Bacteria; Culture media therefor
- C12Y 304/24069: Bontoxilysin (3.4.24.69), i.e. botulinum neurotoxin
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- C07K: PEPTIDES (peptides in foodstuffs A23; obtaining protein compositions for foodstuffs, working-up proteins for foodstuffs A23J; preparations for medicinal purposes A61K; peptides containing beta-lactam rings C07D; cyclic dipeptides not having in their molecule any other peptide link than those which form their ring, e.g. piperazine-2,5-diones, C07D; ergot alkaloids of the cyclic peptide type C07D519/02; macromolecular compounds having statistically distributed amino acid units in their molecules, i.e. when the preparation does not provide for a specific; but for a random sequence of the amino acid units, homopolyamides and block copolyamides derived from amino acids C08G69/00; macromolecular products derived from proteins C08H1/00; preparation of glue or gelatine C09H; single cell proteins, enzymes C12N; genetic engineering processes for obtaining peptides C12N15/00; compositions for measuring or testing processes involving enzymes C12Q; investigation or analysis of biological material G01N33/00)
- C12Y: ENZYMES

## Abstract

The present invention relates to a medium composition for production of botulinum toxin and, more particularly, to a medium composition for culture of Clostridium sp. capable of producing botulinum toxin. The medium composition of the present invention comprises a porcine peptone and at least one plant-derived peptone selected from the group consisting of a garden pea hydrolysate, a cotton seed hydrolysate and a wheat gluten hydrolysate. When the medium according to the present invention, which contains plant-derived peptones, porcine peptones and minerals, is used for culture of Clostridium botulinum, the growth rate of the bacterium in the medium is higher than that in each of the medium that is in current use and the medium comprising plant-derived peptones alone. In addition, when the medium of the present invention is used, a high concentration of botulinum toxin can be produced by culturing the bacterium in a safe manner.

## Claims

### Claim 1

A medium composition for culture of Clostridium botulinum, the medium composition comprising: plant-derived peptones comprising a garden pea hydrolysate, a cotton seed hydrolysate, and a wheat gluten hydrolysate, in a ratio of 1:0.24-43.62:0.01-50.57 by weight, and a porcine peptone.

### Claim 2 (depends on 1)

The medium composition of claim 1, wherein the plant-derived peptones are comprised with a content of 0.1-10% (w/v).

### Claim 3 (depends on 1)

The medium composition of claim 1, wherein the porcine peptone in the medium composition is comprised with a content of 0.2-10% (w/v).

### Claim 4 (depends on 1)

The medium composition of claim 1, wherein the porcine peptone is a hydrolysate comprising about 54.91-60.69 wt % of peptides having a molecular weight of 500 Da or less, or a hydrolysate comprising about 38.48-42.53 wt % of peptides having a molecular weight of 500 Da or less.

### Claim 5 (depends on 1)

The medium composition of claim 1, wherein the plant-derived peptones or the porcine peptones are subjected to an enzyme treatment.

### Claim 6 (depends on 1)

The medium composition of claim 1, further comprising a carbon source and at least one mineral selected from the group consisting of K2HPO4 (dipotassium phosphate), Na2HPO4 (disodium phosphate) and KH2PO4 (monopotassium phosphate).

### Claim 7 (depends on 6)

The medium composition of claim 6, wherein the mineral in the medium composition is comprised with a content of 0.05-3.5% (w/v).

### Claim 8 (depends on 1)

A method for producing botulinum toxin, comprising the steps of:
(a) culturing Clostridium botulinum using the medium composition of claim 1 to produce botulinum toxin; and
(b) recovering the produced botulinum toxin.

### Claim 9 (depends on 8)

The method of claim 8, wherein the culture is performed under anaerobic conditions.

### Claim 10 (depends on 8)

The method of claim 8, wherein the botulinum toxin is selected from the group consisting of botulinum toxin serotypes A, B, C, D, E, F and G.