WO1998026085A1

WO1998026085A1 - Processes for producing vancomycin

Info

Publication number: WO1998026085A1
Application number: PCT/JP1997/004460
Authority: WO
Inventors: Hiroaki Nishiyama; Hajime Asada
Original assignee: Nippon Kayaku Kabushiki Kaisha
Priority date: 1996-12-11
Filing date: 1997-12-05
Publication date: 1998-06-18

Abstract

Decoloring processes appropriate for the economical production of vancomycin on an industrial scale whereby a favorable decoloring effect and a high yield can be both established, characterized by bringing vancomycin into contact with adsorbent phenol resins which can be regenerated without resort to any organic solvents. These processes are typified by one comprising passing an acidic vancomycin solution through an adsorbent phenol resin to thereby give the eluate and another one comprising passing a weakly alkaline vancomycin solution through an adsorbent phenol resin and eluting the vancomycin thus adsorbed with an acidified lower alcohol to thereby give the eluate.

Description

Akira Tsumugi Bankomaishin Manufacturing Method Technical Field

The present invention relates to a method for decolorizing vancomycin, which comprises bringing vancomycin into contact with a phenol-based adsorption resin.

Background art

Noncomimycin is a glycopeptide antibiotic produced in fermentation broth by Nocardia orientalis (see, for example, US Pat. 7 0 9 9). Purification of vancomycin from fermentation broth is carried out by first adsorbing and recovering vancomycin from the fermentation filtrate using an ion exchange resin or the like, then crystallizing the free base, and if necessary It is common practice to perform chromatography, and finally decolorize with activated carbon and then separate as a hydrochloride.

Activated carbon is generally used for decolorization of koncomisin, and its purpose is to remove impurities including dyes coming from the fermentation broth. It is generally said that activated carbon is neutral to weakly acidic and has the highest decolorization ability, but this liquid property has a good decolorization rate.Bancomycin is also adsorbed and the recovery rate decreases. (See Reference Example 1). On the other hand, if the treatment is performed with more acidic or basic to improve the recovery, the recovery rate is better, but the decolorization ability decreases. (See Reference Examples 2 and 3.) Therefore, the desired degree of decolorization cannot be achieved without increasing the amount of activated carbon used. It is difficult to balance the liquidity and the amount of activated carbon used in order to achieve both the recovery rate and the degree of decolorization, and the recovery rate has to be sacrificed. Activated carbon cannot be used repeatedly, so commercial use is not desirable. Therefore, there is a need for an economical decolorization method that balances the degree of decolorization and the recovery rate.

In the present invention, as a result of diligent studies of the above-mentioned problems, the decolorization degree of vancomycin and the recovery of vancomycin were brought into contact by contacting a phenol-based adsorption resin with an aqueous solution containing vancomycin instead of conventional activated carbon A decolorization method that can achieve both yields has been found.

That is, the present invention

1. A method for producing vancomycin, comprising a step of contacting an aqueous solution containing vancomycin with a phenol-based adsorption resin.

2. The production method according to the above-mentioned item 1, wherein the vancomycin solution having an acidic pH is passed through a phenol-based adsorption resin, and the passing solution is used as a decolorizing solution.

3. The production method according to the above 2, wherein the acidic pH is pH 3.0 to 5.0.

4. The method according to the above item 2 or 3, wherein the concentration of the vancomycin solution to be passed is 3 to 15 g ZL.

5. The production method according to the above 2, 3, or 4, wherein the load of noncomicin on the resin is from 150 to 250 g of vancomycin L resin; 6. The vancomycin solution having a weak alkaline pH is passed through a phenol-based adsorption resin, and the adsorbed vancomycin is eluted with an acidic lower alcohol aqueous solution and collected. Manufacturing method described in the paragraph,

7. The production method according to the above 6, wherein the weakly alkaline pH is pH 7.0 to 8.5.

8. The production method according to the above 6 or 7, wherein the concentration of the vancomycin solution to be passed is 3 to 15 g ZL,

9. The above item 6, 7, or 8, wherein the methanol is eluted with an aqueous solution of hydrochloric acid acidic methanol having a pH of from 10 to 50% (V / V) and a pH of from 1.0 to 3.0. Described manufacturing method,

10. The method according to the above 6, 7, 8 or 9, wherein the load of the vancomycin on the resin is 15 to 25 g of the vancomycin L resin.

11. The phenolic adsorption resin has a pore size of 15 to 35 nm and a pore volume of 0.9 to 1.6 ml / g. Manufacturing method described in,

12. The production method according to any one of items 1 to 11 above, wherein the phenol-based adsorption resin is amberlite (Amber 1 ite) 80-761;

13. The production method according to any one of the above items 1 to 12, wherein the aqueous solution containing nocomycin is prepared from a solution in which crystals of vancomycin base are dissolved.

1 4. The aqueous solution containing vancomycin was converted from the vancomycin base crystallized from the aqueous solution containing urea. The production method according to the above item 13, which is prepared,

1 5. Absorbance at 450 nm of the concentration of vancomycin in the solution of vancomycin eluted by the phenol-based adsorption resin (gZ liter) Value (A / C value), the production method according to any one of the above items 1 to 14, wherein

1 6. The vancomycin aqueous solution passed through the phenol-based adsorption resin is passed through the phenol-based adsorption resin again to allow the vancomycin to pass through, or adsorb and elute. The production method according to any one of items 1 to 15,

It is about.

BEST MODE FOR CARRYING OUT THE INVENTION

In the present invention, the phenol-based adsorption resin is a macroporous adsorption resin having a crosslinked structure composed of a phenol-formalin condensate and having a bore of 15 to 15 mm. Those having 35 nm and a pore volume of 0.9 to 1.6 mLZg are preferred.

For example, the trademark is Amber X (Amb er 1 ite) X AD-716, which is a trademark of Rohm and Haas Company of the United States and supplied by Sumitomo Chemical Co., Ltd. in Japan. This resin was previously known as Duolite S-30, Duolite S-761, and Duolite XAD-761. is there.

These phenolic adsorption resins are easy to adsorb and desorb, unlike activated carbon, and can be used repeatedly by regeneration. Also, Styrene vinylbenzene-based adsorption resin, which is currently the most commonly used adsorption resin, is an acidic or alkaline water in which the regenerant contains 50% (V / V) or more of an organic solvent. However, the phenol-based adsorption resin can be regenerated with acid or alkali that does not contain any organic solvent. We focused on the economics of phenol-based adsorbent resin and attempted to use it for decolorizing vancomycin. As a result, we succeeded in exhibiting an excellent decolorizing effect instead of activated carbon.

In the present invention, there are two methods for decolorizing vancomycin using a phenol-based adsorption resin. One method is to pass the vancomycin solution through a cellulose-based adsorption resin packed in a column, and obtain the passing solution as a decolorizing solution. The other is to allow the vancomycin solution to pass through a phenol-based adsorption resin packed in a column, adsorb the vancomycin, and elute with an appropriate eluent. Is obtained as a decolorizing solution.

When a flow-through is obtained, the pH of the bancomicin solution is adjusted to 3.0 to 5.0, preferably to pH 3.5 to 4.5. The concentration of vancomycin at this time is 3 to 15 g / L, and preferably 6 to 12 g / L. Load this solution with 150-250 g noncomycin / L resin, preferably 180-220 g non-commicin ZL resin. Let through. The transit velocity is between 1.0 and 3.0, preferably between 1.5 and 2.5. The degree of coloration of the koncomicin solution is 45 O nm absorption per vancomycin concentration (gZ liter). Evaluate by luminosity (hereinafter abbreviated as AZC) and calculate the dye removal rate from the AZC values before and after treatment. Then the dye removal rate

It is between 85 and 95%. In addition, the recovery rate of

90 to 95%. This method is effective for specifically adsorbing and removing impurities that cannot be removed by the method for obtaining an eluate described below.

To obtain an eluate, adjust the pH of the vancomycin solution to 7.0 to 8.5, preferably 7.5 to 8.0. The concentration of vancomycin at this time is 3 to 15 g ZL, preferably 5 to 11 g ZL. Adsorb 15 to 25 g of this vancomycin L resin with a load of vancomycin L resin. The adsorption speed is less than the space velocity (space Ve1ocity, SV) 2.0. After washing the resin with distilled water, elute it with an acidic lower alcohol aqueous solution. The lower alcohols include, but are not limited to, methanol, ethanol, proso, 'nor' and isopopen, but in the present application, methanol is preferred. Although various substances can be used as the acidic substance to be added, hydrochloric acid, sulfuric acid, and phosphoric acid are preferred, and hydrochloric acid is particularly preferred in the present application. For example, it contains 10% of methanol, 50% to 50%, preferably 25 to 35% (V / V), and has a pH of 1.0 to 3.0, preferably 5 to 2%. Elute with hydrochloric acid-acidic methanol aqueous solution of 5.5. The elution rate is from 0.3 to 0.7 spacevelosity (SV). Dye removal rates range from 90 to 95%. The recovery rate of corn mycin ranges from 85 to 95%. This method is Not only color but also desalting, it can be used in place of other adsorption resins.

By combining the above two methods, decolorization can be further completed. From the obtained decolorized solution, vancomycin is obtained through a step such as concentration, preferably as a hydrochloride.

Further, in the most preferred embodiment of the present invention, the vancomycin aqueous solution to be brought into contact with the phenol-based adsorption resin is prepared by adjusting the pH of the vancomycin aqueous solution containing urea to near the isoelectric point. It is preferably a solution in which a precipitate obtained by crystallizing vancomycin is dissolved.

The pH of aqueous vancomycin solution containing urea is adjusted to near the isoelectric point to crystallize noncomicin as follows.

Add 1 to 15% (wZV) of urea to the vancomycin solution and dissolve in advance. Next, the pH is adjusted from 7.5 to 8.5. The acid or base used for pH adjustment is preferably hydrochloric acid or sulfuric acid, and the base is preferably sodium hydroxide or ammonia. If the stirring is continued after the pH adjustment, precipitation (vancomycin base) occurs for a while. Crystallization recoveries range from 91 to 98%.

The sediment is collected from this batch using a centrifuge or a press filter. This precipitate is filtered at a reasonable speed with a filter cloth (twill weave) with an air permeability (cc / min / cm ² ) of around 100, which is used when filtering a precipitate that is not crystalline. It is possible. The collected precipitate is dissolved, and the above-mentioned crystallization method is repeated to further purify vancomycin. You can also get it.

The solution of the precipitate obtained is passed through a phenol-based adsorption resin, for example, Amberlite (Ambrer1ite) XAD-761, or is adsorbed and eluted to decolorize.

In the present invention, the absorbance value (AC value) at 450 nm per vancomycin concentration (g liter) is not more than 0.005, preferably not more than 0.005. It is possible to obtain a decolorization method with an extremely low degree of coloration of vancomycin. The A / C value can be obtained with a smaller degree of coloring by repeating contact with the phenol-based adsorption resin. Example

Hereinafter, the present invention will be described in detail with reference to examples, but the present invention is not limited to the examples.

Example 1

The bacterial cells were removed from the fermentation broth by using a pre-coated filter and the like to obtain a fermentation filtrate. The fermentation filtrate is passed through a strongly acidic ion-exchange resin Dion PK 208 (Mitsubishi Chemical) at a load of 20 g of Bancoma Eisin ZL resin, washed with water, and washed with 0.25 N Elution was carried out with ammonia water to obtain 60 L of eluate. The eluate had a vancomycin concentration of 18.2 g / L and a pH of 10.3. Add 6 kg (10% W / V) of powdered urea to the eluate and dissolve. Adjust the pH to 8.3 with 6 N hydrochloric acid, keep stirring at 18 ° C for 2 hours, and filter with a press filter.

The concentration of vancomycin in the filtrate was 1.6 g /. The recovery rate of the obtained vancomycin was 91.2%, and the HPLC purity was 94.2%. The obtained nokomycin precipitate was dissolved with hydrochloric acid to obtain a solution. The lysate had a pH of 4.0, A / C = 0.0174, and a concentration of 10 lg / 1. This was passed through Amberlite XAD with a load of 21.6 g vancomycin ZL resin through 761 at SV = 2.5, and the passed liquid was collected. As a result, the recovery rate from the precipitate solution was 94.8%, and the absorption at 450 nm of the vancomycin concentration (gZL) (hereinafter abbreviated as A / C) A / C = 0.02. The pigment removal rate was 87.9%.

Example 2

The vancomycin precipitate obtained in Example 1 was dissolved with hydrochloric acid in the same manner as in Example 1 to obtain a solution. The lysate had a pH of 7.6, A / C = 0.0211, and a concentration of 8.3 g ZL. This is passed through Amberlight XAD — 761 with a load of 18 g of noncomicin resin at SV = 1.0 to adsorb noncomicin. After washing the resin with water, elution is carried out with an aqueous solution of 30% methanolic acid of pH 1.5 with hydrochloric acid at SV = 0.5. The eluate had a recovery of 93.2% and an AZC of 0.0015. The pigment removal rate was 92.9%.

Example 3

The flow-through liquid (AZC = 0.0021, concentration 7.7 g ZL) obtained in Example 1 was adjusted to pH 7.9 with sodium hydroxide, and 22 g With a load of resin, the liquid was passed through Amberlite XAD — 761 at SV = 1.0, and Adsorbs cin. After washing the resin with water, elution is carried out with an aqueous solution of 30% methanol in hydrochloric acid having a pH of 2.0 at SV = 0.5. The eluate had a recovery of 91.3% and A / C = 0.002. The pigment removal rate was 90.5%.

A reference example using activated carbon is shown below.

Reference example 1

The vancomycin precipitate obtained in Example 1 was dissolved with hydrochloric acid in the same manner as in Example 1 to obtain a solution. The lysate had a pH of 2.5, A / C = 0.0168, and a concentration of 11.2 g / L. This is activated carbon with a load of 203 g vancomycin / L resin.

(Shirasaki KL, Takeda Pharmaceutical Co., Ltd.) at SV = 2.5, and the filtrate was collected. As a result, the recovery rate from the solution was 83.0%, and A / C was 0.0101. The dye removal was 39.9%.

Reference example 2

The lysate used in Reference Example 1 was adjusted to pH 6.0. AZC was 0.0173 and the concentration i 0.4 g / L. This was passed through activated carbon (Shirasagi KL, Takeda Pharmaceutical Co., Ltd.) at a load of 180 g of Bancomai Shinno L resin at SV = 2.0, and the passed liquid was collected. As a result, the recovery rate from the solution was 48.2%, and A / C was 0.001. The pigment removal rate was 99.4%.

Reference example 3

The lysate obtained in Example 1 was adjusted to pH 9.0. AC was 0.0178 and concentration was 10.3 g ZL. this Was passed through activated carbon (Shirasagi KL, Takeda Pharmaceutical Co., Ltd.) at a load of 180 g of vancomycinno L resin at SV = 2.0, and the permeate was collected. As a result, the recovery rate from the solution was 94.0%, and A / C was 0.0108. The dye removal was 39.6%.

Reference example 4

A concentration of 10.4 g / A and a pH of 7.8 with a A / C = 0.013 solution of the cocomicin solution at a load of 26 g The solution was passed through activated carbon (Shirasagi KL, Takeda Pharmaceutical), adsorbed vancomycin, washed with water, and eluted with hydrochloric acid-acid 50% ethanol water. The recovery was 79.2% and the dye removal rate was 72.1%.

Industrial applicability

According to the present invention, there is provided a vancomycin decolorizing method suitable for economical industrial production that achieves both a decolorizing effect and a good recovery rate without using activated carbon in the vancomycin production method. You.

Claims

The scope of the claims

2. The production method according to claim 1, wherein the vancomycin solution having an acidic pH is passed through a phenol-based adsorption resin, and the passing solution is used as a decolorizing solution.

3. The production method according to claim 2, wherein the acidic pH is from pH 3.0 to 5.0.

4. The process according to claim 2 or 3, wherein the concentration of the non-comicosin solution to be passed is 3 to 15 g ZL.

5. The production method according to claim 2, wherein the load of the vancomycin on the resin is 150 force, and the amount is 250 g of the vancomycin / L resin.

6. A vancomycin solution having a weak alkaline pH is passed through a phenol-based adsorption resin, and the adsorbed vancomycin is eluted and recovered with an acidic lower alcohol aqueous solution. The manufacturing method described in the item.

7. The method according to claim 6, wherein the weak alkaline pH is from pH 7.0 to 8.5.

8. The method according to claim 6, wherein the concentration of the vancomycin solution to be passed is 3 to 15 g / L.

9. Elute with methanol aqueous hydrochloric acid solution of 10 to 50% (V / V) and pH of 1.0 to 3.0 The manufacturing method according to claim 7 or 8, wherein:

10. The method according to claim 67, wherein the load of the vancomycin on the resin is 15 to 25 g of the vancomycin L resin.

11. The production method according to claim 11, wherein the phenol-based adsorption resin has a pore diameter of 1535 nm and a pore volume of 0.91.6 ml / g. Method.

12. The production method according to any one of claims 11 to 11, wherein the phenol-based adsorption resin is amberlite (Amber1ite) XAD-761.

13. The production method according to claim 11, wherein the aqueous solution containing noncomicin is prepared from a solution in which crystals of vancomycin base are dissolved.

14. The method according to claim 13, wherein the aqueous solution containing vancomycin is produced from a vancomycin base crystallized from an aqueous solution of vancomycin containing urea.

15. The absorbance value (A / C value) at 450 nm per vancomycin concentration (g Z liter) of the vancomycin solution eluted from the phenol-based adsorption resin is 0.0000. 15. The production method according to claim 11, which is 5 or less.

16. The vancomycin aqueous solution that has passed through the phenol-based adsorption resin is passed through the phenol-based adsorption resin again to allow the vancomycin to pass through, or to be adsorbed and eluted. 6. The production method according to any one of items 5.