GB2153823A - Purification of cephalosporins - Google Patents

Abstract

A new process of purifying oxacephalosporins comprises contacting an aqueous solution of an oxacephalosporin carboxylic acid or derivative (at a pH lower than its pKa) with a macroporous polymeric adsorbent (specific gravity: higher than 1.05; and saturated adsorption capacity for cephalosporin C : larger than 60 mg/ml) to achieve separation from non-adsorbable contaminants, and then eluting with an aqueous hydrophilic organic solvent to separate the product desired from non-elutable contaminants.

Description

SPECIFICATION Purification of Cephalosporins This invention relates to a process for purifying oxacephalosporins useful as antibacterials.

In the production of oxacephalosporins, contaminants are generally removed by repeated extraction, precipitation, crystallization, adsorption by oder types of synthetic adsorbents (e.g., Amberlite XAD-2, XAD-4, XAD-7, XAD-8, Diaion HP-i 0, HP-20, HP-30, HP-40, and HP--50), elution, or the like, in the work up procedure.

The present invention is based upon the discovery that the efficiency of purification can be promoted by using a particular macro porous polymeric adsorbent (specific gravity: higher than 1.05; and saturated adsorption capacity for cephalosporin C: larger than 60 mg/ml, i.e., more than 1,2 times as high as adsorbents mentioned in the literature).

It is known to use macroporus polymeric absorbents to remove electro-neutral materials, low molecular weight substances, or the like, from cephalosporin aqueous solutions [e.g., Japanese Patent Publications (Kokoku) SHO-52-50799, 54-16922; and Japanese Patent Application Publications (Kokai) SHO--500-106996, 51-110587].

However, the known methods have several defects (e.g., lower adsorption capacity of the resins, turbidity of eluted solvent, and difficulty in handling due to small specific gravity).

In contrast, the macroporus polymeric adsorbent used in this invention (having a specific gravity of higher than 1.05 and a saturated adsorption capacity for cephalosporin C of larger than 60 mg/ml, i.e. more than 1.2 times as high as adsorbents mentioned in the literature) has no such defects, and is suitable for purifying the unstable oxacephalosporin free carboxylic acids.

Thus, the invention provides a process for purifying an oxacephalosporin, which process comprises contacting an aqueous solution of an oxacephalosporin carboxylic acid or derivative thereof at a pH lower than its pKa with a macroporus polymeric adsorbent of specific gravity higher than 1.05 and saturated adsorption capacity for cephalosporin C larger than 60 mg/ml so as to separate the oxacephalosporin from any non-adsorbable contaminants, and then eluting the adsorbent with an aqueous hydrophilic organic solvent to separate the oxacephalosporin from any nonelutable contaminants.

Typical nonionic macroporus polymeric adsorbents are Amberlite XAD-2000, a styrene divinylbenzene copolymer resin produced by Rohm and Haas Co., and Diaion SP-206 and SP-207, styrene divinylbenzene copolymer resins produced by Mitsubishi Chemical Industries Limited.

The oxacephalosporins which may be purified by the process of this invention are 1-dethia-1- oxa- 3-cephem-4-carboxylic acids or their derivatives having at least one carboxy and having acylamino at position 7 , hydrogen or methoxy at position 7ee, and R or a CH2R group at position 3 (where R is hydrogen, halogen, alkyl, carbamoyl, carboxy, hydroxy, alkoxy, acyloxy, alkylthio, haloalkylthio, arylthio, heterocyclic thio, pyridinium, substituted pyridinium or a like nucleophilic group).

The acyl moiety in the acylamino group may be one found in amido side chains in penicillin or cephalosporin chemistry. The heterocyclic group is a mono- or di-cyclic heterocyclic group having one or more hetero atoms selected from nitrogen, oxygen, and sulfur, and optionally substituted by alkyl or R-substituted alkyl. The said group(s) may have substituent groups R or other substituents convention in cephalosporin chemistry.

The amount of the macroporus polymeric resin in this invention is preferably 5 to 100 times the weight of the oxacephalosporin free carboxylic acid being treated.

The concentration of the oxacephalosporin carboxylic acid is preferably 0.1 to 30 w/w%. Preferably, the solution contains no adsorption inhibiting substance (for example, an organic solvent) over inhibiting concentration. The operating temperature is preferably 0 to 500C.

Adsorption may be effected by contacting an aqueous solution of an oxacephalosporin with the said macroporus polymer adsorbent by a conventional method (e.g., a batch method, column method, or chromatographic method).

In order to remove non-adsorbable materials from the adsorbed material, the resin on which oxacephalosporin has been adsorbed is washed with a liquid which can wash out such non-adsorbable contaminants without eluting the desired substance. Such liquids can be, for example, water, a buffer solution, a salt solution, or an aqueous surfactant solution. Here again, conventional methods (e.g., batch methods, column methods, chromatographic methods, or gradient methods) are applicable. Washing can be continued until no impurity is detectable in the effluents from the resin. The preferred temperature is 0 to 50"C.

For eluting the adsorbed objective material, it is preferred to use an aqueous organic solvent (e.g., an ester, ketone, alcohol, or ether, especially a lower alkanol). This may contain a substance for controlling eluting efficiency or independent of the elution (e.g., an inorganic salt, acid, base, or cation surfactant). The elution may be carried out by conventional operation (e.g., a batch method, column method, chromatographic method, or gradient method), preferably at 0 to 50"C. The eluting procedure is stopped when the objective material has been eluted and the non-elutable contaminants remain on the adsorbent.

The thus-used absorbent can be recovered for using again by conventional operations (with, e.g., aqueous sodium hydroxide, neutral or acidic methanol or isopropanol, or a mixture thereof).

The oxacephalosporin can be recovered from the effluents by conventional operations (e.g., extraction, concentration, precipitation, crystallization, or lyophilization).

Each step of this invention can be carried out at, for example, room temperature or at 0 to 50"C for 30 minutes to 50 hours.

The oxacephalosporin carboxylic acids produced by this invention with low energy consumption are highly pure and may be obtained in high yield.

The invention provides a pharmaceutical or veterinary formulation comprising an oxacephalosporin which has been purified by a process of the invention and formulated for pharmaceutical or veterinary use, respectively. Such formulations are prepared in accordance with standard procedures in the art and may, again in accordance with standard procedures, include acceptable carriers, diluents or excipients and/or be in unit dosage form.

The following Examples describe embodiments of this invention to illustrate but not limit the same.

Example 1 A 0.11% crude latamoxef sodium [i.e., 7(3-(2- p--hydroxyphenyll- 2-carboxyacetamido)- 3--(1- methyltetrazol- 5--yl]thiomethyll- 7cl--methoxyy- 1-dethia- 1-oxa-3-cephem-4-carboxylic acid disodium salt] aqueous solution (produced by hydrolyzing latamoxef dibenzhydryl ester with aluminium chloride and removing organic solvents from the reaction mixture) is adjusted to pH 1.5 with hydrochloric acid.

A nonionic macroporus polymeric adsorbent (Amberlite XAD--2000, a styrene-divinylbenzene copolymer adsorbent resin produced by Rohm and Haas Co.) (200 ml) is packed to 270 mm height in a glass tube which has 32 mm inside diameter and 350 mm length. The said solution (2780 g) is passed through the packed column to adsorb latamoxef. This column is washed with water (1360 ml) and then with 50% methanol (125 ml) to remove non-adsorbing contaminants (e.g., salts).

Then, the latamoxef is eluted with 70% methanol (535 ml). The effluents contain the objective material at a concentration of 0.5% when estimated ultraviolet spectrophotometrically.

The effluents are concentrated under diminished pressure, neutralized with aqueous sodium hydroxide, and lyophilized to obtain pure latamoxef sodium. Yield: 86%.

Example 2 A 0.19% crude latamoxef aqueous solution is adjusted to pH 2.9.

A nonionic macroporus adsorbent (Amberlite XAD-2000, a styrene-divinylbenzene copolymer resin produced by Rohm and Haas Co.) (100 ml) is packed to 255 mm height in a glass tube which has 22 mm inside diameter. The said solution (4800 ml; containing 8.02 g of the said compound) is passed through the packed column to adsorb the objective material. This column is washed with water (2480 ml). Then purified objective material is eluted with 50 to 80% methanol (810 ml). The effluents contain 68.7% of the feeding objective material when estimated ultra-violet spectrophotometrically.

Example 3 A 0.94% crude latamoxef aqueous solution is adjusted to pH 2 to 3.

A nonionic macroporus polymeric adsorbent (Diaion SP-207, a styrene-divinylbenzene copolymer resin produced by Mitsubishi Chemical Industries Limited) (40 ml) is packed to 115 mm height in a glass tube which has 19 mm inside diameter.

The said solution (200 ml; containing 1.88 g of the said compound) is passed through the packed column to adsorb latamoxef. This column is washed with water (200 ml). Then the purified objective material is eluted with 50% acetone (220 ml). The effluents contain 93.3% of the feed objective material when estimated from high precision liquid chromatogram.

Similarly, the eluting solvent, 50% acetone, can be replaced by 70% isopropanol to obtain the same pure product in high yield.

Example 4 A 1.0% crude 7(3- difluoromethylthioacetam- ido-3- [1-(2-hydroxyethyl) tetrazol--55-yl] thiomethyl-7a-methoxy-1- dethia-1-oxa- 3-cephem- 4--carboxylic acid aqueous solution is adjusted to pH about 2.

A nonionic macroporus polymeric adsorbent (Diaion SP-207, a styrene-divinylbenzene copolymer adsorbent resin produced by Mitsubishi Chemical Industries Limited) (40 ml) is packed to 105 mm height in a glass tube which has 22 mm inside diameter. The solution (210 ml; containing 2.0 g of the said compound) is passed through the packed column to adsorb the objective material.

This column is washed with water (100 ml). Then purified objective material is eluted with 70% isopropanol (140 ml). The effluents contain 87.1% of the feed objective material when estimated ultraviolet spectrophotometrically.

Example 5 A 1.0% crude 7P-difluoromethylthioacetamido- 3 -[l-(2-hydroxyethyl)tetrazol-5 --yl] thiome thyl-7-methoxy-1 -dethia-1-oxa-3-ce- phem-4-carboxylic acid aqueous solution is adjusted to pH about 2.

A nonionic macroporus polymeric adsorbent (amberlite XAD--2000, a styrenedivinylbenzene copolymer adsorbent resin produced by Rohm and Haas Co.) (30 ml) is packed to 275 mm height is a glass tube which has a 12 mm inside diameter.

The said solution (206 ml; containing 2.0 g of the said compound) is passed through the packed column to adsorb the objective material. This column is washed with water (300 ml). Then purified objective material is eluted with 70% isopropanol (80 ml). The effluents contain 99.4% of the feed objective material when estimated ultra-violet spectrophotometrically.

Example 6 A 0.15% crude 7(3-[2-(2-carbamoyl-2-fluo- rovinyl-thio) acetamidoj-3- [1-(2-hydroxy- ethyl) tetrazoI-yl]-thiomethyl- 7a- methoxy-1-dethia-1-oxa- 3-cephem4 carboxylic acid aqueous solution is adjusted to pH about 2.

A nonionic macroporus polymeric adsorbent (Diaion SP-207, a styrene-divinylbenzene copolymer resin produced by Mitsubishi Chemical Industries Limited) (30 ml) is packed to 132 mm height in a glass tube which has 16 mm inside diameter.

The said solution (200 ml; containing 0.3 g of the said compound) is passed through the packed column to adsorb the objective material. This column is washed with water (200 ml). Then the purified objective material is eluted with 70% isopropanol (150 ml). The effluents contain 77.5% of the feed objective material when estimated ultra-violet spectrophotometrically.

Claims (11)

1. A process for purifying an oxacephalosporin, which process comprises contacting an aqueous solution of an oxacephalosporin carboxylic acid or derivative thereof at a pH lower than its pKa with a macroporus polymeric adsorbent of specific gravity higher than 1.05 and saturated adsorption capacity for cephalosporin C larger than 60 mg/ml so as to separate the oxacephalosporin from any non-adsorbable contaminants, and then eluting the adsorbent with an aqueous hydrophilic organic solvent to separate the oxacephalosporin from any nonelutable contaminants.

2. A process as claimed in claim 1 and effected at 0 to 50"C.

3. A process as claimed in claim 1 or claim 2, wherein the oxacephalosporin carboxylic acid is 7(3-(2-p-hydroxyphenyl- 2-ca rboxyacetam- ido)-- 3-(l-methyltetrazol-5-yl) thiomethyl- 7o-methoxy-1 -dethia- 1 -oxa-3-cephem- Sca rboxylic acid, 7(3-difluoromethylthioacetam- ido-3- [1-(2-hydroxyethyl) tetrazol--55-yl] thiomethyl-7-m ethoxy-1- dethia-1-oxa- 3-cephem-4-carboxylic acid, or 7(3-[2-(2- carbamoyl-2-fluorovinylthio) acetamido]-3- [1-(2-hydroxyethyl )tetrazol--55-yl] th iomethyl- 7a-methoxy-1-dethia-1 -oxa-3-cephem- 4-carboxylic acid.

4. A process as claimed in any one of claims 1 to 3, wherein the macroporus polymeric adsorbent is Amberlite XAD-2000, Diaion SP-206, or Diaion SP-207.

5. A process as claimed in any one of claims 1 to 4, wherein the amount of the macroporus polymeric adsorbent is 5 to 100 times the weight of the feed oxacephalosporin.

6. A process as claimed in any one of claims 1 to 5, wherein the eluting solvent is 50 to 80% methanol, ethanol, propanol, or isopropanol, or an acetone aqueous solution.

7. A process as claimed in claim 1 and substantially as hereinbefore described.

8. A process as claimed in claim and substantially as hereinbefore described in any one of the Example.

9. A pharmaceutical or veterinary formulation comprising an oxacephalosporin which has been purified by a process as claimed in any one of claims 1 to 8 and formulated for pharmaceutical or veterinary use, respectively.

10. A formulation as claimed in claim 9 and in unit dosage form.

11. A formulation as claimed in claim 9 or claim 10 and also comprising a pharmaceutically acceptable or veterinarily acceptable, respectively, diluent, carrier or excipient.