Organic compounds -- part of the class 532-570 series – Organic compounds – Unsubstituted hydrocarbyl chain between the ring and the -c-...
Reexamination Certificate
2000-02-10
2002-07-09
Berch, Mark L. (Department: 1611)
Organic compounds -- part of the class 532-570 series
Organic compounds
Unsubstituted hydrocarbyl chain between the ring and the -c-...
Reexamination Certificate
active
06417352
ABSTRACT:
The present invention relates to a novel process for the isolation of the pharmaceutically acceptable alkali metal salt of clavulanic acid from a fermentation broth containing impure clavulanic acid comprising the steps of filtration of the fermented broth, extraction of clavulanic acid to a water immiscible or partly water immiscible solvent at a pH from 1.2 to 2, precipitation of the alkali metal salt A of clavulanic acid by addition of a solution of an alkali metal alkylalkanoate.
Clavulanic acid and its alkali metal salts and esters are used in pharmaceutical preparation to prevent the deactivation of &bgr;-lactam antibiotics. Commercial preparations of clavulanic acid contain potassium clavulanate in combination with amoxycillin trihydrate. Clavulanic acid is an unstable hygroscopic oil. Potassium clavulanate is more stable than the free acid or other salts, and is therefore most frequently, used for commercial preparations.
Clavulanic acid and its derivatives are inhibitors of the &bgr;-lactamases. The resistance of &bgr;-lactam antibiotics is associated with inactivation of &bgr;-lactam structure due to the opening of &bgr;-lactam ring by &bgr;-lactamase produced by bacteria. Thus the inactivating enzymes are commonly called as &bgr;-lactamase, they are divided into penicillinase and cephalosporinase.
Furthermore clavulanic acid itself is believed to have an antibacterial activity.
Clavulanic acid is produced from various strains of microorganism by a fermentation process. For this process for example strains belonging to the genus Streptomyces such as
S. clavuligerus
NRRL 3585 (U.S. Pat. No. 4,110,165),
S. jumonjinensis
NRRL 5741 (British Patent 1,563,103),
S. katsurahamanus
IFO 13716 (Japanese Patent 83,009,579), and Streptomyces sp. P6621 FERM 2804 (Japanese Patent 55,162,993) are used. For the preparation of clavulanic acid by a fermentation process the microorganism
Streptomyces clavuligerus
is preferred.
In the state of art different processes for the preparation and purification of the clavulanic acid containing fermentation broth are described.
GB 1 508 977 discloses preparation of clavulanic acid and its salts by filtration of the fermentation broth by passage through an anionic exchange resin.
GB 1 543 563 discloses a fermentation process wherein the pH value of the medium is maintained in the range of 6.3 to 6.7. A pharmaceutically acceptable salt such as potassium clavulanate is prepared by ion exchange process from lithium clavulanate. However lithium clavulanate is not a pharmaceutically acceptable salt. Therefore it is necessary to add an ion exchange process as a further step for preparing a pharmaceutically acceptable form of the compound. Furthermore, the remaining salt lithium chloride is soluble in organic solvents and therefore it is difficult to separate the lithium chloride in the aqueous phase during the extraction process.
Further documents of the state of the art like EP-0 647 229 describe the use of amine salts of clavulanic acid as intermediate compounds for the preparation and purification of clavulanic acid and its alkali metal salts. EP-0 647 229 for example describes a process for the preparation of a purified clavulanic acid or a salt or ester thereof by preparing a diamine salt of clavulanic acid and converting this intermediate compound into clavulanic acid or a pharmaceutically acceptable salt or ester. The conversion is made by adding for example potassium 2-ethylhexanoate and precipitating potassium clavulanate. Many of such amines are either unsuitable for the production of a salt of clavulanic acid or they give rise to amine salts of clavulanic acid which are either hygroscopic or toxic or both and, therefore, are unsuitable for use as intermediates for the preparation of a pharmaceutically acceptable compound.
Other purification processes of the state of the arts are performed without any amine compounds. For example WO 95/34194 A2 describes a process for manufacturing an alkali metal salt of clavulanic acid wherein impure clavulanic acid in aqueous solution is extracted by a solvent mixture of ketone and alkyl acetate under acidic condition. The solution is than treated in a conventional manner and the solution of an alkali metal salt of alkanoic acid dissolved in ketone or alkanol solvent is added to obtain pure alkali metal salt of clavulanic acid. Thus the process according to this state of the art omits the step of formation of amine salts. This process has the advantage that the use of mostly toxic amines is no longer necessary. In a preferred embodiment as alkali metal salt of alkanoic acid sodium or potassium salts are used, especially potassium 2-ethylhexanoate.
A similar process is also described in WO 96/28452 A1. This process comprises the steps of removing solids from a clavulanic acid containing fermentation broth by microfiltration, acidifying the filtrate to a pH between 1 and 3, extracting the acidified filtrate with a water immiscible solvent and separating the clavulanic acid containing extract. This extract is mixed with a metal donor and at least one additional solvent. From the solution the metal clavulanate salt is separated.
As metal donor compounds organic salts, carbonates, bicarbonates or hydroxides of potassium, sodium, lithium or magnesium can be used. The use of carboxylic acid salt is preferred. Further preferred metal donors include potassium 2-ethylhexanoate, potassium acetate, lithium 2-ethylhexanoate and lithium acetate.
EP-0 182 522 B1 also describes a process for the preparation of clavulanic acid and its salts and esters. In this process the fermentation broth is worked up as follows. The solids are removed by filtration or centrifugation. The broth is acidified to a pH of 1 to 3 and clavulanic acid is extracted by adding a water immiscible solvent with two phases being separated for example by centrifugation. This gives the clavulanic acid in the water immiscible phase. The solution is purified by mixing it with the dissolved lithium 2-ethylhexanoate solution isolating lithium clavulanate and optionally converting the lithium salt to other salts or an ester. The conversion of the lithium salt to other salts is carried out by ion exchange procedures using ion exchange resins in the form of the desired cation preferably sodium or potassium.
The processes of the state of the art referred above have the disadvantage, that the alkali metal salts prepared by direct precipitation are not pure enough for a pharmaceutical use. Therefore, further purification steps like recrystallization, purification over a column etc. are necessary.
The object of the invention therefore is to prepare clavulanic acid and its pharmaceutically acceptable alkali metal salts such as potassium clavulanate in a new and simple manner wherein the desired substance is obtained in a very high yield without any additional purification steps and of high purity avoiding the use of toxic amines or lithium compounds.
This technical problem is solved by a process which is characterized by the following steps:
before the filtration of the fermentation broth, the fermentation broth containing clavulanic acid is diluted with water, a flocculating agent is added and the pH is adjusted to pH 3 to 5,
for further purification the alkali metal salt A of clavulanic acid is converted to clavulanic acid by addition of an inorganic acid and is extracted into a water immiscible or partly water immiscible solvent,
and to the solution of clavulanic acid a solution of a different alkali metal alkylalkanoate B is added and the alkali metal salt B of clavulanic acid is precipitated.
Suitable salts according to the present invention are pharmaceutically acceptable alkali metal salts and alkaline earth metal salts like sodium, potassium, calcium and magnesium salts.
Of these compounds potassium clavulanate is the most stable compound which is normally used for pharmaceutical preparations. The clavulanic acid itself is an unstable hygroscopic oil which is not used for the preparation of pharmaceutical compounds.
In a preferred embodiment the al
Berch Mark L.
CIPAN-Companhia Industrial Produtora de Antibioticos, S.A.
Kilpatrick & Stockton LLP
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