SK5232003A3 - Pravastatin sodium substantially free of pravastatin lactone and epi-pravastatin, and compositions containing same - Google Patents

Abstract

The present invention provides pravastatin sodium substantially free of pravastatin lactone and epiprava, the C-6 epimer of pravastatin. The present invention further provides a novel process for recovering pravastatin sodium from a fermentation broth in such high purity. The process includes the stages of forming an solution of the compound by extraction, obtaining an ammonium salt of pravastatin from the solution, purifying the ammonium salt of the compound and transposing the salt of the compound to pravastatin sodium.

Description

Pravastatin sodium substantially free of pravastatin lactone and epi-pravastatin, and compositions based thereon

Technical field

The invention relates to statins, more particularly pravastatin sodium and methods for its isolation from fermentation broth as a product of enzymatic hydroxylation of compactin.

BACKGROUND OF THE INVENTION

Statin drugs are currently the most therapeutically effective drugs available to reduce the level of LDL in the bloodstream of patients at risk for cardiovascular disease. This class of drugs includes pravastatin as well as compactin, lovastatin, simvastatin, fluvastatin and atorvastatin.

Pravastatin is the generic name of the chemical compound, [1S- [1a (/ 3 *, δ *) 2a, 6a, 8/3 (R *), 8aa]] -1, 2,6,7,8,8a- hexahydro- [δ], 6-trihydroxy-2-methyl-8- (2-methyl-1-oxobutyloxy) -1-naphthalene-heptanoic acid (CAS Registry 81093-370). The molecular structure of pravastatin corresponds to the general formula Ia

wherein R is hydroxy. The lactone form corresponds to the general formula Ib

R 5 4 *

(Ib) wherein the numbering of the atoms is indicated.

Pravastatin, compactin (a compound of formula Ia where R is hydrogen), lovastatin (a compound of formula Ia where R is CH ₃ ), simvastatin, fluvastatin and atorvastatin each contain an alkyl chain terminated by a carboxylic acid group bearing two hydroxy groups in the β and δ positions relative to the carboxylic acid group. The carboxylic acid group and the hydroxyl group in the δ position tend to be lactonized (see Formula Ib). Lactonizable compounds, such as statins, may be in the free acid or lactone form, or as an equilibrium mixture of both. Lactonization is a source of processing problems in the manufacture of statin drugs because the free acid form and the lactone form have different polarities. Cleaning of one mold is likely to remove the other mold together with impurities, resulting in lower yields. It is therefore usually necessary to proceed very cautiously when treating the lactonizable compounds to isolate them in high yield.

At present, it is most economical to prepare pravastatin by microbial hydroxylation of compactin at the C-6 position. Although enzymatic methods are highly stereoselective, pravastatin sodium obtained after isolation from fermentation broth is usually contaminated with a considerable amount of pravastatin C-6 epimer (epiprava). The carbon at position 6 is bisalylic and thus tends to epimerize. Minimization of epimerization requires careful control of pH and other conditions during the isolation of pravastatin. However, the known methods for the isolation of pravastatin from the fermentation broth are not suitable for isolating pravastatin in the form of the sodium salt or obtaining the sodium salt of pravastatin, which is contaminated with significant amounts of pravastatin lactone and / or epiprava. The solution according to the invention fulfills the need to develop an efficient process for the isolation of pravastatin sodium from the fermentation broth on a preparative scale to obtain a product of high purity and high yield without the need for chromatographic purification.

SUMMARY OF THE INVENTION

The present invention provides pravastatin sodium substantially free of pravastatin lactone and epiprava, the pravastatin C-6 epimer. It is a further object of the present invention to provide a process for the production of substantially pure pravastatin sodium which can be realized on an industrial scale.

In a preferred embodiment, the process comprises extracting pravastatin from the aqueous fermentation broth into an organic solvent, back-extracting pravastatin into a basic aqueous solution, and reextracting it into the organic solvent to form an organic solution containing pravastatin. The present invention makes it possible to obtain pravastatin from an organic solution in the form of a salt and subsequently to purify the salt by recrystallization. The recrystallized pravastatin salt is then converted to the sodium salt form and any excess sodium ions are captured with an ion exchange resin. The pravastatin sodium can then be isolated from the solution in a highly pure state by recrystallization, lyophilization, or other methods.

which in a way of purity

A more detailed description of the invention follows.

The subject of the invention is pravastatin sodium, substantially free of pravastatin lactone and epipraving the isolation of pravastatin sodium high from the fermentation broth.

Enzymatic hydroxylation of compactin

The enzymatic hydroxylation soil from which pravastatin is isolated can be any aqueous soil known for compactin fermentation on an industrial scale, such as the methods described in U.S. Pat. The enzymatic hydroxylation is preferably carried out using a live culture of Streptomyces and a nutrient mixture of compactin and dextrose. If the soil is neutral or basic upon completion of the fermentation, acid is added thereto to adjust the pH of the soil to about 1 to about 6, preferably to about 1 to about 5.5, and more preferably to 2 to 4. As the acid that is mention may be made of hydrochloric acid, sulfuric acid, trifluoroacetic acid or any other protic acid, preferably an acid having an IM solution in water having a pH of less than 1. By acidifying the fermentation broth, all pravastatin carboxylate salts are converted to the free acid and / or lactone. .

Isolation of substantially pure pravastatin sodium

Preferably, pravastatin is first recovered from the aqueous fermentation broth in a relatively highly concentrated organic solution by a sequence of extraction and recovery steps.

The pravastatin organic solution is an pravastatin organic solution.

extraction. preferably enriched

In the first step, pravastatin is preferably extracted from the fermentation broth. From the aqueous fermentation broth, pravastatin can be effectively extracted with C 2 -C 4 alkyl formates and C 1 -C 4 alkyl esters of C 2 -C 4 carboxylic acids, wherein the alkyl groups may be linear, branched, or cyclic. Preferred esters include ethyl formate, n-propyl formate, isopropyl formate, n-butyl formate, sec-butyl formate, isobutyl formate, tert-butyl formate, methyl acetate, ethyl acetate, n-propyl acetate, isopropyl acetate, n-butyl acetate, sec-butyl acetate, isobutyl acetate, tert-butyl acetate, methyl propionate, ethyl propionate, n-propyl propionate, isopropyl propionate, butyl propionate, methyl butyrate, ethyl butyrate, n-propyl butyrate, isopropyl butyrate, butyl butyrates, methyl isobutyrate, ethyl isobutyrate, propyl isobutyrates and butyl isobutyrates. Among these preferred organic solvents, we have found that ethyl acetate, isobutyl acetate, propyl acetate and ethyl formate are particularly suitable. The most preferred extraction solvent is isobutyl acetate. Other organic solvents may be used instead of esters. Halogenated hydrocarbons, aromatic compounds, such as carbon tetrachloride, 1,2-dichloroethane, ketones and ethers can be used. Such as dichloromethane, chloroform, benzene, methyl butyl ketone, diethyl ether and methyl tert-butyl ether.

Alternatively, pravastatin may be back-extracted into a basic aqueous solution having a pH of about 8.0 to about 9.5. The base is preferably sodium hydroxide, ammonium hydroxide or potassium hydroxide, most preferably sodium hydroxide. The extraction solvent is contacted with the basic aqueous solution until the amount of pravastatin in the organic phase is substantially depleted, as determined by thin layer chromatography or any other method, including subjective assessment that contact has been sufficient to allow complete extraction. . Multiple back extraction can be performed for optimal isolation. However, when the organic phase is isobutyl acetate, a single back extraction is highly efficient. Back extractions can be used to concentrate pravastatin by using an aqueous base in a volume less than the volume of the organic extract. Preferably, the back extraction is performed using a basic aqueous solution in a volume that is less than one-third the volume of the organic extract, preferably less than one-quarter, and most preferably is about one-fifth the volume of the organic extract.

The aqueous solution is preferably acidified with an acid, preferably trifluoroacetic acid, hydrochloric acid, sulfuric acid, acetic acid or phosphoric acid, more preferably sulfuric acid, to a pH in the range of about 1.0 to about 6.5, more preferably from about 2.0 to about 3.7.

solvent fermentation need not use the same extraction of pravastatin

Preferably, pravastatin is re-extracted into one of the organic solvents mentioned above as used to extract pravastatin from the soil. As an organic solvent, however, the solvent as used on the fermentation broth may be. In this reextraction, further enrichment of pravastatin is achieved by re-extracting to an amount of organic solvent which is preferably less than about 50% by volume of the aqueous extract, more preferably about 33% to about 20%, and even more preferably about 25% by volume of the aqueous extract. .

Pravastatin can be concentrated from 100 liters of fermentation broth to 8 liters of organic solution in 89% yield based on the initial organic extract. However, one skilled in the art will be able to achieve a higher yield of purified pravastatin by multiple extraction. However, only the preferred extraction with a single extraction has been described above. This preferred embodiment achieves a balance between solvent costs and high product yield. Deviations from this preferred method, which achieve further improvement of yield by repeated extractions and from which only one embodiment is described above, do not necessarily represent a departure from the spirit of the invention. Prior to treatment to obtain pravastatin from the organic solution by salting out, the organic solution is preferably dried, which may be carried out using conventional drying agents such as magnesium sulfate, sodium sulfate, calcium sulfate, silica, perlite and the like, and optionally decolourised with charcoal. The dried and / or decoloured organic solution is then preferably separated using conventional methods, for example by filtration or decantation.

The present invention makes it possible to obtain solid pravastatin from an organic solution by salting out. The salting-out process preferably comprises steps in which solid pravastatin is obtained in the form of a pravastatin salt. The cation of the pravastatin salt may be ammonium or an amine cation. Alternatively, the pravastatin salt cation may be an alkali metal cation. Preferred alkali metal cations are lithium, sodium and potassium.

In one embodiment of the invention, solid pravastatin from the organic solution is salted out by treatment with ammonium or an amine cation. The amine may be a primary, secondary or tertiary amine. Any alkyl or arylamine which is not hindered in such a way as to prevent the ionic interaction between the amine nitrogen and the carboxy group of pravastatin can be used. Non-limiting examples of amines include methyl-, dimethyl-, trimethyl-, ethyl-, diethyl-, triethylamine and other primary, secondary and tertiary amines having 1 to 6 carbon atoms in each of the hydrocarbon residues, and morpholine, N-methylmorpholine. , isopropylcyclohexylamine, piperidine and the like. Regardless of the absence, presence or multiple substitution at the nitrogen, the salt resulting from the reaction with ammonia or amine is hereinafter referred to as the ammonium salt. Thus, the term both salts with amines and salts with ammonia are meant.

(NH ₄ ) ₂ SO ₄ ;

(NH ₄ ) ₃ PO ₄ ;

Precipitation of the ammonium salt of pravastatin can also be induced by the addition of an ammonium salt, either alone or in combination with ammonia or amine. Among the ammonium salts, the following ammonia salts are preferred: ammonium chloride, NH ₄ Cl; ammonium bromide, NH ₄ Br; ammonium iodide, NH ₄ I; ammonium sulfate, ammonium nitrate, NH ₄ NO ₃ ; ammonium phosphate, ammonium sulfite, (NH ₄ ) ₂ S ₂ O ₄ ; and ammonium acetate,

NH ₄ OAc. Most preferred is ammonium chloride. Ammonium salts and high boiling liquid and solid amines can be added by conventional means, preferably in well ventilated areas, either as solids, liquids in substance or solutions in aqueous or organic solvents. Addition of ammonia gas requires special handling equipment Equipment such as pressure vessels and pipes are easily accessible. In a particularly preferred embodiment, pravastatin is obtained from the organic solution as an ammonium salt by adding ammonia gas and ammonium chloride to the organic solution.

with corrosive gases, regulators, valves

The temperature at which ammonia, amine and / or the ammonium salt was to be added can be determined by routine tests such that a small-scale reaction is carried out and its exothermic is monitored. Preferably, the solution temperature is not allowed to rise above 40 ° C. Although temperatures as high as 80 ° C can be used without significantly decomposing pravastatin, many solvents used for the purposes of the present invention will boil at lower temperatures. When ammonia is used, the temperature is preferably in the range of about -10 to about 40 ° C.

In an alternative embodiment, solid pravastatin can be salted out of the organic solution in the form of an alkali metal salt by adding an alkali metal cation in the form of an alkali metal salt to the organic solution. Preferred alkali metal cations are lithium, sodium and potassium.

The solid pravastatin obtained may be in the form of a crystal or in the form of an amorphous precipitate.

Preferably, as soon as the formation of solid pravastatin is stopped or it is determined by other means that pravastatin has been substantially consumed, the addition of the amine or alkali metal salt should be terminated.

The solid pravastatin of the invention may be in the form of a crystal. Alternatively, the pravastatin solid may be in the form of an amorphous solid.

When ammonia or volatile amine is used, the vessel should preferably be ventilated to disperse excess vapors. Solid pravastatin can then be isolated by filtration, decanting the solvent, evaporating the solvent or otherwise, preferably by filtration. Subsequently, the solid pravastatin can be washed, preferably with isobutyl acetate and acetone.

After optionally washing the solid pravastatin, the ammonium salt of pravastatin is preferably purified by single or multiple, most preferably triple, recrystallization. For purification, pravastatin salt is preferably dissolved in water. The polarity of the solution is preferably reduced by the addition of an anti-solvent. The anti-solvent is preferably a water-soluble organic solvent or solvent mixture in which pravastatin is poorly soluble. Most preferably, the anti-solvent is isobutyl acetate and acetone.

The pravastatin salt may be recrystallized spontaneously or induced by recrystallization by carrying out a further step in which a common ion is added.

In a preferred process wherein pravastatin is purified as an ammonium salt, recrystallization of the ammonium salt is induced by the addition of ammonium chloride.

Recrystallization can be carried out at about -10 ° C to about 40 ° C, preferably at about 0 to about 40 ° C. After the pravastatin salt is substantially recrystallized from the solution, the crystals are isolated and can be washed, for example, with a 1: 1 mixture of isobutyl acetate and acetone, and dried. The drying may be carried out at ambient temperature, but preferably at a slightly elevated temperature below 45 ° C, preferably at about 40 ° C. In order to obtain a good effect, as in the processes of Examples 3 and 4, recrystallization is optionally repeated. Each repetition is carried out with a yield of about 92%.

After purification of the pravastatin salt, the cation of pravastatin salt is preferably converted to form the sodium salt of pravastatin.

Without being limited by any theory, it is believed to be advantageous to convert the cation of pravastatin salt to form pravastatin sodium, since inorganic impurities are removed at this counterion exchange stage.

Preferably, pravastatin is released from its amine or alkali metal salt by dissolving in an aqueous solvent, the aqueous solution is acidified with any protic acid, preferably sulfuric acid, to a pH of about 2 to about 4, more preferably to a pH of about 3.1, and pravastatin is extracted with an organic solvent. The organic solvent, which may be any of the above organic solvents, but preferably isobutyl acetate, is optionally contacted with the acidified solution until pravastatin is substantially completely converted into the organic phase. Preferably, the organic phase is separated from the aqueous phase and after optional washing with water to remove residues, pravastatin is back extracted with aqueous sodium hydroxide solution at a pH of from about 7.4 to about 13.0. The back extraction is preferably carried out at a reduced temperature of from about 8 to about 10 ° C.

After extraction into aqueous sodium hydroxide, excess sodium cations are trapped to achieve an equivalence of sodium cation and pravastatin of almost 1: 1 using a water-insoluble ion exchange resin. Suitable ion exchange resins are those of the cationic and chelate type, preferably strong and weakly acidic ion exchange resins.

Strongly acid cation exchange resins that can be used include cation exchangers containing sulfonic acid groups (SO ₃ H ⁺ ). Examples include the commercial products of Amberlite® IR-118, IR-120, 252H; Amberlyst (R) 15, 36; Amberjet® 1200 (H) (Rohm &Haas); Dowex® 50WX series, Dowex HCR-W2, Dowex 650C, Dowex Marathon C, Dowex DR-2030, and Dowex HCR-S, ion exchange resins (Dow Chemical Co.); Diaion SK 102 through Diaion SK 116 resin and Lewatit SP 120 (Bayer). Preferred strongly acid cation exchange resins are Amberlite® 120, Dowex 50WX and Diaion SK series.

The weakly acid cation exchange resins include resins containing pendant carboxylic acid groups. The weakly acidic cation exchange resins include the commercial products Amberlite CG-50, IRP-64, IRC 50 and C 67, the Dowex CCR series, the Lewatit CNP series (Bayer) and the Diaion WK series (Mitsubishi), the most preferred of which are resins Amberlite® IRC 50, Lewatit CNP 80, and Diaion WK 10. Less preferred are ion exchange resins of the chelate type. Some of the types available on the market include Amberlite® IRC-718 and IRC-467.

The solution containing pravastatin sodium and excess sodium cations can be contacted with the ion exchange resin by any method known in the art, for example by passing the solution through a column or bed of resin or by mixing the solution in the vessel with sufficient resin. The method of contact is not critical. After capture of excess sodium ions, the pH of the pravastatin sodium solution should be in the range of about 7 to about 10, preferably about 7.4 to about 7.8, although the pH will vary depending on dilution. Lowering the pH of the pravastatin sodium solution from higher to lower and stabilizing it at a lower value is evidence that the capture of excess sodium ions is substantially complete. After the scavenging is substantially complete, the pravastatin sodium solution is preferably separated from the resin in the usual manner. The solution may be collected as an eluate from a column or bed, or it may be filtered, decanted, and the like.

Pravastatin sodium can be isolated from the pravastatin sodium solution by crystallization. Effective crystallization may first require partial removal of water, which may be accomplished by vacuum distillation or nanofiltration. In a preferred embodiment, the aqueous solution of pravastatin sodium is concentrated to about 20 to about 50% (w / v) prior to crystallization. If desired, after concentration of the aqueous pravastatin sodium solution, the pH can be adjusted to about 7 to about 10 using an ion exchange resin in H ⁺ form.

Crystallization can be facilitated by the addition of a water-soluble organic solvent or organic solvent mixture to the pravastatin sodium solution. In this context, mention may be made in particular of acetone and mixtures of acetone and acetonitrile, ethanol and acetonitrile, and ethanol and ethyl acetate. One of the most preferred solvent systems for crystallizing pravastatin sodium is a 1: 3: 12 mixture of water, acetone and acetonitrile, which is formed by concentrating the pravastatin sodium solution to about 30% (w / v) and then an appropriate volume of a 1: 4 mixture of acetone and acetonitrile is added thereto. The most preferred crystallization solvent mixture is a 1: 15 mixture of water and acetone.

Pravastatin sodium can also be isolated by lyophilizing an aqueous solution of pravastatin sodium.

Regardless of whether the isolation is carried out by lyophilization or crystallization or by other methods which do not reduce the purity of the product, the pravastatin sodium isolated in the practice of the present invention is substantially free of pravastatin lactone and epiprava. As shown in the examples below, it is possible to isolate pravastatin sodium containing less than 0.5 wt. % of a pravastatin lactone contamination and less than 0.2 wt. pollution generated by epiprava. Using preferred embodiments of the invention, two of which are described in Examples 1 and 3, it is possible to isolate pravastatin sodium containing less than 0.2 wt. pravastatin lactone and 0.1% epiprava.

The highly pure pravastatin sodium obtained by the process of the present invention is preferably used in the treatment of hypercholesterolemia and may be administered to mammals by any means for this purpose. The most preferred prescribed route of administration is a daily oral regimen. In people with normal liver function and moderate body weight, a reduction in serum cholesterol is typically seen when an oral dose of pravastatin sodium is administered at a daily dose of 10 mg or more. The amount of highly pure pravastatin sodium administered can be any effective amount. Preferred oral dosages comprise about 10 mg to about 40 mg pravastatin sodium. Oral dosage forms include tablets, pills, capsules, lozenges, sachets, suspensions, powders, elixirs, and the like. Substantially pure pravastatin sodium may be administered by any route, but oral administration is preferred.

The highly pure pravastatin may be administered alone or in a composition comprising pharmaceutical excipients. Regardless of whether administered alone or in the composition, the highly pure pravastatin sodium of the invention may take the form of a solution or a solid, such as a powder, granules, aggregates or any other solid form.

Compositions of the invention include tableting compositions. Depending on the tableting method used, the desired release rate and other factors, the compositions of the invention may contain fewer or more excipients. For example, the compositions of the invention may include diluents such as cellulose-derived substances such as powdered cellulose, microcrystalline cellulose, microfine cellulose, methylcellulose, ethylcellulose, hydroxyethylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, carboxymethylcellulose derivatives and other substituted and non-substituted cellulose derivatives; starch; pregelatinized starch; inorganic diluents such as calcium carbonate and dibasic phosphate and other diluents known in the pharmaceutical industry. Still other suitable diluents include waxes, sugars and sugar alcohols such as mannitol and sorbitol, acrylate polymers and copolymers, as well as pectin, dextrin and gelatin.

Other tablet excipients are binders such as resin, pregelatinized starch, sodium alginate, glucose and other binders used in wet and dry granulation and direct compression tablet preparation. Excipients which may also be present in solid compositions of novel forms of pravastatin sodium include disintegrants such as sodium starch glycolate, crospovidone, low-substituted hydroxypropylcellulose and the like. Other excipients are tabletting lubricants; such as magnesium and calcium stearate and sodium stearyl fumarate; flavoring and seasonings; sweeteners; preservatives; pharmaceutically acceptable colorants and glidants such as silica.

Capsule dosage forms may comprise a solid composition within the capsule, which is formed of gelatin or other coating material. Tablets and powders may be coated. Tablets and powders may be coated with enteric coatings. The enteric coatings of the powder forms may comprise cellulose acetate phthalate, hydroxypropyl methylcellulose phthalate, polyvinyl alcohol phthalate, carboxymethylcellulose, styrene-maleic acid copolymer, methacrylic acid-methyl methacrylate copolymer and the like and, if desired, suitable or plasticizers may be used. The coated tablet may comprise a coating on its surface or it may be compressed from powder or granules provided with enteric coatings.

The highly pure pravastatin sodium can also be administered in injectable dosage forms, as a solute or suspended solid in a sterile solution or suspension. Suitable carriers for sterile injectable dosage forms include water and oils.

The invention is illustrated by the following examples. These examples are illustrative only and do not limit the scope of the invention in any way.

DETAILED DESCRIPTION OF THE INVENTION

Example 1

Purification of pravastatin

The fermentation broth (100 liters) is acidified to about pH 2.5 to about 5.0 by the addition of sulfuric acid. The acidified fermentation broth was extracted with isobutyl acetate (3 x 50 L). The yield of isobutyl acetate extraction is 95% by HPLC analysis calibrated to an internal standard. The combined isobutyl acetate phases are extracted with water (35 L) at pH from about 7.5 to about 11.0 by the treated addition of concentrated ammonium hydroxide. The aqueous pravastatin solution obtained is again acidified to a pH of about 2.0 to about 4.0 by addition of 5M sulfuric acid and back extracted with isobutyl acetate (8 liters). The resulting pravastatin solution in isobutyl acetate was partially dried with pearlite and sodium sulfate. The pravastatin solution was decanted, freed from the drying agent by filtration and decolorized with activated carbon (1.7 g). The solution is filtered to remove charcoal and placed in a vessel equipped with a gas inlet.

Ammonia gas is introduced into the upper space above the solution with rapid stirring. Precipitated crystals of pravastatin carboxylate ammonium salt were collected by filtration and washed with isobutyl acetate and then acetone to give the ammonium salt of pravastatin of about 94% purity, as determined by HPLC coupled with UV absorbance measured at λ = 238 nm.

The pravastatin ammonium salt is further purified by crystallization from a saturated solution of airionic chloride as follows: The pravastatin salt containing 162 g of the active substance is dissolved in water (960 ml). The resulting solution was diluted with acetone (96 mL) and isobutyl acetate (96 mL) at about 35 ° C to 40 ° C. The resulting solution is cooled to about 30-32 ° C and crystallization of pravastatin ammonium salt is induced by addition of solid ammonium chloride. Ammonium chloride is added until further addition leads to no apparent further crystal growth. After addition of ammonium chloride, the solution is cooled to 0 to 26 ° C. The pravastatin ammonium salt crystals were collected by filtration, washed with isobutyl acetate and acetone as before, and dried at about 40 ° C. Crystals of the ammonium salt of pravastatin (155.5 g) are obtained with about 98% purity as determined by HPLC under the above conditions.

The pravastatin ammonium salt is purified by further recrystallization as follows. The pravastatin ammonium salt (155.5 g active ingredient) was dissolved in water (900 mL). To the resulting solution was added isobutanol (2 mL). The pH of the resulting mixture was raised to about 10 to about 13.7 by addition of concentrated sodium hydroxide solution. The resulting solution was stirred at ambient temperature for 30 minutes and neutralized to about pH 7 by the addition of sulfuric acid. The crystallization of the pravastatin ammonium salt was induced by the addition of solid ammonium chloride. The crystals (150 g) were collected by filtration and washed with acetone. HPLC analysis of the above conditions revealed that the ammonium salt of pravastatin was about 99.3% pure.

The pravastatin ammonium salt is converted to the sodium salt as follows. Pravastatin ammonium salt crystals were dissolved in water (1800 mL). Isobutyl acetate (10.5 L) was added to the aqueous solution. The resulting solution is acidified to about pH 2 to about 4 by the addition of sulfuric acid, whereby pravastatin is converted back to the free acid. The isobutyl acetate phase containing pravastatin is washed with water (5 x 10 mL). Pravastatin is then converted to the sodium salt and back extracted to another aqueous phase by vortexing the isobutyl acetate solution with water (900 to 2700 ml), intermittently adding 8M sodium hydroxide until the pH rises to about

7.4 to about 13.

The pravastatin sodium solution is then mixed with an ion exchange resin to capture excess sodium cations. After separation, the aqueous phase was stirred with IRC 50 ion exchange resin in H ⁺ form for 30 minutes at ambient temperature. Stirring is continued until a pH of about 7.4 to about 7.8 is reached.

The solution was then filtered to remove the resin and partially concentrated to 508 g under reduced pressure. Acetonitrile (480 mL) was added to the partially concentrated solution, and the resulting solution was stirred with activated carbon (5 g) to decolorize. Pravastatin sodium is obtained as crystals (90% yield) by adding additional acetone and acetonitrile to the mixture to form a 1: 3: 12 mixture of water, acetone and acetonitrile (5.9 liters) with cooling to about - 10 to about 0 ° C. Pravastatin sodium is obtained in a total yield of 65% and a purity of about 99.8% based on the starting fermented active substance and measured by HPLC under the conditions described above.

Example 2

The procedure is as described in Example 1, but recrystallization is omitted from a mixture of water, acetone and acetonitrile. Lyophilization of a concentrated solution of pravastatin sodium in water gives pravastatin sodium with about 99% purity in about 72% yield.

Example 3

The procedure of Example 1 is followed, but further purification of the ammonium salt of pravastatin is performed by repeated crystallization of the ammonium salt of pravastatin. Pravastatin sodium is obtained with a purity of 99.8% and a yield of about 68.4%.

Example 4

The procedure of Example 1 is followed, but further purification of the ammonium salt of pravastatin is carried out by repeating the crystallization of the ammonium salt of pravastatin twice. Pravastatin sodium is obtained in about 99.6% purity and in about 53% yield.

Example 5

By the method described in Example 1, the fermentation broth (100 liters) is acidified by the addition of sulfuric acid to a pH of about 2.5 to about 5.0. The acidified fermentation broth was extracted with isobutyl acetate (3 x 50 L). The combined isobutyl acetate phases are then extracted with water (35 liters) at a pH of about 7.5 to about 11.0 by the treated addition of concentrated ammonium hydroxide.

Instead of re-acidifying the aqueous extract and extracting it with isobutyl acetate to produce a further enriched organic solution carried out according to the procedure of Example 1, the aqueous extract is concentrated to a concentration of 140 g / liter under reduced pressure. The resulting concentrated solution is then acidified to pH about 4.0 to about 7.5 by the addition of 1M hydrochloric acid.

Ammonium chloride crystals (405 g) were then added to the concentrated solution and the ammonium salt of pravastatin was allowed to crystallize at ambient temperature. The crystals were collected by filtration, washed with saturated ammonium chloride solution and added to water (1 liter) at 40 ° C. After dissolution, the temperature was lowered to 30 ° C and ammonium chloride (330 g) was added. The solution was then stirred at ambient temperature for 15 hours. The crystals of the pravastatin ammonium salt are isolated by filtration, washed with isobutyl acetate and then with acetone and dried. The crystals obtained are further purified by recrystallization, converted to the sodium salt which is isolated as described in Example 1. Obtain pravastatin sodium with a purity of about 99.9% in a yield of 67.7%.

Example 6

The procedure of Example 1 is followed, but the pravastatin sodium is recrystallized from a 1: 15 mixture of water and acetone.

The product was obtained in a total yield based on the fermented active substance of 64% and a purity of 99.8% by HPLC.

Example 7

The first two paragraphs of Example 5 were followed. A concentrated aqueous extract (140 g / liter) was obtained. This extract is divided into equal parts.

Example 8

Following the procedure of Example 1, the ammonium salt of pravastatin is isolated from the fermentation broth, but the active substance is dissolved and allowed to crystallize after precipitation with ammonia gas.

The enriched pravastatin isobutyl acetate solution (6500 liters) is decolorized with activated carbon (6.5 kg) and then filtered to remove the activated carbon and transferred to a vessel equipped with a gas inlet.

The solution contains 183.2 kg of active ingredient.

The ammonium salt of pravastatin is precipitated with gaseous ammonia as described in Example 1.

The precipitated pravastatin ammonium salt is dissolved by adding water (1099 liters) to the vessel in the presence of isobutyl acetate mother liquor.

The pravastatin ammonium salt was crystallized by adding ammonium chloride (412 kg) to the vessel. Ammonium chloride is added in 31 portions over 5 hours at 30-32 ° C. The resulting suspension was stirred at 24-26 ° C for 1 hour. The crystals are filtered off, suspended in isobutyl acetate, filtered and suspended in a 2: 1 mixture of isobutyl acetate and acetone, filtered, suspended in acetone and filtered off, washed with acetone and dried under reduced pressure.

In this way, the ammonium salt of pravastatin is obtained with about 93% purity as determined by HPLC with UV detection at λ = 238 nm. The crystallized active substance weighs 168.7 kg.

Example 9

Following the procedure of Example 1, the ammonium salt of pravastatin is isolated from the fermentation broth, but crystallization is used instead of ammonia gas precipitation.

The enriched pravastatin isobutyl acetate solution (4150 mL) was decolorized with activated carbon (4.15 g) and then filtered to remove the activated carbon and transferred to a vessel.

To the isobutyl acetate solution was added water (300 mL). The pH was adjusted to 9.36 with concentrated ammonia solution (27 mL).

The pravastatin ammonium salt was crystallized by adding ammonium chloride (121.5 g) to the vessel. Ammonium chloride is added in several portions over 5 hours at 30-32 ° C. The resulting suspension was stirred at 24-26 ° C for 15 hours. The crystals are filtered off, suspended several times, washed and dried.

In this way, the ammonium salt of pravastatin is obtained with about 95% purity as determined by HPLC. The crystallized active substance weighs 42.7 g.

Example 10

By the method described in the example of pravastatin with about 93% purity.

to give the ammonium salt

The active substance (10 g) was dissolved in a mixture of water (60 ml), acetone (6 ml) and isobutyl acetate (6 ml) at 35-40 ° C. The resulting solution was cooled to 30-32 ° C. To the solution was added ammonium chloride (22 g) in several portions over 5 hours.

The resulting suspension was cooled to 24-26 ° C and stirred for an hour. The pravastatin ammonium salt is then filtered off and washed with isobutyl acetate and then acetone.

The pravastatin ammonium salt is dried at 40 ° C. The product is obtained in a yield of 96% and a purity of 97%.

Example 11

The procedure described in Example 8 gave the pravastatin ammonium salt of about 93% purity.

The active substance (10 g) was dissolved in a mixture of water (60 ml), acetone (6 ml) and isobutyl acetate (6 ml) at 35-40 ° C. The resulting solution was cooled to 30-32 ° C. To the solution was added ammonium chloride (11.4 g) in several portions over 3 hours.

The pravastatin ammonium salt is filtered off and washed with isobutyl acetate and then acetone and dried at 40 ° C.

The product is obtained in a yield of 77% and a purity of 97%.

Example 12

The procedure described in Example 8 gave the pravastatin ammonium salt of about 93% purity.

The active substance (10 g) was dissolved in a mixture of water (60 ml), acetone (6 ml) and isobutyl acetate (6 ml) at 35-40 ° C. The resulting solution was cooled to 30-32 ° C. Lithium chloride (9.3 g) was used for salt crystallization.

The pravastatin lithium salt was washed with isobutyl acetate and dried.

The lithium salt of pravastatin is obtained in a yield of 96% with a purity of 89%.

Claims (1)

PATENT CLAIMS Essentially pure pravastatin sodium. less 2. Sodium than 0.5 pravastatin sol according to % pravastatin lactone. claim 1 which contain e less 3. Sodná than 0.2 salt of pravastatin according to % epiprava. claim 1 which contain e 4. Sodium pravastatin sol according to claim 1 which contain e less than 0.5 % pravastatin lactone and less than 0.2% epiprava. less 5. Sodium than 0.2 salt of pravastatin according to % pravastatin lactone. claim 1 which contain e less 6. Sodium as 0.1 pravastatin salt according to % epiprava. claim 1 which It contains 7. Sodná a pravastatin sol according to claim 1 which It contains less than 0.2 % pravastatin lactone and less than 0.1% epiprava. 8. A substantially pure pravastatin sodium prepared by a process comprising a) degree, in who is prepare organic solution statin; b) degree, in who is obtains solid pravastatin in the form pravastatin salts; c) degree, in which sa sol pravastatin purifies recrystallization; d) the step of converting the pravastatin salt cation to form pravastatin sodium; and e) a step of isolating pravastatin sodium substantially free of pravastatin lactone and epiprava. The pravastatin sodium according to claim 8, wherein the organic solution is an enriched organic pravastatin solution. The pravastatin sodium salt of claim 8, wherein the pravastatin salt cation is an ammonium ion or an alkylammonium ion of an amine. The pravastatin sodium of claim 10, wherein the amine is selected from the group consisting of ammonium, primary amine, secondary amine, and tertiary amine. The pravastatin sodium salt of claim 8, wherein the pravastatin salt cation is an alkali metal cation. The pravastatin sodium of claim 12, wherein the alkali metal cation is selected from the group consisting of lithium, sodium, and potassium cations. The pravastatin sodium of claim 8, wherein the solid pravastatin is in the form of a crystal. The pravastatin sodium of claim 8, wherein the solid pravastatin is in the form of an amorphous precipitate. The substantially pure pravastatin sodium according to claim 8, wherein the organic pravastatin solution is prepared by extracting the aqueous fermentation broth with a first organic solvent, re-extracting pravastatin with an aqueous solution at a pH from about 8.0 to about 9.5, basic. the aqueous solution is acidified to a pH of about 2.0 to about 3.7, and the acidified aqueous solution is extracted with a second organic solvent to form an organic pravastatin solution. The substantially pure pravastatin sodium of claim 16, wherein the first and second organic solvents are isobutyl acetate. The substantially pure pravastatin sodium of claim 8, wherein the ammonium salt is purified by at least one crystallization from a mixture of water and an anti-solvent. The substantially pure pravastatin sodium according to claim 18, wherein the anti-solvent is selected from the group consisting of isobutyl acetate and acetone. The substantially pure pravastatin sodium according to claim 18, wherein ammonium chloride is added to the mixture of water and an anti-solvent to induce crystallization of the ammonium salt. The substantially pure pravastatin sodium according to claim 8, wherein the ammonium salt is converted using an acidic or chelating ion exchange resin. The substantially pure pravastatin sodium according to claim 8, wherein the pravastatin sodium is isolated by recrystallization. The substantially pure pravastatin sodium according to claim 8, wherein the pravastatin sodium is isolated by lyophilization. 24. A formulation comprising pravastatin sodium and less than about 0.5% pravastatin lactone. 25. A pharmaceutical composition comprising substantially pure pravastatin sodium according to claim First 26th A pharmaceutical dosage form characterized by by that It contains a pharmaceutical composition according to claim 25. 27th A pharmaceutical dosage form characterized by by that comprises substantially pure pravastatin sodium as claimed