Organic compounds -- part of the class 532-570 series – Organic compounds – Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
2001-02-27
2003-02-18
Owens, Amelia (Department: 1625)
Organic compounds -- part of the class 532-570 series
Organic compounds
Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
active
06521762
ABSTRACT:
FIELD OF THE INVENTION
The invention relates to a process of purifying Lovastatin or Simvastatin, which reduces the level of dimeric impurities in the resulting product.
BACKGROUND OF THE INVENTION
Lovastatin and its analogs, e.g. simvastatin, are potent antihyper-cholesterolemic agents that function by limiting cholesterol biosynthesis. Lovastatin is one of the most important known cholesterol lowering agents. Lovastatin (CAS Registry No. 75330-75-5) is also known as mevinolin or monacolin K and is chemically known as: &bgr;,&bgr;-dihydroxy-7-[1,2,6,7,8,8a-hexahydro-2,6-dimethyl-8-(2-methyl-butyryloxy)-1-napthalen-1-yl]-heptanoic acid &bgr;-lactone of the formula:
Lovastatin, is one member of a class of compounds, which are referred to generally as statins, are known to exist in open ring hydroxy acid and also in lactone form. The lactone form of Lovastatin is shown above.
Lovastatin and its analogs inhibit the enzyme 3-hydroxy-3-methyl-glutarylcoenzyme A reductase (“HMG-CoA reductase”). HMG-CoA reductase catalyzes the formation of mevalonic acid, an early intermediate of cholesterol biosynthesis. Lovastatin is specifically advantageous because, as a result of its application, biosynthetic intermediates that have a toxic steroid skeleton, formed at a later stage of biosynthesis fail to accumulate. Lovastatin also increases the number of LDL-receptors at the surface of the cell membrane, which remove the LDL cholesterol circulating in the blood, thereby inducing the lowering of blood plasma cholesterol level.
Lovastatin is routinely produced via fermentation. GB 2,046,737 discloses that Lovastatin can be produced by some strains belonging to the Monascus genus, e.g., by
M. ruber
1005 cultivated between 7° and 40° C. As a culture medium, an aqueous solution of glucose, peptone, corn steep liquor and ammonium chloride was used. The fermentation was carried out for 10 days in aerobic conditions, and 87 mg Lovastatin was obtained from the filtrate of 5 liters of broth.
U.S. Pat. No. 4,294,926 discloses the biosynthesis of Lovastatin preferably by the application of microorganisms under the deposited numbers ATCC 20541 or 20542 belonging to the
Aspergillus terreus
species on a culture medium containing carbohydrates, e.g., glucose, fructose, maltose, as carbon source; nitrogen sources, e.g., yeast, hydrolyzed yeast, hydrolyzed casein, corn steep liquor, and mineral salts, e.g., calcium carbonate, magnesium sulfate, cobalt, iron, and manganese salts at a temperature of 20-37° C. Similar procedures are described in U.S. Pat. Nos. 4,420,491; 4,342,767; 4,319,039 and 4,294,846 where the fermentations are carried out for 3-5 days on media containing 1-6% carbohydrates and 0.2-6% nitrogen sources.
German Patent No. 4,402,591 discloses biosynthesis of Lovastatin by microorganisms belonging to the Pleurotus genus, e.g.,
P. ostreatus, P. sapidus
and
P. saca
, at 25-35° C. during 7-14 days cultivation time on surface or submerged cultures.
Canadian Patent No. 2,129,416 discloses the preparation of Lovastatin with a microorganism belonging to the Coniothyrium genus, e.g., under the deposited number
Coniothyrium fuckelii
ATCC 74227 on a culture medium containing 3-15% glucose, 0.5-4% peptone, 0.5-5% amylase, 0.2-1% ammonium sulphate, 0.01-0.1% magnesium sulphate, 0.05-0.2% antifoaming agent, 0.2-1.5% L-isoleucine, 0.2-1.5% L-aspartic acid in the pH range of 5-6. According to the examples, the active ingredient concentration of the broth was within 19-430 mg/liter.
Hungarian Patent No. HU 208,997 discloses the application of the holotype strain
Aspergillus obscurus
numbered as MV-1, deposited under the number NCAIM(P)F 001189. The fermentation is preferably carried out on a medium containing yeast extract and/or peptone and/or casein as nitrogen source(s) and glucose and/or maltose or sucrose as carbon source(s). The activity of the broth at the end of the laboratory scale cultivation is between 400-850 mg/liter.
Simvastatin is a synthetic analog of Lovastatin, wherein the 8-acyl moiety is 2,2-dimethylbutyryl. Simvastatin is an even more potent HMG-CoA reductase inhibitor than Lovastatin. Simvastatin is chemically designated as 2,2-dimethylbutanoic acid (4R,6R)-6-[2[1S,2S, 6R,8S,8aR)-1,2,6,7,8,8a-hexahydro-2,6-dimethyl-1-[2-(tetrahydro-4-hydroxy-6-oxo-2H-pyran-2-yl)ethyl]-1-napthalenyl ester (CAS Registry No. 79902-63-9). The chemical structure of Simvastatin is:
Simvastatin is now commercially available as ZOCOR® in some markets. The preparation of Simvastatin was originally described in U.S. Pat. No. 4,444,784. The process involves deacylation of Lovastatin followed by a subsequent acylation with the 2,2-dimethylbutyryl moiety. Simvastatin has also been prepared by the alpha alkylation of the Lovastatin ester moiety as described in U.S. Pat. Nos. 4,582,915 and 4,820,850.
After the fermentation is complete, Lovastatin is present in the broth in both lactone and acid forms. The open hydroxy acid form of the statins is the biologically active form. However, the statins are generally administered to a patient in the lactone form, which is converted to its active metabolite, the hydroxy acid form, in the body. Thus, since only the lactone form is of commercial interest, the acid form is converted into the lactone form through a process called lactonization. The process of lactonization is an equilibrium reaction whereby the open dihydroxy acid form is converted into the closed lactone form. Because lactonization is an equilibrium process, to obtain a high yield of the lactone product, some means must be employed to shift the equilibrium to the lactone side of the equation. This equilibrium equation can be depicted as follows:
Lactonization is an intramolecular esterification. Intermolecular esterification, which leads to dimer formation and higher oligomeric species competes with lactonization:
Lactonization methods are known in the art and many such methods are discussed below. Following lactonization of Lovastatin or following synthesis of Simvastatin, the statins are isolated using crystallization techniques known in the art.
Processes known in the literature for the lactonization of the free, hydroxy acid or its salts are either carried out under high temperature conditions, i.e. refluxing with inert solvents, or catalyzed by strong acids when lactonization is effected at ambient temperature. The process disclosed in U.S. Pat. No. 4,820,850 involves heating the free acid or its salts, e.g. the ammonium salt, to reflux temperature (usually 100-110° C.) in high boiling carbohydrate solvents such as toluene for 7-8 hours. The ambient acidity of the acid is believed to be responsible for the lactonization reaction at these high temperatures. In addition, water that is formed as a by-product of the reaction is continuously removed by azeotropic distillation, which forces the reaction to near completion (shifts the position of equilibrium to the lactone side). The process of lactonization under heat conditions of reflux temperatures is complicated by the formation of dimer impurities which lower the quality of the final lactone product. Once formed, the dimer impurity is difficult to remove and is often present at the levels between 0.4 to 0.08% in the product. To minimize dimerization, high dilutions are often used in the lactonization reaction at the cost of the efficiency of the reaction and process, which is disadvantageous on a commercial manufacturing scale.
U.S. Pat. No. 4,916,239 discloses lactonization at room temperature by treating the free ammonium salt of mevinic acid in a mixture of acetic acid and water, and in the presence of a strong acid catalyst. After the free hydroxy acid-lactone equilibrium is established (reaction has proceeded to 50% conversion), water is gradually added in an amount sufficient to effect crystallization of the lactone from the reaction medium. Removal of the lactone from solution favors lactone formation and thus drives the lactonization to completion. Since the lactone is continuously removed from solution, dimer
Forgas Ilona
Keri Vilmos
Biogal Gyógyszergyar Rt.
Kenyon & Kenyon
Owens Amelia
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