PROCESS FOR PREPARING FORMS OF ATORVASTATIN
CALCIUM SUBSTANTIALLY FREE OF IMPURITIES
This application claims the benefit of U.S. Provisional Patent Application Ser.
No. 60/613,687 filed September 28, 2004, which is incorporated herein by reference.
FIELD OF INVENTION
The present invention relates to atorvastatin calcium impurities and processes
for preparing atorvastatin calcium substantially free of impurities.
BACKGROUND OF THE INVENTION
(pR, SR)-2-(4-fluorophenyl)-J3,6-dihydroxy-5-(l-methylethyl)-3-phenyl-4-
[(phenylamino)carbonyl]-lH-pyrrole-l-heptanoic acid ("atorvastatin") of formula (I)
Mw 558.64
Atorvastatin (I)
is well known in the art, and described, inter alia, in U.S. Patents Nos. 4,681,893,
5,273,995.
Atorvastatin calcium is a member of the class of drugs called statins. Statin
drugs are said to be the most therapeutically effective drugs currently available for
reducing low density lipoprotein (LDL) particle concentration in the blood stream of
patients at risk for cardiovascular disease. A high level of LDL in the bloodstream has
been linked to the formation of coronary lesions which obstruct the flow of blood and
can rupture and promote thrombosis. Goodman and Oilman's Tlie Pharmacological
Basis of Therapeutics 879 (9th ed. 1996). Reducing plasma LDL levels has been
shown to reduce the risk of clinical events in patients with cardiovascular disease and
patients who are free of cardiovascular disease but who have hypercholesterolemia.
Scandinavian Simvastatin Survival Study Group, 1994; Lipid Research Clinics
Program, 1984a, 1984b.
Atorvastatin calcium is marketed under the name LIPITOR® by Pfizer, Inc.
Atorvastatin was first claimed in U.S. Patent No. 4,681,893. The hemi-calcium salt of
atorvastatin is disclosed in U.S. Patent No. 5,273,995. Distinct crystalline forms are
disclosed in several patents and patent applications. Crystalline Forms I, n, HI and IV
of atorvastatin calcium are the subjects of US Patent Nos. 5,959,156 and 6,121,461
assigned to Warner-Lambert and crystalline atorvastatin calcium Forms V and VIII
are disclosed in commonly-owned published application nos. WO 01/36384 and US
2002/0183378, both of which are herein incorporated by reference.
Like any synthetic compound, atorvastatin hemi-calcium salts can contain
extraneous compounds or impurities that can come from many sources. They can be
unreacted starting materials, by-products of the reaction, products of side reactions, or
degradation products. Impurities in atorvastatin hemi-calcium salts or any active
pharmaceutical ingredient (API) are undesirable and, in extreme cases, might even be
harmful to a patient being treated with a dosage form containing the API.
It is also known in the art that impurities in an API may arise from degradation
of the API itself, which is related to the stability of the pure API during storage, and
the manufacturing process, including the chemical synthesis. Process impurities
include unreacted starting materials, chemical derivatives of impurities contained in
starting materials, synthetic by-products, and. degradation products.
In addition to stability, which is a factor in the shelf life of the API, the purity
of the API produced in the commercial manufacturing process is clearly a necessary
condition for commercialization. Impurities introduced during commercial
manufacturing processes must be limited to very small amounts, and are preferably
substantially absent. For example, the ICH Q7A guidance for API manufacturers
requires that process impurities be maintained below set limits by specifying the
quality of raw materials, controlling process parameters, such as temperature,
pressure, time, and stoichiometric ratios, and including purification steps, such as
crystallization, distillation, and liquid-liquid extraction, in the manufacturing process.
The product mixture of a chemical reaction is rarely a single compound with
sufficient purity to comply with pharmaceutical standards. Side products and byproducts
of the reaction and adjunct reagents used in the reaction will, in most cases,
also be present in the product mixture. At certain stages during processing of an API,
such as atorvastatin calcium, it must be analyzed for purity, typically, by HPLC or
TLC analysis, to determine if it is suitable for continued processing and, ultimately,
for use in a pharmaceutical product. The API need not be absolutely pure, as absolute
purity is a theoretical ideal that is typically unattainable. Rather, purity standards are
set with the intention of ensuring that an API is as free of impurities as possible, and,
thus, is as safe as possible for clinical use. As discussed above, in the United States,
the Food and Drug Administration guidelines recommend that the amounts of some
impurities be limited to less than 0.1 percent.
Generally, side products, by-products, and adjunct reagents (collectively
"impurities") are identified spectroscopically and/or with another physical method,
and then associated with a peak position, such as that in a chromatogram, or a spot on
a TLC plate. (Strobel p. 953, Strobel, H.A.; Heineman, W.R., Chemical
Instrumentation: A Systematic Approach, 3rd dd. (Wiley & Sons: New York 1989)).
Thereafter, the impurity can be identified, e.g., by its relative position in the
chromatogram, where the position in a chromatogram is conventionally measured hi
minutes between injection of the sample on the column and elution of the particular
component through the detector. The relative position in the chromatogram is known
as the "retention time."
The retention time can vary about a mean value based upon the condition of
the instrumentation, as well as many other factors. To mitigate the effects such
variations have upon accurate identification of an impurity, practitioners use the
"relative retention time" ("RRT") to identify impurities. (Strobel p. 922). The RRT
of an impurity is its retention tune divided by the retention tune of a reference marker.
It may be advantageous to select a compound other than the API that is added to, or
present in, the mixture in an amount sufficiently large to be detectable and sufficiently
low as not to saturate the column, and to use that compound as the reference marker
for determination of the RRT.
Those skilled in the art of drug manufacturing research and development
understand that a compound in a relatively pure state can be used as a "reference
standard." A reference standard is similar to a reference marker, which is used for
qualitative analysis only, but is used to quantify the amount of the compound of the
reference standard in an unknown mixture, as well. A reference standard is an
"external standard," when a solution of a known concentration of the reference
standard and an unknown mixture are analyzed using the same technique. (Strobel p.
924, Snyder p. 549, Snyder, L.R.; Kirkland, J.J. Introduction to Modem Liquid
Chromatography, 2nd ed. (John Wiley & Sons: New York 1979)). The amount of the
compound in the mixture can be determined by comparing the magnitude of the
detector response. See also U.S. Patent No. 6,333,198, incorporated herein by
reference.
The reference standard can also be used to quantify the amount of another
compound in the mixture if a "response factor," which compensates for differences in
the sensitivity of the detector to the two compounds, has been predetermined. (Strobel
p. 894). For this purpose, the reference standard is added directly to the mixture, and
is known as an "internal standard." (Strobel p. 925, Snyder p. 552).
The reference standard can serve as an internal standard when, without the
deliberate addition of the reference standard, an unknown mixture contains a
detectable amount of the reference standard compound using the technique known as
"standard addition."
In a the "standard addition technique", at least two samples are prepared by adding
known and differing amounts of the internal standard. (Strobel pp. 391-393, Snyder pp.
572). The proportion of the detector response due to the reference standard present in the
mixture without the addition can be determined by plotting the detector response against the
amount of the reference standard added to each of the samples, and extrapolating the plot to
zero concentration of the reference standard. (See, e.g., Strobel, Fig. 11.4 p. 392). The
response of a detector in HPLC (e.g. UV detectors or refractive index detectors) can
be and typically is different for each compound eluting from the HPLC column.
Response factors, as known, account for this difference hi the response signal of the
detector to different compounds eluting from the column.
As is known by those skilled in the art, the management of process impurities
is greatly enhanced by understanding their chemical structures and synthetic
pathways, and by identifying the parameters that influence the amount of impurities in
the final product.
Like any synthetic compound, atorvastatin calcium can contain extraneous
compounds or impurities that can come from many sources. They can be unreacted
starting materials, by-products of the reaction, products of side reactions, or
degradation products.
In this application the reference marker is the impurity N-formyl atorvastatin
calcium in the API. Detection or quantification of the reference marker serves to
establish the level of purity of the API. Use of a compound as a reference marker
requires recourse to a sample of substantially pure compound.
Thus, there is a need in the art for a method for determining the level of
impurities in atorvastatin calcium samples.
SUMMARY OF THE INVENTION
In one aspect the present invention provides the isolated atorvastatin calcium
derivative - atorvastatin calcium epoxy dihydroxy (AED), having the formula:
Mol. Wt: 449.47
The isolated AED of the present invention may be characterized by data selected
from: 1HNMR spectrum having hydrogen chemical shifts at about 1.20,1.21, 2.37,
4.310, 6.032, 7.00,7.06-7.29, 7.30, 7.39,7.41,7.56 ppm; a 13CNMR spectrum having
carbon chemical shifts at about 16.97, 34.66,103.49,106.66,114.72,120.59,125.79,
128.21,128.55,128.74,129.06,129.57,132.38,132.51,135.15, 161.61,163.23 ppm ;
an MS (ESI"1} spectrum having peaks at about having: m/z=472(MNa)+, 454 (MNa-
H2O)+, 432 (MH-H20)+; 344 (FPhCOC(Ph)=C-CONHPh)+ by retention time of about
32 min in HPLC analysis, such as the one described herein below, and by a relative
retention time of about 1.88.
In another aspect, the present invention further provides a process for
preparing AED comprising the steps of:
(a) combining atorvastatin calcium salt and a polar organic solvent or mixtures
thereof with water, with methylene blue, to obtain a solution;
(b) irradiating the obtained solution for about 2 to about 10 hours;
(c) recovering AED.
Preferably, the irradiation of the solution of step (a) is performed in the
presence of oxygen or air, in order to produce a photooxidation reaction. Therefore,
the reaction is conducted, preferably, in an open vessel.
Preferably, the light source for irradiation is selected from the group consisting
of a tungsten lamp, a UV lamp or sun light. More preferably, the light source for
irradiation is a tungsten lamp. Moreover, when using a tungsten lamp as a light
source, the yield is increased.
hi yet another aspect, the present invention also provides a method for
determining the level of AED in atorvastatin calcium comprising
(a) measuring by HPLC the area under a peak corresponding to AED in a
reference standard comprising a known amount of AED;
(b) measuring by HPLC the area under a peak corresponding to AED in a
sample comprising atorvastatin calcium and AED ;
(c) determining the amount of AED in the sample by comparing the area of
step (a) to the area of step (b).
Unless otherwise specified, "atorvastatin calcium" may be either crude
atorvastatin calcium or any form of atorvastatin, including, for example, crystalline
Forms I, H, IV, V, VI, VH, VEI, DC, X, XI, XH and amorphous.
Preferably, the HPLC methodology used in the above method (for the use of
AED as reference standard) includes the steps
(a) combining an atorvastatin calcium sample with a mixture of
acetonitrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain a
solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR. 100 5C-18 (or
similar) column;
(c) eluting the sample from the column at about 50 min using a mixture of
acetonitrile:tetrahydrofuran:buffer (31:9:60) and acetonitrile-.buffer mix
(75:25) as an eluent, and
(d) measuring the AED content in the relevant sample with a UV detector
(preferably at a 254 nm wavelength).
In one aspect, the present invention provides an HPLC method for assaying
atorvastatin calcium comprising the steps
(a) combining an atorvastatin calcium sample with a mixture of
acetomtrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain a
solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR 100 5C-18 (or
similar) column;
(c) eluting the sample from the column at about 50 min using a mixture of
acetonitrile:tetrahydrofuran:buffer (31:9:60) and acetonitrile:buffer mix
(75:25) as an eluent, and
(d) measuring the AED content in the relevant sample with a UV detector
(preferably at a 254 nm wavelength).
Preferably, the buffer contains an aqueous solution of NH^PCu in a
concentration of about 0.05M having a pH of about 5, and ammonium hydroxide.
Preferably, the ratio of the aqueous solution of NFLjHaPC^ and ammonium hydroxide
is of about 1 to 4, respectively.
Preferably, the buffer mix contains the above buffer and tetrahydrofuran.
Preferably, the ratio of the above buffer and tetrahydrofuran is of about 1 to 6.67,
respectively.
In another aspect, the present invention provides a process for preparing a
form of atorvastatin calcium comprising less than about 0.10 w/w of, AED, by HPLC
comprising the steps of
(a) obtaining one or more samples of one or more atorvastatin calcium
batches;
(b) measuring the level of AED in each of the samples of (a);
(c) selecting the atorvastatin calcium batch that comprises a level of AED of
less than about 0.10 w/w by HPLC, based on the measurement or
measurements conducted in step (b); and
(d) using the batch selected in step (c) to prepare said any form of atorvastatin
calcium.
Preferably, the atorvastatin calcium sample of step (a) comprises a sufficiently
low level of AED. More preferably, the atorvastatin calcium sample of step (a)
contains less than about 0.05 w/w by HPLC of AED.
Preferably, said any form of atorvastatin calcium refers to but is not limited to
forms I, II, IV, V, VI, VII, VIE, IX, X, XI, XE and amorphous.
When the atorvastatin calcium sample of step (a) contains more than about
0.10 w/w by HPLC of AED, according to the measurement in step (b), the sample
may be purified, prior to performing step (c).
Preferably, the atorvastatin calcium sample of step (a) obtained after
purification, contains less than about 0.10 w/w by HPLC of AED, more preferably, of
less than about 0.05 w/w by HPLC.
In yet another aspect, the present invention provides a method for reducing the
level of AED in atorvastatin calcium sample by dissolving a selected form of
atorvastatin calcium in an organic solvent, water or mixtures thereof, and crystallizing
to obtain atorvastatin calcium having a reduced level of AED.
Preferably, the atorvastatin calcium sample obtained after purification contains
less than about 0.10 w/w by HPLC of AED, more preferably, of less than about 0.05
w/w by HPLC.
Preferably, the selected form of atorvastatin calcium may be any form of
atorvastatin, such as but not limited to form I, H, IV, V, VI, VII, VIE, IX, X, XI, XII
and amorphous.
Preferably, when the selected form of atorvastatin calcium is the amorphous
form, the crystallization is performed from either a mixture of ester and CS-IQ cyclic or
aliphatic hydrocarbon, from a polar aprotic organic solvent or from a mixture of a Ce-
10 aromatic hydrocarbon and a polar organic solvent, to give atorvastatin calcium
amorphous form. Preferably, the ester is ethylacetate. A preferred Cs-io cyclic or
aliphatic hydrocarbon is hexane. Preferably, the polar organic solvent is either a
ketone or a nitrile. A preferred ketone is acetone. A preferred nitrile is acetonitrile.
Preferably, the Ce-io aromatic hydrocarbon is toluene. A preferred polar organic
solvent is tetrahydrofuran.
Preferably, when the selected form of atorvastatin calcium is form I, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form I. Preferably, the polar organic solvent is a
mixture of CM alcohol and an ether. Preferably, the Ci-4 alcohol is methanol. A
preferred ether is methyltertbutylether.
Preferably, when the selected form of atorvastatin calcium is form II, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form II. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is methanol.
Preferably, when the selected form of atorvastatin calcium is form IV, the
crystallization is performed from a water miscible organic solvent, water and mixtures
thereof, to give atorvastatin calcium form IV. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is methanol, ethanol or 1-
butanol. Preferably, when a mixture of a water miscible organic solvent and water is
used, the water miscible organic solvent is ethanol.
Preferably, when the selected form of atorvastatin calcium is form V, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form V. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form VI, the
crystallization is performed from a mixture of polar aprotic organic solvent and water,
to give atorvastatin calcium form VI. Preferably, the polar aprotic organic solvent is a
ketone. Preferably, the ketone is acetone.
Preferably, when the selected form of atorvastatin calcium is form VII, the
crystallization is performed from a CM alcohol, to give atorvastatin calcium form VII.
Preferably, the CM alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form VHI, the
crystallization is performed from a water miscible organic solvent, water and mixtures
thereof, to give atorvastatin calcium form VHI. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is ethanol, methanol, 1-butanol
or iso-propanol.
Preferably, when the selected form of atorvastatin calcium is form DC, the
crystallization is performed from a water miscible organic solvent, a Cs-io aliphatic
hydrocarbon, water and mixtures thereof, to give atorvastatin calcium form DC
Preferably, the water miscible organic solvent is a CM alcohol. Preferably, the CM
alcohol is ethanol, 1-butanol or iso-propanol. Preferably, the Cs-io aliphatic
hydrocarbon is hexane.
Preferably, when the selected form of atorvastatin calcium is form X, the
crystallization is performed from a mixture of a water miscible organic solvent aid
water, to give atorvastatin calcium form X. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form XI, the
crystallization is performed from a polar aprotic organic solvent or from a water
miscible organic solvent, to give atorvastatin calcium form XI. Preferably, the polar
aprotic organic solvent is a ketone. Preferably, the water miscible organic solvent is a
Ci-4 alcohol. Preferably, the ketone is methylethylketone. A preferred CM alcohol is
isopropanol.
Preferably, when the selected form of atorvastatin calcium is form XII, the
crystallization is performed from a mixture of a water miscible organic solvent and
water, to give atorvastatin calcium form XII. Preferably, the water miscible organic
solvent is a CM alcohol. A preferred CM alcohol is ethanol.
BRIEF DESCRIPTION OF THE DRAWINGS
Figure 1: HPLC chromatogram of AED.
Figure 2:1HNMR spectrum of AED.
Figure 3:13CNMR spectrum of AED.
Figure 4: MS spectrum of AED.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides the isolated atorvastatin calcium derivative -
atorvastatin calcium epoxy dihydroxy (AED), having the formula:
The isolated AED of the present invention may be characterized by data selected
from: 1HNMR spectrum having hydrogen chemical shifts at about 1.20,1.21,2.37,
4.310, 6.032, 7.00, 7.06-7.29, 7.30, 7.39,7.41, 7.56 ppm; a 13CNMR spectrum having
carbon chemical shifts at about 16.97, 34.66,103.49,106.66,114.72,120.59,125.79,
128.21,128.55,128.74, 129.06,129.57,132.38,132.51,135.15, 161.61,163.23 ppm ;
an MS (ESI+) spectrum having peaks at about having: m/z=472(MNa)+., 454 (MNa-
H2O)+ , 432 (MH-H20)+; 344 (FPhCOC(Ph)=C-CONHPh)+ by retention time of about
32 min in HPLC analysis, such as the one described herein below, and by a relative
retention time of about 1.88.
The present invention further provides a process for preparing AED
comprising the steps of:
(a) combining atorvastatin calcium salt and a polar organic solvent or mixtures
thereof with water, with methylene blue, to obtain a solution;
(b) irradiating the obtained solution for about 2 to about 10 hours;
(c) recovering AED.
Preferably, the polar organic solvent is selected from the group consisting of
CM alcohol and nitrile. Preferably, the CM alcohol is either methanol or ethanol. A
preferred nitrile is acetonitrile. Preferably, a mixture of acetonitrile and water is used
in step (a).
Preferably, the irradiation of the solution of step (a) is performed in the
presence of oxygen or air, in order to produce a photooxidation reaction. Therefore,
the reaction is conducted, preferably, in an open vessel.
Preferably, the light source for irradiation is selected from the group consisting
of a tungsten lamp, a UV lamp or sun light. More preferably, the light source for
irradiation is a tungsten lamp. Moreover, when using a tungsten lamp as a light
source, the yield is increased.
Preferably, the solution of step (a) is irradiated for about 2 hours.
Preferably, the crude AED may recovered by evaporating the polar organic
solvent or mixtures thereof with water, more preferably, under vacuum, followed by
filtration and drying to obtain a precipitate, crude AED.
The recovered crude AED may be purified by a process of cliromatography on
a silica-gel column with an eluent of water immiscible polar organic solvent or a
mixture of a polar organic solvent and a Cs-s aliphatic hydrocarbon. Preferably, the
water immiscible polar organic solvent is dichloromethane. A preferred polar organic
solvent is ethyl acetate.
Preferably, AED may be further purified by a process of precipitation from a
water immiscible polar organic solvent or from a mixture of a polar organic solvent
and a CS-IQ aliphatic hydrocarbon. Preferably, the water immiscible polar organic
solvent is dichloromethane. A preferred polar organic solvent is ethyl acetate.
Preferably, the €5-10 aliphatic hydrocarbon is hexane.
The present invention also provides a method for determining the level of
AED in atorvastatin calcium comprising
(a) measuring by HPLC the area under a peak corresponding to AED in a
reference standard comprising a known amount of AED;
(b) measuring by HPLC the area under a peak corresponding to AED in a
sample comprising atorvastatin calcium and AED ;
(c) determining the amount of AED in the sample by comparing the area of
step (a) to the area of step (b).
Unless otherwise specified, "atorvastatin calcium" may be either crude
atorvastatin calcium or any form of atorvastatin, including, for example, crystalline
Forms I, II, IV, V, VI, VII, VIII, K, X, XI, XII and amorphous.
Preferably, the HPLC methodology used in the above method (for the use of
AED as reference standard) includes the steps
(a) combining an atorvastatin calcium sample with a mixture of
acetonitrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain a
solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR100 5C-18 (or
similar) column;
(c) ehiting the sample from the column at about 50 min using a mixture of
acetonitrile:tetrahydrofuran:buffer (31:9:60) and acetonitrile:buffer mix
(75:25) as an eluent, and
(d) measuring the AED content in the relevant sample with a UV detector
(preferably at a 254 nm wavelength).
The present invention further provides an HPLC method for assaying
atorvastatin calcium comprising the steps
(a) combining an atorvastatin calcium sample with a mixture of
acetonitrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain a
solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR100 5C-18 (or
similar) column;
(c) eluting the sample from the column at about 50 min using a mixture of
acetonitrile:tetraliydrofuran:buffer (31:9:60) and acetonitrile:buffer mix
(75:25) as an eluent, and
(d) measuring the AED content in the relevant sample with a UV detector
(preferably at a 254 nm wavelength).
Preferably, the buffer contains an aqueous solution of NKLftPC^ hi a
concentration of about 0.05M having a pH of about 5, and ammonium hydroxide.
Preferably, the ratio of the aqueous solution of NFUHaPC^ and ammonium hydroxide
is of about 1 to 4, respectively.
Preferably, the buffer mix contains the above buffer and tetrahydrofuran.
Preferably, the ratio of the above buffer and tetrahydrofuran is of about 1 to 6.67,
respectively.
The present invention provides a process for preparing a form of atorvastatin
calcium comprising less than about 0.10 w/w of, AED,by HPLC comprising the steps
of
(a) obtaining one or more samples of one or more atorvastatin calcium
batches;
(b) measuring the level of AED hi each of the samples of (a);
(c) selecting the atorvastatin calcium batch that comprises a level of AED of
less than about 0.10 w/w by HPLC, based on the measurement or
measurements conducted hi step (b); and
(d) using the batch selected in step (c) to prepare said any form of atorvastatin
calcium.
Preferably, the atorvastatm calcium sample of step (a) comprises a sufficiently
low level of AED. More preferably, the atorvastatm calcium sample of step (a)
contains less than about 0.05 w/w by HPLC of AED.
Preferably, said any form of atorvastatin calcium refers to but is not limited to
forms I, II, IV, V, VI, VII, VIII, IX, X, XI, XII and amorphous.
When the atorvastatin calcium sample of step (a) contains more than about
0.10 w/w by HPLC of AED, according to the measurement in step (b), the sample
may be purified, prior to performing step (c).
Preferably, the atorvastatin calcium sample of step (a) obtained after
purification, contains less than about 0.10 w/w by HPLC of AED, more preferably, of
less than about 0.05 w/w by HPLC.
The purification may be performed by crystallization from an organic solvent,
water, or mixtures thereof.
The present invention also provides a method for reducing the level of AED in
atorvastatin calcium sample by dissolving a selected fonn of atorvastatin calcium in
an organic solvent, water or mixtures thereof, and crystallizing to obtain atorvastatin
calcium having a reduced level of AED.
Preferably, the atorvastatin calcium sample obtained after purification contains
less than about 0.10 w/w by HPLC of AED, more preferably, of less than about 0.05
w/w by HPLC.
Preferably, the selected form of atorvastatin calcium may be any form of
atorvastatin, such as but not limited to form I, n, IV, V, VI, VII, VIE, IX, X, XI, XH
and amorphous.
Preferably, when the selected form of atorvastatin calcium is the amorphous
form, the crystallization is performed from either a mixture of ester and Cs-io cyclic or
aliphatic hydrocarbon, from a polar aprotic organic solvent or from a mixture of a Ce-
10 aromatic hydrocarbon and a polar organic solvent, to give atorvastatin calcium
amorphous form. Preferably, the ester is ethylacetate. A preferred C5.io cyclic or
aliphatic hydrocarbon is hexane. Preferably, the polar organic solvent is either a
ketone or a nitrile. A preferred ketone is acetone. A preferred nitrile is acetonitrile.
Preferably, the Ce-io aromatic hydrocarbon is toluene. A preferred polar organic
solvent is tetrahydrofuran.
Preferably, when the selected form of atorvastatin calcium is form I, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form I. Preferably, the polar organic solvent is a
mixture of CM alcohol and an ether. Preferably, the Ci_4 alcohol is methanol. A
preferred ether is methyltertbutylether.
Preferably, when the selected form of atorvastatin calcium is form II, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form II. Preferably, the water miscible organic
solvent is a €1.4 alcohol. Preferably, the CM alcohol is methanol.
Preferably, when the selected form of atorvastatin calcium is form IV, the
crystallization is performed from a water miscible organic solvent, water and mixtures
thereof, to give atorvastatin calcium form IV. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is methanol, ethanol or 1-
butanol. Preferably, when a mixture of a water miscible organic solvent and water is
used, the water miscible organic solvent is ethanol.
Preferably, when the selected form of atorvastatin calcium is form V, the
crystallization is performed from a mixture of water miscible organic solvent and
water, to give atorvastatin calcium form V. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form VI, the
crystallization is performed from a mixture of polar aprotic organic solvent and water,
to give atorvastatin calcium form VI. Preferably, the polar aprotic organic solvent is a
ketone. Preferably, the ketone is acetone.
Preferably, when the selected form of atorvastatin calcium is form VII, the
crystallization is performed from a CM alcohol, to give atorvastatin calcium form VH
Preferably, the CM alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form VIII, the
crystallization is performed from a water miscible organic solvent, water and mixtures
thereof, to give atorvastatin calcium form VIE. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the CM alcohol is ethanol, methanol, 1-butanol
or iso-propanol.
Preferably, when the selected fonn of atorvastatin calcium is form DC, the
crystallization is performed from a water miscible organic solvent, a Cs-io aliphatic
hydrocarbon, water and mixtures thereof, to give atorvastatin calcium form IX.
Preferably, the water miscible organic solvent is a CM alcohol. Preferably, the CM
alcohol is ethanol, 1-butanol or iso-propanol. Preferably, the Cs-io aliphatic
hydrocarbon is hexane.
Preferably, when the selected form of atorvastatin calcium is form X, the
crystallization is performed from a mixture of a water miscible organic solvent and
water, to give atorvastatin calcium form X. Preferably, the water miscible organic
solvent is a CM alcohol. Preferably, the €1.4 alcohol is ethanol.
Preferably, when the selected form of atorvastatin calcium is form XI, the
crystallization is performed from a polar aprotic organic solvent or from a water
miscible organic solvent, to give atorvastatin calcium form XI. Preferably, the polar
aprotic organic solvent is a ketone. Preferably, the water miscible organic solvent is a
CM alcohol. Preferably, the ketone is methylethylketone. A preferred CM alcohol is
isopropanol.
Preferably, when the selected form of atorvastatin calcium is form XII, the
crystallization is performed from a mixture of a water miscible organic solvent and
water, to give atorvastatin calcium form XII. Preferably, the water miscible organic
solvent is a CM alcohol. A preferred CM alcohol is ethanol.
Optionally, the crystallization process may be repeated as necessary to obtain
the desired atorvastatin calcium purity.
In order to preserve the purity level of atorvastatin calcium, the sample is
maintained at a temperature of less than about 8°C, preferably the sample is
maintained at a temperature of less than about 4°C.
Having described the invention with reference to certain preferred
embodiments, other embodiments will become apparent to one skilled in the art from
consideration of the specification. The invention is further defined by reference to the
following examples describing in detail the preparation of the composition and
methods of use of the invention. It will be apparent to those skilled in the art that
many modifications, both to materials and methods, may be practiced without
departing from the scope of the invention.
Kromasil KR 100 5C-18 250x4.6mm is suitable.
Acetonitrile:Tetrahydrofuran:Buffer 31:9:60
Acetonitrile:Buffer Mix 75:25
0.05M aqueous NH4H2P04 adjusted to pH 5.0 with
NH4OH (diluted about 1:4)
A mixture of buffer solution and THF 60 volumes

Atorvastatin calcium salt (l.Og) was dissolved in a mixture of acetonitrilewater
(1200ml-800ml) and methylene blue (Img) was added to the solution. The
solution was stirred in an open flask at ambient temperature, and irradiated with
visible light (tungsten lamp, 100W, distance 10cm) for 2 hours. Acetonitrile was
evaporated under vacuum, and precipitated solid was filtered giving, after drying, a
crude product (0.5g) containing impurities at 32 and 33 min. (HPLC control)
The crude product (3.6g) was purified by column chromatography on silica gel
with dichloromethane as eluent, giving the mixture of the impurities at 32 and 33 min
(1.6g). The product was dissolved in dichloromethane (15ml). The solution was
stirred at ambient temperature while a solid was precipitated within a few minutes.
The solid was filtered giving, after drying, the product (80mg).
Example 2; Crystallization of Form VIII
17
Atorvastatin hemi-calcium salt forai V (5g) was added to a boiling solution of
ethanol 96% (150ml) to obtain a solution. The solution was refluxed for 2 hours
(during that time atorvastatin hemi-calcium salt was recrystallized), then cooled to
20°C during 1.5 hours and stirred at this temperature for an additional 16 hours.
Filtration and drying in a vacuum oven at 40°C for 24 hours and then at 60°C for 24
hours gave atorvastatin hemi-calcium salt form VIII.
Example 3: Crystallization of the forms of atorvastatin calcium
Modifying the process in Example 2 by changing the medium of
crystallization results in the following crystal forms
Table Removed

What is claimed is:
1. Isolated atorvastatin epoxy dihydroxy (AED), having the formula:
2. The isolated AED of claim 1, characterized by data selected from the group
consisting of: 1HNMR spectrum having hydrogen chemical shifts at about
1.20, 1.21,2.37, 4.31, 6.032, 7,7.06-7.29, 7.3,7.39, 7.41 and 7.56 ppm;
13CNMR spectrum having carbon chemical shifts at about 16.97, 34.66,
103.49,106.66,114.72, 120.59, 125.79,128.21,128.55, 128.74, 129.06,
129.57,132.38,132.51,135.15,161.61and 163.23 ppm; and by a MS (ESI4)
spectrum having peaks at about: m/z=472(MNa)+, 454 (MNa-ELzO)"1", 432
(MH-H20)+; 344 (FPhCOC(Ph)=C-CONHPh)+.
3. The AED of claim 2, characterized by a 13HNMR spectrum depicted in figure
2.
4. The AED of claim 2, characterized by a 13CNMR spectrum depicted in figure
3.
5. The AED of claim 2, characterized by a MS spectrum depicted in figure 4.
6. A process for the preparation of AED of claim 1, comprising the steps of:
(a) combining atorvastatin calcium salt and a polar organic solvent or mixtures
thereof with water, with methylene blue, to obtain a solution;
(b) irradiating the obtained solution for about 2 to about 10 hours;
(c) recovering AED.
7. The process of claim 6, wherein the organic solvent is selected from the group
consisting of a CM alcohol and nitrile.
8. The process of claim 7, wherein the CM alcohol is either methanol or ethanol.
9. The process of claim 7, wherein the nitrile is acetonitrile.
10. The process of claim 6, wherein a mixture of acetonitrile and water is used in
step (a).
11. The process of claim 6, wherein the irradiation of the solution in step (a) is
performed in the presence of oxygen or air.
12. The process of claim 6, the light source for irradiation is selected from the
group consisting of a tungsten lamp, a UV lamp or sun light.
13. The process of claim 12, wherein the light source for irradiation is a tungsten
lamp.
14. The process of claim 6, wherein the recovered crude AED is purified by
chromatography on a silica gel column.
15. The process of claim 14, wherein the eluent is selected from the group
consisting of a water immiscible polar organic solvent as and a mixture of a
polar organic solvent and a Cs-g aliphatic hydrocarbon.
16. The process of claim 15, wherein the water immiscible polar organic solvent is
dichloromethane.
17. The process of claim 15, wherein the polar organic solvent is emylacetate.
18. The process of claim 15, wherein the Cs.s aliphatic hydrocarbon is hexane.
19. The process of claim 6, wherein the purified crude AED is further purified by
a process of precipitation from a water immiscible polar organic solvent or
from a mixture of a polar organic solvent and a Cs-io aliphatic hydrocarbon.
20. The process of claim 19, wherein the water immiscible polar organic solvent is
dichloromethane.
21. The process of claim 19, wherein the polar organic solvent is ethyl acetate.
22. The process of claim 19, wherein the Cs-io aliphatic hydrocarbon is hexane.
23. AED prepared according to any of claims 6 to 22.
24. A method for detennining the level of AED in atorvastatin calcium
comprising
21
(b) measuring by HPLC the area under a peak corresponding to AED in a
reference standard comprising a known amount of AED;
(c) measuring by HPLC the area under a peak corresponding to AED in a
sample comprising atorvastatin calcium and AED;
(d) determining the amount of AED in the sample by comparing the area
of step (a) to the area of step (b).
25. The method of claim 24, wherein atorvastatin calcium is either crude
atorvastatin calcium or any form of atorvastatin calcium.
26. The method of claim 25, wherein said from atorvastatin calcium is selected
from the group consisting of form I, II, IV, V, VI, VII, VIII, DC, X, XI, XH,
and amorphous.
27. The method of claim 24, wherein the measuring by HPLC in step (a), step (b),
or both, includes the following:
(a) combining an atorvastatin calcium sample with a mixture of
acetonitrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain
a solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR 100 5C-18 (or
similar) column;
(c) eluting the standard or sample from the column at about 50 rain using a
mixture of acetonitrile:tetrahydrofuran:buffer (31:9:60) and
acetonitrile:buffer mix (75:25) as an eluent, and
(d) measuring the AED content in the standard or sample with a UV
detector.
28. The method of claim 27, wherein the UV wavelength is about 254 nm.
29. An HPLC method for assaying atorvastatin calcium comprising the steps
of
(a) combining an atorvastatin calcium sample with a mixture of
acetonitrile:tetrahydrofuran:water in a ratio of about 60:5:35, to obtain a
solution;
(b) injecting the solution of step (a) into a 250X4.6 mm KR 100 5C-18 (or
similar) column;
(c) eluting the standard or sample from the column at about 50 min using a
mixture of acetomtrile:tetrahydrofuran:buffer (31:9:60) and
acetonitrile:buffer mix (75:25) as an eluent, and
(d) measuring the AED content in the standard or sample with a UV detector.
30. The method of claim 29, wherein the UV wavelength is about 254 run.
31. The method of claim 29, wherein the buffer contains an aqueous solution
of NaHPO4 in a concentration of about 0.05M having a pH of about 5, and
ammonium hydroxide.
32. The method of claim 31, wherein the ratio of the said aqueous solution of
NaHPCU and the ammonium hydroxide is of about 1 to 4, respectively.
33. The method of claim 29, wherein the buffer mix contains the buffer-of
claim 31 and tetrahydrofuran.
34. The method of claim 33, wherein the ratio of the said buffer of claim 31
and tetrahydrofuran is of about 1 to 6.67, respectively.
35. A process for preparing a form of atorvastatin calcium comprising less than
about 0.10 w/w of AED, by HPLC comprising the steps of
(a) obtaining one or more samples of one or more atorvastatin calcium
batches;
(b) measuring the level of AED in each of the samples of (a);
(c) selecting the atorvastatin calcium batch that comprises a level of AED
of less than a about 0.10 w/w by HPLC, based on the measurement or
measurements conducted in step (b); and
(d) using the batch selected in step (c) to prepare said any form of
atorvastatin calcium.
36. The process of claim 35, wherein the atorvastatin calcium sample of step
(a) contains less than about 0.05 w/w by HPLC of AED.
37. The process of claim 35, wherein said any form of atorvastatin calcium is
selected form the group consisting of form I, II, IV, V, VI, VII, VIE, DC,
X, XI, XII, and amorphous.
38. The process of claim 35, wherein, if the atorvastatin calcium sample in
step (a) contains more than about 0.10 w/w by HPLC of AED, the sample
may be purified, prior to performing step (c).
39. The process of claim 35, wherein the atorvastatin calcium of step (a)
obtained after purification, contains less than about 0.10 w/w by HPLC of
AED.
40. The process of claim 39, wherein the atorvastatin calcium of step (a)
obtained after purification, contains less than about 0.05 w/w by HPLC of
AED.
41. The process of claim 35, wherein the purification is done by crystallization
from an organic solvent, water, or mixtures thereof.
42. A method for reducing the level of AED in atorvastatin calcium sample by
dissolving a selected form of atorvastatin calcium in an organic solvent,
water or mixtures thereof, and crystallizing to obtain atorvastatin calcium
having a reduced level of AED.
43. The method of claim 42, wherein the atorvastatin calcium obtained after
purification, contains less than about 0.10 w/w by HPLC of AED.
44. The process of claim 43, wherein the atorvastatin calcium obtained after
purification, contains less than about 0.05 w/w by HPLC of AED.
45. The method of claim 42, wherein the selected form of atorvastatin calcium
is selected from group consisting of form I, II, IV, V, VI, VII, VIE, IX, X,
XI, XII, and amorphous.
46. The method of claim 42, wherein the selected form of atorvastatin calcium
is amorphous, the crystallization is performed from a mixture of ester and
Cs-io cyclic or aliphatic hydrocarbon.
47. The method of claim 46, wherein the ester is ethylacetate.
48. The method of claim 46, wherein the Cs-io cyclic or aliphatic hydrocarbon
is hexane.
49. The method of claim 42, wherein the selected form of atorvastatin calcium
is amorphous, the crystallization is performed from a polar aprotic organic
solvent.
50. The method of claim 49, wherein the polar organic solvent is either a
ketone or a nitrile.
51. The method of claim 50, wherein the ketone is acetone.
52. The method of claim 50, wherein the nitrile is acetonitrile.
53. The method of claim 42, wherein the selected form of atorvastatin calcium
is amorphous, the crystallization is performed from a mixture of a Ce-io
aromatic hydrocarbon and a polar organic solvent.
54. The method of claim 53, wherein the Ce-io aromatic hydrocarbon is
toluene.
55. The method of claim 53, wherein the polar organic solvent is
tetrahydrofuran.
56. The method of any of the claims 47 to 54, wherein the obtained
atorvastatin calcium is amorphous.
57. The method of claim 42, wherein the selected form of atorvastatin calcium
is form I, the crystallization is performed from a mixture of a polar organic
solvent and water.
58. The method of claim 58, wherein the polar organic solvent is a mixture of
CM alcohol and an ether.
59. The method of claim 58, wherein the CM alcohol methanol.
60. The method of claim 58, wherein the ether is methyltertbutylether.
61. The method of claim 57, wherein the obtained atorvastatin calcium is
form I.
62. The method of claim 42, wherein the selected form of atorvastatin
calcium is form II, the crystallization is performed from a mixture of water
miscible organic solvent and water.
63. The method of claim 62, wherein the water miscible organic solvent is a
Ci-4 alcohol.
64. The method of claim 63, wherein the CM alcohol is methanol.
65. The method of claim 62, wherein the obtained atorvastatin calcium is form
66. The method of claim 42, wherein the selected form of atorvastatin
calcium is form IV, the crystallization is performed from a water miscible
organic solvent, water and mixtures thereof.
67. The method of claim 66, wherein the water miscible organic solvent is a
alcohol.
25
68. The method of claim 67, wherein the CM alcohol is methanol, ethanol or
1-butanol.
69. The method of claim 66, wherein a mixture of a water miscible organic
solvent and water is used.
70. The method of claim 69, wherein the water miscible organic solvent is
ethanol.
71. The method of claim 66, wherein the obtained atorvastatin calcium is
form IV.
72. The method of claim 42, wherein the selected form of atorvastatin calcium
is form V, the crystallization is performed from a mixture of water
miscible organic solvent and water.
73. The method of claim 72, wherein the water miscible organic solvent is a
CM alcohol.
74. The method of claim 73, wherein the Ci-4 alcohol is ethanol.
75. The method of claim 72, wherein the obtained atorvastatin calcium is form
V.
76. The method of claim 42, wherein the selected form of atorvastatin
calcium is form VI, the crystallization is performed from a mixture of
polar aprotic organic solvent and water.
77. The method of claim 76, wherein the polar aprotic organic solvent is a
ketone.
78. The method of claim 77, wherein the the ketone is acetone.
79. The method of claim 76, wherein the obtained atorvastatin calcium is form
VI
80. The method of claim 42, wherein the selected form of atorvastatin calcium
is form VII, the crystallization is performed from a CM alcohol.
81. The method of claim 80, wherein the CM alcohol is ethanol.
82. The method of claim 70, wherein the obtained atorvastatin calcium is
form VH.
83. The method of claim 42, wherein the selected form of atorvastatin calcium
is form Vffl, the crystallization is performed from a water miscible organic
solvent, water and mixtures thereof.
84. The method of claim 83, wherein the water miscible organic solvent is a
alcohol.
85. The method of claim 84, wherein the CM alcohol is ethanol, methanol, 1-
butanol or iso-propanol.
86. The method of claim 83, wherein the obtained atorvastatin calcium is form
VIII.
87. The method of claim 42, wherein the selected form of atorvastatin
calcium is form IX, the crystallization is performed from a water miscible
organic solvent, a CS-IQ aliphatic hydrocarbon, water and mixtures thereof.
88. The method of claim 87, wherein the water miscible organic solvent is a
CM alcohol.
89. The method of claim 87, wherein the CM alcohol is ethanol, 1-butanol or
iso-propanol.
90. The method of claim 87, wherein the Cs-io aliphatic hydrocarbon is
hexane.
91. The method of claim 87, wherein the obtamed atorvastatin calcium is form
DC.
92. The method of claim 42, wherein the selected form of atorvastatin calcium
is form X, the crystallization is performed from a mixture of a water
miscible organic solvent and water.
93. The method of claim 92, wherein the water miscible organic solvent is a
CM alcohol.
94. The method of claim 93, wherein the the CM alcohol is ethanol.
95. The method of claim 92, wherein the obtained atorvastatin calcium is form
X.
96. The method of claim 42, wherein the selected form of atorvastatin
calcium is form XI, the crystallization is performed from a polar aprotic
organic solvent.
97. The method of claim 96, wherein polar aprotic organic solvent is a ketone.
98. The method of claim 96, wherein the ketone is methylethylketone.
99. The method of claim 96, wherein the selected fonn of atorvastatin calcium
is form XI, the crystallization is performed from a water miscible organic
solvent.
100. The method of claim 99, wherein the water miscible organic solvent is
a CM alcohol.
101. The method of claim 100, wherein the preferred CM alcohol is
isopropanol.
102. The method of any of the claims 96 to 101, wherein the obtained
atorvastatin calcium is form XI.
103. The method of claim 42, wherein the selected form of atorvastatin
calcium is form XII, the crystallization is performed from a mixture of a
water miscible organic solvent and water.
104. The method of claim 103, wherein the water miscible organic solvent
is a CM alcohol.
105. The method of claim 104, wherein the CM alcohol is ethanol.
106. The method of claims 103, wherein the obtained atorvastatin calcium
is form XII.