WO2016055033A1

WO2016055033A1 - A process for preparing glycopyrronium bromide

Info

Publication number: WO2016055033A1
Application number: PCT/CZ2015/000110
Authority: WO
Inventors: Iva OBADALOVA; Tomas Chvojka; Jan Hruby; Ondrej Dammer; Marcela Tkadlecova; Josef Reitmajer; Nadezda NOVOTNA; Petr LEHNERT
Original assignee: Zentiva, K.S.
Priority date: 2014-10-06
Filing date: 2015-09-30
Publication date: 2016-04-14
Also published as: CZ2014680A3; EP3204354A1

Abstract

The invention relates to a process for preparing glycopyrronium bromide in the form of [R,S] and [S.R] diastereoisomers, the method consisting in separation of these desired diastereoisomers from the mixture of [R,S3, [S,R], [R,R] and [S,S] diastereoisomers by means of two or more controlled crystallizations from a solution of a solvent selected from the group consisting of 1-propanol, 2-propanol, 1-butanol, and tert-butanol.

Description

A process for preparing glycopyrronimn bromide

Technical Field The invention relates to a new process for preparing glycopyrronium bromide, (3J?S)-3-[(2S'J?)- (2-cyclopentyl-2-hydroxy-2-phenylacetyl)oxy]-l , 1-dimethylpyrrolidinium bromide (I).

[2S, 3R]-stereoisomer [2R, 3S]-stereoisomer

(I)

In particular it consists in separation of a mixture of four isomers by crystallization from new solvents, a mixture of [25, 3i?] and [2R, 35] isomers being desirable.

Glycopyrronium bromide shows anticholinergic bronchodilating effects. This means that it extends the airways by blocking certain receptors in muscle cells in the lungs, called muscarinic receptors, which control muscular contractions. After inhalation of a drug containing glycopyrronium bromide the airway muscles become released, which helps to keep the airways open and allows the patient to breathe more easily. Glycopyrronium bromide is indicated as an inhalation anticholinergic drug in cooperating patients with the diagnosis of COPD (chronic obstructive pulmonary disease). COPD is defined as a preventable and treatable disease that is characterized by a persistent obstruction of air flow in the bronchi (bronchial obstruction), which usually progresses and is related to an intensified inflammatory response of the airways to harmful particles or gases. The main goal of the treatment of COPD is an improvement of the current control, i.e. elimination of symptoms, improvement of toleration of physical effort, improvement of the health condition and reduction of future risks, i.e. prevention and treatment of exacerbations, prevention of progression of the disease and reduction of mortality. Background Art

The patent EP1856041 Bl describes a process for the preparation of a mixture of two possible diastereomeric pairs of glycopyrronium bromide, the [R,S and [S,R] pair and the [R,R] and [S,S] pair. Separation of these two diastereomeric pairs of glycopyrronium bromide by crystallization is described therein from a mixture of solvents, methanol and methyl ethyl ketone, in ratios varying in the range of 1:5 to 1:8. The patent suggests that with the use of these solvents triple crystallization is necessary to achieve an acceptable optical purity of the product, while the total yield of these three crystallizations is said to be 33%. According to the patent data the particle size of the product should be up to 100 micrometers at most, the particles being then preferably raicronized for use in a dosage form, but the patent does not mention any more detailed information about the particle size distribution, analytic method used etc. However, a reproduction of the preparation method according to the patent has shown that the size of particles obtained with the use of this method varies in the range of 15 to 240 μηι.

Disclosure of Invention The object of this invention is a crystallization process providing the desired diastereomeric pair, [R,S] and [S,R], of glycopyrronium bromide with a high yield and a high diastereomeric purity. The product further exhibits a narrow particle size distribution, which makes the particles suitable for the preparation of the drug form without the need of subsequent micronization in some embodiments.

Detailed description of the invention

The essence of the invention is a crystallization process, wherein the desirable [R,S] and [S,R] pair is separated from a mixture of two diastereomeric pairs of glycopyrronium bromide, the [R,S\ and [S,R] pair and [R,R] and [S,S\ pair, by crystallization with the use of a suitable solvent. 2-Propanol, 1-propanol, 1-butanol and tert-butanol were used as suitable solvents. In a preferred embodiment in accordance with this invention a mixture of two diastereomeric pairs of glycopyrronium bromide is dissolved in a hot state in the selected solvent and the solution is subsequently cooled down, while the desired pair [R,S] and [S,R] is crystallized from the solvent. This cooling should preferably follow a controlled temperature gradient, which ensures a suitable distribution of the particle size. Unlike the use of a system of solvents (solvent - anti-solvent) it is thus convenient to only use one solvent as it facilitates better control of the cooling temperature gradient and control of the entire crystallization.

In the patent EP1856041B1, there are described three recrystallizations with the use of methanol and 2-butanol as the anti-solvent, necessary to achieve an acceptable optical purity, the total yield of these three crystallizations being reported as 33% in the patent. The size of particles obtained with the use of this method varies in the range of 15 to 240 um.

In the case of crystallizations from 2-propanol, 1 -propanol, 1-butanol and /ert-butanol two crystallizations are only sufficient to achieve acceptable diastereomeric purity (not less than 99.9%) and the total yield varies in the range of 40 to 45%. In the case of 2-propanol particles with the size of 8 to 170 μιη are obtained, in the case of 1 -propanol particles with the size of 2 to 11 um are obtained, in the case of 1-butanol particles with the size of 2 to 14 fim are obtained and in the case of teri-butanol particles with the size of 2 to 30 μιη are obtained. In case of 1- propanol, 1-butanol crystals with narrow particle size distributions of less than 20 μηι are provided, which makes them conveniently usable for a dosage form without the need of micronization. All the crystallizations provided glycopyrronium bromide in the crystalline form A, characterized by the XRPD spectrum in Fig. 1 and the carbon solid-state NMR spectrum in Fig. 2.

Brief Description of Drawings

Fig. 1: XRPD pattern of crystalline glycopyrronium bromide prepared according to Example 3 Fig. 2: ssNMR pattern of crystalline glycopyrronium bromide prepared according to Example 3 Fig.3: Microscopic record of the particle size distribution of glycopyrronium bromide

recrystallized from a mixture of methanol - 2-butanone (Example 1)

Fig. 4: Microscopic record of the particle size distribution of glycopyrronium bromide

recrystallized from 2-propanol (Example 3)

Fig. 5: Microscopic record of the particle size distribution of glycopyrronium bromide

recrystallized from 1 -propanol (Example 4)

Fig. 6: Microscopic record of the particle size distribution of glycopyrronium bromide

recrystallized from 1-butanol (Example 5)

Fig. 7: Microscopic record of the particle size distribution of glycopyrronium bromide

recrystallized from iert-butanol (Example 6) Fig.8: Microscopic photograph of particles of glycopyrronium bromide recrystallized from a mixture of methanol - 2-butanone (Example 1)

Fig.9: Microscopic photograph of particles of glycopyrronium bromide recrystallized from 2- propanol (Example 3)

Fig. 10: Microscopic photograph of particles of glycopyrronium bromide recrystallized from 1- propanol (Example 4)

Examples

The starting crude mixture of the two diastereomeric pairs of glycopyrronium bromide for the recrystallizations was prepared in accordance with the patent EP 1 856 041B1. The chemical purity of the mixture prepared this way was 98.1% (HPLC). The mixture contained the [R,S\ and [S,R] pair and [R,R] and [S,S] pair in the ratio of 60 : 40 (CE).

Example 1 - reference

Recrystallization of glycopyrronium bromide from methanol and 2-butanone according to the patent EP1856041B1, step 3

First recrystallization:

The crude mixture of glycopyrronium bromide (4 g, 10.1 mmol) was heated in 3.5 ml of methanol and 24 ml of 2-butanone to 80°C. Subsequently, another 36 ml of 2-butanone was added and heated at 80°C for 30 minutes. Then the mixture was cooled at the rate of 30°C / h to the final -10°C. The resulting crystals were aspirated and washed with cold 2-butanone (12 ml). 1.9 g of the product was obtained, chemical purity 99.1% (HPLC), diastereomeric purity 94.2% (CE).

Second recrystallization:

The product of the first recrystallization (1.9 g, 4.8 mmol) was heated up in 2.8 ml of methanol and 12.2 ml of 2-butanone to 80 °C. Subsequently, another 17.6 ml of 2-butanone was added and heated at 80°C for 30 minutes. Then the mixture was cooled at the rate of 30°C / h to the final -10°C. The resulting crystals were aspirated and washed with cold 2-butanone (8 ml). 1.4 g of the product was obtained, chemical purity 99.9% (HPLC), diastereomeric purity 99.7% (CE). Third recrystallization:

The product of the second recrystallization (1.4 g, 3.5 mmol) was heated up in 1.1 ml of methanol and 8.7 ml of 2-butanone to 80 °C. Subsequently, another 12.4 ml of 2-butanone was added and heated at 80°C for 30 minutes. Then the mixture was cooled at the rate of 30°C / h to the final -10°C. The resulting crystals were aspirated and washed with cold 2-butanone (5 ml). 1.3 g (total yield of three recrystallizations 33%) of a purely white crystalline substance was obtained with the chemical purity of 99.95% (HPLC) and diastereomeric purity of 99.9% (CE). 1H NMR (500 MHz, D₂0): 6 1.24 (m, 1H); 1.50-1.72 (m, 7H); 2.13 (m, 1H); 2.69 (m, 1H); 3.02 (s, 3H); 3.15 (qui, J = 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m, 2H); 3.73 (d, J= 13.9 Hz, 1H); 3.83 (dd, J = 13.9 Hz, J = 6.0 Hz, 1H); 5.50 (m, lH); 7.38 (m, 1H); 7.45 (m, 2H); 7.63 (m, 2H). ¹³C NMR (125,8 MHz, D₂0): δ 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A.

PSD (microscope): 15 - 240 μπι - see Table 1.

Table 1 : Particle size distribution (PSD) - percentage representation of particles of a certain size in the sample

GLP6 ME BU K2

Size V (%) V (%)

(μπι) in the category cumulative

15 0.58% 0.58%

20 2.85% 3.43%

30 12.49% 15.91%

50 29.52% 45.43%

60 13.42% 58.85%

80 19.05% 77.90%

100 10.31% 88.21%

120 4.78% 93.00%

140 3.45% 96.45%

160 1.50% 97.95%

180 0.75% 98.70%

200 0.78% 99.48%

220 0.27% 99.75%

240 0.25% 100.00% Example 2

Recrystallization of glycopyrronium bromide from 2-propanoI

First recrystallization:

A crude mixture of glycopyrronium bromide (4 g, 10.1 mmol) was heated up in 60 ml of 2- propanol to 80°C and stirred at this temperature for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the final -10°C; at 50 to 60 °C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was maintained at -10° C for 15 minutes and the resulting crystals were then aspirated and washed with cold 2-propanol (10 ml). 2 g of the product was obtained, chemical purity 99.3% (HPLC), diastereomeric purity 94.6% (CE).

Second recrystallization:

The product of the first recrystallization (2 g, 5 mmol) was heated in 35 ml of 2-propanol at 80°C for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the final -10°C; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was maintained at -10°C for 30 minutes and the resulting crystals were then aspirated and washed with cold 2-propanol (10 ml). 1.75 g (total yield of two crystallizations 44%) of a purely white crystalline substance was obtained with the chemical purity of 99.95% (HPLC) and diastereomeric purity of 99.9% (CE). 1H NMR (500 MHz, D₂0): δ 1.24 (m, 1H); 1.50-1.72 (m, 7H); 2.13 (m, 1H); 2.69 (m, 1H); 3.02 (s, 3H); 3.15 (qui, J= 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m, 2H); 3.73 (d, J= 13.9 Hz, 1H); 3.83 (dd, J= 13.9 Hz, J = 6.0 Hz, 1H); 5.50 (m, 1H); 7.38 (m, 1H); 7.45 (m, 2H); 7.63 (m, 2H). ¹³C NMR (125,8 MHz, D₂0): δ 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A. PSD (microscope): 8 - 170 um.

Example 3

Recrystallization of glycopyrronium bromide from 2-propanol

First recrystallization:

A crude mixture of glycopyrronium bromide (2 g, 5 mmol) was heated up in 30 ml of 2-propanol to 80°C and stirred at this temperature for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the final -10°C; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was maintained at -10°C for 15 minutes and the resulting crystals were then aspirated and washed with cold 2-propanol (5 ml). 0.95 g of the product was obtained, chemical purity 99.3% (HPLC), diastereomeric purity 95.9% (CE). Second recrystallization:

The product of the first recrystallization (0,95 g, 2,4 mmol) was heated for 30 minutes in 17 ml of 2-propanol at 80°C. Then the solution was cooled at the rate of 20°C / h to the final -10°C; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was maintained at -10°C for 30 minutes and the resulting crystals were then aspirated and washed with cold 2-propanol (4 ml). 0.80 g (total yield of two crystallizations 40%) of a purely white crystalline substance was obtained with the chemical purity of 99.95% (HPLC) and diastereomeric purity of 99.95% (CE). 1H NMR (500 MHz, D₂0): δ 1.24 (m, 1H); 1.50-1.72 (m, 7H); 2.13 (m, 1H); 2.69 (m, 1H); 3.02 (s, 3H); 3.15 (qui, J= 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m, 2H); 3.73 (d, J= 13.9 Hz, 1H); 3.83 (dd, J= 13.9 Hz, J = 6.0 Hz, 1H); 5.50 (m, 1H); 7.38 (m, 1H); 7.45 (m, 2H); 7.63 (m, 2H). ¹³C NMR (125,8 MHz, D₂0): 5 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A.

PSD (microscope): 6 - 160 um - see Table 2.

Table 2: Particle size distribution (PSD) - percentage representation of particles of a certain size in the sample

GLP7 IP K2

Size V (%) V (%)

(um) in the category cumulative

8 0.54% 0.54%

10 0.95% 1.48%

20 19.02% 20.51%

30 26.85% 47.36%

40 17.72% 65.08%

50 13.29% 78.37%

60 7.24% 85.61%

80 9.04% 94.65%

100 2.80% 97.45%

120 1.43% 98.88%

140 0.57% 99.45%

160 0.37% 99.83%

170 0.17% 100.00% Example 4

Recrystallization of glycopyrronium bromide from 1-propanol

First recrystallization:

A crude mixture of glycopyrronium bromide (2 g, 5 mmol) was heated up in 15 ml of 1-propanol to 80°C and stirred at this temperature for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold 1-propanol (2 ml). 0.9 g of the product was obtained, chemical purity 99.4% (HPLC), diastereomeric purity 94.6% (CE).

Second recrystallization:

The product of the first recrystallization (0.9 g, 4.5 mmol) was stirred in 13 ml of 1-propanol at 80°C for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair.

The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold 1-propanol (2 ml).

0.86 g (total yield of two crystallizations 43%) of a purely white crystalline substance was obtained with the chemical purity of 99.9% (HPLC) and diastereomeric purity higher than 99.9%

(CE). Ή NMR (500 MHz, D₂0): δ 1.24 (m, lH); 1.50-1.72 (m, 7H); 2.13 (m, lH); 2.69 (m, 1H);

3.02 (s, 3H); 3.15 (qui, J= 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m, 2H); 3.73 (d, J= 13.9 Hz, 1H);

3.83 (dd, J= 13.9 Hz, J= 6.0 Hz, 1H); 5.50 (m, 1H); 7.38 (m, 1H); 7.45 (m, 2H); 7.63 (m, 2H).

¹³C NMR (125,8 MHz, D₂0): 5 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A.

PSD (microscope): 2 - 11 urn - see Table 3.

Table 3: Particle size distribution (PSD) - percentage representation of particles of a certain size in the sample

GLP3 NP Kl

Size V (%) V (%)

(μιη) in the category cumulative

2 5.32% 5.32% 3 30.48% 35.79%

4 37.96% 73.75%

5 19.33% 93.08%

6 4.64% 97.73%

8 1.97% 99.69%

10 0.15% 99.84%

11 0.16% 100.00%

Example 5

Recrystallization of glycopyrronium bromide from 1 -butanol

First recrystallization:

A crude mixture of glycopyrronium bromide (4 g, 10.1 mmol) was heated up in 30 ml of 1- butanol to 80°C and stirred at this temperature for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold 1 -butanol (4 ml). 1.85 g of the product was obtained, chemical purity 99.5% (HPLC), diastereomeric purity 94.4% (CE).

Second recrystallization:

The product of the first recrystallization (1.85 g, 4.7 mmol) was stirred in 15 ml of 1 -butanol at 80°C for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold 1 -butanol (2 ml). 1.65 g (total yield of two crystallizations 41%) of a purely white crystalline substance was obtained with the chemical purity of 99.9% (HPLC) and diastereomeric purity of 99.9% (CE). Ή NMR (500 MHz, D₂0): δ 1.24 (m, 1H); 1.50-1.72 (m, 7H); 2.13 (m, lH); 2.69 (m, 1H); 3.02 (s, 3H); 3.15 (qui, J= 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m, 2H); 3.73 (d, J= 13.9 Hz, 1H); 3.83 (dd, J = 13.9 Hz, J= 6.0 Hz, 1H); 5.50 (m, 1H); 7.38 (m, 1H); 7.45 (m, 2H); 7.63 (m, 2H). ¹³C NMR (125,8 MHz, D₂0): δ 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A.

PSD (microscope): 2 - 14 um - see table 4. Table 4: Particle size distribution (PSD) - percentage representation of particles of a certain size in the sample

GLP4_NB_K1

Example 6

Recrystallization of glycopyrronium bromide from terf-butanol

First recrystallization:

A crude mixture of glycopyrronium bromide (2 g, 5 mrnol) was heated up in 15 ml of tert- butanol to 80°C and stirred at this temperature for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold iert.butanol (1 ml). 0.85 g of the product was obtained, chemical purity 99.3% (HPLC), diastereomeric purity 94.9%» (CE).

Second recrystallization:

The product of the first recrystallization (0.85 g, 2.1 mrnol) was stirred in 6 ml of tert-butanol at 80°C for 30 minutes. Then the solution was cooled at the rate of 20°C / h to the room temperature; at 50 to 60°C it was inoculated with crystals of the desired diastereoisomeric pair. The mixture was left to crystallize under slow agitation at the room temperature until the next day. The next day the resulting crystals were aspirated and washed with cold fer/.butanol (1 ml). 0.8 g (total yield of two crystallizations 40%) of a purely white crystalline substance was obtained with the chemical purity of 99.8% (HPLC) and diastereomeric purity of 99.9% (CE). 1H NMR (500 MHz, D₂0): δ 1.24 (m, 1H); 1.50-1.72 (m, 7H); 2.13 (m_t 1H); 2.69 (m, 1H); 3.02 (s, 3H); 3.15 (qui, J= 8.5 Hz, 1H); 3.18 (s, 3H); 3.56 (m_s 2H); 3.73 (d, J= 13.9 Hz, 1H); 3.83 (dd, J = 13.9 Hz, J= 6.0 Hz, 1H); 5.50 (m, 1H); 7.38 (m_} 1H); 7.45 (m, 2H); 7.63 (m, 2H). ^I3C NMR (125,8 MHz, D₂0): δ 28.4; 28.7; 28.8; 29.3; 32.6; 47.6; 51.5; 55.6; 56.2; 67.5; 72.8; 76.7; 83.4; 128.7; 131.0; 131.4; 143.3; 177.1. XRPD: crystalline form A.

PSD (microscope): 2 - 30 μιη - see Table 5.

Table 5: Particle size distribution (PSD) - percentage representation of particles of a certain size in the sample

GLP5 TB Kl

Description of analytic methods

Measurement parameters of XRPD: The diffraction patterns were measured using an X'PERT PRO MPD PANalytical diffractometer, used radiation CuKa (λ=0.1542 nm (1.542 A), excitation voltage: 45 kV, anode current: 40 mA, measured range: 2 - 40° 2Θ, increment: 0,01° 2Θ, the measurement was carried out in a flat powder sample that was applied on a Si plate. For the setting of the primary optical equipment programmable divergence slits with the irradiated area of the sample of 10 mm, 0.02 rad Soller slits and a ¼° anti-diffusion slit were used. For the setting of the secondary optical equipment an X'Celerator detector with maximum opening of the detection slot, 0.02 rad Soller slits and a 5.0 mm anti-diffusion slit were used.

Chemical purity was measured with the use of liquid chromatography (HPLC):

Device: Waters Acquity UPLC, PDA detection

Sample preparation: Dissolve 10.0 mg of the tested sample in 10.0 ml of 50% acetonitnle R Column: - dimension : 1 = 0.10 m, 0 = 4.6 mm

- stationary phase: Waters CSH Phenyl-Hexyl, 2.5 μηι particles

- column temperature: 35°C

Mobile phase: A: 0.2% solution of trifluoroacetic acid in water

B; acetonitnle R

Gradient elution:

Detection: spectrophotometer 220

Injected quantity: 2 μ1

Sample temperature: 20°C

Sample concentration: 1 mg ml

Diastereomeric purity was measured by capillary electrophoresis (CE):

Equipment: capillary electrophoresis with an integrated detector (DAD) and a data station

Capillary: quartz, inner diameter 50 μηι, total length 48.5 cm, effective length 40.0 cm

Capillary temperature: 30°C

Used voltage: -10 kV

Detection: UV at 192 nm

Injection: hydrodynamic, 4 s, 50mbar gauge pressure Solution preparation:

Basic electrolyte: 1.08 g of potassium dihydrogen phosphate ( ¾P0₄) is dissolved in 90 ml of water for chromatography R, pH is adjusted to 3.0±0.05 by addition of 85% phosphoric acid R and the solution is diluted with water for chromatography R to 100 ml.

Separation electrolyte: 55 mg of sulfated γ-cyclodextrin (HS-y-CD) (Cyclolab) is dissolved in

0.8 ml of the basic electrolyte. Note: the separation electrolyte must always be freshly prepared- on the day of the analysis.

Comparative solution: 5 mg of each diastereoisomer is dosed into a 5ml volumetric flask.

dissolved in 3 ml of water for chromatography R and topped up with water for chromatography R to 5 ml.

Tested solution: the content of one capsule is dissolved in 3 ml of water for

chromatography R and topped up with water for chromatography R to 5 ml.

Everyday conditioning of the capillary:

The capillary is washed with water for chromatography R for 5 min, 1M sodium hydroxide for 5 min, water for chromatography R for 5 min and separation electrolyte for 5 min (all at the gauge pressure of 950 mbar).

Measurement procedure:

Before every analysis the capillary is washed with the separation electrolyte for 2 min (gauge pressure 950 mbar). Then, the sample (50 mbar, 4 s) is injected into an injection vial. Both the ends of the capillary are immersed into the separation electrolyte and the voltage of -10 kV is applied.

Migration times: Diastereoisomer 1: approx. 13.0 min

Diastereoisomer 2: approx. 14.3 min

Diastereoisomer 3: approx. 14.7 min

Diastereoisomer 4: approx. 15.2 min

The spectra of nuclear magnetic resonance (NMR) were measured using an Avance 500 device made by Bruker. 1H spectra were measured at the frequency of 500.13 MHz, ¹³C at the frequency of 125,8 MHz. The sample was measured in a deuterated solvent specified for the particular analysis, normally at 25°C (unless specified otherwise for a particular analysis). The chemical shift δ is expressed as ppm, the interaction constants J are specified in Hz. The spectra were normally referenced to the residual solvent signal. In the case of D₂0 DSS was used as the internal standard.

Carbon spectra of solid-state nuclear magnetic resonance (ssNMR) were measured with the use of an Avance 400 WB Bruker device, using the CP/MAS method in a 4mm rotor at the speed of 13 kHz, normally at 25°C. Particle size distribution (PSD) was measured with the use of an optical Nikon eclipse N80i microscope, object lens LU PLAN Fluor lOx, eyepiece CFI10/22x_s used suspension media - paraffin oil.

Claims

A process for preparing glycopyrronium bromide in the form of [R,S] and [S,R] diastereoisomers, comprising separation of these desired diastereoisomers from the mixture of [R,S], [S,R], [R,R] and [S,S] diastereoisomers by two or more controlled crystallizations from a solution of a solvent selected from the group consisting of 1- propanol, 2-propanol, 1-butanol, and teri-butanol.

The process according to claim 1 , characterized in that the diastereoisomeric purity of the product is at least 99.9% of R,S / S,R.

The process according to claims 1 to 2, characterized in that said controlled crystallization comprises dissolution of the mixture of diastereoisomers in a hot state and subsequent cooling of the solution to the room temperature at the rate of 20°C / h.

4. The process according to claim 3, characterized in that said mixture of diastereoisomers is dissolved in a solvent at a temperature of from 50°C to the boiling point of the given solvent.

5. The process according to claims 1 to 4, characterized in that said solvent is 1-propanol or 1-butanol.

6. The process according to claim 5, characterized in that the size of all particles of the product is smaller than 20 μιη as measured with an optical microscope.

7. Crystalline glycopyrronium bromide in the form of [R,S] and [S,R] diastereoisomers, obtainable by the process according to claims 1 to 6, with the size of all the particles smaller than 20 μπι, as measured with an optical microscope, and a diastereoisomeric product purity of at least 99.9% R,S / S,R.