FI83516C - PHARMACEUTICAL FORM OF PHARMACOLOGICAL PROPERTIES 2,6-DIMETHYL-4- (3-NITROPHENYL) -1,4-DIHYDROPYRIDINE-3,5-DICARBOXYLSYRA-3- (1-BENZYLPYRROLIDIN-3-YLMETH-3-YL) -ES-3-YL) -DIASTEREOISOMER. - Google Patents

Description

1 83516

The process for the preparation of pharmacologically valuable 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) ester-5-methyl ester, grade A -reoisomeria - For the preparation of pharmacologically active 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) -ester -5-methylester A diastereoisomer

The process for the preparation of pharmacologically valuable 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) ester 5-methyl ester, grade A -reoisomer, a dextrorotatory optical isomer of this A-diastereoisomer, or a pharmaceutically acceptable acid addition salt of this diastereoisomer or optical isomer.

2,6-Dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) ester-5-methyl ester (YM- 09730) has the formula, No. 2 P "CH300C \ A / COO-γ-j 3 1 H ^ 2-0 YM-09730 was first synthesized by us. It has long-lasting vasodilatory and hypotensive activity (U.S. Patent 4,220,649; UK Patent 2,014,134).

YM-09730 has two asymmetric carbon atoms, but no isomers have been described in the above-mentioned patents and the existence of isomers has not been previously confirmed. The difference between the diastereo-isomer pair11a is the different absolute value of the reversibility 2,83516 and many physical and chemical properties (in contrast to the usual optical antipodes11a). Thus, isomers cannot be obtained separately from these differences in properties unless these properties have been elucidated.

We have now succeeded in separating isomers A and B of YM-09730 and their individual optical isomers and have found that "isomer A" (this term is used herein to mean both diastereoisomer A and its dextrorotatory optical isomer) has greater specific pharmaceutical effects than " isomer B "(referred to herein to mean both diastereoisomer B and its optical isomers) or a mixture of diastereoisomers A and B (YM-09730). The hydrochloride of di-astereoisomer A has a melting point of 200 to 206 ° C (decomposes), and the right-turning optical isomer of the hydrochloride has a melting point of 223 to 230 ° C (decomposes). The present invention therefore relates to diastereomer A of YM-09730 and its dextrorotatory isomer (identified by the melting points of its hydrochlorides) and also to their pharmaceutically acceptable acid addition salts.

Pharmaceutically acceptable acid addition salts suitable for this invention include malonates, oxalates, p-nitrobenzoates, 2-ketoglutarates, maleates, 1-malates, hydrochlorides, sulfates, p-toluenesulfonates, phosphates and succinates.

In the pharmacological experiments described below, isomer A of this invention and its pharmaceutically acceptable additives were added. . acid addition salts increase blood flow to the heart 15 to 38 times. as much as diastereoisomer B and 14 to 35 times as much as a mixture of diastereoisomers A and B in equal amounts when administered directly to the cardiac arteries. This indicates the high affinity of isomer A of the present invention for cardiac arteries.

3,83516

On the other hand, the B-isomer11a had almost the same effect as the mixture of isomers. This means that the pharmacological activity of YM-09730 is not a simple average of the effects of the isomer. There is new knowledge in pharmacology that Isomer A and its pharmaceutically acceptable acid addition salts have a high affinity for cardiac arteries. This increases the possibility of using the compounds of this invention as medicaments.

The preparation of the A-isomer and its pharmaceutically acceptable acid addition salts can be carried out as follows: I. Preparation of the A-diastereoisomer YM-09730 can be prepared by the synthesis of Hantzch (Ann. Chim., 215, 1 (1982)) as given in the following Comparative Example 1, to give a 1: 1 mixture of the A-diastereoisomer and the B-diastereoisomer.

The diastereoisomer A can then be obtained as follows: (a) The mixture of diastereoisomers was chromatographed on silica gel using a mixed solution of ethyl acetate and acetic acid as eluent. The A-diastereoisomer was obtained from the previous eluate and the B-diastereoisomer from the later eluate.

Any conventional silica gel suitable for the purpose can be used as the silica gel support in thin layer chromatography. The mixing ratio of the ethyl acetate and the acetic acid used as the eluent is also not limited in any way, but a mixed solvent containing a small amount of acetic acid is usually used. The mixing ratio is, for example, 30 to 50 v / v ethyl acetate and about 1 to 10 v / v acetic acid. If the acetic acid concentration is reduced, the elution time of the desired compound increases. The elution rate and processing temperature can be selected as desired.

4,83516 (b) In another method, an acid addition salt of the A-dia stereoisomer is obtained by converting a mixture of diastereoisomers into an acid addition salt and a fractionally crystallized mixture of salts.

Suitable acid addition salts in this process include, for example, malonate, p-nitrobenzoate, maleate, etc.

These acid addition salts are crystalline salts. The solubility of the A-diastereoisomer and the B-diastereoisomer in an organic solvent is different, making it possible to prepare the A-diastereoisomer by fractional crystallization. A particularly suitable acid addition salt is malonate. When malonate is used, crystals containing a very high content of the diastereoisomer A can be obtained in one recrystallization. Suitable solvents for use in the process include, for example, met a 11 o 1i, ethanol, acetone, acetonitrile, etc.

The A-diastereoisomeric acid addition salt obtained by the above-mentioned method can be converted into another desired acid addition salt by converting it to the free form and reacting with another acid.

II. Preparation of dextrorotatory optical isomer A: (a) The dextrorotatory optical isomer of diastereoisomer A can be obtained by reacting the free bases as a mixture of diastereoisomers A and B or the free base of the acid addition salt of diastereoisomer A of YM-09730 obtained by the above method With L - (-) - maleic acid and then separating the product - - optically in the usual way, as described below.

: * (h) In the preferred method, the A-diastereoisomer: dextrorotatory optical isomer is separated from a mixture (of formula 1 below) of the O-diastereoisomer 5 83516 dextrorotatory optical isomer Q "N ° 2 / H ^ 1) CH 2 OOC ^^ X ^ -COO ^ H Vh

3 3 CH —O

(where the wavy bond means an α-bond or a β-bond, the bond shown by the thick arrow is a β-bond, and the bond at the 3-position of the pyrrolidine ring is a specific β-bond).

The starting compound (I) can be prepared by (i) reacting an m-nitrobenzaldehyde of formula (II) [; V *

CHO

:: S- (3-acetoacetoxy-1-benzylpyrrolidine of formula (III)

CH3COCH7COO

V? m. »-o · '. and the 3-amino acid of formula (IV) is a diacid * 83516

CH C = CHCOOCH

3 | 3 (IV);

N1 (? (Ii) By reacting m-nitrobenzaldehyde of formula (II), acetoacetic acid methyl ester of formula (V) ch3coch2cooch3 (v) and (S) -3- (3-aminocrotonoyloxy) -1 of formula (VI) -benzylpyrrolidine CH 3 C = CHCOC) _

u V

^ O

: (iii) By reacting (S) -i-: ·, - benzyl-3- [2- (m-nitrobenzylidene) acetoacetoxy / pyrrolidine of formula (VII) ςτ ·

CH

s CH 2 COCOO-s__.

(VII),

·· ;; -.- O

previously obtained by reacting the above-mentioned m-nitrobenzaldehyde (II) and the above-mentioned (S) -3-acetoacetoxy-1-benzylpyrrolidine (III) and the above-mentioned 3-aminocrotonic acid methyl ester (IV); or 7,83516 (iv) By reacting 2- (m-nitrobenzylidene) acetoacetic acid methyl ester of formula (VIII) 0no2 (VIII)

CH

tl CH3COCCOOCH3 previously obtained by reacting m-nitrobenzene Idehyde (II) and acetoacetic acid methyl ester (V) and (S) -3- (3-aminocrotonoyloxy) -1-benzylpyrrolidine (V).

These reactions take place without a solvent, but it is preferable to carry out the reaction in a solvent which does not participate in the reaction, such as · alcohol, dioxane, dimethylformamide, dimethylform acid, acetonitrile, water, etc.

The reaction is carried out by heating a mixture of each component in which the components have approximately equal moles of i -; amounts.

:. said compound (IV) or (VI) can be obtained by reacting the compound (V) or (I I) with ammonium acetate and acetic acid in hercene and azeotroping the product. The compound (IV) or (VI) thus obtained is subjected to the reaction (i) or (ii) described above after being isolated from the reaction mixture or without isolating it.

. . Compound (VII) or (VIII), which is the reaction product of the first step of process (iii) or * .. (iv), can be carried out. after isolation or without isolation.

A-d ia s t ereo 1 someer ien dextrorotatory optical isomer; · And a mixture of the levorotatory optical isomer * of the B-diastereoisomer thus obtained is chromatographed using silica gel 8 83516 as a carrier and a mixture of ethyl acetate and acetic acid as eluent to separate the levorotatory optical isomer; or the mixture described above is reacted with an L - (-) - ma 1 non-acid to form a right-turning optical isomer of the A-diastereoisomer 1 - (-) - m »and a left-turning optical isomer L - (-) of the B-diastereoisomer. ) -ma la at, whereby the L - (-) - malate of the dextrorotatory optical isomer of the A-diastereoisomer can be obtained by fractional crystallization of the mixture.

Separation by column chromatography gives the dextrorotatory optical isomer of the A-diastereoisomer from the first eluate and the dextrorotatory optical isomer of the B-diastereoisomer from the subsequent eluate.

Any s’li cinnamon commonly used in pv 1 väsk routa t ogra f ia «can be used as S '1 ikag *' el i kunta jana. The mixing ratio of the mixture used as eluent of ethyl acetate and acetic acid is not particularly limited in any way, but a mixed solvent containing a small amount of acetic acid is preferred. It is preferred that the mixing ratio be 30 to 50 v / v ethyl acetate and about 1 to 10 v / v acetic acid. When the acetic acid content is further reduced,; - increases the elution time of the desired compound. The elution time and processing temperature can be selected as desired.

The method using L - (-) - maleic acid can also be used to separate the dextrorotatory optical isomer of the A-diastereoisomer by crystallization because the dextrorotatory optical isomer of the A-diastereoisomer:: L - (-) - ma la a11i is crystalline. Suitable solvents for use in fractional crystallization include, for example, methanol, ethanol, acetone, acetonitrile, etc.

: 'The dextrorotatory optical isomer of the A-diastereoisomer; **. the L - (-) - mate thus obtained can be used as a drug, but if necessary it can be converted into acetate or 9,83516 other suitable salt by treatment with a base to form the free compound and treatment with a suitable acid.

The levorotatory optical isomer of the B-diastereoisomer separated in the above process can be hydrolyzed to give (S) - (-) - 1-benzyl-3-hydroxypyrrolidine, which can be reused as a starting material in the preparation of a compound of formula (I).

The following tests have been used to illustrate the pharmacological activities, acute toxicity, and clinical doses of the compounds of the invention.

(1) Cardiovascular dilating effect in anesthetized dogs:

In dogs with thoracic opening and anesthetized 3 U mg / kg i.v. with pentobarbital sodium, *: arterial blood was derived from the carotid artery to the left · '· cardiac artery by a extracorporeal loop. The mice were connected to a servo-controlled pump (model 1215D,

Harvard Apparatus) to maintain a perfusion pressure of 120 mmHg (constant by means of a pump control device (SCS-22, Data Graph Co., Tokuya Tukada et al., Folia Pharmacol. Japan, 74, 49, 1978).) In order to record cardiac blood flow, an out-of-body type electromagnetic flow electrode (MF - 29, Nihon Koden) The test compound was administered in a dose of 1 μg directly to the cardiac artery and then the blood flow in the heart was monitored until the blood flow returned to the pre-treatment value, and the area of the curve was calculated. 'percentage increase in blood flow in the heart after intracardiac injection, below and this value was used; V as an index of the total increase in blood flow in the heart.

The results are shown in Table 1.

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Table 1. Cardiovascular dilating activity in anesthetized dogs

Blood flow Exposure time total increase in heart at 1 μg dose (%, min.) A-diastereoisomer (d1-form) hydrochloride 1816 + 377 60 A-dia stereoisomer (d-form) -hydrochloride 4559 + 894 120 A-diastereoisomer (1-form) ) hydrochloride 120 + 21 5 B-diastereoisomer x (dl form) hydrochloride 120+ 28 5

A 1: 1 mixture of Dia Stere isomers A (dl form) and B (dl form) hydrochlorides 129+ 47 10 The total increase in cardiac blood flow caused by isomer A was approximately 15-38 times higher than the 1 increase caused by one high doses of B-isomer ---- hydrochloride and an equal mixture of A and B isomers.

; . This indicates that the A-isomer hydrochloride has a higher affinity for the cardiac artery. In addition, the cardiac vasodilating effect of A-isomer hydrochloride was clearly longer in duration than when B-isomer hydrochloride and a mixture of A- and B-isomers were used. Such oral affinity for the cardiac artery and long duration indicate that the A-isomer hydrochloride is useful in the treatment of cardiac vascular disease such as angina pectoris.

: * ·. (2) Hypotensive effect in anesthetized rats:. * * Blood pressure was measured in urethane anesthetized rats. *** Test compound was administered intravenously at increasing * - 'increasing doses every 20 minutes. From the dose-response curve n 83516, the dose of each compound required to lower blood pressure by 30 mmHg (ED 30 mmHg) was calculated. These doses are shown in Table 2.

Table 2. Hypotensive activity in anesthetized rats ED30 mmHg (mg / kg iv) A-diastereoisomer (d1-inoform) hydroclone 0.002 B-diastereoisomer (d1-form) -hydro chloride 0.14 ΓFrom Table 2 it can be seen that A -diastereoisomer hydrochloride had a hypotensive activity about 70-fold more potent than the effect of diastereoisomer hydrochloride.

(3) Acute toxicity in mice: Six-week-old ICR mice weighing 27 to 29 years old were used. The test compound was suspended in 0.5% methylated saline and administered orally to mice. The LD50 values of the compounds were calculated by the method of Litchfield & Wilcoxon (Journal of Pharmacol. & Ex. Therap., 96, 99-113, 1949). The results are given in Table 3.

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Table 3. Acute toxicity in mice lD50 mg / kg p.o.

Diastereoisomer A (Form dl) - Hydrochloride 293 (242-360) Diastereoisomer A (Form D) - Hydrochloride 190 (138-228) (4) Clinical Dosages: The clinical doses of the compounds of this invention depend on the weight of the patient and the condition of the disease. . Optimal doses for intravenous injection are usually 0.1 to 2 mg and for oral administration 5 to 20 mg once or twice a day.

The following examples and formulation examples illustrate the compounds of this invention, their preparation, and drugs containing them.

• The preparation of a mixture of the A-dia stereoisomer and the B-diastereoisomer used as a raw material is illustrated in Example 1 and also the (S) - (-) - 1- the preparation of benzyl-3-hydroxypyrrolidine is shown in Comparative Examples 2 to 4.

Comparative Example 1 1.51 g (0.01 mol) of 3-nitrobenzladehyde, 2.61 g (0.01 mol) of 1-benzyl-3-acetoacetoxypyrrolidine and 1.15 g (0.01 mol) of 3- Aminochrops were dissolved in 5 ml of isopropanol and the resulting solution was refluxed for 8 hours. The solvent was then distilled off under reduced pressure, the residue thus formed was dissolved in chloroform, and the solution was washed successively with dilute hydrochloric acid, water and saturated aqueous sodium hydrogencarbonate solution, and dried over anhydrous magnesium sulfate. The solvent was distilled off, whereby the solvent was distilled off. 4.91 g of caramel-like 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 5- (1-benzylpyrrolidin-3-yl) ester 5-methyl ester.

The d-diastereomers obtained as a crude base were analyzed by reverse phase high performance ion-pair liquid chromatography (HPLC) under the following conditions:

Column: 4.6 mm x 300 mm, packing material Nucleosil® ^ 1 C1Q, column temperature: 30 ° C, mobile phase: acetonitrile-0.05 m potassium dihydrogen phosphate (pH 3) (20:80 v / v) with tetra-n-n-ethylammonium bromide (3 ml / mole) as counterion, flow rate: 0.9 ml / min, monitor: uitravio1 e detector (λ 254 nm).

The ratio of the A-dia stereoisomer (retention time 28 min.) To the B-: diastereoisomer (retention time 29 min.) Was 1: 1.

Based on NMR analysis, it was confirmed that the A-diastereoisomer and the B-diastereoisomer are different. ‘Chemical shifts with respect to the methylene protons of the N-benzyl group of YM-09730. When measured in heavy methanol d 2 (CD 2 OD) Jeo 1 NMR spectrometer FX-900, A-i soineer i 11 ä and B-i someer i 11 ä have siriglet signals corresponding to 4.40 ppm and 4.30 ppm, respectively.

Comparative Example 2 (1) 17.7 gd 1 -1 - ben t sy y 1 i - 3 - h ^ tfj-ox ip yr ro 1 idi in ia and "· - · 15.2 g D- (-) -mant The acid was dissolved in 66 ml of acetone: V was heated and allowed to stand overnight at 4 ° C to precipitate crystals, 8.5 g of the crystals thus formed were collected and recrystallized from 26 ml of acetone to give 5 , 1 q of (S) - (-) - 1-benzyl-3-hydroxypyrrolidine D - (-) - mandelate, specific rotation [α] D -45.5 ° (c = 1, methanol). recrystallization was repeated, the specific inversion did not change, melting point 101-102 ° C.

In NMR analysis, the signal of N-CJ1 -Ph (the mandelate thus obtained in the (5) - (-) form) was detected at 4.03 ppm (sinalet, 2H). AB-type quartet of the (R) - (-) form (J = 12.5

Hz) signal at 4.01 ppm was not detected.

(2) 22 g of (5) - (-) - 1-Benzyl-3-hydroxypyrrolidine D - (-) - mandelate was dissolved in 50 ml of chloroform, and the chloroform solution thus formed was washed with a solution containing 14.4 g of sodium carbonate in 90 ml. water, and dried over anhydrous magnesium sulfate. After listing the chloroform, the residue was distilled under reduced pressure to give 11.5 g of (S) - (-) - 1-benzyl-3-hydroxide.

O

pyrrolidine. The boiling point was 109 ° C / 0.65 mmHg and / <Vjj "** was -3.77 ° (c = 5, methanol).

Comparative Example 3: 75 g of (S) - (-) - maleic acid and 75 ml of benzylic acid were reacted for 3 hours at 170 ° C to give 52.7 μl of S ) - (-) - 1-benzyl-3-hydroxy-succinimide (melting point 99-101 ° C. Specific rotation / n./. -51.1 °, (c = 1, methanol)). 9.73 g of lithium aluminum hydride were suspended in 340 ml of anhydrous tetrahydrofuran, and a solution of 20.5 g of the imide described above in 200 ml of anhydrous tetra- was added to the suspension under ice-cooling. hydrofuran. The mixture was refluxed for 3 hours, after which 100 g of sodium sulfate i-decahydrate was added to the mixture under ice-cooling, and the resulting mixture was stirred overnight under ice-cooling. The insoluble matter was separated by filtration, the solvent was evaporated from the filtrate under reduced pressure, and the residue was distilled under reduced pressure to give 13.0 g of (S) - (-) - 1-benzyl-3-hydroxypyrrolidine, boiling at 109-115 ° C / 0 , 8 mm H g and specific characteristic / a / j ^ -3.0 ° (c = 5, me ta noli). Analyzed by α N M R using the 3-position as the transfer reagent, it was confirmed that the (S) - (-) form obtained above contained 10% R - (+) - 1-benzyl-3-hydroxypyrrolidine. The product was converted to D - (-) - Mande plate as described in Comparative Example 2. The mandelate (-^ -45.2 ° (c = 1, MeOH) obtained by recrystallization from three times the volume of ethanol and then from six times the volume of ethanol 1o-10 (1: 5 v / v) was treated with chloroform and aqueous sodium carbonate solution as described in Comparative Example 2. in a manner to give 8.6 g of (S) - (-) - 1-benzyl-3-hydroxypyrrolidine; boiling point 115-120 ° C / 1.2-1.5 mmHg, m.p. -3.77 ° (c = 5, methanol).

Comparative Example 4 3.4 g of 2 - (-) - pinene was added to 50 ml of 9-borabicyclo [3.3.1] n / a (0.5M tetrahydrofuran solution) and the mixture was stirred for 5 hours at 60 °. The mixture was cooled to room temperature, then 1.75 g of 1-benzyl-3-pyrrolidinone was added, and the mixture was stirred for 4 days at room temperature, followed by the addition of 1.3 ml of acetaldehyde at 0 ° C. :in.

: The solvent was then distilled off under reduced pressure, and 20 ml of ether was added to the ice. The mixture was cooled to 0 ° C and then 1.5 ml of 2-aminoethanol was added thereto, and the resulting mixture was stirred. The precipitates formed were filtered off. Ether solution recovered as filtrate. . was extracted with 1 N hydrochloric acid and the hydrochloride thus formed. the acid layer was made alkaline with sodium carbonate: and extracted with dichloromethane. The extract thus formed: dried over anhydrous magnesium sulfate and concentrated to give 1.1 g of crude product. The crude product was distilled under reduced pressure to give 0.6 g of 83 51 6 purified product - boiling point, 106 ° C / Q, 9 mmHCl.

The (S) - (-) - 1-benzyl-3-hydroxypyrrolidine thus obtained was 30% as analyzed by NMR spectra with respect to the 3-arsenic proton using tu-TFMC (1 L) transfer reagent.

Preparation of diastereoisomer A Example 1 1.

4.91 g of the crude free base of YM-09730 obtained in Comparative Example 1 was dissolved in 25 ml of chloroform, 15 ml of 10% hydrochloric acid was added thereto, followed by stirring well, and the organic phase thus obtained was separated. The organic layer was re-treated as described above with 10 ml of 1:11 a 10% hydrochloric acid, the treated product was dried over anhydrous magnesium sulfate, and the solvent was distilled off under reduced pressure. The residue thus obtained was dissolved in 10.4 ml of acetone, and the solution was allowed to stand to obtain a 3.5 N crystalline hydrocarbon of YM-U9730. The product was dissolved in 1.8; m 1 of methanol and recrystallized by adding - - 8 ml of acetone. Crystallization once more gave 2.38 g! '. Hydrochloride of YM-09730. The diastereoisomer A; ; the ratio to the B-diastereoisomer in the hydrochloride was 65.6: 34.4 based on the HPL C value. 2.15 g of the above. the salt was dissolved in 25 ml of chloroform, the solution thus obtained was washed twice with 15 ml of saturated aqueous sodium hydrogencarbonate solution, and the formed organic layer was collected and dried over anhydrous magnesium. .. with sulfate. The solvent was distilled off under reduced pressure. 2 a • - the free base thus obtained py 1 v ä skroma t o g ra f o i t i i n silica *. . gel (column LiChroprep® S i 6 0, C size; eluent · ethyl acetate / acetic acid, 30: 5 v / v) to give the oily acetate of the diastereoisomer A.

The product was dissolved in 10 ml of chloroform. The resulting J7 8351 6 L was washed successively with 10 mL of 1: 11 saturated aqueous sodium ethyl carbonate, 10 mL of water and 10 mL of 10% hydrochloric acid and dried over anhydrous Mayn precursor 1 f aa111la. The solvent was then distilled off under reduced pressure, and the residue thus obtained was treated with 0.1 M acetone to give 0.4 g of the hydrochloride of the A-diastereoisomer.

Example 2 4.91 g of the crude free base of YM-09730 obtained in Comparative Example 1 and 1.04 g of malonic acid were dissolved, and the resulting solution was allowed to stand overnight at 0-5 ° C. The crystals thus formed were collected by filtration, washed with a small amount of cold acetonitrile to give 2.03 g of malonate of YM-09730 (the ratio of A-diastereoisomer to B-diastereoisomer was 09.1: 10.9). the fetus was recrystallized twice from 25 times the volume of inethanol to give 1.0 g of malonate of 100% diastereoisomer A of YM-0973Q.

L s i m e r k k i 3 4.91 g of the crude free base of YM-09730 obtained in Comparative Example 1 and 1.04 y of malonic acid were dissolved in 19 ml of methanol under heating, and the solution thus formed was allowed to stand overnight at 0-5 ° C. The formed crystals were collected by filtration, washed with methanol and dried under reduced pressure to give 1.88 g of YM-09730 malonate. The A: B ratio of the diastereoisomers of the crystals thus obtained was 90.7: 9.3. The crystals were crystallized in two. again from methanol to give malonate of the A-diastereoisomer, which did not contain any B-iso- m m o r i; i.

Example 4 1.1 83516

The same procedure as in Example 3 was followed, but acetones were used instead of ethanol. 2.03 q of malonate were obtained. The ratio of diastereomers A: B was 89.1: 10.9. the fetus was recrystallized from a 23-fold volume of methanolite to give 1.57 g of malonate of diastereoisoters A and B in a ratio of 99.5: 0.5. After further recrystallization from 25-fold methanol, 1.27 g of malonate of YM-Q973U (in which no δ-diastereoisomer was detected by HPLC) was obtained. 1.27 g of malonate was suspended in 5 inl of chloroform; The Oro formulation suspension was treated successively twice with saturated aqueous sodium carbonate solution, twice with 2.5 ml of water and then twice with 2.5 ml of 10% hydrochloric acid; the chloroform solution thus obtained was dried over anhydrous magnesium sulfate, filtered and evaporated to dryness under reduced pressure. The residue thus formed was dissolved in 2 ml of acetone. The solution was allowed to stand to precipitate 1.09 g of the hydrochloride of the stereoisomer A: of YM-09730.

: Example 5 595 mg of malonate of diastereoisomer A of compound YM-09730 was suspended in 5 ml of chloroform. The resulting suspension is hand- '.'. was treated twice with 2.5 ml of saturated aqueous sodium hydrogen carbonate solution and then twice with 5 ml of water. The resulting chloroform solution was dried over anhydrous magnesium sulfate and evaporated to dryness under reduced pressure. .. to give 491 mg of the free base of diastereoisomer A. ·. · as a caramel-like substance. The product and 126 mg of oxalic acid dihydrate were dissolved in 3 ml of acetone, the resulting solution was allowed to stand at 4 ° C and the separated crystals "... were collected by filtration to give 400 μm. YM-09 7 id - y hd ι s t. ecu A - cl ias I e re Oiso me er, in uksaluattia.

Example 6 ^ 83516

The procedure of Example 5 was followed, but 146 my 2-ketoglutaric acid was used instead of oxalic acid to give 250 mg of 2-ketoglutarate of diastereoisomer A of YM-09730.

Example 7

The procedure of Example 5 was followed, but 167 mg of p-nitrobenzoic acid was used instead of oxalic acid to give 530 mg of p-nitrobenzoate of diastereoisomer A of YM-09730.

Example B

The procedure of Example 5 was followed, but 116 μm maleic acid was used instead of oxalic acid to give 300 mq of maleate of the Adiastereoisomer of YM-09730.

. f. s i in e r k k i 9 491 mq of diastereoisomer A of YM-09730 was dissolved in -2 ml of acetone, 1 ml of phosphoric acid (1 mol) was added; . methanol solution and the solution was allowed to stand at 4 ° C.

The separated crystals were collected by filtration to give:; 4Θ mq of phosphate of diastereoisomer A of YM-09730 was obtained.

The following table gives the properties of the desired compounds obtained in Examples 1-9.

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Features

Product Formula sp. Elemental analysis CU) HU) NU) Cl (%)

Phosphate C27H29N3 ° 6 216-218 54.39 5.56 6.96 • H3PO4.1 / 2H2O 54.18 5.56 7.02

Hydrochloride G27 ^ 29 ^ 3G6 203-205 61.59 5.71 8.08 6.90 • HCl (dec.) 61.42 5.73 7.96 6.71 p-nitrobenzoate ('27 ^ 29N3 ° 6 150 ~ 151 61.58 5.20 8.32 • C7H5NO4-1 / 4H2O 61.58 5.24 8.32

Maleate G27H29N3 ° 6 168-169 61.40 5.49 6.85 • C4H404 61.28 5 ^ 47 6.92 2-keto-C27H29N3 ° 6 160-161 60.36 5.48 6.52 glutarate · 03Η403 60.28 5.53 6.59

Oxalate C27H29N3O 6 179-180 60.14 5.62 6.93 • C, H, 0. 60.36 5.70 6.88 l 2 4

Malonant ^ 27 ^ 29 ^ 3 ^ 1®1ϊ ^ ~ 69/89 5.37 7.23, c H 0 182.5 60.50 5.58 7.06 344 (naj.) (Upper: Value obtained)

Hydrochloride: NMR (CD OD, (ς (Lower: Calculated value) internal standard, δpprn) 1.80-2.70 (2H, broad m, C 4'-H 2) 2.32.2.34 (6H, s, C 2 F 6 -CH 3) ) jfY ™ 2 3.0-4.0 (4H, m, C2'f5 * -H2) \: 3.63 (3H, s -COOCH3) 4.40 (2H, s, -ch2- φ) V ° 2C ^^ ^ cO2 l__4.

5.08 UH, s, C 4 -H) h C 2 H 3 CH 2 '^ 5' 5.30 (1H, », C 3 -H) 3 3 7.30-8.20 (9H, m, benzene - -HC1 '.

tire h)

Example ID

8351 6 1.5 g of the malorate of the A-diastereoisomer A obtained in Example 3 or 4 were suspended in 5 ml of chloroform, and the suspension was treated successively twice with 3 ml of saturated aqueous sodium hydrogen chloride solution and then twice with 3 ml of water. The chloroform solution was dried over anhydrous magnesium sulfate, and then the solvent was distilled off under reduced pressure. To the obtained residue was added 6 ml of ethanol, and the mixture was allowed to stand overnight at 5 ° C to give 0.86 ° C of the free base of diastereoisomer A of VM-09730 as crystals.

Melting point: 145-148 ° C

Elemental analysis (C H0QN 0,)

C i Lj j O

C (8) H (8) N (8)

Calc .: 65.98 5.95 8.55

Found: 66.04 6.00 8.53 NMR (CDCl 3 TMS internal standard, 6ppm): δ. 1.40 - 2.96 (6H, m, 4 ', 4', H2 ') 2.34, 2.36 (6H, s, C2H2H) 3.65 (5H, s, - 3.10 (1H, s, C 1 H) ly 3.12 (1H, m, C-H) 5.78 (1H, broad s, NH) 7.18 - 8.25 (9H, m, H of the benzene ring), ·V · H3C ° 2C ^ ^ SKCO2v ^ J_4, 2 CH2 '^ n ^ 5 3 1 Λ: V CH2- <p 22 83 5 1 6

Preparation of dextrorotatory isomer A Example 11 (1) 4.91 g of the crude free base obtained in Comparative Example 1 and 1.04 g of malonic acid were dissolved in 15 ml of acetonitrile. The solution was allowed to stand overnight at 0 -5 ° C. The crystals thus separated were collected by filtration (2.03 g) and recrystallized twice from 25 times the volume of methanol to give 1.27 g of malonate of the A-diastereoisomer A of YM-U9730, which did not contain any B-diastereoisomer. The melting point of the product was 181.5-182.5 ° C (decomposes). 1.27 g of the malonate thus obtained were suspended in 5 ml of chloroform, and the suspension was washed successively twice with 2.5 ml of saturated aqueous sodium hydrogencarbonate solution, once with 2.5 ml of 1: 1 water and then twice with 2.5 ml of aqueous 10. p% hydrochloric acid solution. The chloroform solution was dried over anhydrous magnesium sulfate and evaporated to dryness under reduced pressure. The resulting residue was dissolved in 2 ml of acetone and the solution was allowed to stand. 1.09 g of the hydrochloride thus separated from the diastereoisomer A of YM-09730 was recovered.

(2) 4.67 g of YM-09730 compound A-diastereoisomer hydro- *; the chloride was treated with saturated aqueous sodium bicarbonate to give 4.35 g of the free base.

Then, 4.35 g of the free base thus obtained and 1.18 g of L - (-) - maleic acid were dissolved in 28 ml of ethanol under heating, and the solution was allowed to stand overnight at 0-5 ° C. The crystals thus separated were collected by filtration and dried. .. to give 2.43 g of the L - (-) -ma 1 aa 11 of the dextrorotatory optical isomer of the diastereoisomer A of YM-09730.

• The crystals were recrystallized from 85 mL of ethanol to give. · 2.21 g of L - (-) - malate were obtained. The specific inversion (α) ^ + ... was + 82.2 ° (c - 0.5, methanol). When this product was crystallized from ethanol, no change in intrinsic capacity was observed.

23 8351 6

Melting point: 190-191 ° C (decomposes).

Elemental analysis (C27H29N3O6, C4H6O5 ° C (%) H (%) N (S)

Found: 3 9.72 S, 8 0 6.7 3

La s k .: 59.5 1 3.64 6.72

Example 12 4.91 g of the free crude base of YM-09730 obtained in Comparative Example 1 and 1.34 g of L - (-) - maleic acid were dissolved in 23 ml of acetone. The resulting solution was stirred for 48 hours at 0-5 ° C. The separated crystals were collected by filtration and washed with a small amount of cold acetone to give 0.57 g of the dextrorotatory optical isomer I of the A-diastereoisomer A of YM-09730 I. - (-) -mu 1 au 11 ι s. The Um1 female vocal rate was + 78.3 ° (c = 0.5, methanol).

The product was recrystallized from a 50-fold volume of ethanol to give the specific crystals of the obtained crystals (0.44 °) (α) = + 82.2 ° (c = 0.5, methanol).

- Melting point: 190-191 ° C (dec.).

Example 13 2.21 g of the levorotatory optical isomer L - (-) - Mala all of the A-diastereoisomer A of the YM-0 97 3 0 obtained in Example 11 was suspended in 8 ml of chloroform, and the suspension was treated successively twice 4. 3 ml of saturated aqueous sodium hydrogen carbonate solution, once with 4.3 ml of water and then twice with 4.3 ml of 10%. . hydrochloric acid. The chloroform solution was dried over anhydrous magnesium. sulfate. After filtration, the solution was evaporated to dryness under reduced pressure. The resulting residue was dissolved in: 3.5 ml of acetone, and the solution thus obtained was allowed to stand to give 1.64 g of the hydrochloride precipitate of the dextrorotatory optical isomer of the diastereoisomer A-24 8351 of YM-09730.

The specific inversion / α / pU was + 116.5 ° (c z 0.5, methanol). Melting point: 223 - 225 ° C (dec.)

Elemental analysis (Co., HoqN, 0.1HCl)

l i L7 J O

0 (¾) H (5) N (Ä) 01 (¾)

Found: 61.35 5.55 8.01 6.96 L a s k. : 61.42 5.73 7.96 6.71 WMR (CD ^ OD, TM ^ internal standard, <5ppm): 1.80-2.70 (2H, broad m, 0 ^ - - H) 2.32 , 2.34 (6H, s, C 26 -CH 3) 3.0-4.0 (4H, m, C 2,> 5'-H 2) 3.64 (3H, s, -COOCH 2) 4.4 2 (2H, s, -CH (i) 5.08 (1H, s, C 4-11) 5.3 ° (1H, m, 1:,,-11) 7.30-8.20 ( 9 H, m, Benzene ring H)

Example 14 In 5 ml of isopropanol was dissolved (S) -3-acetoacetoxy-1- - benzylyrvalvaline (by reacting 1.77 g of · - · (S) - (-) -1-benzyl-3-hydroxy). rrol ididine ([.alpha.] D @ 20 -3.77 DEG C., = 0 = 5, methanol) and 0.84 .mu.l of diketene for 3 hours at 70-80 DEG C.), 1.51 .mu.l of m-nitrobenzladehyde and 15 g of 3-amino - '' · kr ot is an acidic solution, and the solution is refluxed for 8 hours. The solvent was then distilled off under reduced pressure.

The residue thus obtained was dissolved in chloroform. Formed. The solution was washed successively with dilute hydrochloric acid, water and then with saturated sodium bicarbonate solution and dried over anhydrous magnesium sulfate.

The solvent was then distilled off under reduced pressure to give 4.91 g of a mixture of the levorotatory optical isomer of YM-097 30 .. * to the right-turning optical isomer and the levorotatory optical isomer of B-diastereoisomer 2> 8351 6 as a caramel . The crude free base thus obtained was subjected to silica gel column chromatography (column - Wakogel C-200, 2,000 a; eluent - ethyl acetate / acetic acid, 6: 1 v / v) to give the dextrorotomer of YM-09730 A diastereoisomer. oily acetate of the optical isomer with a retention time of 28 minutes in HPLC chromatography.

The product was treated with a saturated aqueous solution of sodium hydrogencarbonate in chloroform followed by dilute hydrochloric acid to give 1.68 a of the hydrocrone of the dextrorotatory optical isomer 20 of the diastereoisomer A of YM-09730. The omirian inversion capacity was +116.5 (c = 0.5, me La no li).

Example 15 A, 9 y The crude free base obtained in Example JA was dissolved in 25 ml of chloroform, 15 ml of 10% salt was added and the resulting mixture was stirred well. The organic layer thus formed was separated, washed again with 10 ml of 10% hydrochloric acid, and dried over anhydrous magnesium sulfate. This - ·. then the solvent was distilled off under reduced pressure. Formed; * the residue was dissolved in 10% acetone and the solution was administered; stand for 2 days at 4 ° C to give 2.7 a of hydrochloride collecting the formed crystals. To remove the levorotatory optical isomer of the B-diastereoisomer, the product thus obtained was treated with saturated aqueous sodium hydrogencarbonate in chloroform to form * · free base. 0.68 g of L - (-) - malic acid in 15 ml of ethanol was added and the resulting mixture was allowed to stand for 2 days at 4 ° C. The crystals thus separated were collected by filtration and recrystallized from ethanol to give 1.33 g of the right-turning optical isomer L - (-) - malate of the diastereoisomer A of YM-09730, 26 8351 6 having a retention time of 28 minutes in HPLC chromatography. Specific rotation / ol / q® + 82.1 ° (c = 0.5, methanol).

Example 16 1.25 g of the L - (-) malate of the dextrorotatory optical isomer of the diastereoisomer A of YM-09730 was suspended in 5 ml of chloroform, and the suspension thus obtained was treated successively twice with 3 ml of saturated aqueous sodium hydrogencarbonate solution and then twice with 3 ml. ml of water. The chloroform solution was dried over anhydrous magnesium sulfate, and then the solvent was distilled off under reduced pressure. To the residue thus obtained, 6 ml of ethanol was added, and the mixture was allowed to stand overnight at 5 ° C to give 0.8 Å of a crystalline free base of the dextrorotatory optical isomer of the diastereoisomer A of YM-09730.

Melting point: 13B - 1A0 ° C.

Elemental analysis (CHOQN, 0,) L / LJ J 6 C (%) H (») N (%)

Lask. : 65.98 5.95 8.55: Found: 66.0A 5.96 8.51 NMR (CDCl 3, δς internal standard, 6ppm): - 1.40 - 3.0 (6H, m, c2.fa , Δ, -H 2): Y 2.3 A, 2.36 (6H, s, Γ? -CH 3) 3.65 (5H, s, -C00CH 3 and -CH 2) 5.10 (1H, s, (N-H) 5.12 (1H, m, C 3, -H) 5.78 (1H, broad s, -NH) 7.16 - 8.2A (9H, m, benzene ring H) 27 8351 6

Formulation Example 1 (tablets)

One tablet 5000 tablets A-isomer-hydrochloride 10.0mq 50 q

Lactose 101.0 mg 505 g

Maize starch 25.3 mg 126.5 g II hydroxide ι propyl] i. It 1.1 u 1 oosa 3, 0 mg 15 g M Agnes ι umaleura L l: 7, 7 mg 3, 5 a 140 mg 700 g 150 q aqueous 10% hydroxypropylcellulose was added to a single sponge mixture containing 50 g of A-isomer hydrochloride, 505 a of lactose and 126.5 g of corn starch, and the mixture was kneaded and granulated. After drying, 3.5 g of magnesium stearate was added to the granule, the granules were mixed until homogeneous, and then formed into tablets weighing 140 mg.

Formulation Example 2 (capsules) ;; One capsule 1000 capsules A-isomer hydrochloride 10.0 mg 10 g

Crystalline lactose 189.0 mg 189 g

Magnesium stearate a11i 1.0 mg 1 g 200 mg 200 a

The above components were mixed evenly, and 200 mg of the mixture was filled into each capsule to obtain a capsule-modifying drug.

Ormu 1a Sub - example 3 (injection). A-isomer hydrochloride 1 mg D-sorbitol 100 mg

Said components were dissolved in distilled water for injection 28,835,16 and the pH was adjusted to 4 by the addition of hydrochloric acid, after which the total volume was adjusted to 2 ml by adding distilled water for injection.

Claims (2)

29 83 51 6 A process for the preparation of pharmacologically valuable 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) ) A-diastereoisomer of the ester 5-methyl ester, a dextrorotatory optical isomer of this A-diastereoisomer or a pharmaceutically acceptable acid addition salt of this diastereoisomer or optical isomer, characterized in that a) 2,6-dimethyl4- (3-nitrophenyl) A mixture of the A and B diastereoisomers of -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidin-3-yl) -ester-5-methyl ester is flash chromatographed on silica gel using ethyl acetate and ethyl acetate as eluent. and recovering: the eluted acetate of diastereoisomer A, b) fractional crystallization of 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid 3- (1-benzylpyrrolidide). A mixture of the acid addition salt of the diastereoisomers A and B of the 3-yl) -ester-5-methyl ester such that a salt corresponding to the A-diastereoisomer is obtained, c) the A-diastereoisomer is formed according to process options 11 aa) or b) and the resulting A-diastereoisomer is then reacted with L - (-) - maleic acid to give the A - diastereoisomer dextrorotatory optical isomer L - (-) - d), after which L - (-) - maleate is treated with a base, d) 2,6-dimethyl-4- (3-nitrophenyl) -1,4-dihydropyridine-3,5-dicarboxylic acid-3- ( 1-Benzyl-pyrrolidin-3-yl) -ester-5-methyl ester ^ NO, pr ^ H (T) Il ^ C (X) ^ XI Tl ^ H ^ 30 8351 6 wherein the bond represented by the wavy line is α- or β- bond and the bond by the thick arrow fl is a bond, the a diastereoisomer of the right to turn the optical isomer of diastereoisomer B, and the mixture was left to turn the optical isomer of formula I, and (a) a mixture of a carrier pyiväskromatografoidaan using silica gel and eluting with a mixture of ethyl acetate and acetic acid a to the right of the diastereoisomer or (b) reacting said mixture with L - (-) - maleic acid to separate the L-(-) - maleate of the levorotatory optical isomer of the A-diastereoisomer, e) reacting m-nitrobenzate of formula (II) aldehyde n ° 2 CT CHO of formula (III) is (S) -3-acetoacetoxy-1-benzyl-pyrrole-pyrimidine CH3COCH2COCU_ Π (m), wherein NC No binding by the thick arrow is the α-bond, and a compound of formula (IV) 3-aminocrotonic acid methyl ester 3i 83516 CH3C = CHCl2CH3 · ~: I (IV), .3 NH2 f) reacting m-nitrobenzaldehyde of formula (II) above, acetoacetic acid methyl ester of formula (V) - CH3COCH2COOCH3 (V) , and (S) -3- (3-aminocrotonoyloxy) -1-benzylpyrrolidine of formula (VI) ch3c = chcoov NH2 j (vi), -'IN P- g) is reacted with (5) -1- benzyl-3- [2-m-nitrobenzylidene) acetoacetoxy / pyrrolidine of formula (VII) ςρ CH CH 3 COCCOOV_ (VII), = H2 - (3 32 8 351 6 obtained by reacting m-nitrobenzaldehyde of formula (II) above with (S) -3-acetoacetoxy-1-benzylpyrrolidine of formula (III) above and 3- (S) -3-acetoacetoxy-1-benzylpyrrolidine of formula (VI) metyy1iesteri-aminocrotonate; or h) reacting 2- (m-nitrobenzylidene) acetoacetic acid methyl ester of formula (VIII) NO 2 (VIII), CH 2 CH 3 COCCOOCH 3 previously obtained by reacting m-nitrobenzaldehyde of formula (II) above and an acetoacetic acid methyl ester of formula (V) above) and (S) -3- (3-aminocrotonyloxy) -1-benzylpyrrolidine of formula (VI) above, optionally treating the salt with a base and optionally treating the A- diastereoisomer with a pharmaceutically acceptable acid. Process according to Claim Ib, characterized in that the acid addition salt to be crystallized is malonate, p-nitrobenzoate or maleate. 33 8 3 51 6