JPH0145448B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention formula

[expression] or (In the formula, R ¹ represents a hydrogen atom or a methyl group,
R ² represents a hydrogen atom or a methyl group, and R ³ represents a hydroxyl group or a methoxy group. ), a pharmacologically acceptable salt thereof, a lower alkyl ester thereof, or a lactone thereof as an active ingredient. In recent years, there has been renewed strong recognition that coronary artery spasm is involved in the pathogenesis of ischemic heart disease, and in recent years, the above-mentioned spasm has been associated with not only atypical angina among ischemic heart diseases. It is also observed in cases of unstable angina, acute myocardial infarction, etc., and it has been recognized that it accounts for a large proportion of the causes of ischemic heart disease [Circulation 52 33-37 (1975), Southern
Medical Journal 71 729-732 (1978),
Circulation 56 366-374 (1977)]. The present inventors have accomplished the present invention by discovering that administration of a compound selected from the group represented by formulas () and () above significantly suppresses coronary artery spasm. For example, ergonovine, a type of alkaloid, has the effect of causing coronary artery spasm. In particular, significant spasm is observed in patients with heart disease such as the above-mentioned variant angina, and ischemic ST deviation is observed on electrocardiograms. However, by administering a compound selected from the group of compounds having formulas () and () above, such ST deviation on the electrocardiogram does not appear, and the spasm is suppressed. is shown. The compound having the above formula () or () used in the present invention is MB represented by the following formula.
It is derived from -530B or ML-236B as a starting material, and examples of carboxylic acid derivatives include its pharmacologically acceptable salts, its lower alkyl esters, and its lactones. The pharmacologically acceptable salt of the carboxylic acid having the formula () or () is, for example, an alkali metal salt such as sodium or potassium. Lower alkyl esters of carboxylic acids having the formula () or () are, for example, compounds such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl and the like. The lactone form of the compound having the formula () or () is a compound in which the substituent at the 8-position in the hexahydronaphthalene ring has the following partial structural formula. In the above formula (), the compound in which R ¹ is a hydrogen atom is 3-hydroxy-ML-236B,
Compounds whose 3-position substituent is OH are M-4 class,
...OH compounds are abbreviated as M-4's. This compound is disclosed in Japanese Patent Application No. 55-124385 (Japanese Patent Application No. 57-50894).
and Japanese Patent Application No. 105967 (1983)
10572) along with its manufacturing method. In the above formula (), compounds in which R ¹ is a methyl group are hereinafter abbreviated as 3-hydroxy-MB-530Bs. This compound is described in Japanese Patent Application No. 56-134558 (Japanese Unexamined Patent Publication No. 58-35144) together with its manufacturing method. In the formula (), R ² is a hydrogen atom,
Compounds in which R ³ is a hydroxyl group are hereinafter abbreviated as 6-hydroxy-iso ML-236B. This compound is described in Japanese Patent Application No. 55-115483 (Japanese Unexamined Patent Publication No. 57-108039) and Japanese Patent Application No. 55-124385 (Unexamined Japanese Patent Application No. 57-50894).
No.), along with its manufacturing method. In the formula (), R ² is a hydrogen atom,
Compounds in which R ³ is a methoxy group are hereinafter abbreviated as 6-methoxy-iso ML-236Bs. This compound is described in Japanese Patent Application No. 56-114038 (Japanese Unexamined Patent Publication No. 58-15968).
No.), along with its manufacturing method. In the formula (), R ² is a methyl group,
Compounds in which R ³ is a hydroxyl group are hereinafter abbreviated as 6-hydroxy-isoMB-530B. This compound is described in Japanese Patent Application No. 56-134558 (Japanese Unexamined Patent Publication No. 58-35144).
No.), along with its manufacturing method. The compound having the above formula () or () is
It can be obtained by enzymatic hydroxylation using microorganisms using the above-mentioned MB-530B or ML-236B or derivatives thereof as a raw material compound according to the method described in the above-mentioned patent application specification. Examples of producing these compounds will be described below. Production Example 1 M-4' type (1) Twenty 500 ml Erlenmeyer flasks containing 100 ml of a medium with the following composition were inoculated with Synthephalastrum nigricans SANK42372 (Feikoken Bacteria No. 6043), and incubated at 26°C. After 3 days of shaking culture at 220 rpm, the final concentration of ML-236B lactone was 0.05%.
26℃ for another 6 days.
Cultured at 220 rpm. Medium composition Glucose 1.0% Peptone 0.2 Meat extract 0.1 Yeast extract 0.1 Corn steep liquor 0.3 Tap water Remaining (PH uncorrected) After the completion of the culture, the conversion reaction solution was filtered and the pH of the solution was adjusted to 3 with trifluoroacetic acid. Then 1
After extraction three times with ethyl acetate, a fraction containing M-4' carboxylic acid was obtained. Physical properties of M-4′ carboxylic acid 1 TLC TLC plate: Merck Silica Gel Art5715 Solvent: Benzene: Acetone: Acetic acid = 50:50:3 Rf value 0.46 (2) Wash the extract of (1) with saturated saline. After dehydration with sodium sulfate, a catalytic amount of trifluoroacetic acid was added for lactonization. Next, the above extract was washed with a 5% aqueous sodium bicarbonate solution, dehydrated with sodium sulfate, and then dried under reduced pressure. Next, the obtained residue was eluted with benzene:acetone = 7:3 using a Rover column (manufactured by Merck & Co., Si60 size A) to collect the M-4' lactone body, and further eluted with ethyl acetate. After recrystallization, about 180 mg of M-4' lactone was obtained. Physical properties of M-4' lactone 1 NMR spectrum Measured at 100MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 4.25 (1H, multiplet) 4.60 (1H, multiplet) 5.50 (1H, multiplet) 5.75 (1H, multiplet) 5.90 (1H, quartet) 6.01 (1H, multiplet) line) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 230, 237, 245 3 Infrared absorption spectrum (KBr method) cm ^-1 :
3500, 1720 4 Mass spectrum m/e: 406 (M ⁺ ), 304, 286 5 Optical rotation [α] _D : 310.9° (C = 0.66, methanol) 6 Melting point 141-143℃ 7 Elemental analysis value (%) As C ₂₃ H ₃₄ O ₆ Theoretical value C, 67.95; H, 8.43 Experimental value C, 68.05; H, 8.37 8 TLC TLC plate: Merck silica gel Art5715 Solvent: Benzene: Acetone = 1:1 Rf value 0.64 (3) Wash the extract of (1) with saturated saline, then
A fraction containing M-4' carboxylic acid sodium salt was obtained by dissolving the mixture into the aqueous layer using a % sodium bicarbonate aqueous solution. This aqueous layer was adjusted to pH 8.0 with 0.1N hydrochloric acid, and then the fraction containing M-4' carboxylic acid sodium salt was added to Diaion HP
-20 column (manufactured by Mitsubishi Chemical Industries, Ltd.). M-4' carboxylic acid sodium salt was eluted with 50% acetone, the acetone was distilled off, and then freeze-dried to yield M-4' carboxylic acid sodium salt 141
mg was obtained. Physical properties of M-4' carboxylic acid sodium salt 1 NMR spectrum Measured at 60MHz in deuterated methanol using TMS as an internal standard. (CD ₃ OD, δ: _ppn ) 5.50 (1H, broad singlet) 5.70 (1H, broad singlet) 5.95 (1H, quartet) 6.00 (1H, doublet) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 230, 238, 246 3 Infrared absorption spectrum (KBr method) cm ^-1 :
3400, 2900, 1580 4 Elemental analysis value (%) C ₂₃ H ₃₅ O ₇ As Na Theoretical value C, 61.88; H, 7.85 Experimental value C, 61.85; H7, 95 (4) Add the extract of (1) to saturated salt The mixture was washed with water, and then an ether solution of diazomethane was added thereto, left to stand for 30 minutes, and then dried under reduced pressure. Next, the obtained residue was purified using a Lorber column (manufactured by Merck & Co., Si60 size A) in a benzene:acetone = 1:1 system, and 150 mg of the purified product of M-4' carboxylic acid methyl ester was obtained as a colorless product. Obtained as an oil. Physical properties of M-4' carboxylic acid methyl ester 1 NMR spectrum Measured at 60MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 3.70 (3H, singlet) 5.50 (1H, broad singlet) 5.75 (1H, broad singlet) 5.90 (1H, quartet) 6.01 (1H, doublet) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 230, 238, 246 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3400, 1730 4 Mass spectrum After silylation with N,O-bis(trimethylsilyl)trifluoroacetamide, measurement was performed using JEOL D-300 model. m/e: 654 (M ⁺ ) 5 Elemental analysis value (%) C ₂₄ H ₃₈ O ₇ Theoretical value C, 65.73; H, 8.73 Experimental value C: 65.66; H, 8.79 Production example 2 M-4 class (1 ) Absidia coerulea in 20 500 ml Sakaguchi flasks containing 100 ml of medium with the following composition.
Inoculate IFO4423, 26℃, 120s.pm (strokes)
per minute), and after 2 days ML-
236B carboxylic acid sodium salt at final concentration
Added to 0.05% and kept at 26℃ for 5 days.
Cultured at 120s.pm. Medium composition Glucose 2.0% K ₂ HPO ₄ 0.15 MgSO・7H ₂ O 0.15 NH ₄ NO ₃ 0.1 Peptone 0.1 C, S, L 0.2 Yeast extract 0.1 ZnSO ₄・7H ₂ O 0.001 Tap water Remaining (adjusted to PH7.0) After the cultivation was completed, the conversion reaction solution was filtered, and the pH of the solution was adjusted to 3 with trifluoroacetic acid. Then 1
After extraction three times with ethyl acetate, a fraction containing M-4 carboxylic acid was obtained. Physical properties of M-4 carboxylic acid 1 TLC TLC plate: Merck Silica Gel Art5715 Solvent: Benzene: Acetone: Acetic acid = 50:50:3 Rf value 0.45 (2) The extract of (1) was washed with saturated saline. After dehydration with sodium sulfate, a catalytic amount of trifluoroacetic acid was added for lactonization. Next, after washing with a 5% aqueous sodium bicarbonate solution, it was dehydrated with sodium sulfate and concentrated to dryness to obtain a lactone fraction. This was applied to a Rover column (manufactured by Merck & Co., Si60 size A) and eluted with ethyl acetate to separate and collect the M-4 lactone fraction. Furthermore, this was connected to a Rover column (manufactured by Merck, RP-8 size A).
The product was purified using 35% acetonitrile and eluted with 35% acetonitrile. Physical properties of M-4 lactone 1 NMR spectrum Measured at 100MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 4.38 (1H, multiplet) 4.41 (1H, 〃) 4.62 (1H, 〃) 5.41 (1H, 〃) 5.58 (1H, 〃) 5.90 (1H, quartet) 6.01 (1H , double line) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 230, 236.7, 244.6 3 Infrared absorption spectrum (thin film method) cm ^-1 :
Wash the extract of 3400, 2950, 1725 (3) (1) with saturated saline solution, add ethyl solution of diazomethane, and leave for 30 minutes.
It was dried under reduced pressure. The residue was applied to a Rover column (manufactured by Merck & Co., Si60 size A), eluted with a benzene:ethyl acetate ratio of 1:1, and a fraction containing M-4 methyl ester was collected. Add this to the Rover column (Merck, RP-8 size A)
After purification using 35% acetonitrile as elution, 20 mg of purified M-4 carboxylic acid methyl ester was obtained as a colorless oil. Physical properties of M-4 carboxylic acid methyl ester 1 NMR spectrum Measured at 200MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 0.88 (3H, t, J = 7.3Hz) 0.89 (3H, d, J = 6.5Hz) 1.12 (3H, d, J = 6.8Hz) 1.1~1.7 (10H, m) 2.34 (1H, sex, J=7Hz) 2.3~2.5 (2H, m) 2.49 (2H, d, J=6.4Hz) 2.58 (1H, m) 3.72 (3H, s) 3.78 (1H, m) 4.25 (1H, quin, J=7Hz) 4.4 (1H, m) 5.42 (1H, m) 5.56 (1H, m) 5.90 (1H, d, d, J=9.8, 5.6Hz) 5.99 (1H, d, J=9.8Hz) 2 Mass spectrum After silylation with N,O-bis(trimethylsilyl)trifluoroacetamide, measurement was performed using JEOL D-300 model. m/e: 654 (M ⁺ ), 552, 462, 372, 290,
272, 233, 231 3 Ultraviolet absorption spectrum (ethanol solution)
λ _nax (nm): 230.1, 237.3, 246.4 4 Infrared absorption spectrum (thin film method) cm ^-1 :
3400, 2950, 1730 5 TLC TLC plate; Silica gel Art5715 manufactured by Merck & Co. Solvent: Benzene: Acetone = 1:1 Rf value 0.83 (4) Dissolve 100 mg of the M-4 lactone obtained in (2) in a small amount of acetone. , an equivalent amount of aqueous sodium hydroxide solution was added, and the mixture was left at room temperature for 1 hour. Then, after distilling off the acetone, it was freeze-dried to obtain M-4.
Approximately 105 mg of carboxylic acid sodium salt was obtained. Physical properties of M-4 carboxylic acid sodium salt 1 NMR spectrum Measured at 200MHz in heavy methanol using TMS as an internal standard. (CD ₃ OD, δ: _ppn ) 0.91 (3H, t, J = 7.5Hz) 0.92 (3H, d, J = 7Hz) 1.12 (3H, d, J = 7Hz) 1.1~1.8 (10H, m) 2.25 ( 1H, d, d, J = 15, 7.6Hz) 2.34 (1H, d, d, J = 15, 5.5Hz) 2.2 - 2.4 (3H, m) 2.48 (1H, m) 3.68 (1H, m) 4.07 ( 1H, m) 4.28 (1H, m) 5.36 (1H, m) 5.48 (1H, d, d, J = 3, 2Hz) 5.88 (1H, d, d, J = 9.6, 5.3Hz) 5.98 (1H, d , J=9.8Hz) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 230.0, 237.2, 245.0 3 Infrared absorption spectrum (KBr method) cm ^-1 :
3400, 2900, 1725, 1580 4 TLC TLC plate; Silica gel Art5715 manufactured by Merck & Co. Solvent: Benzene; Acetone: Acetic acid = 50:50:3 Rf value 0.45 Production example 3 6-Hydroxy-iso ML-236B class (1) Production example By culturing in 2-(1), the extract contains M-
6-Hydroxy-iso ML with 4 carboxylic acids
-236B A classification containing carboxylic acid is obtained, and 6-
Hydroxy-iso ML-236B carboxylic acid Rf
The value was the same as M-4 carboxylic acid. (2) In the treatment of (2) of Production Example 2, after lactonization with trifluoroacetic acid, it was applied to a Lorber column (manufactured by Merck & Co., Si60 size A), and when eluted with ethyl acetate, 6-hydroxy-isoML- 236B
The lactone body fraction was separated and collected. This was purified in the same manner to obtain 82 mg of a purified 6-hydroxy-iso ML-236B lactone. Physical properties of 6-hydroxy-iso ML-236B lactone 1 NMR spectrum Figure 1 shows the NMR spectrum measured at 100MHz in deuterated chloroform using TMS as an internal standard. 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 229, 234.8, 244.5 3 Infrared absorption spectrum (thin film method) cm ^-1 : 2nd
As shown in the figure. 4 TLC TLC plate; Silica gel Art5715 manufactured by Merck & Co., Ltd. Solvent; Benzene: Acetone: Acetic acid = 50:50:3 Rf value 0.62 (3) In the process of (3) of Production Example 2, after alkylation with diazomethane, Rover column ( 6-Hydroxy-iso ML-236B carboxylic acid methyl ester was separated and collected during elution with a benzene:ethyl acetate system. This was purified in the same manner to obtain 78 mg of a purified sample of 6-hydroxy-iso ML-236B carboxylic acid methyl ester. Physical properties of 6-hydroxy-iso ML-236B carbon methyl ester 1 NMR spectrum Figure 3 shows the NMR spectrum measured at 100 MHz in deuterated chloroform using TMS as an internal standard. 2 Mass spectrum After silylation with N,O-bis(trimethylsilyl)trifluoroacetamide, measurement was performed using JEOL D-300 model. m/e: 654 (M ⁺ ), 552, 462, 372, 272,
233, 231 3 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 229, 234.8, 244.5 4 Infrared absorption spectrum (thin film method) cm ^-1 : 4th
As shown in the figure. 5 TLC TLC plate: Silica gel Art5715 manufactured by Merck & Co. Solvent: Benzene: Acetone = 1:1 Rf value 0.88 (4) 6-Hydroxy-isoML obtained in (2)
100 mg of 236B lactone was dissolved in a small amount of acetone, an equivalent amount of aqueous sodium hydroxide solution was added, and the mixture was left at room temperature for 1 hour. After distilling off the acetone, lyophilization yields 6-hydroxy-iso
Approximately 103 mg of ML-236B carboxylic acid sodium salt was obtained. Physical properties of 6-hydroxy-iso ML-236B carboxylic acid sodium salt 1 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 229 (ah), 235, 245 (sh) 2 Infrared absorption spectrum (KBr method) cm ^-1 :
3400, 2850, 1710, 1580 3 TLC TLC plate; Silica gel Art5715 manufactured by Merck & Co. Solvent; Benzene: Acetone: Acetic acid = 50:50:3 Rf value 0.45 Production example 4 6-methoxy-iso ML-236B class (1) The following composition Abcisia coerulea in 20 500 ml Erlenmeyer flasks containing 100 ml of medium.
IFO4423 was inoculated and cultured with shaking at 26°C and 220 rpm. After 4 days, ML-236B lactone was added to a final concentration of 0.05%, and ML-236B lactone was added for another 6 days.
Cultured at 220 rpm. Medium composition Glucose 2.0% K ₂ HPO ₄ 0.15 MgSO ₄・7H ₂ O 0.15 NH ₄ NO ₃ 0.1 Peptone 0.1 CSL 0.2 Yeast extract 0.1 Z _o SO ₄・7H ₂ O 0.001 Tap water Remainder (adjusted to PH7.0) Culture After completion, the conversion reaction solution was filtered, and the pH of the solution was adjusted to 3 with trifluoroacetic acid. Then 1
After extraction three times with ethyl acetate, a fraction containing 6-methoxy-iso ML-236B carboxylic acid was obtained. The above extract was washed with saturated brine, dehydrated with anhydrous sodium sulfate, and then lactonized by adding a catalytic amount of trifluoroacetic acid. Next, the above extract was washed with a 5% aqueous sodium bicarbonate solution, dehydrated with anhydrous sodium sulfate, and then dried under reduced pressure. The residue was eluted with a benzene-acetone (7:3) system using a Rover column (manufactured by Merck & Co., Si60 size A), and 6-methoxy-iso
ML-236B lactone body was collected. When this was purified, 23 mg of the target product was obtained. Physical properties of 6-methoxy-iso ML-236B lactone 1 NMR spectrum (CDCl ₃ , δ: _ppn ) Measured in deuterated chloroform at 90 MHz using TMS as an internal standard. 6.15 (1H, doublet) 5.70 (1H, quartet) 5.70 (1H, multiplet) 5.40 (1H, 〃 ) 4.60 (1H, 〃 ) 4.40 (1H, 〃 ) 3.45 (1H, doublet) 3.30 (3H, singlet) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 235 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3450, 1730 4 Mass spectrum m/e: 420 (M ⁺ ), 420, 318, 300, 286,
268 5 TLC TLC plate; Silica gel Art5715 manufactured by Merck & Co. Solvent: Benzene: Acetone = 1:1 Rf value 0.7 6 Elemental analysis value (%) C ₂₄ H ₃₆ O ₆ Theoretical value C, 68.54; H, 8.63 Experimental value C, 68.53; H, 8.81 (2) 6-methoxy-isoML-236B lactone body 1
Dissolve g in a small amount of acetone, then add 13 ml of 0.2N aqueous sodium hydroxide solution and stir at 40℃ for 1 hour.
Hydrolyzed for hours. After the reaction was completed, acetone was distilled off from the reaction mixture, and then 5 ml of chloroform was added.
Washed with. Adjust the aqueous layer to PH8.0 with 0.1N hydrochloric acid,
Diaion HP-20 column (Mitsubishi Chemical Industries, Ltd.)
product). Elute the fraction containing 6-methoxy-iso ML-236B carboxylic acid sodium salt with 50% acetone, distill off the acetone, and freeze-dry to obtain 6-methoxy-iso ML.
1.003 g of -236B carboxylic acid sodium salt was obtained. Physical properties of 6-methoxy-iso ML-236B carboxylic acid sodium salt 1 NMR spectrum Measured at 60 MHz in deuterated methanol using TMS as an internal standard. (CD ₃ OD, δ: _ppn ) 3.31 (3H, singlet) 5.42 (1H, multiplet) 5.70 (1H, multiplet) 5.71 (1H, quartet) 6.12 (1H, doublet) 2 Ultraviolet absorption Spectrum (methanol solution)
λ _nax (nm): 235 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3400, 2950, 1580 4 Elemental analysis value (%) C ₂₄ H ₃₇ O ₇ As Na Theoretical value C, 62.61; H, 8.04 Experimental value C, 62.54; H, 8.11 (3) 6-Methoxy-isoML-236B carvone Dissolve 1 g of acid sodium salt in a small amount of methanol,
Next, trifluoroacetic acid was added under cooling to make the mixture acidic, and then a diazomethane solution was immediately added and left for 30 minutes. After the reaction was completed, the solvent was distilled off from the reaction mixture. The obtained residue was purified using a Rover column (manufactured by Merck & Co., RP-8 size B).
When purified in a methanol:water = 6:4 system, 6-
780 mg of methoxy-iso ML-236B carboxylic acid methyl ester was obtained as a colorless oil. Physical properties of 6-methoxy-iso ML-236B carboxylic acid methyl ester 1 NMR spectrum Measured at 60 MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 3.30 (3H, singlet) 3.60 (3H, singlet) 4.37 (1H, multiplet) 4.62 (1H, multiplet) 5.30 (1H, multiplet) 5.70 (1H, multiplet) 5.71 (1H, quartet) 6.13 (1H, doublet) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 235 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3400, 1725 4 Mass spectrum After silylation with N,O-bis(trimethylsilyl)trifluoroacetamide, measurement was performed using JEOL D-300 model. m/e: 668 (M ⁺ ) 5 Elemental analysis value (%) C ₂₅ H ₄₀ O ₇ Theoretical value C, 66.34; H, 8.91 Experimental value C, 66.38; H, 8.98 Production example 5 3-Hydroxy-MB- Class 530B (1) Inoculate 20 500 ml Erlenmeyer flasks containing 100 ml of the medium with the following composition with Mucor hyimaris IFO5834, and heat at 26℃.
Shaking culture at 220rpm, 4 days later, MB-530B
Lactone bodies were added to the final concentration of 0.05%, and the cells were further cultured at 26°C and 220 rpm for 6 days. Medium composition Glucose 1.0% Peptone 0.2 Meat extract 0.1 Yeast extract 0.1 Corn steep liquor 0.3 Tap water Remaining (PH uncorrected) After culturing, filter the conversion reaction solution and transfer the solution to Diaion HP-20 (Mitsubishi Chemical Industries, Ltd.) product) was adsorbed onto a column using the product. Next, elution was performed with 70% methanol, and the eluate was distilled off. It was then extracted twice with 1 portion of ethyl acetate to obtain a fraction containing 3-hydroxy-MB-530B carboxylic acid. Physical properties of 3-hydroxy-MB-530B carboxylic acid 1 TLC TLC plate: Merck silica gel Art5715 Solvent: Benzene: Acetone: Acetic acid = 50:50:3 Rf value 0.47 (2) Extract obtained in (1) An ethereal solution of diazomethane was added to the mixture, and the mixture was left to stand for 30 minutes. The extract was then washed with saturated brine and dried under reduced pressure. The obtained residue was applied to Microbondapak C ₁₈ (manufactured by Waters) and eluted with 53% methanol.
The fraction containing 3-hydroxy-MB-530B carboxylic acid methyl ester was separated and collected. This was purified to obtain 180 mg of the target compound. Physical properties of 3-hydroxy-MB-530B carboxylic acid methyl ester 1 NMR spectrum Measured at 90MHz in deuterated chloroform using TMS as an internal standard. (CDCl ₃ , δ: _ppn ) 3.72 (3H, singlet) 4.28 (1H, quintet) 5.45 (2H, multiplet) 5.93 (1H, quartet) 6.01 (1H, doublet) 2 Ultraviolet absorption Spectrum (methanol solution)
λ _nax (nm): 229, 236, 244.5 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3410, 2975, 1730 4 Elemental analysis value (%) C ₂₅ H ₄₀ O ₇ Theoretical value C, 66.34; H, 8.91 Experimental value C, 66.30; H, 9.02 (3) 3-Hydroxy-MB-530B methyl carboxylate 100 mg of ester was dissolved in 0.1N aqueous sodium hydroxide solution and stirred at 30°C for 1 hour. This solution was then washed with chloroform and freeze-dried to yield 3-hydroxy-MB-530B.
93 mg of carboxylic acid sodium salt was obtained. Physical properties of 3-hydroxy-MB-530B carboxylic acid sodium salt 1 NMR spectrum 90MHz in heavy water using DSS as internal standard
It was measured with (D ₂ O, δ: _ppn ) 5.40 (2H, multiplet) 5.89 (1H, quartet) 6.00 (1H, doublet) 2 Ultraviolet absorption spectrum (aqueous solution) λ _nax
(nm): 229, 236, 245 3 Infrared absorption spectrum (KBr method) cm ^-1 : 3400
~2900, 1580 4 Elemental analysis value (%) C ₂₄ H ₃₇ O ₇ Na Theoretical value C, 62.59; H, 8.097 Experimental value C, 62.37; H, 8.21 Production example 6 6-Hydroxy-isoMB-530B class ( 1) By culturing in (1) of Production Example 5, the extract contains 3-
Hydroxy-MB-530B with carboxylic acid 6
-Hydroxy-isoMB-530B A segment containing carboxylic acid is obtained, and 6-hydroxy-isoMB
-The Rf value of 530B carboxylic acid is 3-hydroxy-
It was the same as MB-530B carboxylic acid. (2) After culturing in (1) of Production Example 5, the extract was washed with saturated saline, dried over sodium sulfate, and then a catalytic amount of trifluoroacetic acid was added. Next, add the extract to 5
% aqueous sodium hydrogen carbonate solution, dried over sodium sulfate, and then dried under reduced pressure. The resulting residue was eluted with ethyl acetate using a Rover column (manufactured by Merck & Co., Si60 size A), and the 6-hydroxy-isoMB-530B lactone was separated and collected. This was purified to obtain 212 mg of the target compound. Physical properties of 6-hydroxy-isoMB-530B lactone 1 NMR spectrum In deuterated acetone, using TMS as an internal standard,
Measured at 90MHz. (CD ₃ COCD ₃ , δ: _ppn ) 3.94 (1H, doublet) 4.35 (1H, multiplet) 4.66 (1H, multiplet) 5.45 (1H, multiplet) 5.62 (1H, doublet) 5.91 (1H , singlet) 2 Ultraviolet absorption spectrum (methanol solution)
λ _nax (nm): 238.4 3 Infrared absorption spectrum (KBr method) cm ^-1 :
3450, 1730 4 Mass spectrum m/e: 420 (M ⁺ ) 6 Elemental analysis value (%) C ₂₄ H ₃₆ O ₆ Theoretical value C, 68.54; H, 8.63 Experimental value C, 68.33; H, 8.81 (3) Dissolve 100 mg of 6-hydroxy-isoMB-530B lactone in a small amount of acetone, then add an equivalent amount of 0.2N sodium hydroxide aqueous solution,
Stir at room temperature. This solution was then freeze-dried to obtain 106 mg of 6-hydroxy-isoMB-530B carboxylic acid sodium salt. Physical properties of 6-hydroxy-isoMB-530B carboxylic acid sodium salt 1 NMR spectrum 90MHz in heavy water using internal standard DSS
It was measured with (D ₂ O, δ: _ppn ) 3.78 (1H, multiplet) 4.01 (1H, multiplet) 4.13 (1H, multiplet) 5.49 (1H, multiplet) 5.62 (1H, multiplet) 6.02 (1H, wide Singlet) 2 Ultraviolet absorption spectrum (aqueous solution) λ _nax
(nm): 238 3 Infrared absorption spectrum (KBr method) cm ^-1 : 3400
~2900, 1575 4 Elemental analysis value (%) C ₂₄ H ₃₇ O ₇ Na Theoretical value C, 62.59; H, 8.097 Experimental value C, 62.28; H, 8.31 (4) In the treatment of (2) of Production Example 5 After alkylation with diazomethane, it was applied to Microbondapak C ₁₈ and eluted with 53% methanol, and the fraction containing 6-hydroxy-isoMB-530B carboxylic acid methyl ester was separated and collected. This was purified to obtain 170 mg of the target compound. Physical properties of 6-hydroxy-isoMB-530B carboxylic acid methyl ester 1 NMR spectrum In deuterated acetone, using TMS as an internal standard,
Measured at 90MHz. (CD ₃ COCD ₃ , δ: _ppn ) 3.70 (1H, singlet) 4.22 (1H, multiplet) 5.42 (1H, multiplet) 5.57 (1H, multiplet) 5.92 (1H, doublet) 2 Ultraviolet absorption Spectrum (methanol solution)
λ _nax (nm): 238.4 3 Infrared absorption spectrum (thin film method) cm ^-1 :
3400, 2960, 1732 4 Elemental analysis value (%) as C ₂₅ H ₄₀ O ₇ Theoretical value C, 66.34; H, 8.91 Experimental value C, 66.28; H, 8.99 Carvone having the above formula () or () of the present invention Acids, their pharmacologically acceptable salts, their lower alkyl esters, or their lactones are active ingredients in therapeutic agents for ischemic heart disease. The administration form includes oral administration using tablets, granules, syrup, etc., subcutaneous injection, intravenous injection,
Examples include parenteral administration using suppositories. These various preparations are made according to the conventional method, and depending on the purpose, solubilizing agents, suspending agents, excipients, binders, disintegrants, lubricants, and flavoring agents are added to the main drug, which are commonly used in the pharmaceutical field. It can be formulated using known adjuvants. The amount used varies depending on the symptoms, age, body weight, etc., route of use, and number of times used, but it is usually 0.05 to 25 mg/day for M-4 class compounds for adults.
Kg/day, preferably 0.2 to 20/mg/Kg/day,
M-4′ compounds and 6-hydroxy-iso ML
-236B class compounds are 0.1 to 50 mg/Kg/day, preferably 0.5 to 25 mg/Kg/day, and 3-hydroxy-
0.05 to 25 mg/Kg/day for MB-530B class compounds and 6-hydroxy-isoMB-530B class compounds;
It is preferably 0.2 to 20 mg/Kg/day, and 0.05 to 50 mg/day for 6-methoxy-iso ML-536B class compounds.
Kg/day, preferably 0.5 to 30 mg/Kg/day. Larger doses can be administered as needed. Acute toxicity For example, the acute toxicity of M-4 carboxylic acid sodium salt to mice is LD ₅₀ (mg/Kg) of 8000 mg/Kg or more for oral administration and 2000 mg/Kg for intravenous injection.
It was over Kg. Also, LD ₅₀ against rats
(mg/Kg) is 12000mg/Kg or more by oral administration,
The dose was 400 mg/Kg or more when administered intravenously. Next, an example in which the drug was administered to a dog will be shown. Example Two groups of beagle dogs (2 males and 2 females per group) were prepared, one group was untreated, and the other group was given 20 mg/Kg/day of M-4 carboxylic acid sodium salt twice a day. The drug was orally administered for 4 consecutive weeks. These beagle dogs were anesthetized, their chests were opened under artificial ventilation, and their coronary arteries were stenosed until no ischemic ST deviation was observed on electrocardiograms (critical stenosis). Electrocardiograms were recorded from the body surface using lead AB, and the above operations were performed under their observation. In this stable state, ergonovine 100 μg/Kg is administered intravenously. Significant ST deviation was observed in the untreated group (see Figure 5).
On the other hand, in the group to which M-4 carboxylic acid sodium salt was administered, no change occurred in the ST region (see Figure 6). In the above experiment, instead of M-4 carboxylic acid sodium salt, M-4' carboxylic acid sodium salt,
In a test using 6-hydroxy-isoML-236B carboxylic acid sodium salt, 6-methoxy-isoML-236B lactone, 3-hydroxy-MB-530B sodium salt, 6-hydroxy-isoMB-530B sodium salt, A similar inhibitory effect on coronary artery spasm was also observed. Formulation Example 1 (Capsules) M-4 carboxylic acid sodium salt 10.0 mg Lactose 151.2 Corn starch 37.8 Magnesium stearate 1.0 200 mg The powder of the above formulation was mixed and passed through a 60 mesh sieve, and 200 mg of this powder was passed through a No. 3 sieve. It was put into a gelatin capsule and made into a capsule. Formulation Example 2 (Tablet) M-4 carboxylic acid sodium salt 5.0mg Lactose 77.4 Corn starch 13.0 Magnesium stearate 0.6 L-HPC (Shin-Etsu Chemical Co., Ltd. product) 24.0 120mg The above formulation was prepared by normal formulation operations. 1 tablet
It was made into a 120 mg tablet.

[Brief explanation of drawings]

Figure 1 shows the nuclear magnetic resonance spectrum of 6-hydroxy-iso ML-236B lactone, and Figure 2 shows the infrared absorption spectrum of the same substance. Figure 3 shows the nuclear magnetic resonance spectrum of 6-hydroxy-iso ML-236B carboxylic acid methyl ester;
The figure shows the infrared absorption spectrum of the same substance. Fifth
The figure shows the electrocardiogram of the untreated group, and FIG. 6 shows the electrocardiogram of the drug-administered group. In FIGS. 5 and 6, A is a diagram before administration of ergonovine, and B is a diagram after administration.

Claims (1)

[Claims] 1 Formula [Formula] or [Formula] (wherein R ¹ represents a hydrogen atom or a methyl group,
R ² represents a hydrogen atom or a methyl group, and R ³ represents a hydroxyl group or a methoxy group. ), a pharmacologically acceptable salt thereof, a lower alkyl ester thereof, or a lactone thereof as an active ingredient.