JP3503904B2 - Method for producing E-type prostaglandins - Google Patents

Description

[Detailed description of the invention]

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for producing E-type prostaglandins by hydrolyzing lower alkyl esters of E-type prostaglandins with enzymes.

[0002]

2. Description of the Prior Art E-type prostaglandins are generally produced by obtaining prostanoic acid from prostanoic acid ester. As a result, it is chemically unstable and prone to dehydration reactions, so that ordinary chemical methods are accompanied by not a little dehydration by-products. Therefore, as the most effective method, a method of hydrolysis using an enzyme is known [JP-A-52-21392.
No. 2004-000001 (Lipase derived from porcine pancreas), N.O. A. Ports
er et al. (Journal of American Chemical Society, 101:4319-4322, 1979) (lipase from porcine pancreas), C-
H. Lin et al. (Journal of the American Chemical Society, Vol. 104, pp. 1621-1628).
Page, 1982) (Lipase derived from porcine pancreas), Atsuo Hazato et al. (Journal of the Chemical Society of Japan, Vol.
2, 1983) (esterase from porcine liver)].

[0003]

However, the enzymatic hydrolysis method used in the above method has the disadvantage that it takes a long time to react depending on the type of E-type prostaglandins. . In particular, a compound in which a double bond is conjugated to a carboxylic acid ester exhibits a very slow hydrolysis reaction. On the other hand, E-type prostaglandins having triple bonds at positions 13 and 14 have the drawback of producing isomers such as 8-β isomers. This 8-β by-product is
It tends to increase as the reaction time becomes longer, but it does not necessarily decrease as the reaction time becomes shorter, and it varies depending on the type of enzyme. Therefore, it was necessary to find an enzyme that does not produce 8-β by-products.

[0004]

Means for Solving the Problems As a result of intensive studies aimed at solving the above-mentioned problems, the present inventors have found that lower alkyl esters of E-type prostaglandins can be obtained by hydrolyzing the esters. The present inventors have found that the use of an enzyme enables the production of desired E-type prostaglandins in a short period of time with a high yield and with extremely low production of isomers, thus completing the present invention.

[0005] That is, the present invention provides the formula

[0006]

[wherein R represents a lower alkyl group, R ¹
and ^R2 are the same or different and represent a hydrogen atom or a hydroxyl-protecting group, A ^, R3 and ^R4 represent any group that does not participate in the hydrolysis reaction, and B represents an ethynylene group. ] to lower alkyl esters of E-type prostaglandins represented by the formula

[0008]

(wherein R ¹ , R ² , A, R ³ , R ⁴ and B
has the same meaning as above. ) is a method for producing E-type prostaglandins represented by

In the present invention, a lower alkyl group means a methyl group, an ethyl group, an n-propyl group, an isopropyl group,
A linear or branched alkyl group such as n-butyl group and isobutyl group. In addition, the hydroxyl-protecting group is one commonly used in the field of prostaglandins,
Examples include t-butyldimethylsilyl group, triethylsilyl group, phenyldimethylsilyl group, t-butyldiphenylsilyl group, tetrahydropyranyl group, tetrahydrofuranyl group, methoxymethyl group, ethoxyethyl group and benzyl group.

Any of A, R ³ and R ⁴ may be used as long as they do not participate in this reaction. If these examples are given, A is, for example, a linear alkylene group having 3 to 8 carbon atoms (e.g., trimethylene group, tetramethylene group, hexamethylene group, octamethylene group, etc.), carbon atom A linear alkenylene group having 3 to 9 numbers [for example, the formula

[0012]

(Wherein, n1 represents an integer of ₁ to ₆ , n2 represents an integer of ₂ to 5, n3 represents an integer of 1 to 6, etc.)], chain A linear alkylene group having 2 to 8 carbon atoms having one oxygen atom or sulfur atom interposed therein [for example, the formula

[0014]

(Wherein, X represents an oxygen atom or a sulfur atom, n4 represents an integer of ₁ to 6, and n5 represents an integer of 1 to _5. ) and the like], the formula

[0016]

( _In the formula, n6 and _n7 are the same or different and represent an integer of 1 to 3.) The group represented by the formula

[0018]

(wherein n ₈ and n ₉ are the same or different and represent an integer of 1 to 3), a group having 3 carbon atoms
~8 linear alkynylene groups [e.g.

[0020]

(In the formula, n ₁₀ represents an integer of 1 to 6.) and the like].

Examples of R ³ include hydrogen atom, methyl group, ethyl group and vinyl group. ^R4 is, for example, a linear or branched alkyl group, alkenyl group or alkynyl group having 1 to 10 carbon atoms (e.g. pentyl group, hexyl group, 1-methylhexyl group, 2-methylhexyl group , 1,1-dimethylhexyl group, 2-pentenyl group, 5-hexenyl group,
1-methyl-3-hexynyl group, 2-methyl-3-hexynyl group, 2-pentynyl group, etc.), C 3-1
0 cycloalkyl groups (e.g., cyclopentyl, cyclohexyl, cycloheptyl, etc.), phenyl, "phenyl, phenoxy, or 5 to
1 to 6 carbon atoms substituted with 6 cycloalkyl groups
four alkyl groups (e.g., cyclopentylmethyl, cyclohexylmethyl, 2-cyclopentylethyl,
3-cyclopentylpropyl group, 3-cyclohexylpropyl group, 4-cyclopentylbutyl group, benzyl group,
phenoxymethyl group, 2-phenylethyl group, 4-phenylbutyl group, etc.).

The enzyme used in the present invention is an ester-hydrolytic enzyme produced by microorganisms belonging to the genus Candida. Specific examples of commercially available products include:
Lipase VII (manufactured by Sigma) [lipase derived from Candida cylindracea], Lipase AY (manufactured by Amano Pharmaceutical) [Candid
a sp.)-derived lipase], Lipase VII-S [purified lipase VII derived from Candida cylindracea], and the like.

The hydrolysis method of the present invention will now be described in detail. In the present invention, an ester-hydrolyzing enzyme produced by a microorganism belonging to the genus Candida and the compound of formula (I) are stirred or shaken in an aqueous solution. At this time, it is preferable to use a buffer solution such as a phosphate buffer solution or a Tris-HCl buffer solution in order to keep the pH of the reaction solution constant. In addition, this reaction can also be carried out without using a buffer solution. In this case, using an aqueous solution such as sodium hydroxide or potassium hydroxide, pH
H should be controlled. In addition, an organic solvent such as acetone can be added to the reaction solution as a substrate dissolution aid. In the case of acetone, 1% to 20% to the reaction solution
can be added up to, preferably 3% to 8%. The pH of the reaction solution may be set according to the enzyme used. For example, various conditions when using Lipase VII are pH 6.0 to 8.0, most preferably pH 7.0 to 7.5. This pH is most favorable for hydrolyzing chemically unstable E-type prostaglandins. The reaction temperature is 25°C to 50°C, preferably 30°C to 35°C. The amount of enzyme to be used may be appropriately determined according to the titer of the enzyme and the amount of substrate, but usually 0.1 of the amount of substrate
times to 20 times. The reaction time was determined by TLC analysis or H
It may be set while checking the progress of the reaction by PLC analysis or the like. As a method for extracting the desired product after the completion of the reaction,
When the reaction solution is acidic or neutral, the reaction solution can be directly extracted with an organic solvent such as ethyl acetate. When the reaction solution is alkaline, dilute hydrochloric acid, dilute phosphoric acid, or the like is added to make it acidic, and the solution is extracted with an organic solvent such as ethyl acetate. After distilling off the solvent, the product can be purified by column chromatography or the like as necessary to isolate the pure target product.

By the way, among the compounds of the formula (I), which are starting materials for the production method of the present invention, novel ones can be produced, for example, according to the production examples given below.

[0026]

INDUSTRIAL APPLICABILITY According to the present invention, it has become possible to produce E-type prostaglandins in a short period of time with high yield. especially,
It has become possible to hydrolyze E-type prostaglandin esters in which a double bond is conjugated to a carboxylic acid ester in an extremely short time. In addition, when the compound of formula (II) is produced from the compound of formula (I) in which B is an ethynylene group, it has become possible to suppress the formation of isomers such as the 8-β isomer.

[0027]

EXAMPLES The present invention will be described in more detail below by way of examples, but the present invention is not limited only by these examples. The progress of hydrolysis was confirmed by the TLC method and HPLC method shown below. TLC-1: TLC plate (manufactured by Merck, Art 57
15) Developing solvent; hexane/ethyl acetate = 1/4, or hexane/benzene/tetrahydrofuran/formic acid = 5/20/
7.5/5 TLC-2: TLC plate RP-18 (manufactured by Merck,
Art 137124) Developing solvent; methanol/water = 8/2 HPLC: Column; ODS 80TM (4.6φ x
150 mm) (manufactured by Tosoh Corporation) Eluent: methanol/water = 7/3 Flow rate: 1 ml/min Temperature: 50°C Detection: UV 210 nm The amount of 8-β produced was determined by the HPLC method shown below. It was measured. HPLC: Column; ODS 80TM (4.6φ×
150 mm) (manufactured by Tosoh Corporation) Eluent: 6.6 mM phosphate buffer (pH 6.3)/methanol = 45/55 Flow rate: 1 ml/min Temperature: 50°C Detection: UV 210 nm

Examples 1 to 3 The enzyme shown in Table 1 [enzyme of Example 3 (pig liver esterase (manufactured by Sigma)) was used for comparison. ]
50 mg was added to 2.25 ml of phosphate buffer (10 mM, p
H7.0) and dissolved in 250 μl of 50% (V/V) acetone-water (2E,17S)-17,20-
5 mg of dimethyl-2,3,13,14 _- tetradehydroprostaglandin E1 methyl ester was added and 30
and shaken at °C. The reaction was continued until TLC showed no spots consistent with starting material. Table 1 shows the time until the end of the reaction. After completion of the reaction, the mixture was extracted with 25 ml of ethyl acetate, and the organic layer was washed with saturated brine, dried over anhydrous sodium sulfate and concentrated. The product was (2E,17S)-17,20-dimethyl-2,
3,13,14-tetradehydroprostaglandin E
₁ matched.

[0029]

[Table 1]

Example 4 Lipase VII, 500 mg, was dissolved in 22.5 ml of phosphate buffer (10 mM, pH 7.0) and 2.5 ml of 5
0% (V/V) dissolved in acetone-water (2E, 17
S)-17,20-dimethyl-2,3,13,14 _- tetradehydroprostaglandin E1 methyl ester (50 mg) was added and stirred at 30°C for 3 hours. The reaction mixture was extracted three times with 30 ml of ethyl acetate, and the organic layer was washed with 30 ml of saturated brine twice, dried over anhydrous magnesium sulfate and concentrated. The crude product was subjected to silica gel column chromatography (developing solvent; ethyl acetate: methanol = 40:
1) to give 41 mg of (2E,17S)-17,2
0-Dimethyl-2,3,13,14 _- tetradehydroprostaglandin E1 was obtained [The production amount of 8-β form measured by HPLC analysis was 1.3% (area %). ]. ¹ H-NMR (CDCl ₃ , 300 Mz) δ (pp
m): 0.86-0.94 (m, 3H), 0.91
(d, J = 6.3 Hz, 3H), 1.10-1.88
(m, 15H), 2.18-2.35 (m, 3H),
2.24 (dd, J = 8.8Hz, 18.6Hz, 1
H), 2.64 (ddd, J = 1.7Hz, 6.1H
z, 11.2Hz, 1H), 2.76 (ddd, J =
1.2Hz, 7.3Hz, 18.6Hz, 1H), 4.
28-4.38 (m, 1H), 4.47 (dt, J =
1.7Hz, 7.1Hz, 1H), 5.84 (dt, J
= 1.5Hz, 15.6Hz, 1H), 7.06(d
t, J=7.1Hz, 15.6Hz, 1H) IR (neat): 3391, 2930, 2859, 1
741, 1698, 1654, 1461, 1381, 1
285,1164,1076,984,757 cm ^-1

Example 5 Lipase VII, 1.95 g, was dissolved in 88 ml of phosphate buffer (10 mM, pH 7.0) and 9.8 ml of 50
% (V/V) acetone-dissolved in water (2E, 16R
S)-16-methyl-2,3,13,14 _- tetradehydroprostaglandin E1 methyl ester 195 m
g was added, and the mixture was stirred at 30° C. for 5 hours. The reaction solution was extracted three times with 50 ml of ethyl acetate, and the organic layer was washed with 50 ml of saturated brine.
1 twice, dried over anhydrous magnesium sulfate and concentrated. The crude product was purified by silica gel column chromatography (developing solvent: ethyl acetate:methanol=40:1), and 160 mg of (2E,16RS)-16-methyl-2,3,13,14-tetradehydroprostaglandin was obtained. _E1 was obtained. ¹ H-NMR (CDCl ₃ , 300 Mz) δ (pp
m): 0.84-0.96 (m, 3H), 0.99
(d, J = 6.7 Hz, 3H), 1.10-1.90
(m, 13H), 2.16-2.36 (m, 3H),
2.24 (dd, J = 9.0 Hz, 18.6 Hz, 1
H), 2.65 (ddd, J = 1.8Hz, 8.3H
z, 11.4Hz, 1H), 2.76 (ddd, J =
1.3Hz, 7.3Hz, 18.6Hz, 1H), 4.
27-4.39 (m, 2H), 5.84 (dt, J =
1.5Hz, 15.6Hz, 1H), 7.05(dt,
J=7.0Hz, 15.6Hz, 1H) IR (neat): 3391, 2931, 2860, 2
237, 1744, 1698, 1653, 1462, 1
418, 1378, 1284, 1158, 1078, 1
032,758 cm ^-1

Example 6 Lipase VII, 1.29 g, was dissolved in 54.45 ml of phosphate buffer (10 mM, pH 7.0) and 6.05 ml of
of 50% (v/v) acetone-water (2E, 1
5RS)-16,16-dimethyl-2,3,13,14
- _{Tetradehydroprostaglandin} E1 methyl ester (129 mg) was added and stirred at 30°C for 4 hours. Treated as in Example 5, 113 mg of (2E,15R)
S)-16,16-dimethyl-2,3,13,14 _- tetradehydroprostaglandin E1 was obtained. ¹ H-NMR (CDCl ₃ , 300 Mz) δ (pp
m): 0.91 (t, J = 6.9 Hz, 3H), 0.9
4 (s, 3H), 0.96 (s, 3H), 1.15~
1.87 (m, 12H), 2.20-2.32 (m, 3
H), 2.23 (dd, J=9.1Hz, 18.5H
z, 1H), 2.66 (ddd, J = 1.7Hz, 8.
2Hz, 11.4Hz, 1H), 2.77(ddd, J
= 1.2Hz, 7.3Hz, 18.5Hz, 1H),
4.08-4.17 (m, 1H), 4.29-4.39
(m, 1H), 5.84 (dt, J=1.5Hz, 1
5.6 Hz, 1 H), 7.05 (dt, J = 7.0 H
z, 15.6 Hz, 1H) IR (neat): 3392, 2932, 2861, 2
235, 1743, 1697, 1653, 1385, 1
284, 1159, 1077, 1024 cm ^-1

Example 7 Lipase VII, 1.90 g, was dissolved in 85.5 ml of phosphate buffer (10 mM, pH 7.0) and 9.5 ml of 5
0% (V/V) dissolved in acetone-water (2E, 17
S)-2a-homo-17,20-dimethyl-2,2a,
13,14 _- tetradehydroprostaglandin E1
190 mg of methyl ester was added and stirred at 30° C. for 5 hours. Treated as in Example 5, 124 mg of (2
E,17S)-2a-homo-17,20-dimethyl-
2,2a,13,14 _- tetradehydroprostaglandin E1 was obtained. ¹ H-NMR (CDCl ₃ , 300 Mz) δ (pp
m): 0.85-0.97 (m, 6H), 1.10-
1.87 (m, 17H), 2.17-2.30 (m, 3
H), 2.24 (dd, J=9.2Hz, 18.5H
z, 1H), 2.64 (ddd, J = 1.7Hz, 8.
3Hz, 11.3Hz, 1H), 2.76 (ddd, J
= 1.2Hz, 7.2Hz, 18.5Hz, 1H),
4.28-4.38 (m, 1H), 4.48 (dt, J
= 1.7Hz, 7.4Hz, 1H), 5.83(dt,
J=1.4Hz, 15.6Hz, 1H), 7.06(d
t, J=7.0Hz, 15.6Hz, 1H) IR (neat): 3369, 2929, 2858, 2
237, 1744, 1697, 1654, 1384, 1
284,1160,1075,983 cm ^-1

Example 8 Lipase VII, 2.16 g, was dissolved in 97.2 ml of phosphate buffer (10 mM, pH 7.0) and dissolved in 10.8 ml of 50% (V/V) acetone-water ( 2E, 17
S)-2a,2b-dihomo-17,20-dimethyl-
216 mg of 2,2a,13,14 _- tetradehydroprostaglandin E1 methyl ester was added and stirred at 30°C for 5.5 hours. Treated as in Example 5, 14
5 mg of (2E,17S)-2a,2b-dihomo-1
216 mg of 7,20-dimethyl-2,2a,13,14 _- tetradehydroprostaglandin E1 were obtained. ¹ H-NMR (CDCl ₃ , 300 Mz) δ (pp
m): 0.84-0.95 (m, 6H), 1.12-
1.84 (m, 19H), 2.17-2.30 (m, 3
H), 2.23 (dd, J=9.2Hz, 18.4H
z, 1H), 2.65 (ddd, J = 1.7Hz, 8.
3Hz, 11.5Hz, 1H), 2.76 (ddd, J
= 1.2Hz, 7.2Hz, 18.4Hz, 1H),
4.28-4.38 (m, 1H), 4.47 (dt, J
= 1.7Hz, 7.1Hz, 1H), 5.83(dt,
J=1.4Hz, 15.6Hz, 1H), 7.06(d
t, J = 7.0Hz, 15.6Hz, 1H) IR (neat): 3382, 2928, 2858, 2
237, 1746, 1697, 1653, 1465, 1
418, 1379, 1284, 1162, 1076, 1
046,984 cm ^-1

Preparation Example (2E,17S)-17,20-dimethyl-2,3,1
3,14-tetradehydro-PGE1 methyl ester (1)(3S,5S)-3-(t-butyldimethylsiloxy)-5-methylnon-1-yne (3.85 g) was dissolved in 28.8 ml of benzene. , n-butyllithium (1.95M, hexane solution, 6.4ml) was added at 0°C,
The mixture was stirred at the same temperature for 30 minutes. Diethylaluminum chloride (0.97 M, hexane solution, 1
4.8 ml) was added, and after the temperature was raised to room temperature, the mixture was stirred for 30 minutes. To this solution at room temperature was added (4R)-2-(N,N-diethylamino)methyl-4-(t-butyldimethylsiloxy)cyclopent-2-en-1-one (0.25 M,
Benzene solution, 38.4 ml) was added and stirred for 15 minutes. The reaction solution was hexane (100 ml)-saturated ammonium chloride aqueous solution (100 ml)-hydrochloric acid aqueous solution (3M, 30
ml) with stirring, the organic layer was separated and washed with a saturated aqueous sodium bicarbonate solution (50 ml). The organic layer was dried and concentrated, and the residue was purified by silica gel column chromatography (developing solvent: hexane:ether = 10:1) to give (3R,4R)-2-methylene-3-[ (3′S,5′S)-3′-(t-butyldimethylsiloxy)-5′-methylnon-1′-ynyl]
3.72 g of 4-(t-butyldimethylsiloxy)cyclopentan-1-one are obtained. ¹ H-NMR (CDCl ₃ , 200 MHz) δppm:
0.09, 0.10 and 0.12 (3s, 12H),
0.89 (s, 18H), 0.80-0.99 (m, 6
H), 1.00-1.72 (m, 9H), 2.32 (d
d, J=7.4Hz, 18.0Hz, 1H), 2.71
(dd, J=6.6 Hz, 18.0 Hz, 1H),3.
47-3.56 (m, 1H), 4.15-4.33
(m, 1H), 4.44 (dt, J = 1.6Hz, 7.
0Hz, 1H), 5.54 (d, J = 2.6Hz, 1
H), 6.13 (d, J = 3.0 Hz, 1H) IR (neat): 2930, 2850, 1740, 1
640, 1460, 1360, 1250, 1120, 1
080,835,770 cm ^-1

(2) (4E)-5-carbomethoxypent-4-enylzinc(II) iodide (0.64M solution in tetrahydrofuran, 2.81m) at -70°C
l) copper (I) cyanide/lithium dichloride (392 m
2.25 ml of the tetrahydrofuran solution of g) was added and stirred at the same temperature for 15 minutes. To this solution, 3 ml of a solution of the compound (434 mg) obtained in the above (1) and 0.21 ml of chlorotrimethylsilane in diethyl ether was added at -70°C, and the temperature was raised to room temperature over about 2 hours while stirring.

15 of saturated ammonium chloride aqueous solution was added to the reaction solution.
ml was added and extracted with hexane. The organic layer was washed with saturated brine, dried and concentrated, and the residue obtained was washed with ether.
Dissolve in 3.5 ml of isopropyl alcohol (1:4) and add p-toluenesulfonic acid pyridine salt (8.8 mg,
0.035 mmol) was added, and the mixture was stirred at room temperature for 12 hours. After extraction with 20 ml of hexane and 10 ml of saturated aqueous sodium bicarbonate solution to the reaction solution, the residue obtained after drying and concentration of the organic layer was subjected to silica gel column chromatography (developing solvent; hexane:ether=4:1) (2E, 17S)-17,20-dimethyl-2,3,
532 mg of 13,14-tetradehydro _- PGE1 methyl ester 11,15-bis(t-butyldimethylsilyl ether) were obtained. ¹ H-NMR (CDCl ₃ , 200 MHz) δppm:
0.10 (s, 6H), 0.11 (s, 3H), 0.1
3 (s, 3H), 0.83-0.98 (m, 6H),
0.89 (s, 9H), 0.90 (s, 9H), 1.0
6-1.82 (m, 15H), 2.11-2.29
(m, 3H), 2.17 (dd, J=7.0Hz, 1
8.0 Hz, 1H), 2.59-2.77 (m, 2
H), 3.73 (s, 3H), 4.23-4.35
(m, 1H), 4.43 (dt, J=1.5Hz, 7.
0Hz, 1H), 5.82 (dt, J = 1.5Hz, 1
5.7Hz, 1H), 6.96 (dt, J = 6.9H
z, 15.7 Hz, 1 H) IR (neat): 2954, 2930, 2858, 2
234, 1748, 1728, 1660, 1463, 1
436, 1362, 1326, 1259, 1198, 1
124, 1092, 1006, 838, 779, 670
cm ^-1

(3) The compound (532m
g, 0.857 mmol) to acetonitrile (29 m
l) and dissolved in 50% aqueous hydrofluoric acid solution (6.
9 ml) was added. After stirring for 90 minutes at 0° C., the reaction was poured into ethyl acetate (40 ml) and saturated aqueous sodium bicarbonate (230 ml). Extract with ethyl acetate, wash with saturated aqueous sodium bicarbonate solution and saturated brine,
The residue obtained after drying and concentration was subjected to silica gel column chromatography (developing solvent; ethyl acetate: methanol = 4
0:1) to give 305 mg of the title compound. ¹ H-NMR (CDCl ₃ , 300 MHz) δ ppm:
0.83-0.97 (m, 6H), 1.08-1.90
(m, 15H), 2.12-2.30 (m, 4H),
2.62 (dd, J = 9.0 Hz, 10.5 Hz, 1
H), 2.75 (dd, J = 7.3Hz, 18.5H
z, 1H), 2.92 (br.s, 2H), 3.72
(s, 3H), 4.27-4.36 (m, 1H), 4.
47 (dt, J = 1.0Hz, 6.7Hz, 1H),
5.68 (d, J = 15.7Hz, 1H), 6.96
(dt, J = 7.4 Hz, 15.7 Hz, 1H) IR (neat): 3380, 2910, 2230, 1
720, 1700, 1650, 1435, 1270, 1
040 cm ^-1

(3S, 5S)-3 in Production Example (1)
-(t-butyldimethylsiloxy)-5-methylnona-
Using the corresponding starting material instead of 1-yne, and (4E)-5-carbomethoxypento-
The following compounds were obtained in the same manner as in the Production Examples, except that the corresponding raw materials were used in place of 4-enylzinc(II) iodide.

(2E,16RS)-16-methyl-2,
3,13,14-tetradehydroprostaglandin E
1 methyl ester ¹ H-NMR (CDCl ₃ , 300 MHz) δppm:
0.85-0.96 (m, 3H), 0.99 (d, J =
6.7 Hz, 3H), 1.10-1.87 (m, 13
H), 2.17-2.30 (m, 3H), 2.24 (d
d, J=9.1Hz, 18.5Hz, 1H), 2.64
(ddd, J = 1.7Hz, 8.2Hz, 9.9Hz,
1H), 2.76 (dd, J=7.3Hz, 18.5H
z, 1H), 3.73 (s, 3H), 4.25-4.3
8 (m, 2H), 5.83 (d, J=15.7Hz, 1
H), 6.96 (dt, J=7.0Hz, 15.7H
z, 1H) IR (neat): 3420, 2931, 2860, 2
236, 1746, 1728, 1657, 1438, 1
275, 1203, 1158, 1079, 1036 cm
^-1

(2E,15RS)-16,16-dimethy
Ru-2,3,13,14-tetradehydroprostagla
Carcassin E1 methyl ester ¹ H-NMR ( _CDCl3 , 300 MHz) δppm:
0.91 (t, J=7.0Hz, 3H), 0.94
(s, 3H), 0.96 (s, 3H), 1.19-1.
87 (m, 12H), 2.15-2.35 (m, 3
H), 2.24 (dd, J=9.2Hz, 18.5H
z, 1H), 2.65 (ddd, J = 1.8Hz, 8.
3Hz, 11.4Hz, 1H), 2.76 (ddd, J
= 1.3Hz, 7.3Hz, 18.5Hz, 1H),
3.73 (s, 3H), 4.07-4.17 (m, 1
H), 4.27-4.40 (m, 1H), 5.82 (d
t, J = 1.5Hz, 15.6Hz, 1H), 6.95
(dt, J = 7.0 Hz, 15.6 Hz, 1H) IR (neat): 3435, 2932, 2861, 2
233, 1746, 1728, 1657, 1438, 1
385, 1275, 1202, 1159, 1079, 1
034 cm ^-1

(2E,17S)-2a-homo-17,2
0-dimethyl-2,2a,13,14-tetradehydro
Prostaglandin E 1 methyl ester ¹ H-NMR (CDCl ₃ , 300 MHz) δppm:
0.86-0.95 (m, 6H), 1.10-1.83
(m, 17H), 2.16-2.29 (m, 3H),
2.23 (dd, J = 9.0 Hz, 18.5 Hz, 1
H), 2.64 (ddd, J = 1.7Hz, 8.2H
z, 11.3Hz, 1H), 2.76 (ddd, J =
1.3Hz, 7.2Hz, 18.5Hz, 1H),3.
73 (s, 3H), 4.28-4.38 (m, 1H),
4.48 (dt, J = 1.7 Hz, 7.1 Hz, 1
H), 5.82 (dt, J=1.5Hz, 15.7H
z, 1H), 6.96 (dt, J = 7.0Hz, 15.
7Hz, 1H) IR (neat): 3419, 2930, 2858, 2
237, 1747, 1729, 1658, 1462, 1
438, 1319, 1276, 1201, 1162, 1
078, 1044, 853, 727 cm ^-1

(2E,17S)-2a,2b-dihomo-
17,20-dimethyl-2,2a,13,14-tetra
Dehydroprostaglandin E 1 methyl ester ¹ H-NMR (CDCl ₃ , 300 MHz) δppm:
0.85-0.96 (m, 6H), 1.06-1.86
(m, 19H), 2.15-2.27 (m, 3H),
2.23 (dd, J = 9.2 Hz, 18.5 Hz, 1
H), 2.65 (ddd, J = 1.7Hz, 8.3H
z, 11.5Hz, 1H), 2.75 (ddd, J =
1.2Hz, 7.2Hz, 18.5Hz, 1H),3.
73 (s, 3H), 4.28-4.38 (m, 1H),
4.47 (dt, J = 1.7 Hz, 7.1 Hz, 1
H), 5.82 (dt, J=1.5Hz, 15.6H
z, 1H), 6.96 (dt, J = 7.0Hz, 15.
6Hz, 1H) IR (neat): 3413, 2929, 2858, 2
236, 1746, 1728, 1657, 1438, 1
273, 1201, 1172, 1044, 984 cm
^-1

──────────────────────────────────────────────────── ──Continued from the front page (72) Inventor Kazunori Hanada 3-24-1 Takada, Toshima-ku, Tokyo Taisho Pharmaceutical Co., Ltd. (72) Inventor Toru Tanami 3-24-1 Takada, Toshima-ku, Tokyo Inside Taisho Pharmaceutical Co., Ltd. (72) Inventor Fumie Sato 3-1-219 Kugenuma Higashi, Fujisawa City, Kanagawa Prefecture (56) References JP-A-52-21392 (JP, A) JP-A-55-13282 ) US Patent 3875003 (US, A) (58) Searched field (Int.Cl. ⁷ , DB name) C12P 31/00 C07C 405/00 C12R 1:72 CA/REGISTRY/WPIDS/B IOSIS (STN)

Claims (1)

(57) [Claims] [Claim 1] Formula [Chemical 1] [In the formula, R represents a lower alkyl group, R ¹ and R ² are the same or different and represent a hydrogen atom or a hydroxyl-protecting group, and A, R ³ and R ⁴ represent any group that does not participate in the hydrolysis reaction. and B represents an ethynylene group . ] To lower alkyl esters of E-type prostaglandins represented by
It is hydrolyzed by the action of an ester-hydrolyzing enzyme produced by microorganisms belonging to the genus Candida to give the following formula: (wherein R ¹ , R ² , A, R ³ , R ⁴ and B have the same meanings as defined above).