JPH0720965B2 - 1,3,4-Tridehydro-phanthinolium hydroxide - Google Patents

Description

[Detailed Description of the Invention] Powder control (Stephania tetrandra S.MOORE) is a herbal medicine based on the roots of the family Menispermaceae,
As a result of pharmacological studies in China, analgesic, anti-inflammatory, anti-allergic, antihypertensive effects, etc. have been proved. It is known that the roots of powder control self contain about 1.2% of alkaloids such as tetrandrine, funtinorin, menisin, menisidine and cyclanoline, and the present inventors have conducted a search for this alkaloid component, They have found a new alkaloid and completed the present invention.

That is, the present invention has the following formula 1,3,4-tridehydro-phenantholinium hydroxide represented by the following (hereinafter referred to as the compound of the present invention).

For example, the following method can be used to obtain the compound of the present invention.

Powdery self is extracted with water, alcohols or a mixed solvent of water and alcohols, and the residue obtained by removing the solvent from the extract as it is, or if necessary, dissolved in water or a mixed solvent of water and alcohols, n -Distribution with an organic solvent such as hexane and petroleum ether, after removing the fat-soluble components that migrate to the organic solvent, it is dissolved in pH 9 ammonia water and further extracted with chloroform to extract the ammonia water extract and the chloroform extract. obtain. Next, the aqueous ammonia extract is used as an elution solvent for at least one selected from water, methanol, chloroform, ether, hexane, benzene, and ethyl acetate, and Sephadex such as Sephadex LH 20 and a porous polymer such as Diaion HP 20.
It can be obtained by subjecting to column chromatography using a carrier such as alumina or silica gel several times and fractionating while confirming the target component by thin layer chromatography.

The above-mentioned aqueous ammonia extract may be directly subjected to column chromatography, but it may be added after adding a suitable acid such as hydrochloric acid to form a salt. When converted to a salt, it is freed by subjecting it to alumina (basic) column chromatography using chloroform, methanol, ethyl acetate, etc. as the elution solvent, or making it alkaline with ammonia water and extracting with chloroform, etc. be able to.

Further, upon purification, recrystallization may be carried out using an appropriate solvent such as acetone, methanol, ethanol and the like.

The production examples of the compound of the present invention are as follows.

Production Example 1 7.76 kg of powder powder was extracted with 30 l of methanol, and methanol was removed from the extract to obtain 410 g of methanol extract. This methanol extract was dissolved in 1.5 l of 90% methanol-water mixture, extracted with 1.5 l of n-hexane three times to remove fat-soluble components, and then 90% methanol-water fraction was concentrated under reduced pressure to obtain 1.5 l of ammonia water having a pH of 9 was added to the residue to dissolve the residue, and the mixture was extracted 5 times with 1.5 l of chloroform. The ammonia water layer and the chloroform layer were concentrated under reduced pressure to obtain an ammonia water extract and a chloroform extract. Then, the aqueous ammonia extract was subjected to column chromatography using water and methanol as eluents, and the methanol eluate was concentrated under reduced pressure, and chloroform, chloroform-methanol and methanol were used as eluents. Fr-1 attached to alumina column chromatography (aluminum oxide 90 manufactured by Merck)
(Meaning a fraction. The same applies below), divided into Fr-2 and Fr-3 fractions, and separating Fr-2 into silica gel (Kieselgel).
60, manufactured by Merck)
Chloroform-methanol-ammonia water (70: 10: 1)
Was eluted with Fr2-1 and Fr2-2. Next, Fr2-2 was dissolved in methanol, 10% hydrochloric acid-methanol was added to adjust the pH to 5, then methanol was concentrated under reduced pressure, and then recrystallized using methanol-acetone to give an Rf value of 0.41 [thin layer Plate: Kieselgel 60 F ₂₅₄ (Merck), developing solvent: lower layer of chloroform-methanol-water (65:35:10), color reagent: Dragendorff reagent] The hydrochloride salt was obtained.

Melting point: 300 ° C or less 3400,2932,2548,1610,1558,1512,1266,1226,1204,1124,
840 Proton Nuclear Magnetic Resonance Spectrum (δppm in CD ₃ OD): 3.04 (3H, s), 3.45 (6H, s), 3.90 (3H, s), 4.06 (3H, s), 4.13 (1H, d, J = 15.9Hz), 4.48 (3H, s), 5.44 (1H, d, J = 15.9Hz), 5.96 (1H, s), 6.62 (1H, d-like), 6.65 (1H, s), 6.81 (1H, dd, J = 8.06,2.2Hz), 6.86 (1H, s-like), 6.94 (1H, dd, J = 8.3,2.44Hz) 6.98 (1H, d, J = 8.06Hz), 7.17 (1H, dd, J = 8.3,2.44Hz), 7.42 (1H, s), 7.63 (1H, dd, J = 8.06,2.2Hz), 8.10 (1H, d, J = 6.8Hz), 8.36 (1H, d, J = 6.8) Hz), and then the yellow needle crystals were subjected to alumina column chromatography, and the solvent was removed from the solution eluted with chloroform-methanol (20: 1) to obtain an orange powder having the following physicochemical properties.

3412,2924,2836,1610,1568,1530,1506,1484,1424,1372,
1352,1290,1262,1204,1126,838 Proton Nuclear Magnetic Resonance Spectrum (δppm in CDCl ₃ ): 2.50 (3H, s), 3.56 (3H, s), 3.87 (3H, s), 3.89 (3H, s) , 3.96 (3H, s), 4.08 (1H, d, J = 16Hz), 5.67 (1H, s), 6.06 (1H, d, J = 16Hz), 6.31 (1H, d, J = 1.95Hz), 6.48 (1H, s), 6.53 (2H), 6.60 (1H, s), 6.79 (1H, dd, J = 8.3,2.44Hz), 6.84 (1H, d, J = 8.3Hz), 6.87 (1H, d- like), 7.07 (1H, d-like), 7.16 (1H, d, J = 6Hz), 7.21 (1H, d, J = 6Hz), ▲ [α] ²³ _D ▼ -59.7 ° (c = 0.32, EtOH ) FD-MSm / z: 605 (M ⁺ ) In the infrared absorption spectrum of this physicochemical property,
The absorption at 12 cm ^-1 suggests the presence of hydroxyl group, and in the proton nuclear magnetic resonance spectrum, 3.56, 3.87 and 3.89 pp
Signals of 3 methoxy groups at m, 2 N-methyl groups at 2.50 and 3.96 ppm, and 2 olefin protons characteristic at 7.16 and 7.21 ppm were observed. In addition, M ⁺ was observed at m / z 605 in the FD-MS spectrum. Based on these findings, it was determined to be 1,3,4, -tridehydro-phanthinolium hydroxide.

Further, the compound of the present invention has the following formula It can also be obtained by oxidizing Fangchinoline represented by Futaninolin is a powdery self-extracting substance extracted with water, alcohols or a mixed solvent of water and alcohols, and the residue obtained by removing the solvent from the extract is n-hexane or petroleum ether with water or a mixed solvent of water and alcohols. After partitioning with an organic solvent such as the above to remove the fat-soluble component that migrates to the organic solvent, it is dissolved in aqueous ammonia having a pH of 9 and further extracted with chloroform to obtain an aqueous ammonia extract and a chloroform extract. Then, the chloroform extract is used as an elution solvent with at least one selected from water, methanol, chloroform, ether, hexane, benzene, and ethyl acetate, and Sephadex LH.
Obtained by fractionating while confirming the target component by thin layer chromatography, which is applied several times to column chromatography using a Sephadex such as 20 and a porous polymer such as Diaion HP20, a carrier such as alumina or silica gel, etc. be able to. In some cases, it may be purified by recrystallization using a suitable solvent such as acetone, methanol or ethanol.

Production Example 2 7.76 kg of powder powder was extracted with 30 l of methanol, and methanol was removed from the extract to obtain 410 g of methanol extract. This methanol extract was dissolved in 1.5 l of 90% methanol-water mixture, extracted with 1.5 l of n-hexane three times to remove fat-soluble components, and then 90% methanol-water fraction was concentrated under reduced pressure to obtain 1.5 l of ammonia water having a pH of 9 was added to the residue to dissolve the residue, and the mixture was extracted 5 times with 1.5 l of chloroform. The ammonia water layer and the chloroform layer were concentrated under reduced pressure to obtain an ammonia water extract and a chloroform extract. Then extract the chloroform extract with alumina (aluminum oxide 90
It was subjected to column chromatography (manufactured by Mekuru Co., Ltd.) and eluted with chloroform to obtain Fr-1 and Fr-2. Then Fr
-2 was subjected to silica gel (Kieselgel 60 Meckl) column chromatography and eluted with chloroform-methanol (20: 1) to obtain Fr2-1, Fr2-2, Fr2-3 and Fr2-4, and Fr2 Rf value 0.34 [thin layer plate: Kieselgel] by removing the eluting solvent from -1 and recrystallizing with acetone
60 F ₂₅₄ , developing solvent: chloroform-methanol (3:
1), color reagent: Dragendorff reagent] was obtained.

Next, ethanol (7 ml) and excess manganese dioxide (204 mg) were added to futanolin (204 mg), and the mixture was allowed to stand at 4 ° C for 3 days. The residue was filtered and concentrated under reduced pressure. The residue was subjected to silica gel column chromatography, and chloroform-methanol-ammonia water (40 10:10) to obtain the compound of the present invention.

The compound of the present invention has an angiotensin I converting enzyme inhibitory action and is useful as an antihypertensive agent. This will be described with reference to experimental examples.

Experimental example 50 g of 2 g of Rabitutrang acetone powder (manufactured by Sigma)
Suspend in 30 ml of mM phosphate buffer (pH 8.3) at 34,000 g
Centrifuge for 40 minutes and add the supernatant to a dialysis tube.
It was dialyzed with 3 l of 10 mM phosphate buffer and further appropriately diluted with 50 mM phosphate buffer to obtain an angiotensin I converting enzyme solution.

The compound obtained in Preparation Example 1 was used at a final concentration of 1 mM.
Dissolve in 25% dimethylsulfoxide, add 25 μl of each solution to each test tube, add 59 μl of hypryl histidyl leucine (final concentration 5 mM) as a substrate, and further add 25 μl of 3 M sodium chloride solution, 500 mM phosphoric acid. After adding 50 μl of buffer solution (pH 8.3) and incubating at 37 ° C for 10 minutes, add 100 μl of enzyme solution obtained as above and add 37 ° C.
And reacted for 60 minutes. After that, 100 μl of 1N hydrochloric acid was added to stop the reaction, 25 μl of benzoylalanine (1 mg / ml) was added as an internal standard, 1 ml of ethyl acetate was added, and the amount of hypuric acid extracted in ethyl acetate was rapidly adjusted. Liquid chromatography [column, μ-Bondapak C ₁₈ (diameter
4 mm, length 30 cm); mobile phase, acetonitrile: methanol: 1% acetic acid (1: 1: 8); flow rate, 1 ml / min; detection, ultraviolet light (25
4 nm)] and used as the enzyme activity.

With respect to this result, the inhibition rate was calculated by the following formula.

C: Hypuric acid peak area when the compound obtained in the specific example is not included (corrected by internal standard) S: Hypuric acid peak area when the compound obtained in the specific example is added (corrected by internal standard) As a result, The inhibition rate of the compound of the present invention obtained in Production Example 1 is 8
It was 0.4%, and an angiotensin I converting enzyme inhibitory action was confirmed.

In addition, the compound obtained in Production Example 1
It was confirmed that the compound of the present invention is relatively safe since no deaths were observed even after oral administration of g / kg.

The compound of the present invention can also be used as a commonly used salt of pharmaceuticals such as hydrochloric acid, sulfuric acid, succinic acid, etc. in order to achieve the effect as a pharmaceutical.

The compound of the present invention can be administered to animals and humans as it is or together with a conventional pharmaceutical carrier. The dosage form is not particularly limited and may be appropriately selected and used as needed, and examples thereof include oral agents such as tablets, capsules and granules, and parenteral agents such as injections and suppositories. Oral preparations such as tablets, capsules and granules are manufactured according to a conventional method. Tablets are produced by mixing and shaping the compound of the present invention with pharmaceutical excipients such as gelatin, starch, lactose, magnesium stearate, talc, gum arabic, etc. It is produced by mixing with an active pharmaceutical filler or diluent and filling it into a hard gelatin capsule, a soft gelatin capsule or the like. Syrups and elixirs are produced by mixing the compound of the present invention with sweeteners such as sucrose, preservatives such as methyl and propylparabens, colorants, seasonings, fragrances, and auxiliary agents.

The parenteral preparation is produced according to a conventional method, and distilled water for injection, physiological saline, dextrose aqueous solution, propylene glycol or the like can be generally used as a diluent. Further, if necessary, a bactericide, a preservative, and a stabilizer may be added.
In addition, from the viewpoint of stability, this parenteral preparation is frozen after filling into ampoules and the like, and water is removed by a normal freeze-drying technique.
It is also possible to reconstitute a solution from a lyophilized product just before use.

Other parenteral agents include liquid preparations for external use, coating agents such as ointments, suppositories for rectal administration, and the like, and they are manufactured by a conventional method.

Claims (1)

[Claims] 1. The following formula 1,3,4-tridehydro-phenantholinium hydroxide represented by.