KR950005737B1 - Separation of monocomponents from ginkgoride mixtures - Google Patents

Abstract

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Description

Separation of monocomponents from ginkgoride mixtures

The present invention relates to a method for effectively separating each single component from the jinkoride mixture, and more particularly, to change the polarity difference by substituting jinkolife B for derivatives of other structures in the gyrolide mixture. The present invention relates to a method for the simple separation of ginkgo lide monosaccharides by separation and recovery of the substituted derivatives in the original structure of zyloride B.

Ginkgolide, a terpenoid-based compound found in the roots and leaves of ginkgo biloba, is a substance with a very unique chemical structure. It is ginkgo according to its chemical structure among ginkgolides separated from natural substances by Nakanishi in 1967. Four kinds of substances were identified and identified, including Laad A, B, C, and M. Then, in 1987, Weing discovered another derivative 0, jinkoride J. The general chemical structure of jinkoride is represented by same.

In the above structural formula, R ₁ , R ₂ and R ₃ are the same or different and each is a hydrogen atom or a hydroxyl group.

The ginkgolide compounds of the above formula (A) are classified according to the substituents R ₁ , R ₂ and R ₃ of the formula as shown in the following table.

These ginkgorides are classified by the difference in the hydroxy group (-OH group) of carbons 1, 3, and 7 in terms of their chemical structure, but their chemical properties are very similar to each other. .

Therefore, after separating fractional recrystallization several times using the difference in solubility according to the difference of hydroxyl group as a method of separating each component of the jinkoride, a pure single substance is separated using a silica gel column [K , Nakanish, Pure. Appl. Chem. 1967. 89-113), or Weing et al. Used an ion exchange resin such as Sephadex-LH-20 two to three times to separate each component. Liebigs. Ann. Chem. 1987. 521-526

However, in the conventional separation method, the separation efficiency is greatly reduced due to the similar properties of each component in the use of fractional recrystallization. In particular, the zinc chromatography A and B have very similar properties, and thus thin plate chromatography (silica gel thin film, toluene : When acetone = 7: 3 using a developing solvent), there is almost no difference in the Rf value between 0.32 and 0.30 for B, so the silica gel column can hardly be separated to be repeated several times. Because of the need to use expensive ion exchange resins such as Sephadex-LH-20, it is very difficult to separate them.

Moreover, recently, since the early 1980s, research by the Institute of Henri Beaufour (IHB), France, has revealed that zincoride has a PAF- antagonistic activity. Although it has been found that the drug is very good compared to the kinds of substances, an effective separation method has not been developed, and thus, it is attempting to pharmaceutical as a mixture of jinkoride A, B and C. [Refer to British Patent Application No. 22062062A]

However, in this case, since ginkgorides A and C, which are relatively less effective than ginkgoride B, are mixed, it is necessary to separate only pure single ingredient ginkgoride B, and in order to use the properties according to each single ingredient of ginkgoride. If it is possible to separate the simple, easy and economical separation of ginkgoide monosaccharides because of the need for separation, this opens up a new way to the use of ginkgoride, which is very useful.

Accordingly, the present inventors have steadily studied to solve such difficulties in separation of the conventional jingolide components. As a result, the present inventors simply substitute hydroxy groups by using the chemical structural differences of the xingolides A, B, and C components to replace each component. It has been found that the present invention can be effectively separated to complete the present invention.

It is an object of the present invention to provide a method for economically and effectively separating a single jinkoride constituent by changing the polarity difference during the awakening by substituting a hydroxyl group from the jinkoride mixture.

Hereinafter, the present invention will be described in detail.

In the present invention, in the separation of a zinc core component from a zinc core mixture, the hydroxy group of carbon 1 and carbon 10 in zinc core B and C is substituted with a cyclic acetal or ketal compound represented by the following structural formula (I) , To separate unsubstituted unreacted ginkgoride and the substituted compound of formula (I), wherein the compound of formula (I) is hydrolyzed with acidic aqueous solution to recover the original ginkgoride B and C, respectively. It is done.

In the above formula, when R is H, it is zinc cord B, and when OH is zinc cord C, R ₁ and R ₂ are the same as or different from each other and are a hydrogen atom, a methyl group or an ethyl group.

Referring to the present invention in more detail as follows.

In the present invention, the hydroxy group attached to the carbon 1 and the hydroxyl group of the carbon 1 and the zinc 1 in the cholcoride mixture of cyclic acetal (Cyclic Acetal) or ketal (Ketal) of the structure (I) Substituting in the form of), it can be easily separated from the reactant substituted by using a cheap silica gel column simply by using a cheap silica gel column by changing the polar difference with the unsubstituted jinkoride A, which does not have a hydroxyl group on carbon 1 The superposition of the above formula (I), which is a derivative of jinkoride B and C, is hydrolyzed with an acid solution such as HCI and replaced with the original jinkoride B and C, thereby effectively separating each jinkoride single component. It is.

According to the present invention, for example, the mixture of ginkgorides A, B, and C is slowly dissolved in a mixture of acetic acid and sulfuric acid, and then an excess amount of formaldehyde or paraformaldehyde is added and stirred at room temperature for 3 to 8 hours. Or by heating to 70 to 90 ° C. and stirring for 1 to 3 hours to form a 1,10-dioxy-zincolide derivative represented by the above formula (I) in the mixture.

The mixture of the structural formula (I) thus obtained has a significant polarity difference with the unreacted residual zinc cord A. The reaction mixture is slowly diluted with 0 ° C. cooling water and then chloroform, ethyl acetate and ethyl ether as a nonpolar solvent insoluble in water. The organic solvent layer was dried under reduced pressure by fractional extraction with a solvent selected from chloroform, and then chloroform black was easily reacted using a silica gel column using a mixed solvent of chloroform methanol. Deoxy-jincolide derivatives and pure ginkgoride A are each obtained quantitatively.

The thus-obtained ginkgoride derivative compound of the structural formula (I) is hydrolyzed with an acidic aqueous solution. The compound of the structural formula (I) is added with an acidic solution selected from, for example, 2-4N hydrochloric acid, sulfuric acid, or acetic acid, and then 2-3. Heated to a temperature of 90-120 ° C. for a period of time, diluted with 0 ° C. cooling water, fractionated with a nonpolar solvent selected from water and insoluble solvents such as chloroform, ether or acetate, and the organic layer was dried under reduced pressure, and then ethyl alcohol, methyl alcohol or When recrystallized from its aqueous solution, pure ginkgoride B precipitates.

In this case, since the glycolic acid C is dissolved in the alcohol solution without precipitation, the zincoid C is also naturally separated.

As such, according to the present invention, each single component can be very efficiently and economically separated from the mixture of zincide A, B and C, in particular. In the present invention, the separation of the hydroxyl groups at positions 1 and 10 in the jinkoride B and C with a cyclic structure to facilitate the separation, it is substituted with a cyclic acetal or ketal as in the present invention or similar cyclic Even if the structure is substituted, it is expected to be easily separated due to the polarity difference with the jinkoride A.

As described above, according to the present invention, since only a part of the jinkoride mixture is substituted with a hydroxyl group to convert the polarity, it is simple even if the conventional ion exchange resin is not repeatedly used many times even after fractional recrystallization. It is possible to separate the jinkoride of the component, and recover the jinkoride by the simple and economical method by recovering the separated component to the original jinkoride component through a single hand hydrolysis treatment.

Hereinafter, the present invention will be described in detail with reference to Examples, but the present invention is not limited by Examples.

[Production example]

Synthesis of 1,10-methylenedioxy-zincolide B

10 g of zorolide B is added to a mixture of 250 ml of acetic acid and 200 ml of sulfuric acid, which is then dissolved by heating to 50 ° C. 20 g of paraformaldehyde is added thereto and stirred at room temperature for 8 hours.

The reaction was slowly diluted with 2 L of 0 ° C. cooling water, extracted three times with 10 L of dichloromethane, and the organic layers were collected, dried under reduced pressure, and recrystallized from 100 ml of ethyl alcohol to give 9.3 g (90%) of 1,10-dimethylenedioxy. Zingkoride B is obtained.

Rf value = 0.65 (silica gel thin plate, developing solvent (toluene: acetone = 7: 3))

¹ H-NMR (300 MHz, CDCl ₃ ); 5.93 (s, 1 H); H ₁₂ , 5.36 (s, 1 H); H ₁₀ , 5.22 (brs, 1 H); H ₆ , 5.15 (q, 2H); Methylene, 4.71 (d, 1H) H ₂ , 3.96 (d, 1H); H ₁ , 3.07 (q, 1 H); H ₁₄ , 2.36-2.31 (m, 2H); H ₇ , 2.0 (dd, 1H); H ₈ , 1.31 (d, 3H); CH ₃ , 1.10 (s, 9H); t-Bu.

Melting point; 380 ℃ decomposition

Example 1

10 g of the mixture containing ginkgorides A and B in a ratio of 1: 1 (weight ratio) are added to a mixture of 200 ml of acetic acid and 150 ml of sulfuric acid, which is then dissolved by heating to 50 ° C. 10 g of paraformaldehyde is added thereto and stirred at room temperature for 8 hours.

The reaction was slowly diluted with 1.5 L of 0 ° C. cooling water, and extracted three times with 1 L of chloroform to collect the organic layers and dried under reduced pressure to obtain a mixture of 10 g. 10 g of the obtained mixture was passed through a 500 g silica gel packed column with a mixed solvent of chloroform and methanol (10: 1), and 4.7 g of 4.8 g (yield 93%) of pure 1,10-methylenedioxy-zincolide B was obtained. Pure ginkgoride A (yield 94%) is obtained respectively.

[Production of Silica Gel Column]

500 g of silica gel (Merck, Cat. No. 9385, 230-400 mesh) is filled in a column 5 cm in diameter and 1.2 m high, flowing with a mixed solvent of chloroform and methanol (10: 1).

Example 2

10 g of the mixture containing ginkgorides A, B and C, each at 2: 2: 1 (weight ratio), was added to a mixture of 200 ml of acetic acid and 150 ml of sulfuric acid, heated to 50 ° C., and dissolved in 10 g of paraformaldehyde, followed by heating to 90 ° C. Stir for 3 hours.

The reaction was slowly diluted with 1.5 L of 0 ° C. cooling water, repeatedly extracted three times with 1 L of ethyl acetate, and the organic layers were collected and dried under reduced pressure to obtain a mixture of 10 g.

When 10 g of the obtained mixture was passed through a silica gel column prepared in the same manner as in Example 1, 3.8 g (92%) of 1,10-methylenedioxy-gencolide B and 3.8 g (95%) of ginkgoride A and 1, 1.7 g (85%) of 10-methylenedioxy-zincolide C are obtained respectively.

Example 3

4 g of 1,10-methylenedioxy-zincolide B obtained in Example 1 was added to 40 ml of 2N HCl, stirred at 100 ° C. for 3 hours, cooled to room temperature, diluted with 300 ml of distilled water, and then diluted with 3 ml of 200 ml of dichloromethane. After repeated extraction, the organic layer was collected, dried under reduced pressure, 30 ml of anhydrous ethyl alcohol was added thereto, and the resultant was heated to 60 ° C. to coagulate. The mixture was left at 0 ° C. for 3 hours, and the resulting crystals were filtered and dried under reduced pressure to obtain 3.7 g (yield 95%). Pure ginkgoide B is obtained.

Example 4

3.8 g of 1,10-methylenedioxy-gencolide B and 1.7 g of 1,10-methylenedioxy-gencolide C obtained in Example 2 were added to 50 ml of 3N H ₂ SO ₄ and stirred at 100 ° C. for 3 hours. After cooling to room temperature, diluted with 300ml of distilled water, and repeatedly extracted with 300ml of ethyl acetate. The organic layers were collected and dried under reduced pressure. 40 ml of anhydrous ethyl alcohol was added thereto, heated to 60 ° C for dissolution, and left at 0 ° C for 5 hours. The crystals thus obtained are filtered and dried under reduced pressure to yield 3.6 g (97%) of pure jinkoride B.

The remaining filtrate was concentrated to 10ml and 20ml of distilled water was added and left at 0 ° C. for 2 hours to obtain 1.4 g (83%) of pure glycolide C by filtering and drying the resulting precipitate.

Claims (9)

In the separation of the jinkoride monocomponent from the jinkoride mixture, the hydroxy groups of carbons 1 and 10 in jinkoride B and C are replaced with cyclic acetals or ketal compounds represented by the following structural formula (I), and not substituted. Unreacted ginkgoride and the substituted compound of formula (I), wherein the compound of formula (I) is hydrolyzed by acidic aqueous solution to recover ginkgo B and C, respectively. A method of separating short components from a ride mixture. In the above formula, when R is H, it is jinkoride B, and when OH is jinkoride C, R ₁ and R ₂ are the same as or different from each other and are hydrogen, metal or ethyl. The method of claim 1, wherein R ₁ and R ₂ of the formula (I) are each methyl. The method according to claim 1, wherein the substitution with Structural Formula (I) is performed by dissolving the zinc chloride mixture in a mixed solution of acetic acid and sulfuric acid and then adding an excess of formaldehyde or paraformaldehyde. The separation method according to claim 1, wherein the separation of the mixture of unreacted ginkgoride and the structural formula (I) is diluted with water and extracted with water and an insoluble nonpolar solvent. The method of claim 4, wherein the nonpolar solvent is selected from chloroform, ethyl acetate and ethyl ether. The method of claim 1, wherein the hydrolysis using the acidic aqueous solution is carried out by heating using an aqueous solution selected from 2 to 8N hydrochloric acid, sulfuric acid or acetic acid. The method according to claim 1, wherein the recovery of the original zinc cords B and C after the hydrolysis is performed by diluting the hydrolyzed zinc cords with water, extracting them with a nonpolar solvent that is insoluble with water, and recrystallization. 8. The method of claim 7, wherein the nonpolar solvent is selected from chloroform, ether or acetate. 8. The method of claim 7, wherein the recrystallization is to precipitate using ethyl alcohol, methyl alcohol or an aqueous solution thereof.