JP7773177B2 - Method for producing 18α-oleanane triterpene diglucuronides - Google Patents

Description

Application of Article 30, Paragraph 2 of the Patent Act August 31, 2021 67th Annual Meeting of the Japanese Society of Herbal Pharmaceutical Sciences [Tokyo] Collection of Lecture Abstracts (Page 250) 1

The present invention relates to a method for producing 18α-oleanane-type triterpene diglucuronides.

Licorice (Glycyrrhiza sp.) has many useful pharmacological activities, including antiulcer activity, and is an extremely important medicinal plant that is included in many herbal medicines.

The main component of licorice is the sweet saponin glycyrrhizin, which chemically contains an α,β-unsaturated ketone as a characteristic functional group in the C-ring, and is a member of the oleanane-type triterpene diglucuronide family, with the 18th position in the β-configuration.

On the other hand, licorice also contains minor structural analogues of glycyrrhizinic acid. For example, compounds with a hydroxymethyl group at the 23-position and compounds with different stereochemistry at the 20-position are known.

It has been reported that 18α-glycyrrhizinic acid, in which the hydrogen at the 18th position is inverted, can be produced as the major product in a thermodynamic equilibrium mixture of both isomers by heating and refluxing 18β-glycyrrhizinic acid in an aqueous alkali hydroxide solution (potassium or sodium) (see, for example, Patent Document 1 below).

Furthermore, Patent Document 2 below discloses that purified 18-isomerized glycyrrhizin methyl ester is heated with a dilute aqueous alkali hydroxide solution in ethanol, and then added to an acidic aqueous solution to obtain isomerized glycyrrhizin as a precipitate.

Special Publication No. 2-22080 Chinese Patent Application Publication No. 104861031

With the aim of isolating and producing high-purity 18α-glycyrrhizinic acid, the inventors retried an isomerization reaction using an aqueous alkali hydroxide solution, referring to the technology described in Patent Document 1, and attempted to purify 18α-glycyrrhizinic acid from the resulting reaction mixture. However, the desired 18α-glycyrrhizinic acid exhibited nearly identical physicochemical behavior to the raw material 18β-glycyrrhizinic acid, and they were unsuccessful in isolating it with a high degree of purity. Furthermore, even using the technology described in Patent Document 2 was not sufficient to isolate it with a high degree of purity.

In view of the above problems, an object of the present invention is to provide a method for separating and producing 18α-oleanane type triterpene diglucuronides with higher purity.

After investigating the above-mentioned issues, the inventors concluded that one of the reasons for the difficulty in separating and purifying the reaction mixture may be that the target compound has a triterpene carboxylic acid structure containing two glucuronic acid residues, making it extremely polar, and therefore limiting the solvents that can be used for purification using chromatography, etc.

Therefore, the mixture of alkali salts of glycyrrhizinate obtained after the isomerization reaction was treated with dimethyl sulfate (or methyl iodide) in dimethyl sulfoxide (or N,N-dimethylformamide) to convert it into a methyl ester derivative. As a result, no change in the chromatographic behavior of 18β-glycyrrhizinic acid trimethyl ester and 18α-glycyrrhizinic acid trimethyl ester was observed, but their solubilities in methanol differed significantly, and it was found that the latter could be isolated and purified as a poorly soluble compound by fractional recrystallization.

That is, a method for producing 18α-oleanane-type triterpene diglucuronides according to one aspect of the present invention comprises the steps of: (1) mixing and heating an oleanane-type triterpene diglucuronide with an aqueous alkali hydroxide solution to isomerize the hydrogen at the 18-position of the oleanane-type triterpene diglucuronide, thereby obtaining an equilibrium mixture of the 18α-isomer and the 18β-isomer, and adding the resulting mixture dropwise to an alcoholic solution containing acetic acid to obtain a precipitate of an alkali salt derived from the alkali hydroxide used; (2) treating the equilibrium mixture with an alkylating agent in an aprotic polar solvent to convert it into an 18α-trialkyl ester, which is then separated and purified; and (3) hydrolyzing the separated and purified 18α- trialkyl ester with an alcoholic aqueous alkali hydroxide solution, and adding the resulting mixture dropwise to an alcoholic solution containing acetic acid to obtain a precipitate of an alkali salt derived from the alkali hydroxide used.

In addition, in this respect, the oleanane-type triterpene diglucuronide is preferably represented by the following formula:

As described above, the present invention provides a method for separating and producing 18α-oleanane-type triterpene diglucuronides with higher purity.

The ¹ H NMR chart is shown below. The ¹³ C NMR chart is shown.

Embodiments of the present invention are described in detail below with reference to the accompanying drawings. However, the present invention can be implemented in many different forms and is not limited to the specific examples shown in the following embodiments and examples.

The method for producing 18α-oleanane triterpene diglucuronides according to this embodiment (hereinafter referred to as "this method") comprises the steps of: (1) mixing and heating an oleanane triterpene diglucuronide with an aqueous alkali hydroxide solution to isomerize the hydrogen at the 18-position of the oleanane triterpene diglucuronide, thereby obtaining an equilibrium mixture of the 18α-isomer and the 18β-isomer, and then adding the resulting mixture dropwise to an alcoholic solution containing acetic acid to obtain an alkali salt precipitate derived from the alkali hydroxide used; (2) treating the alkali salt mixture with an alkylating agent in an aprotic polar solvent to convert it into an 18α-trialkyl ester, which is then separated and purified; and (3) hydrolyzing the separated and purified 18α- trialkyl ester with an alcoholic aqueous alkali hydroxide solution, and adding the hydrolyzed mixture dropwise to an alcoholic solution containing acetic acid to obtain an alkali salt precipitate derived from the alkali hydroxide used.

As described above, this method first includes the step of (1) mixing and heating an oleanane-type triterpene diglucuronide with an aqueous alkali hydroxide solution to isomerize the hydrogen at the 18-position of the oleanane-type triterpene diglucuronide, thereby obtaining an equilibrium mixture of the 18α-isomer and the 18β-isomer as a precipitate of an alkali salt derived from the alkali hydroxide used.

In this method, the "oleanane triterpene diglucuronide" is a compound having an oleanane triterpene aglycone and two glucuronic acid units in its structure. The specific structure of the oleanane triterpene diglucuronide is not limited, but is preferably, for example, one represented by the following formula:

Furthermore, the alkali hydroxide used in the alkali hydroxide aqueous solution in this step is not limited as long as it can achieve the objective of this step, but examples include potassium hydroxide and sodium hydroxide.

In addition, the concentration of the aqueous alkali hydroxide solution in this step is not particularly limited, but is preferably 10% by weight or more and 50% by weight or less, and more preferably 15% by weight or more and 25% by weight or less.

In addition, in this step, the amount of alkali hydroxide relative to the oleanane-type triterpene diglucuronide is not limited as long as the objective of this step can be achieved, but it is preferably 5 mol or more and 40 mol or less, and more preferably 20 mol or more and 30 mol or less, relative to 1 mol of oleanane-type triterpene diglucuronide.

In addition, in this step, as described above, the oleanane-type triterpene diglucuronide and the aqueous alkali hydroxide solution are mixed and then heated. This heating is performed to promote the reaction, and although there are no limitations, it is preferably in the range of 80°C to 150°C, and more preferably in the range of 100°C to 120°C.

The reaction time between the oleanane-type triterpene diglucuronide and the aqueous alkali hydroxide solution in this step varies greatly depending on the reaction temperature and concentration, and is not limited as long as it allows the reaction to proceed sufficiently. However, it is preferably, for example, 5 to 15 hours, more preferably 8 to 10 hours.

Furthermore, in this step, after the above reaction, the equilibrium mixture of the 18α-isomer and 18β-isomer of oleanane-type triterpene diglucuronide can be added dropwise to an alcohol solution containing acetic acid, thereby directly obtaining the alkali salt of the alkali hydroxide used.

In this step, the alcohol solution containing acetic acid is used to efficiently obtain a precipitate, and although not limited thereto, the alcohol is preferably at least one of methanol, ethanol, and propanol. Furthermore, the alcohol concentration in the final alcoholic aqueous solution is preferably 80% or higher, and more preferably 90% or higher. The acetic acid may be replaced by an organic acid such as formic acid or propionic acid. In other words, an alcoholic solution containing an organic acid can be used.

In this step, the alcohol solution containing acetic acid is used to efficiently obtain a precipitate, and although there are no limitations, the amount of acetic acid is preferably at least the amount needed to neutralize the amount of alkali hydroxide used, and more preferably 3 to 5 times the amount.

In this step, the hydrogen at the 18th position of the oleanane-type triterpene diglucuronide is isomerized by the above reaction, allowing the equilibrium mixture of the 18α-isomer and the 18β-isomer to be directly obtained as an alkali salt precipitate derived from alkali hydroxide using alkali hydroxide.

Following the above step, this method also includes (2) treating the equilibrium mixture with an alkylating agent in an aprotic polar solvent to convert it into an 18α-trialkyl ester, which is then separated and purified, as described above.

In this step, "aprotic polar solvent" refers to a solvent that does not have an ionizable proton. As long as the objective of this step can be achieved, it is not limited to this solvent, but examples include dimethyl sulfoxide and dimethylformamide.

In this step, the "alkylating agent" refers to a compound used for alkylation, and examples thereof include, but are not limited to, at least one of dimethyl sulfate, methyl iodide, diethyl sulfate, and ethyl iodide.

The amount of alkylating agent relative to the equilibrium mixture in this step is not limited as long as the reaction proceeds sufficiently, but it is preferable to use 3 mol or more and 8 mol or less of alkylating agent, and more preferably 4.5 mol or more and 6 mol or less, per 1 mol of the equilibrium mixture.

The temperature for the reaction between the equilibrium mixture and the alkylating agent in this step is not limited as long as the reaction proceeds sufficiently, but is preferably 15°C or higher and 40°C or lower, and more preferably 20°C or higher and 30°C or lower, although the reaction will proceed satisfactorily even at room temperature.

Furthermore, the reaction time between the equilibrium mixture and the alkylating agent in this step is not limited as long as the reaction proceeds sufficiently, but is preferably 2 hours or more and 24 hours or less, and more preferably 4 hours or more and 6 hours or less.

In addition, in this step, it is preferable to add a base to complete the reaction. Specific examples include, but are not limited to, potassium carbonate and sodium carbonate.

In this step, the equilibrium mixture is alkylated to yield the corresponding trialkyl ester mixture.

In addition, in this step, it is necessary to separate and purify the 18α-trialkyl ester. Specifically, it is preferable to filter and wash the precipitate that forms, and then use a purification method such as chromatography. Although there are no limitations on the type of chromatography, silica gel chromatography can be used.

In this step, the mixture of trimethyl esters of isomerized oleanane-type triterpene diglucuronides exhibits differences in solubility in methanol. Therefore, by utilizing this difference in solubility, it becomes possible to obtain the poorly soluble 18-isomerized trimethyl esters as highly pure crystals.

As described above, this method also includes the step of (3) hydrolyzing the separated and purified 18α -trialkyl ester with an alcoholic aqueous alkali hydroxide solution and adding the hydrolysis product dropwise to an alcohol solution containing acetic acid to obtain a precipitate of an alkali salt derived from the alkali hydroxide used.

In this step, "alcoholic aqueous alkali hydroxide solution" refers to an aqueous alkali hydroxide solution containing alcohol.

Here, the alkali hydroxide is not limited as long as it can achieve the objective of this step, but examples include potassium hydroxide and sodium hydroxide.

Here, the alcohol is not limited as long as it can achieve the objective of this step, but examples include methanol, ethanol, and propanol.

In addition, in this step, the concentration of the aqueous alkali hydroxide solution is not limited as long as the objective of this step can be achieved, but it is preferably 5% by weight or more and 25% by weight or less in the aqueous solution, and more preferably 10% by weight or more and 15% by weight or less.

Furthermore, the amount of alkali hydroxide relative to the 18α -trialkyl ester in this step is not limited as long as the reaction proceeds sufficiently, but it is preferable that the amount of alkali hydroxide be 5 mol or more and 20 mol or less, and more preferably 8 mol or more and 12 mol or less, per 1 mol of the 18α- trialkyl ester.

In this step, the amount of alcohol added to the aqueous alkali hydroxide solution is not limited as long as the 18α- trialkyl ester is dissolved therein, and it is preferable that the volume ratio of the alcohol to the alkali hydroxide used is 1.

Furthermore, in this step, after the above reaction, the 18α-oleanane triterpene diglucuronide can be directly obtained as the alkali salt of the alkali hydroxide used by adding it dropwise to an alcohol solution containing acetic acid. This alkali salt can be further dissolved in the solution again and recrystallized, making it possible to further increase its purity.

In this step, the alcohol solution containing acetic acid is used to efficiently obtain a precipitate, and is not limited thereto. However, the alcohol is preferably at least one of methanol, ethanol, and propanol. Furthermore, the alcohol concentration in the final alcoholic aqueous solution is preferably 80% or higher, and more preferably 90% or higher. The acetic acid may be replaced with an organic acid such as formic acid or propionic acid. In other words, an alcoholic solution containing an organic acid can be used.

In this step, the alcohol solution containing acetic acid is used to efficiently obtain a precipitate, and although there are no limitations, it is preferable that the amount of acetic acid be equal to or greater than the amount of alkali hydroxide used to neutralize it, and more preferably 3 to 5 times the amount.

In this step, the obtained alkali salt of 18α-oleanane type triterpene diglucuronide can be converted into 18α-oleanane type triterpene diglucuronide by acidifying it, and the 18α-oleanane type triterpene diglucuronide can be converted into the ammonium salt of 18α-oleanane type triterpene diglucuronide by treating it with aqueous ammonia solution.

As described above, this method provides a method for separating and producing 18α-oleanane triterpene diglucuronides with higher purity. The compounds obtained by this method are expected to have higher pharmacological activity than 18β-oleanane triterpene diglucuronides before isomerization.

Here, we actually implemented the manufacturing method according to the above embodiment and confirmed its effectiveness. The details are explained below.

(A) Isomerization reaction First, 18β-glycyrrhizinic acid ammonium salt (5 g, 5.95 × 10 ⁻³ M) was heated under reflux for 8 hours together with an aqueous solution of potassium hydroxide (8 g, 143 × 10 ⁻³ M) dissolved in water (32 mL), and then added dropwise to a solution of acetic acid (10 mL, 175 × 10 ⁻³ M) dissolved in 99.5% ethanol (300 mL) with stirring at room temperature, and the precipitated crystals were filtered by suction to obtain the potassium salt (4.6 g) of the crude isomerization mixture.

(a) Methylation Reaction Next, the potassium salt of the crude isomerized mixture (6.26 g, 7.274 × 10 ^-3 M) was added to dimethyl sulfoxide (18 mL) containing dimethyl sulfate (3.5 mL, 36.4 × 10 ^-3 M) and potassium carbonate (5 g, 36.4 × 10 ^-3 M), and the mixture was stirred overnight at room temperature. Then, the mixture was poured into a 2% aqueous hydrochloric acid solution (75 mL), and the precipitate was filtered and washed with water to obtain a reaction product (6.03 g). Of this, 3.29 g was subjected to silica gel column chromatography using a chloroform:methanol (10:1) mixture as the elution solvent to obtain a crystalline fraction (1.0 g, 29%) corresponding to a mixture of 18α- and 18β-trimethyl esters. The fraction was washed repeatedly with methanol to obtain 18α-glycyrrhizinic acid trimethyl ester (0.7 g, 20%) as colorless fine needles, mp 285-288° C. The HRMS, specific rotation ([α] _D ), ¹ H NMR, and ¹³ C NMR data for the obtained trimethyl ester are as follows:

HRMS: m/z 865.4574 (Calcd for [C ₄₅ H ₆₉ O ₁₆ ] ⁺ : 865.4580), 899.4223 (Calcd for [C ₄₅ H ₆₈ O ₁₆ Cl] ^- : 899.4201)

Specific optical rotation [α] _D ²⁷ +47 (c 0.4, EtOH)

^1H NMR (600MHz, pyridine- _d5 ) δ: 0.66 (3H, s, 28-Me), 0.78 (1H, d, J = 11Hz, 5-H), 1.06 (1H, brt, J = 13Hz, 1-H), 1.11 (3H, s, 26-Me), 1.14 (1H, brt, J = 13Hz, 1 5-H), 1.19 (3H, s, 24-Me), 1.26 (1H, dd, J = 13,7Hz, 16-H), 1.30 (6H, s, 27-Me, 30-Me), 1.31 (1H, s, 22-H), 1.35 (3H, s, 23-Me), 1.36 (3H, s, 25-Me), 1.36-1.42 (1H, m, 6-H), 1.38 (1H, d, J = 3Hz, 7-H), 1 .45 (1H, dt, J=14,4Hz, 16-H), 1.52-1.60 (4H, m, 6-H, 7-H, 21-H, 22-H), 1.63 (1H,brd,J=12Hz,19-H),1.76(1H,t,J=12Hz,19-H),1.84(1H,dt,J=14,5Hz,15-H),2 .01 (1H, dt, J=14,4Hz, 21-H), 2.05-2.13 (1H, m, 2-H), 2.20-2.24 (1H, m, 2-H), 2.27 (1H, br d, J = 12Hz, 18-H), 2.35 (1H, s, 9-H), 2.98 (1H, dt, J = 13,3Hz, 1-H), 3.33 (1H, dd, J = 11,4Hz, 3-H), 3.71 (6H, s, 31-Me, 7'-Me), 3.86 (3H, s, 7''-Me), 4.20 (1H, t, J = 9Hz, 2'-H), 4.21 (1H, t, J=9Hz, 2''-H), 4.29 (1H, t, J=9Hz, 3''-H), 4.34 (1H, t, J=9Hz, 3'-H), 4.44 (1H, t, J =9Hz, 4'-H), 4.50 (1H, d, J = 10Hz, 5'-H), 4.53 (1H, d, J = 10Hz, 5''-H), 4.54 (1H, t, J = 1 0Hz, 4''-H), 5.01 (1H, d, J = 7Hz, 1'-H), 5.43 (1H, d, J = 7Hz, 1''-H), 5.70 (1H, s, 12-H)

^13C NMR (150MHz, pyridine- _d5 ) δ: 16.3 ( _C28 ), 16.9 ( _C24 ), 17.3 ( _C25 ), 18.0 ( _C6 ), 19.0 ( _C26 ), 20.9 ( _C30 ), 21.1 ( _C27 ) ), 27.0 (C ₂ ), 27.2 (C ₁₅ ), 28.2 (C ₂₃ ), 29.3 (C ₂₁ ), 32.4 (C ₁₉ ), 34.3 (C ₇ ), 35.9 (C ₁₇ ), 36.3 (C ₁₆ ), 37.4 (C ₁₀ ), 37.9 (C ₂₂ ), 39.8 (C ₁ ), 40.2 (C ₄ ), 40.7 (C ₁₈ ), 43.2 (C ₂₀ ), 44.3 (C ₈ ), 45.4 (C ₁₄ ), 52.2 (C ₇ ''), 52.3 (C ₃₁ ), 52.4 (C ₇ '), 55.8 (C ₅ ), 61.3 (C ₉ ), 73.0 (C ₄ ′), 73.3 (C ₄ ″), 76.9 (C ₂ ″), 77.1 (C ₅ ′), 77.8 (C ₃ ″), 77.9 (C ₃ ′), 78.0 (C ₅ ″), 84.8 (C ₂ ′), 89.7 (C ₃ ), 105.4 (C ₁ '), 107.2 (C ₁ ''), 124.6 (C ₁₂ ), 165.8 (C ₁₃ ), 170.7 (C ₆ '), 170.8 (C ₆ '') 179 (C ₂₉ ), 199.5 (C ₁₁ )

(c) Hydrolysis Reaction 18α-Glycyrrhizic acid trimethyl ester (0.302 g, 0.35 × 10 ^-3 M) was dissolved in 95% ethanol (2 mL), 10% KOH (2 mL, 3.5 × 10 ^-3 M) was added, and the mixture was heated under reflux for 3 hours. The mixture was then added dropwise to a 99.5% ethanol (16 mL) solution containing acetic acid (0.3 mL, 17.5 × 10 ^-3 M) with stirring at room temperature. The resulting precipitate was filtered, yielding potassium 18α-glycyrrhizinate quantitatively as crude crystals. This was then recrystallized twice from 60% aqueous propionic acid to yield potassium 18α-glycyrrhizinate (0.273 g, 91%), colorless fine needles, mp 218-222°C (dec). Treatment of this potassium with 10% hydrochloric acid yielded 18α-glycyrrhizinic acid, a colorless powder, mp 203-206°C, and treatment of the 18α-glycyrrhizinic acid with ammonia yielded ammonium 18α-glycyrrhizinate, colorless, fine needles, mp 204-208°C (dec). HRMS, specific rotation ([α] _D ), ^1H NMR, and ^13C NMR data are as follows. Figures 1 and 2 show ^1H and ^13C NMR charts for 18β-glycyrrhizinic acid and 18α-glycyrrhizinic acid. Figure 1 is a ^1H NMR chart, with the top chart showing 18β-glycyrrhizinic acid and the bottom chart showing 18α-glycyrrhizinic acid. FIG. 2 is a ¹³ C NMR chart, the top of which shows 18β-glycyrrhizinic acid, and the bottom of which shows 18α-glycyrrhizinic acid.

HRMS (18α-ammonium glycyrrhizinate): m/z 821.3984 (Calcd for [C ₄₂ H ₆₁ O ₁₆ N—NH ₃ —H] ⁻ : 821.3965), 845.3931 (Calcd for [C ₄₂ H ₆₂ O ₁₆ N—NH ₃ +Na] ⁺ : 845.3930).

Specific rotation (18α-glycyrrhizinic acid) [α] _D ²⁸ +23 (c 0.5, EtOH)

¹ H NMR (18α-glycyrrhizinic acid: 600 MHz, pyridine-d ₅ ) δ: 0.70 (3H, s, 28-Me), 0.74 (1H, t, J = 10 Hz, 5-H), 1.02-1.06 (1H, m, 1H), 1.05 (3H, s, 26-Me), 1.12 (1H, dd, J = 12, 5 Hz, 15-H), 1.20-1.35 (4H, m, 6-H, 7-H, 16-H, 22-H), 1.27 (3H, s, 25-Me), 1.28 (3H, s, 24-Me), 1.31 (3H, s, 27-Me), 1.42 (3H, s, 23-Me), 1.46 (3H, s, 30-Me), 1.46-1.60 (4H, m, 6-H, 7-H, 1 6-H, 22-H), 1.71 (1H, d, J = 13Hz, 21-H), 1.76-1.85 (2H, m, 15-H, 19-H), 1.97 (1H, t, J = 13Hz, 19-H), 2.13 (1H, dt, J = 14, 14 Hz, 2-H), 2.23 (1H, dt, J = 14, 4Hz, 21-H), 2.32 (1H, brd, J = 9Hz, 2-H, 18-H), 2.33 (1H, s, 12-H), 2.94 (1H, dt, J = 14, 3Hz, 1-H), 3.37 (1H, dd, J = 12,5Hz, 3-H), 4.29 (1H, t, J = 8Hz, 2''-H), 4.32 (1H, td, J = 8,7Hz, 2'-H), 4.36 (1H, t, J = 9Hz, 3''-H), 4.44 (1H, t, J = 9Hz, 3'-H), 4.60 (1H, t, J = 10Hz, 4'-H), 4.64 (1H, d, J = 10Hz, 5'-H), 4.66 (1H, d, J = 10H z, 5''-H), 4.68 (1H, t, J = 10Hz, 4''-H), 5.08 (1H, d, J = 7Hz, 1'-H), 5.47 (1H, d, J = 8Hz, 1''-H), 5.78 (1H, s, 12-H).

^13C NMR (18α-glycyrrhizic acid: 150 MHz, pyridine-d ₅ ) δ: 16.4 (C ₂₈ ), 17.27 (C ₂₄ ), 17.28 (C ₂₅ ), 18.0 (C ₆ ), 18.9 (C ₂₆ ), 21.0 (C ₂₇ ), 21.5 (C ₃₀ ), 27.0 (C ₂ ), 27.3 (C ₁₅ ), 28.4 (C ₂₃ ), 29.7 (C ₂₁ ), 32.8 (C ₁₉ ), 34.3 (C ₇ ), 36.0 (C ₁₇ ), 36.7 (C ₁₆ ), 37.3 (C ₁₀ ), 38.0 (C ₂₂ ), 39.8 (C ₁ ), 40.2 (C ₄ ), 40.9 (C ₁₈ ), 43.1 (C ₂₀ ), 44.3 (C ₈ ), 45.4 (C ₁₄ ), 55.8 (C ₅ ), 61.2 (C ₉ ), 73.4 (C ₄ '), 73.6 (C ₄ ''), 77.2 (C ₂ ''), 77.8 (C ₅ ''), 78.0 (C ₃ ''), 78.1 (C ₃ ''), 78.8 (C ₅ ''), 84.9 (C ₂ ''), 89.5 (C ₃ ), 105.5 (C ₁ ''), 107.3 (C ₁ ''), 124.7 (C ₁₂ ), 166.1 (C ₁₃ ), 172.5 (C ₆ ''), 172.7 (C ₆ '), 181.3 (C ₂₉ ), 199.5 (C ₁₁ ).

As described above, this example confirms that it is possible to provide a method for separating and producing 18α-oleanane-type triterpene diglucuronides with higher purity.

The present invention has industrial applicability as a method for producing stereoisomers, 18α-glycyrrhizinic acid and its analogues, from 18β-glycyrrhizinic acid and its analogues.

Claims (2)

mixing and heating an oleanane-type triterpene diglucuronide with an aqueous alkali hydroxide solution to isomerize the hydrogen at position 18 of the oleanane-type triterpene diglucuronide , thereby obtaining an equilibrium mixture of 18α-isomers and 18β-isomers as an alkali salt precipitate derived from the alkali hydroxide;
treating the equilibrium mixture with an alkylating agent in an aprotic polar solvent to convert it into an 18α-trialkyl ester, which is then separated and purified;
and hydrolyzing the separated and purified 18α-trialkyl ester with an alcoholic aqueous alkali hydroxide solution. The method for producing 18α-oleanane-type triterpene diglucuronides according to claim 1, wherein the oleanane-type triterpene diglucuronide is represented by the following formula: