A preparation method of chrysogenine D

Abstract

The present invention belongs to the field of organic synthesis technology and specifically relates to a method for preparing sucralose D. This invention provides the first chemical preparation method for sucralose D. Using commercially available and structurally simple serine methyl ester as the starting material, this method prepares sucralose D for the first time via a 10-step chemical reaction, providing a reliable material foundation for the further development and utilization of sucralose D.

Description

Technical Field

[0001] The invention belongs to the technical field of organic synthesis, and particularly relates to a method for preparing dapoxetine D. Background Art

[0002] Zhugeline D is a new natural alkaloid compound recently extracted and isolated from the seeds of Zhugeline. In an in vitro screening experiment for antiradiation activity, human umbilical vein epithelial cells (HUVECs) were used as a cell model and irradiated with 60Co-generated gamma rays at a dose of 8 Gy at a dose rate of 0.8 Gy / min. The radiation protection effect of Zhugeline D was calculated by comparing it with a positive control group (523) and a negative control plate. The results show that Zhugeline D has good antiradiation activity in vitro and in vivo. In the in vitro antiradiation activity experiment, the survival rate of umbilical vein endothelial cells in the Zhugeline D group after irradiation was significantly higher than that in the control group. In the in vivo antiradiation activity experiment, the survival rate of mice in the high-dose Zhugeline D group after irradiation was 100%, while the survival rate in the control group was 0%, demonstrating that Zhugeline D has good antiradiation activity in vitro and in vivo. However, current methods for preparing Zhugeline D generally involve extraction and isolation from plants, which is associated with limited raw material sources, poor purity, and complex extraction methods. Summary of the Invention

[0003] The object of the present invention is to provide a method for preparing chrysoprine D. The preparation method provided by the present invention uses serine methyl ester with a simple structure as an initial raw material and prepares chrysoprine D for the first time by a chemical synthesis method.

[0004] In order to achieve the above object, the present invention provides the following technical solutions:

[0005] The present invention provides a method for preparing chrysoprine D, comprising the following steps:

[0006] (1) mixing a compound of formula I, an organic solvent, triethylamine, benzaldehyde, acetic acid, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula II;

[0007] (2) mixing a compound of formula II, an organic solvent, formaldehyde, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula III;

[0008] (3) mixing the compound represented by formula III, an organic solvent, an inorganic strong base, and water to carry out a hydrolysis reaction to obtain a compound represented by formula IV;

[0009] (4) mixing the compound of formula IV, methyl 4-aminobenzoate, an organic solvent, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula V;

[0010] (5) mixing the compound of formula V, an organic solvent, and a reducing agent to perform a reduction reaction to obtain a compound of formula VI;

[0011] (6) In a hydrogen atmosphere, mixing the compound represented by Formula VI, an organic solvent, and a catalyst to carry out a catalytic hydrogenation reaction to obtain a compound represented by Formula VII;

[0012] (7) mixing a compound of formula VII, an organic solvent, N,N-diisopropylethylamine, 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula VIII;

[0013] (8) mixing the compound represented by the structure of Formula VIII, an acid, and an organic solvent to carry out a demethylation reaction to obtain the compound represented by the structure of Formula IX;

[0014] (9) mixing a compound of the structure represented by Formula IX, triphenylphosphine, an organic solvent, and an azo reagent to carry out a substitution reaction to obtain a compound of the structure represented by Formula X;

[0015] (10) mixing a compound represented by formula X, an organic solvent, and hydrochloric acid to carry out a hydrolysis reaction to obtain scutellarine D;

[0016]

[0017] Preferably, in step (1), the molar ratio of benzaldehyde to the compound of formula I is 0.5:1 to 1.5:1;

[0018] The molar ratio of the reducing agent to the compound of the structure shown in formula I is 1:1 to 5:1.

[0019] Preferably, in step (2): the reducing agent comprises sodium cyanoborohydride or sodium borohydride;

[0020] The molar ratio of the formaldehyde to the compound represented by the structure of formula II is 1:1 to 5:1;

[0021] The molar ratio of the reducing agent to the compound of the structure shown in Formula II is 1:1 to 5:1.

[0022] Preferably, in step (3): the inorganic strong base includes sodium hydroxide, lithium hydroxide or potassium hydroxide;

[0023] The molar ratio of the inorganic strong base to the compound represented by formula III is 1:1 to 5:1.

[0024] Preferably, in step (4): the condensation reagent includes carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole;

[0025] The molar ratio of the condensation reagent to the compound represented by formula IV is 1:1 to 5:1.

[0026] Preferably, in step (5): the reducing agent is BH3;

[0027] The molar ratio of the reducing agent to the compound of the structure shown in Formula V is 2:1 to 10:1.

[0028] Preferably, in step (6): the catalyst comprises palladium carbon or palladium hydroxide carbon.

[0029] Preferably, in step (7): the condensation reagent includes 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate, carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole;

[0030] The molar ratio of the condensation reagent to the structural compound represented by formula VII is 1:1 to 5:1.

[0031] Preferably, in step (8): the acid comprises HCl or trifluoroacetic acid.

[0032] Preferably, in step (9): the azo reagent includes diethyl azodicarboxylate or diisopropyl azodicarboxylate;

[0033] The molar ratio of the azo reagent to the compound having the structure represented by formula IX is 1:1 to 5:1.

[0034] The present invention provides a preparation method of scutellarine D, comprising the following steps: (1) mixing a compound of a structure shown in formula I, an organic solvent, triethylamine, benzaldehyde, acetic acid and a reducing agent for a reductive amination reaction to obtain a compound of a structure shown in formula II; (2) mixing a compound of a structure shown in formula II, an organic solvent, formaldehyde and a reducing agent for a reductive amination reaction to obtain a compound of a structure shown in formula III; (3) mixing a compound of a structure shown in formula III, an organic solvent, an inorganic strong base and water for a hydrolysis reaction to obtain a compound of a structure shown in formula IV; (4) mixing a compound of a structure shown in formula IV, methyl 4-aminobenzoate, an organic solvent and a condensation reagent for a condensation reaction to obtain a compound of a structure shown in formula V; and (5) mixing a compound of a structure shown in formula V, an organic solvent and a reducing agent for a reduction reaction to obtain a compound of a structure shown in formula VI. (6) in a hydrogen atmosphere, the compound of formula VI, an organic solvent and a catalyst are mixed to carry out a catalytic hydrogenation reaction to obtain a compound of formula VII; (7) the compound of formula VII, an organic solvent, N,N-diisopropylethylamine, 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid and a condensation reagent are mixed to carry out a condensation reaction to obtain a compound of formula VIII; (8) the compound of formula VIII, an acid and an organic solvent are mixed to carry out a demethylation reaction to obtain a compound of formula IX; (9) the compound of formula IX, triphenylphosphine, an organic solvent and an azo reagent are mixed to carry out a substitution reaction to obtain a compound of formula X; (10) the compound of formula X, an organic solvent and hydrochloric acid are mixed to carry out a hydrolysis reaction to obtain chrysoprine D. The present invention provides a chemical preparation method of chrysoprine D for the first time. The present invention uses commercially available chemical serine methyl ester with a simple structure as the starting raw material, and prepares scutellarine D through 10 steps of chemical reaction for the first time, thus providing a reliable material basis for the in-depth development and utilization of scutellarine D. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 The synthetic flow chart of scutellarine D provided in Example 1 of the present invention;

[0036] Figure 2 The scutellarine D prepared in Example 1 of the present invention 1 H NMR spectrum;

[0037] Figure 3 The scutellarine D prepared in Example 1 of the present invention 13 C NMR spectrum;

[0038] Figure 4 This is a high-resolution mass spectrum of tricholamine D prepared in Example 1 of the present invention. DETAILED DESCRIPTION

[0039] The present invention provides a method for preparing chrysoprine D, comprising the following steps:

[0040] (1) mixing a compound of formula I, an organic solvent, triethylamine, benzaldehyde, acetic acid, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula II;

[0041] (2) mixing a compound of formula II, an organic solvent, formaldehyde, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula III;

[0042] (3) mixing the compound represented by formula III, an organic solvent, an inorganic strong base, and water to carry out a hydrolysis reaction to obtain a compound represented by formula IV;

[0043] (4) mixing the compound of formula IV, methyl 4-aminobenzoate, an organic solvent, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula V;

[0044] (5) mixing the compound of formula V, an organic solvent, and a reducing agent to perform a reduction reaction to obtain a compound of formula VI;

[0045] (6) In a hydrogen atmosphere, mixing the compound represented by Formula VI, an organic solvent, and a catalyst to carry out a catalytic hydrogenation reaction to obtain a compound represented by Formula VII;

[0046] (7) mixing a compound of formula VII, an organic solvent, N,N-diisopropylethylamine, 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula VIII;

[0047] (8) mixing the compound represented by the structure of Formula VIII, an acid, and an organic solvent to carry out a demethylation reaction to obtain the compound represented by the structure of Formula IX;

[0048] (9) mixing a compound of the structure represented by Formula IX, triphenylphosphine, an organic solvent, and an azo reagent to carry out a substitution reaction to obtain a compound of the structure represented by Formula X;

[0049] (10) mixing a compound represented by formula X, an organic solvent, and hydrochloric acid to carry out a hydrolysis reaction to obtain scutellarine D;

[0050]

[0051] In the present invention, unless otherwise specified, all preparation raw materials / components are commercially available products well known to those skilled in the art.

[0052] The chemical formula structure of the lycine D prepared by the present invention is as follows:

[0053]

[0054] The preparation method provided by the present invention is Figure 1 The preparation process is as shown in the following. Figure 1 The preparation process shown is a detailed description of the preparation method provided by the present invention.

[0055] In the present invention, a compound having a structure represented by Formula I, an organic solvent (hereinafter referred to as the first organic solvent), triethylamine (TEA), benzaldehyde, acetic acid, and a reducing agent (hereinafter referred to as the first reducing agent) are mixed (hereinafter referred to as the first mixture) to perform a reductive amination reaction (hereinafter referred to as the first reductive amination reaction) to obtain a compound having a structure represented by Formula II.

[0056] In the present invention, the compound of Formula I is preferably subjected to the first reductive amination reaction using the hydrochloride salt of Formula I. The first organic solvent is preferably methanol. The first reducing agent is preferably NaBH4. The molar ratio of benzaldehyde to the compound of Formula I is preferably 0.5:1 to 1.5:1, more preferably 0.9:1 to 1.1:1. The molar ratio of the compound of Formula I to TEA is preferably 1:1 to 1.1. The molar ratio of the compound of Formula I to acetic acid is preferably 1:1.5 to 2. The molar ratio of the first reducing agent to the compound of Formula I is 1:1 to 5:1, preferably 2:1 to 2.1:1. The present invention has no specific requirements for the amount of the first organic solvent used, as long as it ensures the smooth progress of the first reductive amination reaction. In the present invention, TEA is used to neutralize the hydrochloric acid in the hydrochloride salt of Formula I, freeing the amino groups in the substrate and facilitating the reaction. The first mixing preferably comprises: dissolving the compound of Formula 1 in hydrochloric acid in a first organic solvent, then adding TEA, and stirring at room temperature for 10 minutes to obtain a first mixture; then sequentially adding benzaldehyde and acetic acid to the mixture, and stirring at room temperature for 30 minutes to obtain a second mixture. The second mixture is cooled to 0°C and mixed with the first reducing agent. The temperature of the first reductive amination reaction is preferably room temperature. The first reductive amination reaction is carried out under stirring. After the first reductive amination reaction, a first reductive amination reaction liquid is obtained. In the present invention, the first reductive amination reaction liquid is preferably post-treated. The post-treatment preferably comprises: solid-liquid separation of the first reductive amination reaction liquid, and concentrating the resulting liquid to obtain an oily product, which is the compound of Formula II. The compound of Formula II can be used directly in the following reaction without further purification. The solid-liquid separation is preferably filtration. The concentration is preferably concentrated under reduced pressure.

[0057] After obtaining the compound of formula II, the present invention mixes the compound of formula II, an organic solvent (hereinafter referred to as the second organic solvent), formaldehyde, and a reducing agent (hereinafter referred to as the second reducing agent) (hereinafter referred to as the second mixture) and performs a reductive amination reaction (hereinafter referred to as the second reductive amination reaction) to obtain a compound of formula III.

[0058] In the present invention, the second organic solvent is preferably methanol. The formaldehyde is preferably used in the form of an aqueous formaldehyde solution. The formaldehyde content of the aqueous formaldehyde solution is preferably 37% by weight. The second reducing agent preferably comprises sodium cyanoborohydride or sodium borohydride, more preferably sodium cyanoborohydride. The molar ratio of formaldehyde to the compound represented by Formula II is preferably 1:1 to 5:1, more preferably 1.4:1 to 1.5:1. The molar ratio of the second reducing agent to the compound represented by Formula II is preferably 1:1 to 5:1, more preferably 1.5:1 to 3:1, and most preferably 2:1. The present invention does not require a specific amount of the second organic solvent; it only requires that the second reductive amination reaction proceeds smoothly. In the present invention, the second mixing step is preferably: dissolving the compound represented by Formula II in the second organic solvent, then adding the aqueous formaldehyde solution under stirring, exchanging and mixing the resulting mixture at room temperature for 30 minutes, cooling to 0°C, and mixing with the second reducing agent. The temperature of the second reductive amination reaction is preferably room temperature. The second reductive amination reaction is carried out under stirring. The second reductive amination reaction is preferably quenched with an aqueous sodium bicarbonate solution. After the second reductive amination reaction is completed, a second reductive amination reaction liquid is obtained. The present invention preferably performs a post-treatment on the second reductive amination reaction liquid. The post-treatment preferably comprises: extracting the second reductive amination reaction liquid with ethyl acetate (EA) to obtain an organic phase; drying the organic phase and concentrating it to obtain an oily product, which is the compound represented by Formula III. The compound represented by Formula III is used directly in the following reaction without further purification. The extraction is preferably performed three times. The present invention preferably combines the extracted organic phases obtained each time and then dries them. The reagent used for drying is preferably sodium sulfate. The concentration is preferably concentrated under reduced pressure.

[0059] After obtaining the compound of formula III, the present invention mixes the compound of formula III, an organic solvent (hereinafter referred to as the third organic solvent), an inorganic strong base and water (hereinafter referred to as the third mixture) and performs a hydrolysis reaction (hereinafter referred to as the first hydrolysis reaction) to obtain a compound of formula IV.

[0060] In the present invention, the third organic solvent is preferably methanol and tetrahydrofuran (THF). The inorganic strong base is preferably sodium hydroxide, lithium hydroxide, or potassium hydroxide, more preferably lithium hydroxide. The molar ratio of the inorganic strong base to the compound represented by Formula III is preferably 1:1 to 5:1, more preferably 2:1 to 2.1:1. The present invention does not have any specific requirements for the amounts of the third organic solvent and water, as long as they ensure the smooth progress of the first hydrolysis reaction. In the present invention, the third mixing preferably comprises: dissolving the compound represented by Formula III in the third organic solvent; dissolving the inorganic strong base in water; and then mixing the organic and aqueous phases. The temperature of the first hydrolysis reaction is preferably room temperature. After the first hydrolysis reaction is completed, a first hydrolysis reaction liquid is obtained. In the present invention, the first hydrolysis reaction liquid is preferably post-treated. The post-treatment preferably comprises: concentrating the first hydrolysis reaction liquid to obtain a concentrate; acidifying the concentrate with hydrochloric acid to a pH of 6 to 7 to obtain an acidified reaction liquid; and purifying the acidified reaction liquid by reverse-phase flash chromatography. The resulting purified reaction liquid is then desolventized to obtain a pure compound represented by Formula IV. The concentration is preferably concentrated under reduced pressure. The molar concentration of the hydrochloric acid is preferably 1 M. The mobile phase used in the reversed-phase flash chromatography is preferably an aqueous solution of CH3CN and trifluoroacetic acid (TFA), and the mass content of TFA in the aqueous TFA solution is preferably 0.05%. The elution procedure is preferably a continuous gradient elution, and the continuous gradient elution procedure is preferably: the volume content of CH3CN in the mobile phase increases linearly from 5% to 35% over 40 minutes.

[0061] After obtaining the compound of the structure shown in Formula IV, the present invention mixes the compound of the structure shown in Formula IV, methyl 4-aminobenzoate, an organic solvent (hereinafter referred to as the fourth organic solvent) and a condensation reagent (hereinafter referred to as the first condensation reagent) (hereinafter referred to as the fourth mixing) to carry out a condensation reaction (hereinafter referred to as the first condensation reaction) to obtain a compound of the structure shown in Formula V.

[0062] In the present invention, the fourth organic solvent is preferably pyridine. The first condensation reagent preferably includes carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole, more preferably carbodiimide hydrochloride (EDCI). The molar ratio of the compound of the structure shown in Formula IV and methyl 4-aminobenzoate is preferably 1:1 to 1.2. The molar ratio of the first condensation reagent to the compound of the structure shown in Formula IV is preferably 1:1 to 5:1, more preferably 2:1 to 2.1:1. The present invention has no special requirements for the amount of the fourth organic solvent, as long as the first condensation reaction is carried out smoothly. The fourth mixing preferably includes: dissolving the compound of the structure shown in Formula IV and methyl 4-aminobenzoate in a fourth organic solvent, and then mixing with the first condensation reagent under stirring. The first condensation reaction is carried out at room temperature, and the first condensation reaction is carried out under stirring. After the first condensation reaction is completed, a first condensation reaction liquid is obtained. In the present invention, the first condensation reaction liquid is preferably subjected to post-treatment, and the post-treatment preferably includes: concentrating the first condensation reaction liquid to obtain a concentrate; purifying the concentrate by reverse-phase flash chromatography, removing the solvent from the obtained purified reaction liquid, and obtaining a pure product of the compound represented by Formula V. The concentration is preferably concentrated under reduced pressure. The mobile phase used in the reverse-phase flash chromatography is preferably an aqueous solution of CH3CN and formic acid (FA), and the mass content of FA in the aqueous solution is preferably 0.1%. The elution procedure is preferably continuous gradient elution, and the continuous gradient elution procedure is preferably: the volume content of CH3CN in the mobile phase is linearly increased from 5% to 65% over 40 minutes.

[0063] After obtaining the compound of formula V, the present invention mixes the compound of formula V, an organic solvent (hereinafter referred to as the fifth organic solvent) and a reducing agent (hereinafter referred to as the third reducing agent) (hereinafter referred to as the fifth mixture) for reduction reaction to obtain a compound of formula VI.

[0064] In the present invention, the fifth organic solvent is preferably THF. The third reducing agent is preferably BH3. The molar ratio of the third reducing agent to the compound of the structure represented by Formula V is preferably 2:1 to 10:1, more preferably 4:1 to 4.5:1. The present invention has no special requirements for the amount of the fifth organic solvent, as long as the first reduction reaction is carried out smoothly. The fifth mixing preferably includes: dissolving the compound of the structure represented by Formula V in part of the fifth organic solvent to obtain a solution of the compound of the structure represented by Formula V; dissolving the third reducing agent in the remaining fifth organic solvent to obtain a third reducing agent solution, and the molar concentration of the third reducing agent solution is preferably 1M. At 0°C, the third reducing agent solution is added dropwise to the solution of the compound of the structure represented by Formula V. The temperature of the first reduction reaction is preferably 40°C, the insulation time is preferably 16h, and the reduction reaction is carried out under stirring. The present invention preferably uses methanol to quench the reaction at 0°C. After the reduction reaction is completed, a reduction reaction solution is obtained. The present invention preferably post-treats the reduction reaction solution, which preferably includes: concentrating the reduction reaction solution to obtain a concentrate; purifying the concentrate by reverse-phase flash chromatography, removing the solvent from the purified reaction solution, and obtaining a pure compound of the structure represented by Formula VI. The concentration is preferably concentrated under reduced pressure. The mobile phase used in the reverse-phase flash chromatography is preferably an aqueous solution of CH3CN and formic acid (FA), wherein the mass content of FA in the aqueous solution is preferably 0.1%. The elution procedure is preferably continuous gradient elution, wherein the volume content of CH3CN in the mobile phase is preferably linearly increased from 5% to 65% over 40 minutes.

[0065] After obtaining the compound of formula VI, the present invention performs a catalytic hydrogenation reaction on the compound of formula VI, an organic solvent (hereinafter referred to as the sixth organic solvent) and a catalyst mixture (hereinafter referred to as the sixth mixture) in a hydrogen atmosphere to obtain a compound of formula VII.

[0066] In the present invention, the sixth organic solvent is preferably methanol. The catalyst preferably comprises palladium-carbon or palladium hydroxide-carbon, more preferably palladium hydroxide-carbon, wherein the palladium hydroxide-carbon preferably contains 10% by mass of palladium hydroxide. The mass ratio of the compound represented by Formula VI to the catalyst is preferably 8.9:4.5. The pressure of the hydrogen is preferably 3 atmospheres. The present invention has no specific requirements for the amount of the sixth organic solvent used, as long as the catalytic hydrogenation reaction proceeds smoothly. The sixth mixing preferably comprises: dissolving the compound represented by Formula VI in the sixth organic solvent, and then adding the catalyst. The temperature of the catalytic hydrogenation reaction is preferably room temperature, the duration is preferably 5 hours, and the catalytic hydrogenation reaction is carried out under stirring. After the catalytic hydrogenation reaction, a catalytic hydrogenation reaction liquid is obtained. In the present invention, the catalytic hydrogenation reaction liquid is preferably post-treated, and the post-treatment preferably comprises: filtering the catalytic hydrogenation reaction liquid through a diatomaceous earth pad, mixing the obtained filtrate with a hydrochloric acid solution, and then concentrating. The hydrochloric acid solution is preferably a solution of HCl in 1,4-dioxane, and the molar concentration of the hydrochloric acid solution is preferably 4M. The concentration is preferably concentrated under reduced pressure.

[0067] After preparing the compound of the structure shown in formula VII, the present invention mixes the compound of the structure shown in formula VII, an organic solvent (hereinafter referred to as the seventh organic solvent), N,N-diisopropylethylamine (DIEA), 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid and a condensation reagent (hereinafter referred to as the second condensation reagent) (hereinafter referred to as the seventh mixture) to carry out a condensation reaction (hereinafter referred to as the second condensation reaction) to obtain a compound of the structure shown in formula VIII.

[0068] In the present invention, the seventh organic solvent is preferably dimethylformamide. The second condensation reagent preferably includes 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate (HATU), carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole, more preferably HATU. The molar ratio of the compound of the structure shown in Formula VII to DIEA is preferably 1:4 to 4.2. The molar ratio of the compound of the structure shown in Formula VII to 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid is preferably 1:1 to 1.1. The molar ratio of the second condensation reagent to the compound of the structure shown in Formula VII is preferably 1:1 to 5:1, more preferably 1:1 to 2:1. The present invention has no special requirements for the amount of the seventh organic solvent used, as long as it ensures that the second condensation reaction proceeds smoothly. The seventh mixing step preferably comprises dissolving the compound represented by Formula VII in a seventh organic solvent, and then sequentially mixing with DIEA, 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid, and a second condensation reagent under stirring. The second condensation reaction is conducted at room temperature and under stirring. After the second condensation reaction, a second condensation reaction liquid is obtained. In the present invention, the second condensation reaction liquid is preferably post-treated, and the post-treatment preferably comprises purifying the second condensation reaction liquid by reverse-phase flash chromatography, removing the solvent from the purified reaction liquid, and obtaining a pure compound represented by Formula VIII. The mobile phase used in the reverse-phase flash chromatography is preferably an aqueous solution of CH3CN and formic acid (FA), and the FA content in the aqueous solution is preferably 0.1%. The elution procedure is preferably a continuous gradient elution, and the continuous gradient elution procedure is preferably such that the CH3CN content in the mobile phase increases linearly from 5% to 35% by volume over 40 minutes. The concentration step is preferably concentrated under reduced pressure.

[0069] After preparing the compound of formula VIII, the present invention mixes the compound of formula VIII, an acid and an organic solvent (hereinafter referred to as the eighth organic solvent) (hereinafter referred to as the eighth mixture) for demethylation to obtain a compound of formula IX.

[0070] In the present invention, the eighth organic solvent is preferably 1,4-dioxane. The acid preferably comprises HCl or trifluoroacetic acid, more preferably HCl. When the acid is preferably HCl, the product obtained from the second hydrolysis reaction is the hydrochloride salt of the compound represented by structure IX. In the present invention, the hydrochloride salt of the compound represented by structure IX is preferably used for the substitution reaction. The eighth mixing preferably comprises dissolving the acid in the eighth organic solvent to obtain an acid solution, wherein the molar concentration of the acid solution is preferably 4 M. The compound represented by structure VIII is dissolved in the acid solution. The ratio of the mass of the compound represented by structure VIII to the volume of the acid solution is preferably 5.3 g:200 mL. The temperature of the demethylation reaction is preferably room temperature. After the demethylation reaction, a demethylation reaction solution is obtained. In the present invention, the demethylation reaction solution is preferably post-treated, and the post-treatment preferably comprises: concentrating the demethylation reaction solution to obtain a concentrate; and dissolving the concentrate in an alcohol solvent for recrystallization. The concentration is preferably concentrated under reduced pressure. The alcohol solvent is preferably ethanol.

[0071] After obtaining the compound represented by Formula IX, the present invention mixes the compound represented by Formula IX, triphenylphosphine, an organic solvent (hereinafter referred to as the ninth organic solvent) and an azo reagent (hereinafter referred to as the ninth mixture) to perform a substitution reaction to obtain a compound represented by Formula X.

[0072] In the present invention, the ninth organic solvent is preferably THF and N,N-dimethylformamide (DMF); the volume ratio of THF to DMF is preferably 10:3. The azo reagent preferably includes diethyl azodicarboxylate or diisopropyl azodicarboxylate (DIAD), more preferably DIAD. The molar ratio of the compound represented by Formula IX to triphenylphosphine is preferably 11:3. The molar ratio of the azo reagent to the compound represented by Formula IX is preferably 1:1 to 5:1, preferably 2:1. The present invention does not require a specific amount of the ninth organic solvent, as long as it ensures the substitution reaction proceeds smoothly. The ninth organic solvent preferably comprises: dissolving the compound represented by Formula IX and triphenylphosphine in a portion of the ninth organic solvent to obtain a first mixture; dissolving the azo reagent in the remaining ninth organic solvent to obtain a second mixture; and then adding the second mixture to the first mixture under stirring. The portion of the ninth organic solvent is preferably a portion of THF and all of DMF; the remaining ninth organic solvent is preferably raw THF. The substitution reaction is carried out under a protective gas atmosphere, preferably nitrogen. The temperature of the substitution reaction is preferably 50°C, and the substitution reaction is carried out under stirring. After the substitution reaction is completed, a substitution reaction liquid is obtained. The present invention preferably performs post-treatment on the substitution reaction liquid, and the post-treatment preferably includes: concentrating the substitution reaction liquid and purifying it by reverse phase flash chromatography, removing the solvent from the purified reaction liquid to obtain a pure product of the compound with the structure shown in Formula X. The mobile phase used in the reverse phase flash chromatography is preferably CH3CN and formic acid (FA) aqueous solution, and the mass content of FA in the FA aqueous solution is preferably 0.1%. The elution procedure is preferably continuous gradient elution, and the procedure of the continuous gradient elution is preferably: the volume content of CH3CN in the mobile phase increases linearly from 5% to 70% in 40 minutes. The concentration is preferably concentrated under reduced pressure.

[0073] After obtaining the compound represented by formula X, the present invention mixes the compound represented by formula X, an organic solvent (hereinafter referred to as the tenth organic solvent) and hydrochloric acid (hereinafter referred to as the tenth organic mixture) and performs a hydrolysis reaction (hereinafter referred to as the second hydrolysis reaction) to obtain sucralose D.

[0074] In the present invention, the tenth organic solvent is preferably acetonitrile (ACN). The molar concentration of hydrochloric acid (HCl aqueous solution) is preferably 0.5M. The ratio of the mass of the compound of the structure shown in Formula X to the volume of the hydrochloric acid is preferably 4g:4mL. The present invention has no special requirements for the amount of the tenth organic solvent, as long as the second hydrolysis reaction is carried out smoothly. In the present invention, the tenth mixing preferably includes: dissolving the compound of the structure shown in Formula X in the tenth organic solvent and then mixing it with hydrochloric acid. The temperature of the second hydrolysis reaction is preferably room temperature. After the second hydrolysis reaction is completed, a second hydrolysis reaction liquid is obtained. The present invention preferably post-treats the second hydrolysis reaction liquid, and the post-treatment preferably includes: concentrating the second hydrolysis reaction liquid and performing a first purification by reverse phase flash chromatography to obtain a crude product; performing a second purification by Prep-HPLC on the crude product, and drying the purified product to obtain a pure product of scutellarin D. During the first purification: the catalog number of the column used in the reverse phase flash chromatography is SO230120-2, C18, 120g, 20~45μm, Batch: BP0002P2503; the mobile phase is preferably CH3CN and aqueous HCl, with the HCl content in the aqueous HCl solution preferably being 0.1% by weight. The elution procedure is preferably continuous gradient elution, with the CH3CN content in the mobile phase preferably increasing linearly from 5% to 55% by volume over 40 minutes. Detectors: UV 220nm and 254nm. The second purification is performed using Prep-HPLC, with mobile phase A preferably being an aqueous FA solution with an FA content of 0.1% by weight, and mobile phase B preferably being ACN. The elution procedure is preferably continuous gradient elution, with the ACN content in the mobile phase preferably increasing linearly from 5% to 50% by volume over 30 minutes. The drying step is preferably lyophilization.

[0075] In order to further illustrate the present invention, the technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0076] The following examples adopt Figure 1 The preparation was carried out according to the flow chart shown.

[0077] Example 1

[0078] a) To a solution of (2R)-methyl 2-amino-3-hydroxypropanoate hydrochloride (the hydrochloride salt of the compound of Formula I, 50 g, 322.331 mmol, 1 equivalent) in methanol (1 L) was added TEA (34.90 g, 344.886 mmol, 1.07 equivalent) with stirring. The resulting mixture was stirred at room temperature for an additional 10 minutes. Benzaldehyde (37.63 g, 354.564 mmol, 1.1 equivalent) and acetic acid (38.71 g, 644.662 mmol, 2 equivalents) were added to the above mixture. The resulting mixture was stirred at room temperature for an additional 30 minutes. NaBH4 (25 g, 660.851 mmol, 2.05 equivalents) was added to the above mixture at 0°C. The resulting mixture was stirred at room temperature overnight. The resulting mixture was filtered. The filtrate was concentrated under reduced pressure to obtain (2R)-2-(benzylamino)-3-hydroxypropionic acid methyl ester (compound represented by formula II, 40 g, 45.54%) as a yellow oil. The product was used directly in the next step without further purification. MS (ESI) calculation. For C 11 H 15 NO3209.11 m / z, found 210.10 [m+H] + .

[0079] b) To a solution of methyl (2R)-2-(benzylamino)-3-hydroxypropanoate (40 g, 191.163 mmol, 1 equivalent) in methanol (500 mL) was added HCHO (23 g, 283.666 mmol, 1.48 equivalent, 37% in H₂O) at room temperature. The resulting mixture was stirred at room temperature for an additional 30 minutes. NaBH₃CN (24 g, 381.922 mmol, 2.00 equivalent) was added to the mixture at 0°C. The resulting mixture was stirred at room temperature overnight. The reaction was quenched with NaHCO₃ / H₂O (200 mL). The resulting mixture was extracted with EA (3 x 500 mL) and dried over Na₂SO₄. After filtration, the filtrate was concentrated under reduced pressure to afford methyl (2R)-2-[benzyl(methyl)amino]-3-hydroxypropanoate (compound represented by Formula III, 30 g, 70.29%) as a yellow oil. The product was used directly in the next step without further purification. MS (ESI) calculations. 12 H 17 NO3, 223.12 m / z, found 224.15 [m+H] + .

[0080] c) To a stirred solution of (2R)-2-[benzyl(methyl)amino]-3-hydroxypropionic acid methyl ester (30 g, 134.365 mmol, 1 equivalent) in methanol (200 mL) and THF (200 mL) was added a solution of LiOH (6.5 g, 271.399 mmol, 2.02 equivalents) in water (200 mL). The resulting mixture was stirred at room temperature overnight. The resulting mixture was concentrated under reduced pressure. The mixture was acidified to a pH of 6-7 with aqueous HCl (1 M). The residue was purified by reverse phase flash chromatography under the following conditions: chromatographic column, mobile phase CH3CN:H2O (0.05% TFA) = 5% increased to 35% within 40 minutes; after concentration, a white solid (2R)-2-[benzyl(methyl)amino]-3-hydroxypropionic acid (compound represented by the structure of Formula IV, 27 g, 96.03%) was obtained. MS (ESI) calculation. For C 11 H 15 NO3209.11 m / z, found 210.15 [m+H] + .

[0081] d) To a stirred solution of (2R)-2-[benzyl(methyl)amino]-3-hydroxypropionic acid (27g, 129.035mmol, 1 equivalent) and methyl 4-aminobenzoate (21.6g, 142.890mmol, 1.11 equivalents) in pyridine (400mL) was added EDCI (50g, 260.824mmol, 2.02 equivalents). The resulting mixture was stirred at room temperature for another 3 hours. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase flash chromatography under the following conditions: mobile phase, CH3CN:H2O (0.1% FA) = 5% increased to 65% in 40 minutes. After concentration, brown oily 4-[(2R)-2-[benzyl(methyl)amino]-3-hydroxypropionamide]benzoic acid methyl ester (structural compound shown in Formula V, 24g, 54.32%) was obtained. MS (ESI) calculation. For C 19 H 22 N2O4 342.16 m / z, found 343.10 [m+H] + .

[0082] e) To a stirred solution of methyl 4-[(2R)-2-[benzyl(methyl)amino]-3-hydroxypropionamide]benzoate (24 g, 70.094 mmol, 1 equivalent) in THF (50 mL) was added borane (300 mL, 300.0 mmol, 4.28 equivalents, 1 M in THF) dropwise at 0°C. The resulting mixture was stirred at 40°C for an additional 16 hours. The reaction was quenched with MeOH (100 mL) at 0°C. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase flash chromatography under the following conditions: mobile phase, CH3CN:H2O (0.1% FA) = 5% increasing to 65% over 40 minutes. After concentration, methyl 4-{[(2S)-2-[benzyl(methyl)amino]-3-hydroxypropyl]amino}benzoate (compound represented by formula VI, 8.9 g, 38.66%) was obtained as a yellow solid. MS (ESI) calculations. 19 H 24 N2O3, 328.18 m / z, found 329.20 [m+H] + .

[0083] f) Pd(OH)2 / C (4.5 g, 10% w / w) was added to a solution of methyl 4-{[(2S)-2-[benzyl(methyl)amino]-3-hydroxypropyl]amino}benzoate (8.9 g, 27.100 mmol, 1 equivalent) in MeOH (500 mL). The mixture was stirred at room temperature for 5 hours under a hydrogen atmosphere (3 atmospheres). The reaction mixture was filtered through a pad of celite. HCl (10 mL, 4 M in 1,4-dioxane) was added to the solution and concentrated under reduced pressure to give methyl 4-{[(2S)-3-hydroxy-2-(methylamino)propyl]amino}benzoate HCl (structural compound shown in Formula VII, 6.7 g, 85%) as an off-white solid. MS (ESI) calculation. For C 12 H 18 N2O3, 238.13 m / z, found 239.10 [m+H] + .

[0084] g) To a stirred solution of methyl 4-{[(2S)-3-hydroxy-2-(methylamino)propyl]amino}benzoate HCl (6.7 g, 24.452 mmol, 1 eq) in dimethylformamide (100 mL) was added N,N-diisopropylethylamine (DIEA) (13 g, 100.583 mmol, 4.11 eq), 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid (4.3 g, 25.275 mmol, 1.03 eq), and HATU (12.83 g, 33.740 mmol, 1.2 eq). The resulting mixture was stirred at room temperature for an additional 3 hours. The residue was purified by reverse phase flash chromatography under the following conditions: mobile phase: CH3CN:H2O (0.1% FA) = 5%, increasing to 35% over 40 minutes. After concentration, methyl 4-{[(2S)-2-[1-(4-amino-6-methoxy-1,3,5-triazin-2-yl)-N-methylformamido]-3-hydroxypropyl]amino}benzoate (compound represented by formula VIII, 5.3 g, 48.28%) was obtained as a yellow oil. MS (ESI) calculation. For C 17 H 22 N6O5 390.17 m / z, found 391.20 [m+H] + .

[0085] h) A solution of methyl 4-{[(2S)-2-[1-(4-amino-6-methoxy-1,3,5-triazine-2-yl)-N-methylformamido]-3-hydroxypropyl]amino}benzoate (5.3 g, 13.576 mmol, 1 equivalent) in HCl (200 mL, 4 M in 1,4-dioxane) was stirred at room temperature overnight. The resulting mixture was concentrated under reduced pressure. The crude product was recrystallized from Et2O (200 mL) to give methyl 4-{[(2S)-2-[1-(4-amino-6-hydroxy-1,3,5-triazine-2-yl)-N-methylformamido]-3-hydroxypropyl]amino}benzoate HCl (compound represented by formula IX, 4.3 g, 84.16%) as a yellow solid. MS (ESI) calculated. For C 16 H 20 N6O5 376.15 m / z, found 377.20 [m+H] + .

[0086] i) To a stirred solution of methyl 4-{[(2S)-2-[1-(4-amino-6-hydroxy-1,3,5-triazin-2-yl)-N-methylformamido]-3-hydroxypropyl]amino}benzoate HCl (4.3 g, 11.425 mmol, 1 eq.) and triphenylphosphine (5.99 g, 22.850 mmol, 2 eq.) in THF (50 mL) and DMF (15 mL) at 0° C. under an N atmosphere was added a solution of DIAD (4.62 g, 22.850 mmol, 2 eq.) in tetrahydrofuran (5 mL). The resulting mixture was stirred at 50° C. overnight. The resulting mixture was concentrated under reduced pressure. The residue was purified by reverse phase flash chromatography under the following conditions: mobile phase CH 3 CN:H 2 O (0.1% FA) = 5% increasing to 70% over 40 minutes. After concentration, methyl 4-({[(7S)-8-methyl-4,9-dioxo-2-[(triphenyl-λ5-phosphinyl)amino]-6H,7H-pyrazino[1,2-a][1,3,5]triazin-7-yl]methyl}amino)benzoate (compound represented by formula X, 4 g, 56.59%) was obtained as a yellow oil. MS (ESI) calculation. For C 34 H 31 N6O4P, 618.21 m / z, found 619.25 [m+H] + .

[0087] j) To a stirred solution of methyl 4-({[(7S)-8-methyl-4,9-dioxo-2-[(triphenyl-λ5-phosphinylidene)amino]-6H,7H-pyrazino[1,2-a][1,3,5]triazin-7-yl]methyl}amino)benzoate (4 g, 6.466 mmol, 1 eq) in ACN (40 mL) was added hydrogen chloride (4 mL, 0.5 M in H2O). The resulting mixture was stirred at room temperature overnight. The residue was purified by reverse phase flash chromatography under the following conditions (column, catalog number: SO230120-2, C 18 , 120g, 20~45μm, Batch: BP0002P2503; mobile phase, CH3CN:H2O (0.1% HCl) = 5%, increased to 55% within 40 minutes; detector, UV 220nm and 254nm) to obtain a crude product. The crude product was purified by Prep-HPLC under the following conditions: mobile phase A: water (0.1% FA), mobile phase B: ACN, ACN: water (0.1% FA) = 5%, increased to 50% within 30 minutes. After lyophilization, 4-({[(7S)-2-amino-8-methyl-4,9-dioxo-6H,7H-pyrazino[1,2-a][1,3,5]triazin-7-yl]methyl}amino)benzoic acid methyl ester (Zhuogeline D, 616.6 mg, 26.29%) was obtained as an orange solid. MS (ESI) calculation. For C 16 H 18 N6O4 358.14 m / z, found 359.1410 [m+H] + . 1 HNMR (600MHz, DMSO-d6) δ (ppm): 7.75 (s, 1H), 7.65 (d, J = 8.8Hz, 2H), 7.51 (s, 1H), 6.72 (t, J = 6.2Hz, 1H), 6.60 (d, J = 8.8Hz, 2H), 4.39 (d, J=13.2Hz,1H),3.92(d,J=5.2Hz,1H),3.80–3.75(m,1H),3.74(s,3H),3.40(dd,J=12.7,7.2Hz,1H),3.36(d,J=6.8Hz,1H),3.07(s,3H). 13 C NMR (151 MHz, DMSO) δ 166.73, 166.34, 155.78, 154.43, 154.11, 152.55, 131.40, 117.00, 111.52, 54.40, 51.71, 42.88, 41.05, 35.13. Figure 2 and Figure 3 As shown, the overall yield was 0.33%.

[0088] Although the above embodiment provides a detailed description of the present invention, it is only a part of the embodiments of the present invention, not all of the embodiments. Other embodiments can be obtained based on this embodiment without creativity, and these embodiments all fall within the scope of protection of the present invention.

Claims

1. A method for preparing chrysoprine D, characterized in that: The following steps are involved: (1) mixing a compound of formula I, an organic solvent, triethylamine, benzaldehyde, acetic acid, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula II; (2) mixing a compound of formula II, an organic solvent, formaldehyde, and a reducing agent to perform a reductive amination reaction to obtain a compound of formula III; (3) mixing the compound represented by formula III, an organic solvent, an inorganic strong base, and water to carry out a hydrolysis reaction to obtain a compound represented by formula IV; (4) mixing the compound of formula IV, methyl 4-aminobenzoate, an organic solvent, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula V; (5) mixing the compound of formula V, an organic solvent, and a reducing agent to perform a reduction reaction to obtain a compound of formula VI; (6) In a hydrogen atmosphere, mixing the compound represented by Formula VI, an organic solvent, and a catalyst to carry out a catalytic hydrogenation reaction to obtain a compound represented by Formula VII; (7) mixing a compound of formula VII, an organic solvent, N,N-diisopropylethylamine, 4-amino-6-methoxy-1,3,5-triazine-2-carboxylic acid, and a condensation reagent to carry out a condensation reaction to obtain a compound of formula VIII; (8) mixing the compound represented by the structure of Formula VIII, an acid, and an organic solvent to carry out a demethylation reaction to obtain the compound represented by the structure of Formula IX; (9) mixing a compound of the structure represented by Formula IX, triphenylphosphine, an organic solvent, and an azo reagent to carry out a substitution reaction to obtain a compound of the structure represented by Formula X; (10) mixing a compound represented by formula X, an organic solvent, and hydrochloric acid to carry out a hydrolysis reaction to obtain scutellarine D; 2. The preparation method according to claim 1, characterized in that In step (1), the molar ratio of benzaldehyde to the compound represented by formula I is 0.5:1 to 1.5:1; The molar ratio of the reducing agent to the compound of the structure shown in formula I is 1:1 to 5:

1.

3. The preparation method according to claim 1, in step (2): the reducing agent comprises sodium cyanoborohydride or sodium borohydride; The molar ratio of the formaldehyde to the compound represented by the structure of formula II is 1:1 to 5:1; The molar ratio of the reducing agent to the compound of the structure shown in Formula II is 1:1 to 5:

1.

4. The preparation method according to claim 1, wherein in step (3): the inorganic strong base comprises sodium hydroxide, lithium hydroxide or potassium hydroxide; The molar ratio of the inorganic strong base to the compound represented by formula III is 1:1 to 5:

1.

5. The preparation method according to claim 1, wherein in step (4): the condensation reagent comprises carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole; The molar ratio of the condensation reagent to the compound represented by formula IV is 1:1 to 5:

1.

6. The preparation method according to claim 1, wherein in step (5): the reducing agent is BH3; The molar ratio of the reducing agent to the compound of the structure shown in Formula V is 2:1 to 10:

1.

7. The preparation method according to claim 1, wherein in step (6): the catalyst comprises palladium carbon or palladium hydroxide carbon.

8. The preparation method according to claim 1, wherein in step (7): the condensation reagent comprises 2-(7-azabenzotriazole)-N,N,N',N'-tetramethyluronium hexafluorophosphate, carbodiimide hydrochloride, 1-propylphosphoric acid cyclic anhydride or carbonyldiimidazole; The molar ratio of the condensation reagent to the structural compound represented by formula VII is 1:1 to 5:

1.

9. The preparation method according to claim 1, wherein in step (8): the acid comprises HCl or trifluoroacetic acid.

10. The preparation method according to claim 1, wherein in step (9): the azo reagent comprises diethyl azodicarboxylate or diisopropyl azodicarboxylate; The molar ratio of the azo reagent to the compound having the structure represented by formula IX is 1:1 to 5:1.

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