Method for efficiently synthesizing aristolochic acid

Abstract

The invention discloses a method for efficiently synthesizing aristolochic acid, and belongs to the technical field of heterocyclic compound synthesis. The invention relates to a method for efficiently synthesizing aristolochic acid, which comprises the following steps: in a phosphate buffer system, stirring and reacting aristolochic acid serving as a raw material under the action of a reducing agent until a reaction solution is clear and colorless, and purifying to obtain aristolochic acid, the reducing agent is sodium hydrosulfite or thiourea dioxide. The synthesis method disclosed by the invention has the characteristics of few by-products, simplicity and convenience in operation, mild reaction, low cost of a reducing reagent, environment friendliness and good repeatability, and has a wide application prospect.

Description

Technical Field

[0001] This invention belongs to the field of heterocyclic compound synthesis technology, specifically relating to a method for the efficient synthesis of aristolochic acid. Background Technology

[0002] Aristolochic acids (AAs), as typical representatives of plant-derived natural toxins, have become an important issue in the global public health field due to their nephrotoxicity and carcinogenicity. AAs are a class of structurally similar natural nitrophenanthrene carboxylic acid compounds, mainly including aristolochic acid I (AAI), aristolochic acid II (AAII), aristolochic acid IVa (AAIVa), and other analogues, among which AAI and AAII are the main toxic substances.

[0003] AAs are mainly found in the Aristolochiaceae family ( Aristolochiaceae Among plants, especially in the genus Aristolochia ( Aristlolochia ) and Asarum ( Asarum These plants are commonly found in plants. However, they also have a long history of use in traditional medicine systems, possessing multiple pharmacological effects such as antiviral, antibacterial, anti-inflammatory, diuretic, and metabolic regulation. Related preparations are widely used in traditional and local medicines around the world.

[0004] Metabolic studies have shown that the transformation pathways of aristolochic acid (AAs) in organisms vary across species. Analysis of urine and feces from different species revealed that the main metabolites of AAs are nitro-reduced aristolochic acid (ALs) and other small amounts of decarboxyl and denitrification metabolites. In humans, only ALs were detected, indicating that humans primarily metabolize AAs via the nitro-reduction pathway, and this metabolic characteristic is directly related to their toxicity expression mechanisms. Therefore, aristolochic acid obtained through denitrification is considered a key attenuated metabolite of AAs and an important target for studying structural toxicity and the mechanisms of action of traditional Chinese medicine. However, aristolochic acid is currently not readily available in the market due to its high price and long synthesis cycle, failing to meet market demand.

[0005] Meanwhile, the 2020 edition of the Pharmacopoeia of the People's Republic of China has removed many traditional Chinese medicine preparations made from plants of the Aristolochiaceae family, but still retains a number of traditional Chinese medicine preparations containing Asarum. The Pharmacopoeia has made strict regulations on the use of Asarum, explicitly requiring the testing of its AAI content, stipulating that the AAI content in dried Asarum must not exceed 0.001%; ​​for a small number of traditional Chinese medicine preparations containing a higher proportion of Asarum, such as Xinqin Granules and Jiuwei Qianghuo Pills, the Pharmacopoeia also requires the testing of AAI content, and stipulates that when tested by high performance liquid chromatography, the AAI content must not exceed the standard.

[0006] Therefore, it is of great importance to develop an efficient, simple and low-cost method for preparing aristolochic acid, to lay the foundation for its application as a scientific research material, promote scientific research and practical application in related fields, and at the same time achieve efficient and safe removal of AAI from traditional Chinese medicine preparations containing Asarum, so that the AAI content in related products meets the regulations. Summary of the Invention

[0007] The problem this invention aims to solve is to provide an efficient method for synthesizing aristolochic acid, thereby addressing the issues of low preparation efficiency, low product purity, and high cost in existing aristolochic acid preparation methods.

[0008] The technical solution adopted to solve the technical problem is to provide an efficient method for synthesizing aristolochic acid, including the following steps: In a phosphate buffer system, aristolochic acid is used as a raw material and stirred under the action of a reducing agent until the reaction solution is clear and colorless. After purification, aristolochic acid is obtained. The reducing agent is sodium dithionite or thiourea dioxide.

[0009] The beneficial effects of the above technical solution adopted in this invention are as follows: The synthesis path is as follows: In the synthesis process of this invention, the reducing agent's effect on aristolochic acid is presumably mainly derived from the sulfur dioxide anion radicals (SO2) released by sodium dithionite (Na2S2O4) or thiourea dioxide (CH4N2O2S, abbreviated as TUO2). •- Through the action of sodium dithionite or thiourea dioxide, aristolochic acid can be efficiently converted into aristolochic acid.

[0010] More preferably, the reducing agent is sodium dithionite.

[0011] Preferably, the aristolochic acid is aristolochic acid I, aristolochic acid II, or aristolochic acid IVa, and the corresponding synthesized aristolochic acid is aristolochic acid I, aristolochic acid II, or aristolochic acid IVa, in sequence.

[0012] More preferably, aristolochic acid is added in the form of an aristolochic acid solution, which is prepared by the following steps: dissolving aristolochic acid in a mixed solvent containing methanol and sodium bicarbonate solution.

[0013] More preferably, the volume ratio of methanol to sodium bicarbonate solution in the mixed solvent is (1~2):(1~2); and the mass concentration of sodium bicarbonate solution is 2~5%.

[0014] More preferably, the volume ratio of methanol to sodium bicarbonate solution in the mixed solvent is 1:1; and the mass concentration of sodium bicarbonate solution is 3%.

[0015] More preferably, the molar concentration of the aristolochic acid solution is 0.3~0.6 mM.

[0016] More preferably, the molar concentration of the aristolochic acid solution is 0.5 mM.

[0017] Preferably, the phosphate buffer system is a phosphate buffer solution with a pH of 7.4-10. The phosphate buffer solution is prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and is treated with Chelex 100 resin to remove impurity metal ions.

[0018] Preferably, the pH of the phosphate buffer solution is 7.4.

[0019] Preferably, the molar ratio of aristolochic acid to reducing agent in the reaction solution is (0.03~0.06):(10~100).

[0020] More preferably, the molar ratio of aristolochic acid to reducing agent in the reaction solution is 0.05:(10~100).

[0021] Preferably, the stirring reaction is carried out at room temperature for 20 min to 6 h.

[0022] Preferably, the purification process includes sequential organic phase extraction, rotary evaporation concentration, rotary drying, recrystallization, and freeze drying of the reaction solution.

[0023] More preferably, organic phase extraction is performed using ethyl acetate; recrystallization is performed using a mixed solution of dimethyl sulfoxide and deionized water.

[0024] The present invention has the following beneficial effects: (1) The synthesis method of the present invention can rapidly remove the nitro group in the molecular structure of aristolochic acid under mild conditions by using inexpensive and readily available reducing reagents and simple operating procedures, thereby efficiently synthesizing the target product aristolochic acid.

[0025] (2) The synthesis method of the present invention has the characteristics of few by-products, simple operation, low cost of reducing reagent, environmental friendliness and good reproducibility, and is suitable for scientific research on aristolochic acid and large-scale preparation of drugs. Attached Figure Description

[0026] Figure 1 This is a reaction chromatogram monitored by HPLC in Example 1; Figure 2 This is a reaction chromatogram monitored by HPLC in Example 2; Figure 3 The following are reaction diagrams monitored by HPLC in Example 3; (a) is the monitored reaction diagram; (b) is the absorption spectrum of aristolochic acid IVa; and (c) is the absorption spectrum of aristolochic acid IVa. Figure 4 This is a reaction chromatogram monitored by HPLC in Example 4; Figure 5 This is a reaction chromatogram monitored by HPLC in Example 5; Figure 6 This is a reaction chromatogram monitored by HPLC in Example 6; Figure 7 The reaction chromatogram in Comparative Example 1 was monitored by HPLC. Figure 8 The reaction diagrams for Comparative Examples 2-4 are monitored by HPLC. Detailed Implementation

[0027] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are only for explaining the invention and are not intended to limit the invention; that is, the described embodiments are only a part of the embodiments of this invention, and not all of them.

[0028] Therefore, the following detailed description of the embodiments of the present invention is not intended to limit the scope of the claimed invention, but only to illustrate selected embodiments of the invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention.

[0029] The features and performance of the present invention will be further described in detail below with reference to embodiments.

[0030] Example 1 A method for the efficient synthesis of aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a sodium dithionite solution with a molar concentration of 100 mM using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid I solution to the phosphate buffer solution with pH 7.4 prepared in step (1), mix well, and then add sodium dithionite solution to make the molar concentration of aristolochic acid I solution 0.05 mM and the molar concentration of sodium dithionite solution 10 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid I is monitored in real time by high performance liquid chromatography (HPLC) to determine the reaction progress. After 20 min of reaction, the chromatographic peak of aristolochic acid I completely disappeared and no by-product chromatographic peaks other than aristolochic acid I were detected. At this time, the formation rate of aristolochic acid I was 100%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 to the saturated solution and mix well for recrystallization; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator overnight and freeze dry it to obtain aristolochic acid I (light yellow powder, yield >80%, purity 100%, without aristolochic acid I).

[0031] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 1 (Detection wavelength: Sig=275 nm).

[0032] Example 2 A method for the efficient synthesis of aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid II as raw material and prepare a 0.5 mM aristolochic acid II solution using the mixed solvent. (3) Prepare a sodium dithionite solution with a molar concentration of 100 mM using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid II solution to the pH 7.4 phosphate buffer solution prepared in step (1), mix well, and then add sodium dithionite solution to make the molar concentration of aristolochic acid II solution 0.05 mM and the molar concentration of sodium dithionite solution 10 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid II is monitored in real time by HPLC to determine the reaction progress. After 20 min of reaction, the chromatographic peak of aristolochic acid II basically disappeared. At this time, the formation rate of aristolochic acid II was 96%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 of saturated solution to deionized water and mix well for recrystallization; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator to stand overnight and freeze dry it to obtain aristolochic acid II (light yellow powder, yield >80%, purity >96%, without aristolochic acid II).

[0033] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 2 (Detection wavelength: Sig=275 nm).

[0034] Example 3 A method for the efficient synthesis of aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid IVa as raw material and prepare a 0.5 mM aristolochic acid IVa solution using the mixed solvent. (3) Prepare a sodium dithionite solution with a molar concentration of 100 mM using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid IVa solution to the pH 7.4 phosphate buffer solution prepared in step (1), mix well, and then add sodium dithionite solution to make the molar concentration of aristolochic acid IVa solution 0.05 mM and the molar concentration of sodium dithionite solution 50 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid IVa is monitored in real time by HPLC to determine the reaction progress. After 20 min of reaction, the chromatographic peak of aristolochic acid IVa basically disappeared. At this time, the aristolochic acid IVa formation rate was 85%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 of saturated solution to deionized water and mix well for recrystallization; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator to stand overnight and freeze dry it using a freeze dryer to obtain aristolochic acid IVa (light yellow powder, yield >80%, purity >85%, does not contain aristolochic acid IVa).

[0035] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 3 (Detection wavelength: Sig=275 nm).

[0036] Example 4 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a 100 mM thiourea dioxide solution using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid I solution to the phosphate buffer solution with pH 7.4 prepared in step (1), mix well, and then add the thiourea dioxide solution so that the molar concentration of the aristolochic acid I solution is 0.05 mM and the molar concentration of the thiourea dioxide solution is 50 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid I is monitored in real time by HPLC to determine the reaction progress. After 6 h of reaction, the chromatographic peak of aristolochic acid I completely disappeared and no by-product chromatographic peaks other than aristolochic acid I were detected. At this time, the formation rate of aristolochic acid I is 100%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 of saturated solution to deionized water, mix well and recrystallize; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator overnight and freeze dry it to obtain aristolochic acid I (light yellow powder, yield >80%, purity 100%, does not contain aristolochic acid I).

[0037] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 4 (Detection wavelength: Sig=275 nm).

[0038] Example 5 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid II as raw material and prepare a 0.5 mM aristolochic acid II solution using the mixed solvent. (3) Prepare a 100 mM thiourea dioxide solution using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid II solution to the pH 7.4 phosphate buffer solution prepared in step (1), mix well, and then add the thiourea dioxide solution so that the molar concentration of the aristolochic acid II solution is 0.05 mM and the molar concentration of the thiourea dioxide solution is 50 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid I is monitored in real time by HPLC to determine the reaction progress. After 6 h of reaction, the chromatographic peak of aristolochic acid II basically disappeared. At this time, the formation rate of aristolochic acid II was 96%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 of saturated solution to deionized water and mix well for recrystallization; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator to stand overnight and freeze dry it to obtain aristolochic acid II (light yellow powder, yield >80%, purity >96%, without aristolochic acid II).

[0039] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 5 (Detection wavelength: Sig=275 nm).

[0040] Example 6 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid IVa as raw material and prepare a 0.5 mM aristolochic acid IVa solution using the mixed solvent. (3) Prepare a 200 mM thiourea dioxide solution using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid IVa solution to the phosphate buffer solution with pH 7.4 prepared in step (1), mix well, and then add the thiourea dioxide solution so that the molar concentration of the aristolochic acid IVa solution is 0.05 mM and the molar concentration of the thiourea dioxide solution is 100 mM. Stir the reaction at room temperature until the reaction solution is clear and colorless. During the reaction, the chromatographic peak change of aristolochic acid IVa is monitored in real time by HPLC to determine the reaction progress. After 6 h of reaction, the chromatographic peak of aristolochic acid IVa basically disappears. At this time, the formation rate of aristolochic acid IVa is 85%. (5) Extract the reaction solution obtained in step (4) twice with ethyl acetate and collect the organic phase; concentrate the collected organic phase to 5 mL by rotary evaporation, and then place it in a centrifuge tube and use a vacuum centrifuge to evaporate it until only light yellow powder remains in the centrifuge tube; collect the light yellow powder and dissolve it in dimethyl sulfoxide until a saturated solution is formed, add deionized water at a volume ratio of 1:5 of saturated solution and deionized water, mix well and recrystallize; centrifuge at 8000 rpm for 10 min, collect the precipitate and place it in a -40℃ refrigerator overnight and freeze dry it to obtain aristolochic acid IVa (light yellow powder, yield >80%, purity >85%, does not contain aristolochic acid IVa).

[0041] The reaction chromatogram monitored by HPLC in step (4) is shown below. Figure 6 (Detection wavelength: Sig=275 nm).

[0042] Comparative Example 1 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a 200 mM L-cysteine ​​solution using the pH 7.4 phosphate buffer solution obtained in step (1); (4) Add the aristolochic acid I solution to the pH 7.4 phosphate buffer solution prepared in step (1), mix well, and then add L-cysteine ​​solution so that the molar concentration of aristolochic acid I solution is 0.05 mM and the molar concentration of L-cysteine ​​solution is 100 mM. Stir the reaction at room temperature. During the reaction, HPLC is used to monitor the chromatographic peak change of aristolochic acid I in real time to determine the reaction progress.

[0043] The reaction chromatogram monitored by HPLC is shown below. Figure 7 (Detection wavelength: Sig=275 nm) The disappearance of the chromatographic peak of aristolochic acid I was observed at 0 h, 4 h, 8 h, 12 h and 16 h of reaction. The results showed that under the action of L-cysteine, the chromatographic peak of aristolochic acid I disappeared slowly, the formation of aristolochic acid I was slow, and the reaction efficiency was significantly lower than that of the example.

[0044] Comparative Example 2 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a 1 M sodium sulfite (Na2SO3) solution using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid I solution to the phosphate buffer solution with pH 7.4 prepared in step (1), mix well, and then add sodium sulfite solution so that the molar concentration of aristolochic acid I solution is 0.05 mM and the molar concentration of sodium sulfite solution is 100 mM. Stir the reaction at room temperature. During the reaction, HPLC is used to monitor the chromatographic peak change of aristolochic acid I in real time to determine the reaction progress.

[0045] The reaction chromatogram monitored by HPLC is shown below. Figure 8 (Detection wavelength: Sig=275 nm) After 12 h of reaction, the chromatographic peak of aristolochic acid I completely disappeared, but the chromatographic peaks of byproducts other than aristolochic acid I were detected. At this time, the formation rate of aristolochic acid I was 5%. It can be seen that when sodium sulfite is used as the reducing agent, there are more reaction byproducts and the effect of generating aristolochic acid I is poor.

[0046] Comparative Example 3 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a 1 M sodium bisulfite (NaHSO3) solution using the phosphate buffer solution with pH 7.4 obtained in step (1); (4) Add the aristolochic acid I solution to the phosphate buffer solution with pH 7.4 prepared in step (1), mix well, and then add sodium bisulfite solution so that the molar concentration of aristolochic acid I solution is 0.05 mM and the molar concentration of sodium bisulfite solution is 80 mM. Stir at room temperature, and use HPLC to monitor the chromatographic peak change of aristolochic acid I in real time to determine the reaction progress.

[0047] The reaction chromatogram monitored by HPLC is shown below. Figure 8 (Detection wavelength: Sig=275 nm) After 12 h of reaction, the chromatographic peak of aristolochic acid I completely disappeared, but the chromatographic peaks of byproducts other than aristolochic acid I were detected. At this time, the formation rate of aristolochic acid I was 4%. It can be seen that when sodium bisulfite is used as the reducing agent, there are more reaction byproducts and the effect of generating aristolochic acid I is poor.

[0048] Comparative Example 4 A method for synthesizing aristolochic acid includes the following steps: (1) A phosphate buffer solution was prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and treated with Chelex 100 resin to remove impurity metal ions, resulting in a phosphate buffer solution with a pH of 7.4 and a concentration of 100 mM. (2) Measure out methanol and sodium bicarbonate solution with a mass concentration of 3% and mix them in a volume ratio of 1:1 to obtain a mixed solvent; use aristolochic acid I as raw material and prepare a 0.5 mM aristolochic acid I solution using the mixed solvent. (3) Prepare a 1 M sodium metabisulfite (Na2S2O5) solution using a phosphate buffer solution with a pH of 7.4; (4) Add aristolochic acid I solution to phosphate buffer solution, mix well, and then add sodium metabisulfite to make the molar concentration of aristolochic acid I solution 0.05 mM and the molar concentration of sodium metabisulfite solution 40 mM. Stir at room temperature and use HPLC to monitor the chromatographic peak changes of aristolochic acid I in real time to determine the reaction progress.

[0049] The reaction chromatogram monitored by HPLC is shown below. Figure 8 (Detection wavelength: Sig=275 nm) After 12 h of reaction, the chromatographic peak of aristolochic acid I completely disappeared, but the chromatographic peaks of byproducts other than aristolochic acid I were detected. At this time, the formation rate of aristolochic acid I was 5%. It can be seen that when sodium metabisulfite is used as the reducing agent, there are more reaction byproducts and the effect of generating aristolochic acid I is poor.

[0050] The present invention has been described according to the above embodiments. It should be understood that the above embodiments do not limit the present invention in any way. All technical solutions obtained by equivalent substitution or equivalent transformation fall within the scope of the present invention.

Claims

1. A method for the efficient synthesis of aristolochic acid, characterized in that, Includes the following steps: In a phosphate buffer system, aristolochic acid is used as a raw material and stirred under the action of a reducing agent until the reaction solution is clear and colorless. After purification, aristolochic acid is obtained. The reducing agent is sodium dithionite or thiourea dioxide.

2. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The aristolochic acid is aristolochic acid I, aristolochic acid II, or aristolochic acid IVa, and the corresponding synthesized aristolochic acid is aristolochic acid I, aristolochic acid II, or aristolochic acid IVa, in sequence.

3. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The aristolochic acid is added in the form of an aristolochic acid solution, which is prepared by the following steps: dissolving the aristolochic acid in a mixed solvent containing methanol and sodium bicarbonate solution.

4. The method for efficiently synthesizing aristolochic acid as described in claim 3, characterized in that, The volume ratio of methanol to sodium bicarbonate solution in the mixed solvent is (1~2):(1~2); the mass concentration of sodium bicarbonate solution is 2~5%.

5. The method for efficiently synthesizing aristolochic acid as described in claim 3, characterized in that, The molar concentration of the aristolochic acid solution is 0.3~0.6 mM.

6. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The phosphate buffer system is a phosphate buffer solution with a pH of 7.4-10. The phosphate buffer solution is prepared using sodium hydroxide, phosphoric acid, sodium dihydrogen phosphate dihydrate and disodium hydrogen phosphate dodecahydrate, and is treated with Chelex 100 resin to remove impurity metal ions.

7. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The molar ratio of aristolochic acid to reducing agent in the reaction solution is (0.03~0.06):(10~100).

8. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The stirring reaction was carried out at room temperature for 20 min to 6 h.

9. The method for efficiently synthesizing aristolochic acid as described in claim 1, characterized in that, The purification process includes sequential organic phase extraction, rotary evaporation concentration, rotary drying, recrystallization, and freeze drying of the reaction solution.

10. The method for efficiently synthesizing aristolochic acid as described in claim 9, characterized in that, The organic phase extraction was performed using ethyl acetate; the recrystallization was performed using a mixed solution of dimethyl sulfoxide and deionized water.

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