Synthetic method of acetyl bromo-uridine triphosphate
By reacting compound I with phosphorus oxychloride and tri-n-butylammonium pyrophosphate in an aqueous solvent, the synthesis conditions of acetylbromourea triphosphate were optimized, solving the problem of poor reaction selectivity in large-scale preparation. This resulted in the preparation of high-purity and high-conversion acetylbromourea triphosphate, suitable for industrial production.
Patent Information
- Application Number
- CN202410763595.X
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-06-13
- Publication Date
- 2025-12-16
AI Technical Summary
In existing technologies, the large-scale preparation of acetylbromourea triphosphate suffers from poor reaction selectivity, resulting in numerous byproducts and making it difficult to achieve efficient and stable synthesis.
Compound I was reacted with phosphorus oxychloride in an aqueous solvent to generate compound II, which was then reacted with tri-n-butylammonium pyrophosphate in an inert solvent. The synthesis conditions were optimized by controlling the temperature and solvent composition to improve the purity and conversion rate.
The efficient synthesis of acetylbromoureanine triphosphate was achieved with few byproducts, high purity, and a conversion rate of about 80%, making it suitable for large-scale industrial production.
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Figure CN121135772A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of biochemistry, specifically relating to a method for synthesizing acetylbromourea triphosphate. Background Technology
[0002] Acetyl bromide triphosphate (acyBrUTP), with its furanose moiety replaced by a 2-hydroxyethoxymethyl group, can be used as a PCR terminator. Experimental data show that the error rate of acyBrUTP as a PCR substrate is significantly lower than that of the corresponding dUTP or ddUTP substrates. Therefore, it is very important in applications such as DNA sequencing and SNP detection. Furthermore, acyBrUTP can also be used as a PCR terminator for time-of-flight mass spectrometry (TOF-MS) detection.
[0003] Currently, large-scale preparation of acetylbromoureanine triphosphate still faces many challenges. Because the acetylbromoureanine molecule has multiple chemical groups with varying reactivity in the synthetic reaction, the reaction selectivity differs, leading to a large number of synthetic byproducts.
[0004] Therefore, those skilled in the art are dedicated to developing synthetic methods for molecules with high reaction rates, low by-product content, and stable processes. Summary of the Invention
[0005] The purpose of this invention is to provide a method for synthesizing acetylbromoureanyl triphosphate (acyBrUTP), which is suitable for large-scale preparation of acetylbromoureanyl triphosphate (acyBrUTP).
[0006] In a first aspect of the present invention, a method for preparing acetylbromoureidine triphosphate (acyBrUTP) is provided, the method comprising the steps of:
[0007] (S1) In an aqueous solvent, compound I (N-acetylbromourean (acyBrU)) reacts with phosphorus oxychloride to form compound II, as shown in the following reaction formula:
[0008]
[0009] In another preferred embodiment, the method further includes the step of:
[0010] (S2) Compound II reacts with tri-n-butylammonium pyrophosphate to form compound III, as shown in the following reaction formula:
[0011]
[0012] In another preferred embodiment, the aqueous solvent in step (S1) is an inert aqueous solvent.
[0013] In another preferred embodiment, the aqueous solvent in step (S1) includes: trimethyl phosphate, water, and proton sponge.
[0014] In another preferred embodiment, the aqueous solvent is composed of trimethyl phosphate, water, and a proton sponge.
[0015] In another preferred embodiment, the aqueous solvent includes:
[0016] 50-150 parts by volume of trimethyl phosphate;
[0017] 1-5 parts by volume of water;
[0018] Furthermore, the volume-to-mass ratio of trimethyl phosphate to proton sponge is 50-150 ml: 50-100 g.
[0019] In another preferred embodiment, the aqueous solvent includes:
[0020] 100 parts by volume of trimethyl phosphate;
[0021] 3 parts by volume of water;
[0022] Furthermore, the volume-to-mass ratio of trimethyl phosphate to proton sponge is 100 ml: 80 g.
[0023] In another preferred embodiment, step (S1) is carried out under the protection of nitrogen or an inert gas.
[0024] In another preferred embodiment, the reaction temperature of step (S1) is -10°C to 5°C; preferably -10°C to 0°C; more preferably about -5°C to 0°C.
[0025] In another preferred embodiment, the reaction time for step (S1) is 0.5-3 hours; preferably about 1-2 hours.
[0026] In another preferred embodiment, in step (S1), compound I is first dissolved with trimethyl phosphate, and then water and proton sponge are added; preferably, the mass-to-volume ratio of compound I to trimethyl phosphate is 30-80 g: 50-150 ml; more preferably, about 50 g: 100 ml.
[0027] In another preferred embodiment, the mass-to-volume ratio of compound I and phosphorus oxychloride in step (S1) is 30-80 g: 40-90 mL; preferably about 50 g: 60 mL.
[0028] In another preferred embodiment, step (S2) is carried out in an inert solvent; preferably, the inert solvent comprises anhydrous acetonitrile and tri-n-butylamine; preferably, the volume ratio of anhydrous acetonitrile to tri-n-butylamine is 500-1000:500 (e.g., 900:500).
[0029] In another preferred embodiment, the reaction temperature of step (S2) is -10°C to 20°C; preferably about -5°C to 20°C; more preferably about -5°C to 15°C.
[0030] In another preferred embodiment, in step (S2), tributylammonium pyrophosphate is first dissolved in an inert solvent; preferably, the mass-volume ratio of tributylammonium pyrophosphate to inert solvent is 100-200 g: 1000-2000 mL; more preferably, it is 150 g: 1400 mL; more preferably, it is pre-cooled at -20°C after dissolution.
[0031] In another preferred embodiment, after the reaction in step (S2) is completed, water is added to quench the reaction.
[0032] In another preferred embodiment, the method further includes a step of purifying compounds I and II.
[0033] In another preferred embodiment, the method includes the steps of:
[0034] (1) Weigh 50g of N-acetylbromourean (acyBrU), add it to the reactor, and insert a thermometer to monitor the temperature;
[0035] (2) Add 100ml of trimethyl phosphate to dissolve N-acetylbromourean (acyBrU), then add 3ml of water and 80g of proton sponge. After N2 replacement, pre-cool at -5℃ for 10min.
[0036] (3) Slowly add 60 ml of phosphorus oxychloride, control the reaction temperature to not exceed 0℃, and react for 1.5 h;
[0037] (4) Weigh 150g of tri-n-butylammonium pyrophosphate, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine, and after the tri-n-butylammonium pyrophosphate is dissolved, pre-cool it at -20℃ for 1.5h. After (3) the reaction is completed, quickly add the dissolved mixed solution into the reactor and react for 10min.
[0038] After the reaction (4) is completed, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with DCM, separate the liquid and obtain an aqueous solution. Then add 600 ml of 2M TEAB solution to the aqueous solution and rotary evaporate to obtain the target compound.
[0039] It should be understood that, within the scope of this invention, the above-described technical features of this invention and the technical features specifically described below (such as in the embodiments) can be combined with each other to form new or preferred technical solutions. Due to space limitations, they will not be described in detail here. Attached Figure Description
[0040] Figure 1The HPLC detection results of the acetylbromourean triphosphate prepared in Example 1 are shown.
[0041] Figure 2 The HPLC detection results of the acetylbromourean triphosphate prepared in Example 2 are shown.
[0042] Figure 3 The HPLC detection results of the acetylbromourea triphosphate prepared in Example 3 are shown. Detailed Implementation
[0043] This invention provides a chemical synthesis method for preparing acetylbromoureanidine triphosphate. The method of this invention produces acetylbromoureanidine triphosphate with higher synthesis efficiency, fewer side reactions, lower product impurity content, better process stability, and the purity of the target compound can reach over 70%, with a synthesis conversion rate of approximately 80%.
[0044] Before describing this invention, it should be understood that the invention is not limited to the specific methods and experimental conditions described, as such methods and conditions can be varied. It should also be understood that the terminology used herein is intended only to describe particular embodiments and is not intended to be limiting; the scope of the invention will be limited only by the appended claims.
[0045] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. As used herein, when referring to a specifically enumerated numerical value, the term “about” means that the value can vary from the enumerated value by no more than 1%. For example, as used herein, the expression “about 100” includes all values between 99 and 101 (e.g., 99.1, 99.2, 99.3, 99.4, etc.).
[0046] While any methods and materials similar to or equivalent to those described herein may be used in the implementation or testing of this invention, preferred methods and materials are exemplified herein.
[0047] In a preferred embodiment of the present invention, the method for synthesizing acetylbromourea triphosphate according to the present invention includes the following steps:
[0048] (1) Weigh 50g of N-acetylbromourean (acyBrU), add it to the reactor, and insert a thermometer to monitor the temperature;
[0049] (2) Add 100ml of trimethyl phosphate to dissolve N-acetylbromourean (acyBrU), then add 3ml of water and 80g of proton sponge. After N2 replacement, pre-cool at -5℃ for 10min.
[0050] (3) Slowly add 60 ml of phosphorus oxychloride, control the reaction temperature to not exceed 0℃, and react for 1.5 h;
[0051] (4) Weigh 150g of tri-n-butylammonium pyrophosphate, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine, and after the tri-n-butylammonium pyrophosphate is dissolved, pre-cool it at -20℃ for 1.5h. After (3) the reaction is completed, quickly add the dissolved mixed solution into the reactor and react for 10min.
[0052] After the reaction (4) is completed, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with DCM, separate the liquid and obtain an aqueous solution. Then add 600 ml of 2M TEAB solution to the aqueous solution and rotary evaporate to obtain the target compound.
[0053] The main advantages of this invention are:
[0054] (1) The method of the present invention has high reaction efficiency, few side reactions, low product impurity content, and high purity of target compound. The conversion rate can reach about 80% and the purity of target compound can reach more than 70%.
[0055] (2) The method of the present invention has good reaction stability and high reproducibility in the preparation of acetylbromourea triphosphate in multiple batches.
[0056] (3) The method of the present invention is suitable for the large-scale preparation of acetylbromourean triphosphate, and is therefore suitable for large-scale industrial production.
[0057] The present invention will be further described in detail below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Experimental methods in the following embodiments, unless otherwise specified, are generally performed under conventional conditions as described in *Molecular Cloning: A Laboratory Manual* by Sambrook J. et al. (translated by Huang Peitang et al., Beijing: Science Press, 2002), or as recommended by the manufacturer. Percentages and parts are by weight unless otherwise stated. Unless otherwise specified, all experimental materials and reagents used in the following embodiments are commercially available.
[0058] Example 1
[0059] At room temperature, weigh 50g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 100ml of trimethyl phosphate to dissolve acyBrU, then add 4ml of water and 80g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0060] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0061] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine. After the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixture to a three-necked flask using a syringe and react for 10min.
[0062] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0063] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0064] HPLC analysis showed a purity of 67.68%. The calculated yield was 70.3%. The HPLC results are as follows: Figure 1 As shown.
[0065] Example 2
[0066] At room temperature, weigh 50g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 100ml of trimethyl phosphate to dissolve acyBrU, then add 3ml of water and 80g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0067] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0068] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine. After the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixture to a three-necked flask using a syringe and react for 10min.
[0069] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0070] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0071] HPLC analysis showed a purity of 82.91%. The calculated yield was 81.7%. The HPLC results are as follows: Figure 2 As shown.
[0072] Example 3
[0073] At room temperature, weigh 60g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 100ml of trimethyl phosphate to dissolve acyBrU, then add 3ml of water and 80g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0074] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0075] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine. After the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixture to a three-necked flask using a syringe and react for 10min.
[0076] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0077] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0078] HPLC analysis showed a purity of 77.82%. The calculated yield was 80.1%. The HPLC results are as follows: Figure 3 As shown.
[0079] Example 4
[0080] At room temperature, weigh 50g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 100ml of trimethyl phosphate to dissolve acyBrU, then add 3ml of water and 100g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0081] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0082] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 900ml of anhydrous acetonitrile and 600ml of tri-n-butylamine, and after the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixed solution to a three-necked flask using a syringe, and react for 10min.
[0083] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0084] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0085] HPLC analysis showed a purity of 83.09%. The calculated yield was 82.2%.
[0086] Example 5
[0087] At room temperature, weigh 50g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 150ml of trimethyl phosphate to dissolve acyBrU, then add 3ml of water and 60g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0088] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0089] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 900ml of anhydrous acetonitrile and 500ml of tri-n-butylamine. After the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixture to a three-necked flask using a syringe and react for 10min.
[0090] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0091] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0092] HPLC analysis showed a purity of 78.36%. The calculated yield was 78.3%.
[0093] Example 6
[0094] At room temperature, weigh 60g of N-acetylbourneourea (acyBrU) and add it to a 5000ml three-necked flask. Insert a thermometer to monitor the temperature, add a magnetic stir bar, then add 150ml of trimethyl phosphate to dissolve acyBrU, then add 4ml of water and 80g of proton sponge. After N2 replacement, place it in a low-temperature cold bath magnetic stirrer, set the temperature to -5℃, and pre-cool for 10min. When the temperature on the thermometer reads -5℃, use a syringe to draw 60ml of phosphorus oxychloride and slowly add it to the reaction system. Observe the thermometer and control the reaction temperature to not exceed 0℃. After the addition is complete, react for 1.5h.
[0095] The reaction phenomenon is that the reaction solution changes from colorless and transparent to pale yellow, and white mist is generated above the solution in the bottle. As the reaction time increases, the white mist becomes less dense and eventually disappears.
[0096] Take a 2000ml single-necked flask, weigh 150g of tri-n-butylammonium pyrophosphate into the flask, add 1000ml of anhydrous acetonitrile and 500ml of tri-n-butylamine. After the tri-n-butylammonium pyrophosphate is completely dissolved, pre-cool it at -20℃ for 1.5h. After the above pale yellow solution has reacted, quickly add the dissolved tri-n-butylammonium pyrophosphate mixture to a three-necked flask using a syringe and react for 10min.
[0097] The reaction phenomenon is that the temperature of the reaction system rises rapidly from -5℃ to about 15℃, the pale yellow solution turns into a yellow solution, and white fumes reappear above the solution in the bottle. As the reaction time increases, the white fumes gradually become fainter and eventually disappear.
[0098] After the reaction is complete, take 6000 ml of aqueous solution to quench the reaction. After quenching, extract with 1000 ml of DCM (dichloromethane), separate the solution using a separatory funnel, and then add 600 ml of 2M TEAB (triethylamine-carbonate buffer solution) to the aqueous solution. After rotary evaporation, the target product is obtained.
[0099] HPLC analysis showed a purity of 75.64%. The calculated yield was 79.1%.
[0100] All documents mentioned in this invention are incorporated herein by reference as if each document were individually incorporated by reference. Furthermore, it should be understood that after reading the foregoing teachings of this invention, those skilled in the art can make various alterations or modifications to this invention, and these equivalent forms also fall within the scope defined by the appended claims.
Claims
1. A method for preparing acetylbromoureidine triphosphate (acyBrUTP), the method comprising the steps of: (S1) In an aqueous solvent, compound I (N-acetylbromourean (acyBrU)) reacts with phosphorus oxychloride to form compound II, as shown in the following reaction formula:
2. The method as described in claim 1, characterized in that, The method further includes the following steps: (S2) Compound II reacts with tri-n-butylammonium pyrophosphate to form compound III, as shown in the following reaction formula:
3. The method as described in claim 1, characterized in that, The aqueous solvent in step (S1) is an inert aqueous solvent.
4. The method as described in claim 3, characterized in that, The aqueous solvent in step (S1) includes: trimethyl phosphate, water, and proton sponge.
5. The method as described in claim 3, characterized in that, The aqueous solvent includes: 50-150 parts by volume of trimethyl phosphate; 1-5 parts by volume of water; Furthermore, the volume-to-mass ratio of trimethyl phosphate to proton sponge is 50-150 ml: 50-100 g.
6. The method as described in claim 5, characterized in that, The aqueous solvent includes: 100 parts by volume of trimethyl phosphate; 3 parts by volume of water; Furthermore, the volume-to-mass ratio of trimethyl phosphate to proton sponge is 100 ml: 80 g.
7. The method as described in claim 1, characterized in that, In step (S1), compound I is first dissolved with trimethyl phosphate, and then water and a proton sponge are added.
8. The method as described in claim 1, characterized in that, The mass-to-volume ratio of compound I to trimethyl phosphate is 30-80 g : 50-150 ml.
9. The method as described in claim 1, characterized in that, In step (S1), the mass-to-volume ratio of compound I and phosphorus oxychloride is 30-80 g: 40-90 ml.
10. The method as described in claim 1, characterized in that, The reaction is carried out in an inert solvent in step (S2).