A method for the synthesis of 2'-ome adenosine

Abstract

The application discloses a synthesis method of 2'-OMe adenosine, comprising the following steps: S1. dissolving adenosine in DMF, and reacting with a methylation reagent in the presence of a base to obtain a mixture of 2'-OMe adenosine and 3'-OMe adenosine; S2. reacting the mixture of 2'-OMe adenosine and 3'-OMe adenosine with acetic anhydride in a solvent to obtain an acetylated mixture; S3. deacetylating the acetylated mixture under the action of ammonia, and crystallizing to obtain 2'-OMe adenosine; the application uses cheap and easily available raw materials and a safe alkylating reagent, synthesizes 2'-OMe adenosine through a shorter route, avoids column chromatography operation, and provides a new synthesis method of 2'-OMe adenosine which is easy to process.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of drug and intermediate preparation, in particular to a synthesis method of 2'-OMe adenosine. BACKGROUND

[0002] 2'-OMe ribonucleic acid is widely distributed in RNA, and since 1950, the research on its synthesis method has become the focus of attention. The application of 2'-OMe oligonucleotide in the study of precursor messenger RNA splicing and spliceosome structure has further promoted the research on the synthesis method of 2'-OMe nucleoside.

[0003] At present, there are three kinds of synthesis methods of 2'-OMe adenosine. The first one is SnCl2-promoted direct methylation of adenosine, but the selectivity of 2' and 3'-OH is not good, and it needs to be purified by column chromatography, the yield is 38%, the alkylating agent is diazomethane, the process safety risk is high, and it is difficult to scale up production; the second one is to realize the selective methylation of 2'-OH of 2-amino-6-chloropurine nucleoside under the condition of MeI / NaH, the yield is 65%, then the hydroxyl group is acetylated and protected, the free radical is deaminated, and finally the deprotection / amination is carried out to obtain 2'-OMe adenosine, the overall yield is 37%. The whole process does not have a column chromatography step, and it is easy to scale up the process, but the disadvantages are that the raw material price is high and the route is long. The third one is to protect the 3', 5'-OH of adenosine by TIPDS, then methylate the 2'-OH, and finally remove TIPDS to obtain 2'-OMe adenosine, but the cost of 1,3 dichloro-1,1,3,3-tetraisopropyl disiloxane in this route is high, and the advantage is not obvious. In general, the existing synthesis methods have the problems of low safety of raw materials, high cost of raw materials, long process route or difficult scaling up production. SUMMARY

[0004] Based on the above problems, the present application uses cheap and easily available raw materials and safe alkylating agents to synthesize 2'-OMe adenosine through a shorter route, avoids column chromatography operation, and provides a new synthesis method of 2'-OMe adenosine which is easy to scale up the process.

[0005] The present application provides a synthesis method of 2'-OMe adenosine, which adopts the following technical scheme:

[0006] A synthesis method of 2'-OMe adenosine, comprising the following steps:

[0007] S1. Dissolve adenosine in DMF, and react with a methylating agent in the presence of a base to obtain a mixture of 2'-OMe adenosine and 3'-OMe adenosine;

[0008] S2. React the mixture of 2'-OMe adenosine and 3'-OMe adenosine with acetic anhydride in a solvent to obtain an acetylated mixture;

[0009] S3. The acetylated mixture is deacetylated under the action of ammonia, and crystallized to obtain 2'-OMe adenosine.

[0010] By adopting the technical scheme, the 2'-OH methylation is achieved, and then the inorganic salt is derivatized and removed, and then the protecting group is removed, and the product is obtained by crystallization.

[0011] Preferably, in step S1, the base is any one or a combination of lithium hydroxide, sodium hydroxide, potassium hydroxide, cesium hydroxide, and sodium hydride. The above-mentioned bases have strong alkalinity, which ensures the reactivity of methylation; in addition, they are low in price and easy to obtain.

[0012] Preferably, in step S1, the methylation reagent is any one or a combination of methyl iodide, dimethyl sulfate, dimethyl carbonate, and methyl p-toluenesulfonate. The above-mentioned methylation reagents are common methylation reagents, and are low in price and easy to obtain.

[0013] Preferably, in step S2, the solvent is any one or a combination of dichloromethane, trichloromethane, and dichloroethane. These solvents are chlorinated hydrocarbon solvents, which have good solubility for the product of step S2, and can ensure a fast reaction rate, and also have high extraction efficiency in post-processing.

[0014] Preferably, in step S3, the ammonia is any one or a combination of aqueous ammonia, ammonia methanol solution, ammonia ethanol solution, and ammonia dioxane solution. The above-mentioned ammonia is a common aqueous ammonia solution or organic solution, and has a moderate concentration, and also has a fast deprotection reaction rate.

[0015] Preferably, the solvent used for crystallization includes 95-100wt% ethanol, 95-100wt% methanol, or a combination of the two.

[0016] Preferably, in step S1, the amount of the base is 1.2-2.5 times the molar amount of adenosine, and the amount of the methylation reagent is 1.0-2.0 times the molar amount of adenosine.

[0017] Preferably, in step S2, the amount of acetic anhydride is 2.2-4.0 eq of the molar amount of adenosine in step S1.

[0018] In summary, the present application has at least the following beneficial effects:

[0019] 1. The present application discloses a method for synthesizing 2'-OMe adenosine by a shorter route using low-cost and easily available raw materials and suitable reagents, and avoiding column chromatography operation;

[0020] 2, the reaction yield of 2'-OMe adenosine prepared by the synthesis method of the application is 26-34%; the liquid phase purity reaches 98.6%-99.4%; 3, the cost of 2'-OMe adenosine prepared by the synthesis method of the application is significantly reduced; because in the second synthesis method "2-amino-6-chloropurine nucleoside realizes selective methylation of 2'-OH under the condition of MeI / NaH", the raw material used is 20-30 times the cost of adenosine of the application, so the raw material cost is greatly reduced; and compared with the third synthesis method "3', 5'-OH of adenosine is protected by TIPDS, then 2'-OH is methylated, and finally TIPDS is removed to obtain 2'-OMe adenosine", the cost is reduced by 30%. DETAILED DESCRIPTION

[0021] In order to make the purpose, technical scheme and beneficial technical effects of the application of the application more clear, the application will be described in detail below. It should be noted that the aspects, features, embodiments and advantages described in the application can be compatible and / or combined together.

[0022] Unless otherwise specified, the meanings of the scientific and technical terms in the specification are the same as those generally understood by those skilled in the art.

[0023] The application relates to a synthesis method of 2'-OMe adenosine.

[0024] The application will be specifically described below.

[0025] The application provides a synthesis method of 2'-OMe adenosine, which adopts the following technical scheme:

[0026] The synthesis method of 2'-OMe adenosine comprises the following steps:

[0027] S1: adenosine is dissolved in DMF and reacted with a methylation reagent in the presence of a base to obtain a mixture of 2'-OMe adenosine and 3'-OMe adenosine;

[0028] S2: the mixture of 2'-OMe adenosine and 3'-OMe adenosine is reacted with acetic anhydride in a solvent to obtain an acetylated mixture; S3: the acetylated mixture is deacetylated under the action of ammonia, and crystallization is performed to obtain 2'-OMe adenosine.

[0029] The reaction structural formula of the synthesis method of 2'-OMe adenosine of the application is as follows:

[0030] S1:

[0031]

[0032] S2:

[0033]

[0034] S3:

[0035]

[0036] In some embodiments, in step S1, the base is any one or a combination of lithium hydroxide, sodium hydroxide, potassium hydroxide, cesium hydroxide, sodium hydride; preferably potassium hydroxide or lithium hydroxide; further preferably potassium hydroxide.

[0037] In some embodiments, in step S1, the methylating agent is any one or a combination of iodomethane, dimethyl sulfate, dimethyl carbonate, methyl p-toluenesulfonate; preferably iodomethane or methyl p-toluenesulfonate; further preferably methyl p-toluenesulfonate.

[0038] In some embodiments, in step S2, the solvent is any one or a combination of dichloromethane, trichloromethane, dichloroethane; preferably dichloromethane or dichloroethane; further preferably dichloromethane.

[0039] In some embodiments, in step S3, the ammonia is any one or a combination of aqueous ammonia, ammonia in methanol, ammonia in ethanol, ammonia in dioxane; preferably aqueous ammonia or ammonia in methanol; further preferably aqueous ammonia.

[0040] In some embodiments, in step S3, the solvent used for crystallization includes 95-100 wt% ethanol, 95-100 wt% methanol, or a combination of the two; preferably 98-100 wt% ethanol, 98-100 wt% methanol; further preferably 98-100 wt% ethanol.

[0041] In some embodiments, in step S1, the amount of the base is 1.2-2.5 times the molar amount of adenosine, and the amount of the methylating agent is 1.0-2.0 times the molar amount of adenosine.

[0042] In some embodiments, in step S2, the amount of acetic anhydride is 2.2-4.0 eq of the molar amount of adenosine in step S1.

[0043] Examples

[0044] In order to make the purposes, technical solutions and advantages of the present application clearer, the present application will be further described in detail below with reference to examples. It should be understood that the specific examples described herein are only used to explain the present application and not intended to limit the present application.

[0045] Example 1:

[0046] A method for synthesizing 2'-OMe adenosine, comprising the following steps:

[0047] S1: 100 g of adenosine (Formula I) and 900 mL of DMF were added to a reaction bottle, stirred, heated to dissolve, and cooled to 20-30 °C. 35.7 g of potassium hydroxide was added and stirred for 10 min. The temperature was adjusted to 10-15 °C, and a previously prepared solution of methyl p-toluenesulfonate in DMF (the amount of methyl p-toluenesulfonate was 104.5 g, and the amount of DMF was 100 mL) was added dropwise to the reaction bottle. The reaction was stirred at 10-20 °C for 3-4 h. After the reaction was completed, the temperature was controlled at 5-10 °C, 50 mL of water was added dropwise, and then 6N HCl was added to adjust the pH to 6-8. The reaction solution was reduced to remove the solvent until no obvious fraction was obtained, a light yellow paste was obtained, 2 L of DCM was added to disperse the paste, and the obtained suspension was directly used for the next step;

[0048]

[0049] S2: Synthesis of Formula III compound: The temperature was controlled at 5-15 °C, 121.1 g of triethylamine was added dropwise to the DCM suspension obtained in the first step, and then 114.6 g of acetic anhydride was added dropwise. The reaction was incubated for 3-5 h. After the reaction was completed, 1 L of water was added, stirred for 10 min, and then allowed to stand to separate the liquid. The aqueous phase was extracted with 500 mL of DCM, and the organic phases were combined and washed with water twice, each time with 1 L of water. The organic phase was reduced to remove the solvent until no obvious fraction was obtained, and a yellow oil was obtained, which was directly used for the next step;

[0050]

[0051] S3: Synthesis of Formula IV compound: The temperature was controlled at 15-25 °C, 1 L of ethanol and 500 mL of ammonia water were added to the yellow oil obtained in the second step, and stirred at 15-25 °C for 12-15 h. After the reaction was completed, the solvent was removed until no obvious fraction was obtained, and then 1 L of ethanol was added and evaporated once. 500 mL of ethanol was added to the obtained yellow oil, heated to reflux to obtain a clear solution, and then slowly cooled to 20 °C to precipitate solids. The solids were stirred for 6 h, filtered to obtain a white solid. The solid was dispersed in 500 mL of 98 wt% ethanol, heated to reflux to dissolve, and then slowly cooled to 20 °C to precipitate solids. The solids were stirred for 6 h, filtered to obtain a white solid. The wet filter cake was measured for purity, and the purity was required to be >98.0%, and the single impurity was required to be <1.0%. If it was unqualified, the above recrystallization operation was repeated until it was qualified. The wet filter cake with qualified purity was dried to obtain 33.7 g of white solid;

[0052]

[0053] Example 2:

[0054] A method for synthesizing 2'-OMe adenosine, which is different from Example 1 in that the base in S1 is sodium hydroxide, and the amount used is 37.4 g.

[0055] Example 3:

[0056] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the base in S1 is lithium hydroxide, used in an amount of 10.8 g.

[0057] Example 4:

[0058] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the methylating agent used in S1 is methyl iodide, used in an amount of 79.6 g.

[0059] Example 5:

[0060] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the methylating agent used in S1 is dimethyl sulfate, used in an amount of 70.7 g.

[0061] Example 6:

[0062] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the solvent used in S2 is chloroform.

[0063] Example 7:

[0064] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the solvent used in S2 is dichloroethane.

[0065] Example 8:

[0066] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the ammonia used in S3 is an ethanol solution of ammonia.

[0067] Example 9:

[0068] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the ammonia used in S3 is a methanol solution of ammonia.

[0069] Example 10:

[0070] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the crystallization solvent used in S3 is methanol, used in an amount of 400 mL.

[0071] Example 11:

[0072] A method of synthesizing 2'-OMe adenosine, which differs from Example 1 in that the crystallization solvent used in S3 is ethanol, used in an amount of 1000 mL.

[0073] Example 12:

[0074] A method for synthesizing 2'-OMe adenosine, which is different from example 1 in that in step S1, the amount of base is 1.2 times the molar amount of adenosine, and the amount of methylating agent is 1 times the molar amount of adenosine; in step S2, the amount of acetic anhydride is 2.2 eq of the molar amount of adenosine in step S1.

[0075] Example 13:

[0076] A method for synthesizing 2'-OMe adenosine, which is different from example 1 in that in step S1, the amount of base is 2.5 times the molar amount of adenosine, and the amount of methylating agent is 2 times the molar amount of adenosine; in step S2, the amount of acetic anhydride is 4.0 eq of the molar amount of adenosine in step S1.

[0077] Detection means:

[0078] (1) Liquid phase purity: detected by HPLC, wavelength is 260 nm;

[0079] (2) Reaction yield: the mass of white solid obtained after drying / theoretical mass.

[0080] Table 1. Influence of selection of base in step S1 of examples 1-3 on liquid phase purity

[0081] Base Liquid phase purity Example 1 Potassium hydroxide 52.1％ Example 2 Sodium hydroxide 43.7％ Example 3 Lithium hydroxide 48.3％

[0082] Table 2. Influence of selection of methylating agent in step S1 of examples 1, 4-5 on liquid phase purity

[0083]

[0084]

[0085] Table 3. Influence of selection of solvent in step S2 of examples 1, 6-7 on liquid phase purity

[0086] Solvent Liquid phase purity Example 1 Dichloromethane 48.4％ Example 6 Trichloromethane 38.4％ Example 7 Dichloroethane 43.2％

[0087] Table 4. Influence of selection of ammonia in step S3 of examples 1, 8-9 on liquid phase purity

[0088] Ammonia Liquid phase purity Example 1 Aqueous ammonia 45.2％ Example 8 Ethanol solution of ammonia 43.1％ Example 9 Methanol solution of ammonia 44.0％

[0089] Table 5. Influence of selection of crystallization solvent in step S3 of examples 1, 10-11 on total reaction yield

[0090] Crystallization solvent Liquid phase purity Overall reaction yield Example 1 Ethanol 98 wt% 98.6％ 34％ Example 10 Methanol 99.4％ 26％ Example 11 Ethanol 98.8％ 30％

[0091] Table 6. Liquid phase purity in example 1, example 12, and example 13

[0092]

[0093] From the detection results of Examples 1-3 and Table 1, it can be seen that when the base is potassium hydroxide, sodium hydroxide or lithium hydroxide, the liquid phase purity varies in the range of 43.7%-52.1%, and in particular, when the base is potassium hydroxide, the liquid phase purity can reach 52.1%, indicating that potassium hydroxide has a promoting effect on the reactivity of the methylation reagent; therefore, the base is preferably potassium hydroxide or lithium hydroxide.

[0094] From the detection results of Examples 1, 4-5 and Table 2, it can be seen that the methylation reagent has the greatest influence on the liquid phase purity of the product of step S2, and the methylation reagent can be selected from methyl p-toluenesulfonate, iodomethane and dimethyl sulfate, wherein when the methylation reagent is methyl p-toluenesulfonate, the liquid phase purity can reach 52.1%, and when the methylation reagent is dimethyl sulfate, the liquid phase purity is only 33.2%, which indicates that the electrophilicity of the sulfonyl group in methyl p-toluenesulfonate is stronger, thereby improving the methylation reactivity and significantly improving the liquid phase purity; therefore, the methylation reagent is preferably methyl p-toluenesulfonate or iodomethane.

[0095] From the detection results of Examples 1, 6-7 and Table 3, it can be seen that compared with trichloroalkane, dichloroalkane as the solvent significantly improves the liquid phase purity of step S2, which indicates that dichloroalkane has moderate polarity, which promotes the reaction while avoiding the generation of impurities; in addition, when dichloroalkane is used as the solvent, the odor of the solution is also relatively small.

[0096] From the detection results of Examples 1, 8-9 and Table 4, it can be seen that the selection of ammonia as aqueous ammonia or ammonia methanol solution is helpful for the removal of acetyl group, thereby improving the liquid phase purity of step S3, which indicates that water and methanol have relatively large polarity, which promotes the rate of ammonolysis reaction.

[0097] From the detection results of Examples 1, 10-11 and Table 5, it can be seen that when 98% ethanol or ethanol is used for crystallization and purification in step S3, a higher yield can be obtained; because compared with methanol, ethanol has a poorer solubility for the product, which makes more product precipitate, which is conducive to improving the yield.

[0098] In addition, the synthesis method of the present application can significantly reduce the cost of 2'-OMe adenosine; by comparing the raw materials used in the present application with those used in the prior art, for example, in the second synthesis method "2-amino-6-chloropurine nucleoside realizes selective methylation of 2'-OH under MeI / NaH conditions", the cost of the raw material is 20-30 times that of the adenosine of the present application; in the third synthesis method "3', 5'-OH of adenosine is protected by TIPDS, then 2'-OH is methylated, and finally TIPDS is removed to obtain 2'-OMe adenosine", the cost of the raw material is reduced by 30%, therefore, the present application also has excellent economic value.

Claims

1. A method for synthesizing 2'-OMe adenosine, characterized in that, Includes the following steps: S1. Adenosine is dissolved in DMF and reacted with a methylating agent in the presence of a base to obtain a mixture of 2'-OMe adenosine and 3'-OMe adenosine; A mixture of S2,2'-OMe adenosine and 3'-OMe adenosine was reacted with acetic anhydride in a solvent to give an acetylated mixture; S3. The acetylated mixture is deacetylated under the action of ammonia and crystallized to give 2'-OMe adenosine; In step S1, the amount of alkali added is 1.2 to 2.5 times the molar amount of adenosine, and the amount of methylation reagent added is 1.0 to 2.0 times the molar amount of adenosine. In step S2, the amount of acetic anhydride fed is 2.2 to 4.0 eq of the molar amount of adenosine in step S1; In step S1, the base is potassium hydroxide; In step S1, the methylating agent is methyl p-toluenesulfonate; In step S2, the solvent is dichloromethane; The solvent used for crystallization was 98 wt% ethanol.

2. The method for synthesizing 2'-OMe adenosine according to claim 1, characterized in that: In step S3, ammonia is any one or a combination of several of ammonia water, a methanol solution of ammonia, and an ethanol solution of ammonia.

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