Preparation method of carbohydrate compound

Abstract

The invention discloses a preparation method of a carbohydrate compound. The invention provides a preparation method of a carbohydrate compound, which comprises the following steps: in an organic solvent, in the presence of alkali, carrying out hydroxyl protection reaction on a compound 1 and 4, 4-dimethoxytriphenylchloromethane to obtain the carbohydrate compound as shown in a compound 2, the molar ratio of the 4, 4-dimethoxy triphenyl chloromethane to the compound 1 is (0.8 to 1.5): 1. The method has the advantages that by-products are turned into wealth from wastes, the synthesis efficiency is high, the reaction conditions are mild, the specific reaction of target sites can be realized, the conversion rate is high, the by-products are few, the purification method is simple and convenient, and the like. The preparation method has the advantages of simplicity and convenience in operation and post-treatment, high yield, high product purity and the like. # imgabs0 #

Description

Technical Field

[0001] The invention relates to a method for preparing a carbohydrate compound. Background Art

[0002] GalNAc (N-acetylgalactosamine) coupling modification is mainly used for drug delivery of oligonucleotide drugs to the liver. GalNAc has a good affinity for asialoglycoprotein (ASGPR), which only exists in liver parenchymal cells. Therefore, the mechanism of oligonucleotide drug delivery involving GalNAc has great application potential in the treatment of liver-related diseases involving gene overexpression.

[0003] The traditional method for synthesizing DMTr-protected GalNAc intermediates is as follows:

[0004]

[0005] or:

[0006]

[0007] The GalNAc intermediate is usually produced by an acid-amine condensation reaction of a compound as shown in Formula 1 and a compound as shown in Formula 3, or a compound as shown in Formula 4 and a compound as shown in Formula 5. During the post-treatment and storage of the GalNAc intermediate, the DMTr protecting group is easily removed, thereby generating a byproduct without a DMTr structure. Since it is relatively late in the route, it affects the yield and cost. Summary of the invention

[0008] The technical problem to be solved by the present invention is to provide a method for preparing a saccharide compound in view of the fact that by-products are easily produced in the prior art, which affects the yield and cost. The preparation method of the present invention has the advantages of simple operation and post-treatment, high yield, high product purity, etc.

[0009] The present invention provides a method for preparing a carbohydrate compound, which comprises the following steps: in an organic solvent, in the presence of a base, compound 1 and 4,4-dimethoxytriphenylmethane undergo a hydroxyl protection reaction to obtain a carbohydrate compound as shown in compound 2; the molar ratio of the 4,4-dimethoxytriphenylmethane to the compound 1 is (0.8-1.5):1;

[0010]

[0011] In a certain embodiment, the organic solvent may be one or more of a halogenated hydrocarbon solvent, an amide solvent, a nitrile solvent, an ether solvent and an ester solvent; the halogenated hydrocarbon solvent may be dichloromethane; the amide solvent may be N,N-dimethylformamide (DMF); the nitrile solvent may be acetonitrile; the ether solvent may be tetrahydrofuran; the ester solvent may be ethyl acetate; the organic solvent is preferably dichloromethane; more preferably anhydrous dichloromethane.

[0012] In a certain embodiment, the volume mass ratio of the organic solvent to the compound 1 can be (5-30):1 mL / g, for example 10:1 mL / g.

[0013] In a certain embodiment, the base can be an organic base, such as triethylamine, pyridine, 2,6-lutidine or N-methylmorpholine (NMM), preferably triethylamine.

[0014] In one embodiment, the molar ratio of the base to the compound 1 can be (1-2):1, for example 1.23:1.

[0015] In one embodiment, the molar ratio of the 4,4-dimethoxytriphenylmethane to the compound 1 is 1:1.

[0016] The reaction temperature of the hydroxyl protection reaction is a conventional temperature for such reactions in the art, for example, 25°C.

[0017] The progress of the hydroxyl protection reaction is monitored by conventional monitoring methods in the art (eg, TLC, HPLC, LCMS or NMR), and the end point of the reaction is generally when 4,4-dimethoxytriphenylmethane disappears or stops reacting.

[0018] In a certain embodiment, the preparation method of the saccharide compound may further include post-treatment, and the post-treatment operation is a conventional post-treatment operation in the preparation method of this type in the art. For example, the preparation method may further include the following steps after the hydroxyl protection reaction is completed: quenching, extraction, drying, and concentration.

[0019] The quenching is preferably carried out with a saturated sodium bicarbonate aqueous solution, the extraction is preferably carried out with dichloromethane, and the drying is preferably carried out with anhydrous sodium sulfate.

[0020] In a certain embodiment, the preparation method of the carbohydrate compound can also be carried out in the presence of the carbohydrate compound shown as compound 2. The mass content of the compound 1 in the substrate of the hydroxyl protection reaction is 60% to 99.99%, for example 85%.

[0021] The preparation method of the carbohydrate compound may further include a refining method, which may further include the following steps: using preparative high performance liquid chromatography to gradient elute the product of the hydroxyl protection reaction; wherein the mobile phase of the gradient elution is a water-acetonitrile system.

[0022] In a certain embodiment, in the preparative high performance liquid chromatography, the chromatographic column may be Welch XtimateC18 250×100mm#10μm.

[0023] In a certain embodiment, in the gradient elution, based on the total volume of the mobile phase as 100%, the volume fraction of acetonitrile in the mobile phase is 20% to 80%, and more preferably 30% to 60%; preferably, at 20-25 minutes, the volume fraction of acetonitrile in the mobile phase increases from 25-35% to 55-65%; optimally, at 23 minutes, the volume fraction of acetonitrile in the mobile phase increases from 30% to 60%.

[0024] The present invention accurately calculates the content of the GalNAc intermediate by mass nuclear magnetic resonance, and then calculates the required amount of DMTrCl, and then completes the DMTr protection of the 6-hydroxyl group in the GalNAc intermediate in a one-step method, thereby realizing a specific reaction of the target site. The preparation method has the advantages of simple operation and post-processing, high yield, and high product purity.

[0025] Without violating the common sense in the art, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present invention.

[0026] The reagents and raw materials used in the present invention are commercially available.

[0027] The positive progress of the present invention is that the present invention has the advantages of turning by-products into treasures, high synthesis efficiency, mild reaction conditions, the ability to achieve a specific reaction of a target site, high conversion rate, few by-products, and a simple purification method. The preparation method has the advantages of simple operation and post-processing, high yield, and high product purity. DETAILED DESCRIPTION

[0028] The present invention is further described below by way of examples, but the present invention is not limited to the scope of the examples. The experimental methods in the following examples without specifying specific conditions are carried out according to conventional methods and conditions, or selected according to the product specifications.

[0029] Example 1

[0030]

[0031] Compound 1 (21 g, 85% by mass, 1 eq) was added to anhydrous dichloromethane (210 mL), triethylamine (1.06 g, 10.47 mmol, 1.23 eq) was added to the reaction, and then 4,4-dimethoxytriphenylmethane (2.78 g, 8.48 mmol, 1 eq) was added to the reaction. After the addition was completed, the reaction solution was controlled to stir and react for 16 hours at 25°C. LCMS monitored the reaction to be complete, and the reaction solution was poured into a saturated sodium bicarbonate aqueous solution (200 mL) to quench the reaction, and the extraction was separated. The aqueous phase was extracted twice with dichloromethane (210 mL), the organic phases were combined, dried and filtered with sodium sulfate, and concentrated with a rotary evaporator to obtain a yellow oily crude product.

[0032] The crude product was separated by preparative high performance liquid chromatography {stationary phase: Welch Xtimate C18 250×100mm#10μm; mobile phase: [water-acetonitrile system]; gradient: 30%-60% acetonitrile (23.0min)} to obtain a white solid, which was the pure product of the carbohydrate compound shown as compound 2 (19.0g, yield 93%).

[0033] Structural characterization of carbohydrate compounds as shown in compound 2:

[0034] MS-ESI: m / 2z = 1202.2 [M / 2-H] - .

[0035] 1 H NMR(400MHz DMSO-d6): δ7.86-7.79(m,6H),7.73(t,J=5.6Hz,3H),7.34-7.26(m,4H),7.25-7.13(m,5H),6.98(s,1H),6.90-6.83(m,4H),5. 21(d,J=3.2Hz,3H),5.00-4.88(m,4H),4.49(d,J=8.4Hz,3H),4.43-4.35(m,1H),4.19-4.09(m,1H),4.06-3.98(m,9H),3.95-3 .81(m,3H),3.75-3.67(m,9H),3.60-3.48(m,13H),3.44-3.37(m,3H),3.20-3.12(m,1H),3.09-2.96(m,14H),2.27(t,J=6.4Hz ,6H),2.23-2.15(m,2H),2.10(s,9H),2.04(t,J=7.2Hz,8H),1.99(s,9H),1.77(s,9H),1.55-1.37(m,22H),1.33-1.09(m,12H).

Claims

1. A method for preparing a carbohydrate compound, characterized in that: The method comprises the following steps: in an organic solvent, in the presence of a base, compound 1 and 4,4-dimethoxytriphenylmethane undergo a hydroxyl protection reaction to obtain a carbohydrate compound as shown in compound 2; the molar ratio of the 4,4-dimethoxytriphenylmethane to the compound 1 is (0.8-1.5):1; 2. The preparation method according to claim 1, characterized in that It meets one or more of the following conditions: (1) The organic solvent is one or more of a halogenated hydrocarbon solvent, an amide solvent, a nitrile solvent, an ether solvent and an ester solvent; (2) The volume mass ratio of the organic solvent to the compound 1 is (5-30): 1 mL / g; (3) the base is an organic base; and (4) The molar ratio of the base to the compound 1 is (1-2):

1.

3. The preparation method according to claim 2, characterized in that: It meets one or more of the following conditions: (1) The halogenated hydrocarbon solvent is dichloromethane; (2) The amide solvent is N,N-dimethylformamide; (3) The nitrile solvent is acetonitrile; (4) The ether solvent is tetrahydrofuran; (5) The ester solvent is ethyl acetate; (6) The volume mass ratio of the organic solvent to the compound 1 is 10:1 mL / g; (7) The base is triethylamine, pyridine, 2,6-lutidine or N-methylmorpholine; and (8) The molar ratio of the base to the compound 1 is 1.23:

1.

4. The preparation method according to claim 3, characterized in that: It meets one or both of the following conditions: (1) The organic solvent is dichloromethane; and (2) The base is triethylamine.

5. The preparation method according to claim 1, characterized in that: The preparation method further comprises the following steps after the hydroxyl protection reaction is completed: quenching, extraction, drying and concentration.

6. The preparation method according to claim 1, characterized in that: The preparation method further comprises the following steps: using preparative high performance liquid chromatography to gradient elute the product of the hydroxyl protection reaction; wherein the mobile phase of the gradient elution is a water-acetonitrile system.

7. The preparation method according to claim 6, characterized in that: In the gradient elution, based on the total volume of the mobile phase being 100%, the volume fraction of acetonitrile in the mobile phase is 20% to 80%, or 30% to 60%.

8. The preparation method according to claim 7, characterized in that: In the gradient elution, based on the total volume of the mobile phase as 100%, at 20-25 min, the volume fraction of acetonitrile in the mobile phase increases from 25-35% to 55-65%; optimally, at 23 min, the volume fraction of acetonitrile in the mobile phase increases from 30% to 60%.

9. The preparation method according to claim 1, characterized in that: The reaction is also carried out in the presence of the carbohydrate compound shown as compound 2. At this time, the mass content of compound 1 in the substrate of the hydroxyl protection reaction is 60% to 99.99%.

10. The preparation method according to claim 9, characterized in that: The mass content of the compound 1 in the substrate of the hydroxyl protection reaction is 85%.

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