CN101974051B - Method for synthesizing azacitidine - Google Patents
Method for synthesizing azacitidine Download PDFInfo
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Abstract
The invention relates to the field of pharmaceutical synthesis process and discloses a method for synthesizing azacitidine. The method comprises the following steps: performing silanization reaction on bis(trimethyl disilylamine) and 5-azacytosine by taking toluene as solvent under the catalysis of phase transfer catalyst of quaternary ammonium compounds under a moisture-isolated condition to generate a compound with structure of formula II; performing condensation reaction between the compound and ribose tetracetate to generate a compound with a structure of formula IV; and performing alcoholysis on the compound in an alkaline environment to generate the azacitidine. The synthesis method of the invention shortens the time of silanization reaction, reduces the waste amount of the bis(trimethyl disilylamine) and avoids influence of the moisture in methanol and ethanol in the purification stage on the purity of the finished product of azacitidine; moreover, the synthesis method has the advantages of cheap reagents, few reaction steps and mild reaction conditions and is favorable for industrial production.
Description
Technical field
The present invention relates to the medicine synthesising process field, be specifically related to a kind of compound method of azacitidine.
Background technology
Azacitidine, molecular formula are C
8H
12N
4O
5, English Azacitidine by name, chemistry 1-(β-D-ribofuranosyl) by name-4-amino-1,3,5-triazines-2 (1H)-ketone has another name called 5-azacytidine, azacytidine, has suc as formula structure shown in the V:
Azacitidine is a white, needle-shaped crystals, belongs to the cytosine nucleoside medicine, is by the dnmt rna suppressor factor of U.S. Pharmion company exploitation, is mainly used in the treatment myelodysplastic syndrome clinically.Azacitidine is first antitumor drug based on the unusual exploitation of tumour expressing gene, can directly mix among the DNA, and it is synthetic to suppress DNA and RNA, can kill and wound the cell that is in the S phase, for mammary cancer, intestinal cancer, melanoma etc. certain curative effect is arranged.Its mechanism of action is that azacitidine is through being attached to after the phosphorylation on the dna molecular; Dnmt rna and azacitidine generation methylation reaction; Form the covalent attachment product, the activity inhibited of dnmt rna is also degraded, and causes that the dna methylation level reduces in the tumor tissues; Hyper-methylation cancer suppressor gene demethylation suppresses tumour cell thereby make gene recover to express.
At present; The compound method that azacitidine is commonly used is that 5-azepine cytosine(Cyt) carries out Silanization reaction with two trimethylammonium two silicon amine (HMDS) under the catalysis of ammonium sulfate, the N-of generation is trimethyl silicon based-and the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine and tetrem acyl ribose carry out condensation under the effect of catalyzer; The 1-(2,3,5-three-O-ethanoyl-β-D-ribofuranosyl) that generates-4-amino-1; 3; 5-triazine-2 (1H)-ketone generates the azacitidine bullion with the methanol solution of sodium methylate or the methanol solution hydrolysis deprotection of logical ammonia, and bullion obtains the azacitidine finished product through the organic solvent recrystallization, like the described compound method of patent US7038038, US3819780 and US55591012014.
Above-mentioned compound method general efficient when carrying out Silanization reaction is lower, consuming time longer.Simultaneously, synthetic middle a large amount of HMDS that adopt have produced a large amount of HMDS waste liquids.In addition, all adopted methyl alcohol or alcohol solvent to carry out purifying in above-mentioned each method of purification phase, and methyl alcohol or ethanol are hydrophilic solvent, azacitidine is very easily destroyed in water, has influenced azacitidine finished product purity.
Summary of the invention
In view of this, the object of the present invention is to provide a kind of compound method of azacitidine, this compound method can shorten the Silanization reaction time, improves reaction efficiency, reduces two trimethylammonium two silicon amine (HMDS) waste liquid amount simultaneously.
The present invention provides a kind of compound method of azacitidine, may further comprise the steps:
Step 1, under isolated moisture condition; Two trimethylammonium two silicon amine and 5-azepine cytosine(Cyt) are solvent with toluene; Under quaternary ammonium salt-type phase transfer catalyst catalysis, carry out Silanization reaction; Generation has the compound of formula II structure, and the mol ratio of said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.005-0.5;
Step 2, formula II compound and tetrem acyl ribose carry out condensation reaction and generate the compound with formula IV structure, and the alcoholysis under alkaline environment of formula IV compound generates azacitidine;
The said Silanization reaction of step 1 is by two trimethylammonium two silicon amine (HMDS) amino and hydroxyl on the triazine ring of 5-azepine cytosine(Cyt) with formula I structure to be protected, generate N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; The 5-triazine, promptly said formula II compound, temperature of reaction is 70-125 ℃; Be preferably 100-125 ℃; Reaction times is 4-48h, is preferably 6-12h, and reaction equation is following:
Wherein, HMDS Chinese two trimethylammonium two silicon amine or hmds by name, molecular formula is C
6H
19NSi
2, be a kind of basic silane protective material.
In existing compound method; Silanization reaction generally adopts a large amount of HMDS to carry out, and is wherein a part of as reaction raw materials, and the overwhelming majority is as reaction solvent; So just make and produce a large amount of HMDS waste liquids in the Silanization reaction process; Therefore the present invention all replaces the HMDS as reaction solvent with toluene, has so both reduced the waste liquid amount of HMDS, has practiced thrift cost again.Say that from reaction principle reaction solvent is with a kind of in acetonitrile, toluene, ethylene dichloride, the trichloroethane or wherein plant mixed solution arbitrarily and substitute the HMDS as the solvent part, all can reduce the waste liquid amount of HMDS.But, in the existing feasible solvent, have only the boiling point of toluene higher, relatively be beneficial to Silanization reaction, consuming time shorter, the boiling point of mixed solution all is lower than the optimal temperature of Silanization reaction between other solvents or all kinds of SOLVENTS, and the reaction times is prolonged.
Simultaneously, Silanization reaction adopts ammonium sulfate as catalyzer usually in the existing compound method, but catalytic efficiency (is not high, so the present invention adopts quaternary ammonium salt-type phase transfer catalyst to replace ammonium sulfate, is shortened in the reaction times greatly.Phase-transfer catalysis is called for short PT, is in organic synthesis, to have used a kind of widely day by day new synthetic technology since the seventies in 20th century.In organic synthesis, often run into heterogeneous organic reaction, the common speed of this type reaction is very slow, and yield is low.If but use water-soluble inorganic salt; With the little organic solvent dissolution organism of polarity, and add the quaternary ammonium salt or the quaternary alkylphosphonium salt of (below the 0.05mol) on a small quantity, reaction then is easy to carry out; This type can impel and improve speed of response and at the salt of two alternate transfer negative ions, be called phase-transfer catalyst.
The present invention adopts toluene as solvent, with quaternary ammonium salt-type phase transfer catalyst catalysis, not only can shorten the Silanization reaction time, improve reaction efficiency, has also reduced the waste liquid amount of HMDS.
Wherein, as preferably, said quaternary ammonium salt-type phase transfer catalyst is 4-butyl ammonium hydrogen sulfate, Tetrabutyl amonium bromide, tetrabutylammonium chloride, tetrabutylammonium iodide, benzyltriethylammoinium chloride or benzyl triethyl ammonium bromide.
Wherein, the said toluene consumption of step 1 is that every 1g 5-azepine cytosine(Cyt) adds toluene 10-25ml, and said 5-azepine cytosine(Cyt) is 1 with the mol ratio of two trimethylammonium two silicon amine: 2.5-5.
As preferably, the said 5-azepine of step 1 cytosine(Cyt) is 1: 2.6 with the mol ratio of two trimethylammonium two silicon amine.
As preferably, the mol ratio of step 1 said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.01-0.1.
Step 2 Chinese style II compound carries out condensation reaction with the tetrem acyl ribose with formula III structure, generates 1-(2,3,5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1; 3,5-triazine-2 (1H)-ketone, i.e. formula IV compound; The alcoholysis deacetylation generates the azacitidine with formula V structure under the formula IV compound alkaline condition, and temperature of reaction is-20-50 ℃, is preferably-5-30 ℃; Reaction times is 4-24h, is preferably 4-12h, and reaction equation is following:
In existing compound method; Condensation reaction catalyzer commonly used is trifluoromethyl trimethylammonium silicone grease (TMS-triflate) and tin tetrachloride; But tin tetrachloride is difficult for Ex-all at subsequent reactions with in handling; Very easily cause the finished product heavy metals exceeding standard, so the present invention adopts the catalyzer of trifluoromethyl trimethylammonium silicone grease as condensation reaction.
In the prior art; The alcoholysis deacetylation generally adopts the methanol solution of sodium methylate or the methanol solution of logical ammonia; Through further investigation; Hydrolysis deprotection of the present invention adopts the methanol solution of NaOH to carry out the alcoholysis deacetylation, can reach the deacetylation effect of methanol solution of methanol solution and the logical ammonia of sodium methylate equally, and azacitidine bullion purity is also higher.Therefore; The present invention carries out the deacetylation except the methanol solution that can adopt sodium methylate or the methanol solution of logical ammonia, can also adopt the methanol solution of NaOH to carry out the alcoholysis deacetylation, and temperature of reaction is-20-50 ℃; Be preferably-5-30 ℃; Reaction times is 0.5-10h, is preferably 0.5-4h, adopts the latter on cost, can obtain saving.
Moisture can influence whole building-up reactions, therefore in entire synthesis process, needs isolated moisture.The present invention adopts and feeds the isolated moisture of nitrogen.
Compound method according to the invention also comprises with N-Methyl pyrrolidone and hexanaphthene purifying azacitidine.At present, all adopt methyl alcohol or alcohol solvent to carry out purifying in existing each method of purification phase, and methyl alcohol or ethanol are hydrophilic solvent, azacitidine is very easily destroyed in water, has influenced azacitidine finished product purity.Use hexanaphthene to replace methyl alcohol or ethanol during purifying of the present invention, hexanaphthene is a hydrophobic solvent, uses the hexanaphthene purifying that product is reduced the loss during with the baking material at purifying, improves product purity.In addition, N-Methyl pyrrolidone and hexanaphthene are two kind solvents in ICH (the human drug legislation technology) guide, and limit is higher, uses the absolute ethanol washing filter cake during suction filtration, can guarantee that dissolvent residual is qualified.Wherein, said N-Methyl pyrrolidone and hexanaphthene consumption are that every 1g azacitidine adds N-Methyl pyrrolidone 5-20ml and hexanaphthene 5-20ml.
The compound method of azacitidine according to the invention; Shortened the Silanization reaction time; Improved reaction efficiency, reduced the HMDS waste liquid amount simultaneously, avoided in purification phase methyl alcohol and the ethanol moisture the influence of azacitidine finished product purity; And agents useful for same is cheap, reactions step is few, reaction conditions is gentle, helps suitability for industrialized production.
Embodiment
The invention discloses a kind of compound method of azacitidine, those skilled in the art can use for reference this paper content, suitably improve processing parameter and realize.Special needs to be pointed out is that all similarly replace and change apparent to those skilled in the art, they all are regarded as and are included in the present invention.Compound method of the present invention is described through preferred embodiment; The related personnel obviously can change or suitably change and combination methods and applications as herein described in not breaking away from content of the present invention, spirit and scope, realizes and use technology of the present invention.
According to the present invention, the compound method of said azacitidine may further comprise the steps:
Step 1, under isolated moisture condition; Two trimethylammonium two silicon amine and 5-azepine cytosine(Cyt) are solvent with toluene; Under quaternary ammonium salt-type phase transfer catalyst catalysis, carry out Silanization reaction; Generation has the compound of formula II structure, and the mol ratio of said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.005-0.5;
Step 2, formula II compound and tetrem acyl ribose carry out condensation reaction and generate the compound with formula IV structure, and the alcoholysis under alkaline environment of formula IV compound generates azacitidine;
Wherein, as preferably, said quaternary ammonium salt-type phase transfer catalyst is 4-butyl ammonium hydrogen sulfate, Tetrabutyl amonium bromide, tetrabutylammonium chloride, tetrabutylammonium iodide, benzyltriethylammoinium chloride or benzyl triethyl ammonium bromide.
Wherein, the said toluene consumption of step 1 is that every 1g 5-azepine cytosine(Cyt) adds toluene 10-25ml, and said 5-azepine cytosine(Cyt) is 1 with the mol ratio of two trimethylammonium two silicon amine: 2.5-5.
As preferably, the said 5-azepine of step 1 cytosine(Cyt) is 1: 2.6 with the mol ratio of two trimethylammonium two silicon amine.
As preferably, the mol ratio of step 1 said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.01-0.1.
Wherein, according to the invention under alkaline environment alcoholysis for carrying out alcoholysis with the methanol solution of NaOH; Said exclusion of water is divided into the isolated moisture of feeding nitrogen.
Compound method according to the invention also comprises with N-Methyl pyrrolidone and hexanaphthene purifying azacitidine.
Wherein, said N-Methyl pyrrolidone and hexanaphthene consumption are that every 1g azacitidine adds N-Methyl pyrrolidone 5-20ml and hexanaphthene 5-20ml.
The present invention adopts quaternary ammonium salt-type phase transfer catalyst catalysis Silanization reaction than adopting the ammonium sulfate catalysis used time of Silanization reaction to shorten about 50%; Owing to adopt toluene to replace most of HMDS, make the HMDS waste liquid amount reduce 65-85%; Use hexanaphthene to replace methyl alcohol or ethanol during purifying of the present invention, avoid in purification phase methyl alcohol and the ethanol moisture the influence of azacitidine finished product purity.
Below in conjunction with embodiment, further set forth the present invention, raw materials used and reagent is the commercially available prod among the embodiment.
Embodiment 1: the compound method of azacitidine according to the invention
Under nitrogen protection, 25.8g (0.23mol) 5-azepine cytosine(Cyt), 1.0g (0.003mol) 4-butyl ammonium hydrogen sulfate are put in the 1L there-necked flask, add the stirring of 125ml HMDS and 500ml toluene and be warming up to 125 ℃; The solution clarification of the 6h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 65.0g (0.25mol).
Under nitrogen protection, 65.0g (0.25mol) formula II compound with after the 500ml THF dissolving, is dropped in the 1L there-necked flask and stirs, add 120.0g (0.38mol) tetrem acyl ribose again, wash feed hopper with the 100ml THF again; Normal temperature slowly drips 64.5ml (0.36mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 4h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 500ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.2% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 70.2g (0.19mol).
Under nitrogen protection, the formula IV compound 70.2g (0.19mol) that obtains is dropped in the 2L there-necked flask, add 1000ml methyl alcohol stirring and dissolving; 4.0g sodium hydroxide is dissolved in the 20ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 1h, the cooling suction filtration gets azacitidine bullion 34.0g; With 80 ℃ of dissolvings of 400ml N-Methyl pyrrolidone; Drip 400ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 26.5g with absolute ethanol washing.
Embodiment 2: the compound method of azacitidine according to the invention
Under nitrogen protection, 25.8g (0.23mol) 5-azepine cytosine(Cyt), 0.4g (0.001mol) Tetrabutyl amonium bromide are put in the 1L there-necked flask, add the stirring of 120ml (0.575mol) HMDS and 258ml toluene and be warming up to 125 ℃; The solution clarification of the 5h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 62.0g (0.24mol).
Under nitrogen protection, 62.0g (0.24mol) formula II compound with after the 500ml THF dissolving, is dropped in the 1L there-necked flask and stirs, add 115.0g (0.36mol) tetrem acyl ribose again, wash feed hopper with the 100ml THF again; Normal temperature slowly drips 64.5ml (0.36mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 4h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 450ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.2% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 67.0g (0.18mol).
Under nitrogen protection, the formula IV compound 67.0g (0.18mol) that obtains is dropped in the 2L there-necked flask, add 900ml methyl alcohol stirring and dissolving; 3.8g sodium hydroxide is dissolved in the 20ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 0.5h, the cooling suction filtration gets azacitidine bullion 29.0g; With 80 ℃ of dissolvings of 400ml N-Methyl pyrrolidone; Drip 400ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 24.8g with absolute ethanol washing.
Embodiment 3: the compound method of azacitidine according to the invention
Under nitrogen protection, 25.8g (0.23mol) 5-azepine cytosine(Cyt), 38.2g (0.115mol) tetrabutylammonium chloride are put in the 1L there-necked flask, add the stirring of 150ml (0.72mol) HMDS and 650ml toluene and be warming up to 125 ℃; The solution clarification of the 4h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 85.0g (0.33mol).
Under nitrogen protection, 85.0g (0.33mol) formula II compound with after the 500ml THF dissolving, is dropped in the 1L there-necked flask and stirs, add 120.0g (0.38mol) tetrem acyl ribose again, wash feed hopper with the 100ml THF again; Normal temperature slowly drips 64.5ml (0.36mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 7h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 500ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.2% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 74.0g (0.20mol).
Under nitrogen protection, the formula IV compound 74.0g (0.20mol) that obtains is dropped in the 2L there-necked flask, add 950ml methyl alcohol stirring and dissolving; 5.0g sodium hydroxide is dissolved in the 20ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 1h, the cooling suction filtration gets azacitidine bullion 34.0g; With 80 ℃ of dissolvings of 400ml N-Methyl pyrrolidone; Drip 400ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 28.5g with absolute ethanol washing.
Embodiment 4: the compound method of azacitidine according to the invention
Under nitrogen protection, 100.0g (0.89mol) 5-azepine cytosine(Cyt), 3.0g (0.009mol) tetrabutylammonium iodide are put in the 3L there-necked flask, add the stirring of 480ml (2.3mol) HMDS and 1900ml toluene and be warming up to 125 ℃; The solution clarification of the 8h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 300.0g (1.15mol).
Under nitrogen protection, 300.0g (1.15mol) formula II compound with after the 1500ml THF dissolving, is dropped in the 3L there-necked flask and stirs, add 480.0g (1.52mol) tetrem acyl ribose again, wash feed hopper with the 300ml THF again; Normal temperature slowly drips 260.0ml (1.45mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 8h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 1500ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.5% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 320.7g (0.87mol).
Under nitrogen protection, the formula IV compound 320.7g (0.87mol) that obtains is dropped in the 5L there-necked flask, add 3000ml methyl alcohol stirring and dissolving; 20.0g sodium hydroxide is dissolved in the 300ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 1.5h, the cooling suction filtration gets azacitidine bullion 112.6g; With 80 ℃ of dissolvings of 2000ml N-Methyl pyrrolidone; Drip 2000ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 90.5g with absolute ethanol washing.
Embodiment 5: the compound method of azacitidine according to the invention
Under nitrogen protection, 25.8g (0.23mol) 5-azepine cytosine(Cyt), 7.7g (0.023mol) benzyltriethylammoinium chloride are put in the 1L there-necked flask, add the stirring of 144ml (0.69mol) HMDS and 570ml toluene and be warming up to 125 ℃; The solution clarification of the 5h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 68.0g (0.25mol).
Under nitrogen protection, 68.0g (0.26mol) formula II compound with after the 500ml THF dissolving, is dropped in the 1L there-necked flask and stirs, add 120.0g (0.38mol) tetrem acyl ribose again, wash feed hopper with the 100ml THF again; Normal temperature slowly drips 64.5ml (0.36mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 6h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 500ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.2% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 72.1g (0.20mol).
Under nitrogen protection, the formula IV compound 72.1g (0.20mol) that obtains is dropped in the 2L there-necked flask, add 1000ml methyl alcohol stirring and dissolving; 4.0g sodium hydroxide is dissolved in the 20ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 0.5h, the cooling suction filtration gets azacitidine bullion 36.0g; With 80 ℃ of dissolvings of 360ml N-Methyl pyrrolidone; Drip 360ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 28.4g with absolute ethanol washing.
Embodiment 6: the compound method of azacitidine according to the invention
Under nitrogen protection, 100.0g (0.89mol) 5-azepine cytosine(Cyt), 15.0g (0.044mol) benzyl triethyl ammonium bromide are put in the 3L there-necked flask, add the stirring of 593ml (3.65mol) HMDS and 1500ml toluene and be warming up to 125 ℃; The solution clarification of the 6h left and right sides; Decompression is steamed solvent to constant weight then, N-trimethyl silicon based-the trimethyl silicon based oxygen base of 4--2-amine-1,3; 5-triazine, i.e. formula II compound 284.3g (1.09mol).
Under nitrogen protection, 284.3g (1.09mol) formula II compound with after the 1500ml THF dissolving, is dropped in the 1L there-necked flask and stirs, add 420.0g (1.33mol) tetrem acyl ribose again, wash feed hopper with the 300ml THF again; Normal temperature slowly drips 260.0ml (1.45mol) trifluoromethyl trimethylammonium silicone grease down, and stirring reaction behind the reaction 8h, is concentrated into dried at 55 ℃ reaction solution; Oily matter dissolves with vinyl trichloride, and adding a large amount of saturated sodium carbonate solutions to pH value of solution then is 9-10; Separatory, water layer extracts with the 500ml vinyl trichloride at twice; Merge organic layer, organic layer spends the night with anhydrous magnesium sulfate drying, and diatomite filtration is evaporated to dried; 70 ℃ of dry 6h dry by the fire to moisture and are lower than 0.5% (moisture low more be beneficial to down more step reaction), 1-(2,3; 5-three-O-ethanoyl-β-D-ribofuranosyl)-4-amino-1,3,5-triazines-2 (1H)-ketone, i.e. formula IV compound 314.6g (0.85mol).
Under nitrogen protection, the formula IV compound 314.6g (0.85mol) that obtains is dropped in the 5L there-necked flask, add 3000ml methyl alcohol stirring and dissolving; 16.0g sodium hydroxide is dissolved in the 270ml methyl alcohol, under agitation sodium hydroxide-methanol solution is slowly dripped in the there-necked flask stirring reaction; Behind the reaction 0.7h, the cooling suction filtration gets azacitidine bullion 136.3g; With 80 ℃ of dissolvings of 1500ml N-Methyl pyrrolidone; Drip 1500ml hexanaphthene crystallization, suction filtration, filter cake obtains azacitidine finished product 112.5g with absolute ethanol washing.
Embodiment 7: compound method Silanization reaction according to the invention efficiency analysis
Respectively embodiment 1 to embodiment 6 synthetic formula II compound is taken a sample; The HPLC external standard method detects its purity; And according to embodiment 1 to embodiment 6 Silanization reaction condition; Substitute quaternary ammonium salt-type phase transfer catalyst with ammonium sulfate, required time when writing down formula II compound that ammonium sulfate catalysis generates respectively and reaching embodiment 1 to embodiment 6 synthetic formula II compound equal extent purity, the result is referring to table 1.
Wherein, the HPLC chromatographic column is anti-phase C18 post (5 μ m, 250 * 4.6mm); Mobile phase A is the potassium hydrogenphosphate of 15mM and the ammonium acetate mixing solutions of 15mM, and pH is adjusted to 6.5-6.8 with the phosphoric acid (10ml 85% phosphoric acid and 100ml water) of dilution; Mobile phase B is that the mixing solutions mobile phase A of mobile phase A and acetonitrile and the volume ratio of acetonitrile are 3: 2, and concrete reaction conditions is referring to table 2.
The efficiency analysis of table 1 Silanization reaction
Table 2HPLC reaction conditions
Working time | Flow velocity | Detect wavelength | Column temperature | Sample size | Diluent | Mobile phase A | Mobile phase B |
50min | 0.7ml/min | 235nm | 15℃ | 2μl | DMSO | 55% | 45% |
The result shows; According to embodiment 1 to embodiment 6 Silanization reaction condition; Substituting quaternary ammonium salt-type phase transfer catalyst with ammonium sulfate reacts; Required time was about 13h when the formula II compound purity of its generation reached each embodiment synthetic formula II compound equal extent purity, and each embodiment required time is about 6-7.The Silanization reaction time that shows compound method according to the invention has obviously shortened about 50%.
Embodiment 8:HMDS waste liquid amount is analyzed
According to formula: the HMDS waste liquid percentage amounts of minimizing=solvent section H MDS input amount/total HMDS input amount * 100%=toluene input amount/(raw material HMDS input amount+toluene input amount) * 100%; HMDS waste liquid amount to the embodiment of the invention 1 to embodiment 6 reduces is added up; Owing to adopt toluene to replace most of HMDS; Make the input amount of HMDS greatly reduce, then HMDS waste liquid percentage amounts reduces accordingly, and the result is referring to table 3.
Table 3HMDS waste liquid amount is analyzed
Embodiment 1 | Embodiment 2 | Embodiment 3 | Embodiment 4 | Embodiment 5 | Embodiment 6 | |
The HMDS waste liquid percentage amounts that reduces | 80% | 68.25% | 81.25% | 79.83% | 79.83% | 71.67% |
The result shows that compound method according to the invention can effectively reduce the HMDS waste liquid amount.
Embodiment 9: the purity detecting of compound method synthetic azacitidine according to the invention
HPLC detection method with reference to embodiment 7 is carried out purity detecting to embodiment 1 to embodiment 6 synthetic azacitidine bullion and finished product, and the result shows each embodiment synthetic azacitidine bullion purity all more than 85%, and finished product purity is all more than 98%.Show that compound method according to the invention shortening the Silanization reaction time, improving on the basis of reaction efficiency and reduction HMDS waste liquid amount, can guarantee product purity simultaneously.In addition,, avoid the influence of purification phase methyl alcohol and ethanol, make higher purity of compound method synthetic azacitidine finished product maintenance according to the invention azacitidine finished product purity owing to use hexanaphthene to replace methyl alcohol or ethanol during purifying of the present invention.
The above only is a preferred implementation of the present invention; Should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the principle of the invention; Can also make some improvement and retouching, these improvement and retouching also should be regarded as protection scope of the present invention.
Claims (9)
1. the compound method of an azacitidine is characterized in that, may further comprise the steps:
Step 1, under isolated moisture condition; Two trimethylammonium two silicon amine and 5-azepine cytosine(Cyt) are solvent with toluene; Under quaternary ammonium salt-type phase transfer catalyst catalysis, carry out Silanization reaction; Generation has the compound of formula II structure, and the mol ratio of said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.005-0.5;
Step 2, formula II compound and tetrem acyl ribose carry out condensation reaction and generate the compound with formula IV structure, and the alcoholysis under alkaline environment of formula IV compound generates azacitidine;
Wherein, the said quaternary ammonium salt-type phase transfer catalyst of step 1 is 4-butyl ammonium hydrogen sulfate, Tetrabutyl amonium bromide, tetrabutylammonium chloride, tetrabutylammonium iodide, benzyltriethylammoinium chloride or benzyl triethyl ammonium bromide.
2. according to the said compound method of claim 1, it is characterized in that the said 5-azepine of step 1 cytosine(Cyt) is 1 with the mol ratio of two trimethylammonium two silicon amine: 2.5-5.
3. according to the said compound method of claim 1, it is characterized in that the said 5-azepine of step 1 cytosine(Cyt) is 1: 2.6 with the mol ratio of two trimethylammonium two silicon amine.
4. according to the said compound method of claim 1, it is characterized in that the said toluene consumption of step 1 is that every 1g 5-azepine cytosine(Cyt) adds toluene 10-25ml.
5. according to the said compound method of claim 1, it is characterized in that the mol ratio of step 1 said 5-azepine cytosine(Cyt) and quaternary ammonium salt-type phase transfer catalyst is 1: 0.01-0.1.
6. according to the said compound method of claim 1, it is characterized in that the said exclusion of water of step 1 is divided into the isolated moisture of feeding nitrogen.
7. according to the said compound method of claim 1, it is characterized in that, step 2 said under alkaline environment alcoholysis for carrying out alcoholysis with the methanol solution of NaOH.
8. according to the said compound method of claim 1, it is characterized in that, also comprise with N-Methyl pyrrolidone and hexanaphthene purifying azacitidine.
9. said according to Claim 8 compound method is characterized in that, said N-Methyl pyrrolidone and hexanaphthene consumption are that every 1g azacitidine adds N-Methyl pyrrolidone 5-20ml and hexanaphthene 5-20ml.
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CN103524584A (en) * | 2012-07-05 | 2014-01-22 | 天津九海医药科技有限公司 | Synthetic method of azacitidine |
CN103450303A (en) * | 2013-09-04 | 2013-12-18 | 重庆泰濠制药有限公司 | Azacitidine crystal form A and azacitidine crystal form B as well as preparation method thereof |
CN104672289A (en) * | 2013-11-29 | 2015-06-03 | 南京圣和药业股份有限公司 | Preparation method of azacitidine impurity |
CN104297361A (en) * | 2014-08-29 | 2015-01-21 | 四川汇宇制药有限公司 | Detection method for tetraacetylribose in azacitidine raw material |
CN110092807A (en) * | 2018-01-30 | 2019-08-06 | 中国医学科学院药物研究所 | A kind of method for preparing azacitidine of high-purity, low ignition residue |
CN112279881A (en) * | 2020-10-12 | 2021-01-29 | 福建南方制药股份有限公司 | Method for preparing antitumor drug azacitidine |
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