Preparation method of methylene blue

Abstract

The invention discloses a preparation method of methylene blue, which comprises the following steps: by taking phenothiazine as a raw material, carrying out bromination reaction to obtain 3, 7-dibromo-10H-phenothiazine, then carrying out acetylation reaction to obtain 3, 7-dibromo-10-acetyl phenothiazine, then carrying out ammonification reaction to obtain 3, 7-bis (dimethylamino)-10-acetyl phenothiazine, and finally, carrying out amination reaction to obtain 3, 7-bis (dimethylamino)-10-acetyl phenothiazine. And finally hydrolyzing and oxidizing to obtain methylene blue. The method is used for solving the technical problem of low purity of methylene blue.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceuticals, and in particular relates to a method for preparing methylene blue. Background Art

[0002] Methylene blue (Methylthioninium Chloride), chemical name: 3,7-bis(dimethylamino)phenothiazin-5-ium chloride trihydrate, its structural formula is as follows:

[0003]

[0004] Methylene blue, also known as methylene blue, methylene blue, and methylene blue, was first synthesized in the form of zinc oxide salt. Since 1876, methylene blue has been mainly used to treat methemoglobinemia caused by nitrites, chlorates, quinones, quinoneimines, aniline, and nitrobenzene. In addition, high concentrations of methylene blue can also be used to treat cyanide poisoning. In recent years, the clinical application of methylene blue has also been extended to the treatment of infectious and traumatic shock, cancer, herpes zoster, etc. Pharmacological studies have shown that methylene blue has detoxification, analgesia, antibacterial, antiviral, and anti-tumor effects, and can correct hemodynamic disorders in patients with septic shock and treat abnormal penile erections.

[0005] The use of methylene blue as a drug is not limited to its original synthesis year of 1876, but its use has been continuously expanded and optimized with the deepening of medical research and clinical application. From the initial treatment of methemoglobinemia, to the later treatment of cyanide poisoning, to a wider range of clinical applications, methylene blue plays an important role in the medical field. With the advancement of scientific research, the application prospects of methylene blue may be further expanded, providing new possibilities for the treatment of more diseases.

[0006] Methylene blue was first synthesized by German chemist H. Caro in 1876. It was first commercially produced and sold by BASF in 1876. It used dimethylaniline for nitrosation, then reduced to N,N-dimethyl-1,4-diaminobenzene, and used hydrogen sulfide and ferric chloride for oxidative coupling. Later, based on this process, after multiple rounds of evolution, dimethylaminoaniline was formed using dimethylaniline as the starting material, through nitrosation and reduction reactions. Dimethylaminoaniline was oxidized with thiosulfonic acid in sulfuric acid under the catalysis of zinc chloride to produce the thiosulfonic acid derivative of p-aminodimethylaniline, which was then oxidized twice to form methylene blue. The specific reaction process is as follows:

[0007]

[0008] The methylene blue produced by this process contains multiple impurities such as Azure A and Azure B, which have similar structures and polarities to methylene blue. It is impossible to obtain high-purity compounds through crystallization purification. In addition, a large amount of metal oxidants and excipients are used in this process, and methylene blue has a complexing effect on metal ions. It is also difficult to obtain methylene blue that meets the elemental impurity limits required by the pharmacopoeia through other treatment methods. Therefore, this route is difficult to provide high-quality API-grade methylene blue.

[0009] In 1997, N. Leventis et al. published an article for preparing MTC analogs from phenothiazine in Tetrahedron, Vol. 53, N°29, pp. 10083-10092. Leventis et al. described a two-step process, firstly converting phenothiazine into 3,7-dibromophenothiazin-5-ium bromide by bromine in acetic acid, and then using the above product to react with a disubstituted amine in ethanol or chloroform to form the bromide salt of the MTC analog. When dimethylamine is used to prepare methylene blue, the reported yield is 70% to 85%. However, this process also has some disadvantages: 1) This process uses 20 times the molar amount of bromine. If this process is used for industrial production of raw materials, the cost is high and the treatment of excess bromine is difficult, which puts great pressure on environmental protection; 2) Chloroform and ethanol are used as solvents, among which chloroform is highly toxic and not suitable for industrial production, while ethanol easily competes with dimethylamine in nucleophilic substitution reaction to form by-products. This method also has certain advantages. Its raw materials, phenothiazine, dimethylamine and bromine, are large chemical products with low prices.

[0010] Based on the work of N.Leventis et al., CN110418784A discloses a method for preparing 3,7-bis(dimethylamino)-phenothiazin-5-ium chloride or bromide. The patent converts phenothiazine into 3,7-dihalogenated-phenothiazin-5-ium chloride or bromide in a one-step method, and then adds dimethylamine to convert it into the target product. The method does not separate the intermediate bromide or chloride, which reduces the process operation and production cost. However, due to the poor stability of 3,7-dihalogenated-phenothiazin-5-ium chloride or bromide itself, the one-step method is not convenient for controlling the impurities in the halogenation reaction step, and the halogenated product will also decompose, resulting in poor quality and high impurities of the product, which cannot meet the requirements of medicinal use.

[0011] CM Wischik et al. published a method for purifying methylene blue, which can purify methylene blue with a commercial purity of less than 95% to a high purity level. First, methylene blue is reduced to leuco methylene blue by sodium borohydride (or hydrazine hydrate and methyl hydrazine can be selected), followed by acylation reaction using acetic anhydride, crystallization and purification of the acylated product, and then hydrolysis and oxidation to obtain high-purity methylene blue. We actually carried out a repetitive experiment of the preparation scheme and found that the yield of 3,7-di(dimethylamino)-10-acetyl-phenothiazine prepared by the reaction and the yield of methylene blue prepared by hydrolysis and oxidation were both about 50%, which is close to the embodiment in the patent. The total yield of the purification step is about 25%, which is a low yield. And leuco methylene blue is easily oxidized to methylene blue by air due to its strong reducing property, so it is not suitable for extraction as a separate intermediate. The main advantage of this method is that the prepared 3,7-di(dimethylamino)-10-acetyl-phenothiazine loses the complexing effect of methylene blue on metal ions, which has a certain positive effect on removing elemental impurities, and the polarity of the acetylated product becomes smaller, which is easy to be crystallized and purified to obtain high-purity methylene blue; the main disadvantage is that the yield is low, and the molar yield of the acetylation step is only about 50%.

[0012]

[0013] At present, methylene blue has been used in the form of tablets or injections. In order to adapt to a wider range of applications, it is necessary to provide raw materials with high chemical purity and low heavy metal content to meet the efficacy and safety of the drug as much as possible. Summary of the invention

[0014] The invention provides a method for preparing methylene blue, which is used to solve the technical problem of low purity of methylene blue.

[0015] In view of this, the present invention provides a method for preparing methylene blue, comprising the following steps: using phenothiazine as a raw material, performing a bromination reaction to obtain 3,7-dibromo-10H phenothiazine, performing an acetylation reaction to obtain 3,7-dibromo-10-acetyl phenothiazine, performing an amination reaction to obtain 3,7-di(dimethylamino)-10-acetyl phenothiazine, and finally performing hydrolysis and oxidation to obtain methylene blue.

[0016] Further, the process route of methylene blue is as follows:

[0017]

[0018] Optionally, the methylene blue is prepared by the following method: adding 3,7-bis(dimethylamino)-10-acetylphenothiazine to a hydrochloric acid aqueous solution, heating to react, cooling after the reaction is completed, adding an aqueous solution of ferric chloride, continuing the reaction, and then filtering, washing, and drying to obtain methylene blue.

[0019] Furthermore, the methylene blue is prepared by the following method: 3,7-bis(dimethylamino)-10-acetylphenothiazine and water are added to a 2000mL reaction bottle, and concentrated hydrochloric acid is added, heated to 70-100°C, reacted for 0.8-1.2h, cooled to 0-30°C, and then 1.2 or 2.0eq of ferric chloride aqueous solution is added, and the reaction is continued for 0.8-1.2h, and then filtered, washed with a 1% hydrochloric acid aqueous solution by mass, and then washed with water, and dried to obtain methylene blue;

[0020] Among them, the amount of water added to every 6g of 3,7-bis(dimethylamino)-10-acetylphenothiazine is 10-80mL, the amount of hydrochloric acid added to every 1g of 3,7-bis(dimethylamino)-10-acetylphenothiazine is 1.5-3mL, 1.2eq of ferric chloride aqueous solution is prepared by dissolving 59.2g of ferric chloride hexahydrate in 177.6g of water, and 2.0eq is prepared by dissolving 98.7g of ferric chloride hexahydrate in 296g of water.

[0021] Optionally, the 3,7-dibromo-10H phenothiazine is prepared by the following method: adding phenothiazine to acetonitrile, adding bromine at a certain temperature, reacting after the addition, adding sodium bisulfite solution or sodium sulfite solution to the reaction system after the reaction, filtering, washing, and obtaining 3,7-dibromo-10H phenothiazine.

[0022] Furthermore, the 3,7-dibromo-10H phenothiazine is prepared by the following method: adding phenothiazine and acetonitrile to a 2000 mL reaction bottle, adding bromine at a certain temperature, reacting for 1-2 hours after the addition, adding sodium bisulfite solution or sodium sulfite solution to the reaction system, filtering, washing with water, and obtaining 3,7-dibromo-10H phenothiazine;

[0023] The amount of acetonitrile added to 1g of phenothiazine is 10-20mL, and the amount of sodium bisulfite added to 1g of phenothiazine is 2-4g or 2.5-4.5g of sodium sulfite, which are prepared into an aqueous solution.

[0024] Optionally, the weight ratio of phenothiazine to bromine is 1:(1.6-3.2).

[0025] Optionally, the temperature is 20-30°C.

[0026] Optionally, the 3,7-dibromo-10-acetyl phenothiazine is prepared by the following method: 3,7-dibromo-10H phenothiazine is added to a first solvent, and then an acylating agent is added to react. After the reaction is completed, water is added, filtered, washed, and dried to obtain 3,7-dibromo-10-acetyl phenothiazine.

[0027] Optionally, the first solvent is any one of DMF and acetonitrile, and the acylating agent is any one of acetyl chloride and acetic anhydride.

[0028] Further, the 3,7-dibromo-10-acetylphenothiazine is prepared by the following method: 3,7-dibromo-10H phenothiazine and DMF are added to a 2000mL reaction bottle, and then triethylamine is added, and acetyl chloride is added at a temperature of -10-0°C, and after completion, the current temperature is maintained for reaction for 1.5-2.5h, water is added, filtered, washed with water, and dried to obtain 3,7-dibromo-10-acetylphenothiazine; wherein, The amount of DMF added to each 1g of 3,7-dibromo-10H-phenothiazine is 4-6mL, the weight ratio of 3,7-dibromo-10H-phenothiazine and triethylamine is 1:(0.3387-0.3388), the weight ratio of 3,7-dibromo-10H-phenothiazine and acetyl chloride is 1:(0.23-0.25), and the amount of water added to each 1g of 3,7-dibromo-10H-phenothiazine is 18-22mL.

[0029] Furthermore, the 3,7-dibromo-10-acetylphenothiazine is prepared by the following method: 3,7-dibromo-10H phenothiazine and acetonitrile are added to a 2000mL reaction bottle, and then acetic anhydride is added. After completion, the temperature is raised to 90-110°C, reacted for 4-8h, water is added, filtered, washed with water, and dried to obtain 3,7-dibromo-10-acetylphenothiazine; wherein the amount of acetonitrile added to each 1g of 3,7-dibromo-10H phenothiazine is 3.7-3.8mL, the weight ratio of 3,7-dibromo-10H phenothiazine to acetic anhydride is 1:(1.42-1.43), and the amount of water added to each 1g of 3,7-dibromo-10H phenothiazine is 10-15mL.

[0030] Optionally, the 3,7-bis(dimethylamino)-10-acetylphenothiazine is prepared by the following method: adding 3,7-dibromo-10-acetylphenothiazine to a second solvent, then adding a dimethylamine aqueous solution and cuprous iodide, heating and heating, reacting, cooling after completion, adding water, crystallizing, filtering, washing, and drying.

[0031] 3,7-bis(dimethylamino)-10-acetylphenothiazine was obtained.

[0032] Furthermore, the 3,7-bis(dimethylamino)-10-acetylphenothiazine is prepared by the following method: 3,7-dibromo-10-acetylphenothiazine and a second solvent are added to a 2000 mL reactor, and then a dimethylamine aqueous solution and cuprous iodide are added, and the temperature is raised to 100-150° C., and the reaction is performed for 10-20 hours. After the reaction is completed, the temperature is lowered to 5-50° C., water is added, and crystallization is performed for 0.8-1.2 hours, and the reaction is performed by filtering, washing with water, and drying to obtain 3,7-bis(dimethylamino)-10-acetylphenothiazine;

[0033] The amount of the second solvent added to each gram of 3,7-dibromo-10-acetylphenothiazine is 5-6.25 mL, the mass fraction of the dimethylamine aqueous solution is 40%, the weight ratio of 3,7-dibromo-10-acetylphenothiazine to cuprous iodide is 1:(0.0075-0.01), and the amount of water added to each gram of 3,7-dibromo-10-acetylphenothiazine is 10-15 mL.

[0034] Optionally, the weight ratio of the 3,7-bis(dimethylamino)-10-acetylphenothiazine to the dimethylamine aqueous solution is 1:(1.14-2.28).

[0035] Optionally, the second solvent is any one of NMP and DMF.

[0036] It can be seen from the above technical solutions that the embodiments of the present invention have the following advantages:

[0037] 1. The total yield of methylene blue with pharmaceutical-grade quality level prepared by this process using phenothiazine as raw material can reach more than 94%, the purity can reach more than 99.6%, and the content of azure B is reduced to less than 0.3%. In terms of comprehensive quality and yield, it is much higher than the existing process level.

[0038] 2. The use of bromine and dimethylamine is reduced, which not only reduces production costs, but also reduces environmental pressure. No highly toxic solvents such as chloroform are used, which is environmentally friendly.

[0039] 3. Since methylene blue has the function of complexing metal ions, if more materials containing metal ions are used in the process, it is easy to cause the elemental impurities in the finished product to exceed the pharmacopoeia limit (the European Pharmacopoeia has clear provisions). In the steps of this application, only two metal element-containing materials, ferric chloride and cuprous iodide, are used, and it is found through testing that the metal elements of this product meet the pharmacopoeia quality standards. Therefore, it is shown that the methylene blue prepared by this process is suitable for medicinal use. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments.

[0041] Figure 1 is the hydrogen spectrum of the prepared methylene blue.

[0042] Figure 2 This is a liquid phase diagram of the prepared methylene blue. DETAILED DESCRIPTION

[0043] In order to make the technical scheme of the present invention better understood by the personnel of the technical field, the technical scheme in the embodiment of the present invention is clearly and completely described below. Obviously, the described embodiment is only a part of the embodiment of the present invention, rather than all the embodiments. Based on the embodiment in the present invention, all other embodiments obtained by ordinary technicians in the field without making creative work are within the scope of protection of the present invention. Unless otherwise specified, various raw materials, reagents, instruments and equipment used in the present invention can be purchased from the market or can be prepared by existing methods.

[0044] Preparation Example

[0045] Preparation Example 1

[0046] A 3,7-dibromo-10H phenothiazine prepared by the following method:

[0047] Add 50 g of phenothiazine and 500 mL of acetonitrile to a 2000 mL reaction bottle, add 80 g of bromine at 25 ° C, react for 1 hour, and after the reaction, add 208 g of an aqueous solution of sodium bisulfite to the reaction system, filter, and wash with water to obtain 3,7-dibromo-10H phenothiazine; the yield is 86.5 g, and the yield is 96%.

[0048] Preparation Example 2

[0049] A 3,7-dibromo-10H phenothiazine prepared by the following method:

[0050] Add 50 g of phenothiazine and 1000 mL of acetonitrile to a 2000 mL reaction bottle, add 160 g of bromine at 25°C, react for 2 h, add 500 g of an aqueous solution of sodium sulfite to the reaction system after the reaction, filter, and wash with water to obtain 3,7-dibromo-10H phenothiazine; yield 87.1 g, yield rate 96.7%.

[0051] Preparation Example 3

[0052] A 3,7-dibromo-10-acetyl phenothiazine, which is prepared by the following method:

[0053] In a 2000mL reaction bottle, add 80g of 3,7-dibromo-10H phenothiazine prepared in Preparation Example 2 and 400mL of DMF, then add 27.1g of triethylamine, and add 19.2g of acetyl chloride at a temperature of -5°C. After completion, maintain the current temperature for 2h, add 1600mL of water, filter, wash with water, and dry to obtain 3,7-dibromo-10-acetylphenothiazine, a total of 83.1g, with a yield of 93.0%.

[0054] Preparation Example 4

[0055] A 3,7-dibromo-10-acetyl phenothiazine, which is prepared by the following method:

[0056] In a 2000mL reaction bottle, add 80g of 3,7-dibromo-10H phenothiazine prepared in Preparation Example 2 and 300mL of acetonitrile, and then add 114g of acetic anhydride. After completion, heat to 100°C, react for 6h, add 1000mL of water, filter, wash with water, and dry to obtain 3,7-dibromo-10-acetylphenothiazine, a total of 80.7g, with a yield of 90.3%.

[0057] Preparation Example 5

[0058] A 3,7-bis(dimethylamino)-10-acetyl phenothiazine, which is prepared by the following method:

[0059] In a 2000mL reactor, 80g of 3,7-dibromo-10-acetylphenothiazine prepared in Preparation Example 3 and 400mL of NMP were added, and then 91.2g of a 40% aqueous solution of dimethylamine by mass and 0.8g of cuprous iodide were added. The mixture was heated to 135°C and reacted for 20h. After completion, the mixture was cooled to 30°C, 1000mL of water was added, and crystallization was performed for 1h. The mixture was filtered, washed with water, and dried to obtain 3,7-bis(dimethylamino)-10-acetylphenothiazine, a total of 62.5g, with a yield of 95.1%.

[0060] Preparation Example 6

[0061] A 3,7-bis(dimethylamino)-10-acetyl phenothiazine, which is prepared by the following method:

[0062] In a 2000mL reactor, 80g of 3,7-dibromo-10-acetylphenothiazine prepared in Preparation Example 3 and 500mL of DMF were added, and then 182.4g of a 40% aqueous solution of dimethylamine by mass and 0.6g of cuprous iodide were added. The mixture was heated to 135°C and reacted for 10h. After completion, the mixture was cooled to 30°C, 1000mL of water was added, and crystallization was performed for 1h. The mixture was filtered, washed with water, and dried to obtain 3,7-bis(dimethylamino)-10-acetylphenothiazine, a total of 61.6g, with a yield of 93.7%.

[0063] Example

[0064] Example 1

[0065] A method for preparing methylene blue comprises the following steps:

[0066] In a 2000mL reaction bottle, 60g of 3,7-bis(dimethylamino)-10-acetylphenothiazine prepared in Preparation Example 5 and 600mL of water were added, and then 180mL of concentrated hydrochloric acid was added. The mixture was heated to 90°C and reacted for 1h. The mixture was cooled to 25°C, and then 2eq of ferric chloride solution (98.7g of ferric chloride hexahydrate dissolved in 296g of water) was added. The reaction was continued for 1h. The mixture was filtered, washed with a 1% hydrochloric acid aqueous solution, then washed with water, and dried to obtain methylene blue, a total of 67.4g, a yield of 99%, a purity of 99.67%, and a Azure B content of 0.28%.

[0067] Example 2

[0068] A method for preparing methylene blue comprises the following steps:

[0069] In a 2000mL reaction bottle, 60g of 3,7-bis(dimethylamino)-10-acetylphenothiazine prepared in Preparation Example 5 and 800mL of water were added, and then 90mL of concentrated hydrochloric acid was added. The mixture was heated to 75°C, reacted for 1h, cooled to 5°C, and then 1.2eq of ferric chloride solution (59.2g of ferric chloride hexahydrate dissolved in 177.6g of water) was added. The reaction was continued for 1h, filtered, washed with a 1% hydrochloric acid aqueous solution, then washed with water, and dried to obtain methylene blue, a total of 64.5g, a yield of 94.7%, a purity of 99.71%, and a Azure B content of 0.25%.

[0070] Performance testing

[0071] The finished product methylene blue in Example 1 was subjected to the following performance tests and compared with the EP 11.0 limit. The test results are shown in Table 1.

[0072] Table 1 Test results

[0073]

[0074]

[0075] In combination with Examples 1-2 and Table 1, it can be seen that the methylene blue prepared by the preparation method of the present application reduces the use of bromine and dimethylamine, which not only reduces the production cost, but also reduces the environmental pressure, and does not use highly toxic solvents such as chloroform, which is environmentally friendly; and the methylene blue prepared by the present application has better quality, higher purity, and higher yield, among which the yield can reach more than 94%, the purity can reach more than 99.6%, and the azure B content can be reduced to less than 0.3%. In terms of comprehensive quality and yield, it is much higher than the existing process level.

[0076] As described above, the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the same. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features thereof may be replaced by equivalents. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing methylene blue, characterized in that: The method comprises the following steps: using phenothiazine as a raw material, performing a bromination reaction to obtain 3,7-dibromo-10H phenothiazine, performing an acetylation reaction to obtain 3,7-dibromo-10-acetyl phenothiazine, performing an amination reaction to obtain 3,7-di(dimethylamino)-10-acetyl phenothiazine, and finally performing hydrolysis and oxidation to obtain methylene blue.

2. The method for preparing methylene blue according to claim 1, wherein: The methylene blue is prepared by the following method: adding 3,7-di(dimethylamino)-10-acetylphenothiazine to a hydrochloric acid aqueous solution, heating to react, cooling after the reaction is completed, adding a ferric chloride aqueous solution, continuing the reaction, and then filtering, washing, and drying to obtain methylene blue.

3. The method for preparing methylene blue according to claim 1, wherein: The 3,7-dibromo-10H phenothiazine is prepared by the following method: adding phenothiazine to acetonitrile, adding bromine at a certain temperature, reacting after the addition, adding sodium bisulfite solution or sodium sulfite solution to the reaction system after the reaction, filtering, washing, and obtaining 3,7-dibromo-10H phenothiazine.

4. The method for preparing methylene blue according to claim 3, wherein: The weight ratio of phenothiazine to bromine is 1:(1.6-3.2).

5. The method for preparing methylene blue according to claim 3, wherein: The temperature is 20-30°C.

6. The method for preparing methylene blue according to claim 1, wherein: The 3,7-dibromo-10-acetylphenothiazine is prepared by the following method: 3,7-dibromo-10H phenothiazine is added to a first solvent, and then an acylating agent is added to react. After the reaction is completed, water is added, and the reaction is filtered, washed, and dried to obtain 3,7-dibromo-10-acetylphenothiazine.

7. The method for preparing methylene blue according to claim 6, wherein: The first solvent is any one of DMF and acetonitrile, and the acylating agent is any one of acetyl chloride and acetic anhydride.

8. The method for preparing methylene blue according to claim 1, wherein: The 3,7-bis(dimethylamino)-10-acetylphenothiazine is prepared by the following method: 3,7-dibromo-10-acetylphenothiazine is added to a second solvent, and then a dimethylamine aqueous solution and cuprous iodide are added, the temperature is raised, the reaction is carried out, the temperature is lowered after completion, water is added, crystallization is carried out, filtering, washing, and drying are carried out to obtain 3,7-bis(dimethylamino)-10-acetylphenothiazine.

9. The method for preparing methylene blue according to claim 8, wherein: The weight ratio of the 3,7-bis(dimethylamino)-10-acetylphenothiazine to the dimethylamine aqueous solution is 1:(1.14-2.28).

10. The method for preparing methylene blue according to claim 8, characterized in that: The second solvent is any one of NMP and DMF.

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