A kind of preparation method of L-carnosine

Abstract

The present invention relates to the field of medical technology, and in particular to a method for preparing L-carnosine, the method comprising the following steps: using L-aspartic acid as a starting material, reacting with cyanoacetyl chloride to obtain (S)-2-(2-cyanoacetylamino)succinic acid, (S)-2-(2-cyanoacetylamino)succinic acid under the action of a dehydrating agent to obtain (S)-2-(2-cyanoacetylamino)succinic anhydride, (S)-2-(2-cyanoacetylamino)succinic anhydride reacting with trimethylsulfoxide iodide and hydrochloric acid, then cyclizing with formamidine acetate to obtain cyanoacetyl-L-histidine, and finally reducing with hydrogen to obtain the target product L-carnosine. The L-carnosine prepared by the preparation method of the present invention has the characteristics of high purity, high yield, low raw material cost and low energy consumption, and is suitable for industrial production.

Description

Technical Field

[0001] The invention relates to a preparation method of L-carnosine and belongs to the technical field of medicines. Background Art

[0002] L-Carnosine is a dipeptide compound formed by the dehydration condensation of β-alanine and L-histidine, connected by peptide bonds. Its chemical name is β-alanyl-L-histidine. Subsequent studies have shown that L-carnosine is widely present in the brain, muscle tissue and nervous system of mammals, and has the effects of anti-oxidation, delaying aging, improving memory, promoting wound healing, reducing free radicals in the body, removing chelated transition metals in the blood, preventing ulcers, and anti-tumor, and has a very broad application prospect.

[0003] The existing L-carnosine preparation methods are mainly divided into chemical synthesis and biotechnology synthesis:

[0004] 1) The advantages of chemical synthesis are: the target product is synthesized through a predetermined route, and the types and contents of impurities generated during the process are relatively stable. By adjusting the reaction conditions and the refining method, the impurity level can be stably controlled to obtain a product that meets the pharmaceutical quality standards; the disadvantages of chemical synthesis are: the main chemical synthesis methods currently use β-alanine and L-histidine as starting materials, and amino acid compounds are amphoteric compounds containing both amino groups and carboxyl groups, with more reaction sites. Therefore, in the process of preparing L-carnosine, it is necessary to introduce a protective group to selectively protect the reaction sites and synthesize the target product. Since the protective group itself does not enter the chemical structure of the product L-carnosine, the use of the protective group, the reagents used to introduce the protective group, and the reagents used to remove the protective group all increase the raw material cost for preparing L-carnosine.

[0005] 2) The advantages of biotechnology synthesis are: through microbial fermentation or bioenzymatic method, under certain conditions, the target product L-carnosine can be directly synthesized from β-alanine and L-histidine, and then separated and refined to obtain a pure product, saving the related costs of using protecting groups in chemical synthesis; the disadvantages of biotechnology synthesis are: during the microbial fermentation production process, the types of impurities, impurity content and yield are unstable; the bioenzymatic method can obtain products with stable impurity levels, but different substrates and reaction conditions require different enzymes, which are expensive and costly, and are only suitable for scientific research, resulting in certain risks in product quality control.

[0006] In its Chinese patent application CN111051289A, Tokuyama Co., Ltd. of Japan disclosed a method for synthesizing L-carnosine, using β-alanine and L-histidine as main raw materials, and protecting them with protective groups respectively: protecting the amino group of β-alanine (protecting group: benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, trifluoroacetyl, tert-butyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl or formyl), and then preparing the corresponding acid anhydride to obtain compound (1); protecting the carboxyl group, nitrogen on the imidazole, and amino group of L-histidine to obtain compound (2); compound (1) and compound (2) are used to prepare compound (3), and then removing the protective group to obtain the target product L-carnosine. The specific process route is as follows.

[0007] ;

[0008] ;

[0009] .

[0010] R2 is: ; R3 is benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, trifluoroacetyl, tert-butyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl or formyl. In formula (1b), m is an integer of 0 or 1, R4, R5 and R6 are hydrogen atoms or alkyl groups with 1 to 6 carbon atoms, wherein when m is 0, at least two of R4, R5 and R6 are alkyl groups with 1 to 6 carbon atoms. R7 and R8 are independently selected from hydrogen atoms or trialkylsilyl groups; R9 is a hydrogen atom or a protecting group for a carboxyl group.

[0011] The protecting groups (R3, R7, R8, R9) used in the method and the R2 group in the structure of compound (1) do not constitute the skeleton structure of the target product, and the process of introducing and removing the protecting groups is required, resulting in waste of auxiliary materials and labor. The method for preparing compound (1) described in the patent application is: using acyl chloride and carboxylic acid, or triphosgene and carboxylic acid. Although triphosgene is relatively stable in nature, if it is used in the production process, it is easy to have incomplete reaction and generate by-products including phosgene (a highly toxic drug), which has high requirements for production equipment, operating level and waste treatment. The total yield of the target product L-carnosine obtained by this method is less than 80% (based on N-benzyloxycarbonyl-β-alanine as raw material), which is relatively low.

[0012] Chinese patent application CN112480206A discloses a method for synthesizing L-carnosine, which uses β-alanine and L-histidine as main raw materials, firstly protects the amino group and carboxyl group of β-alanine: protects the carboxyl group of β-alanine to obtain alanine methyl ester, then protects the amino group (protection reagent: BOC anhydride, benzyl chloroformate) to obtain the corresponding amino-protected alanine methyl ester (1), and condenses the compound with L-histidine to obtain the corresponding amino-protected L-carnosine, which is then deprotected to obtain the target product L-carnosine. This method also requires the introduction and removal of the protecting group, resulting in waste of auxiliary materials and labor. The protecting group tert-butyloxycarbonyl (BOC) is usually not completely reacted when deprotected with hydrochloric acid, while trifluoroacetic acid is highly irritating and corrosive, and will corrode the surface of the general glass-lined reactor. The composite material of silicon dioxide and trace elements and metals is therefore required to be more corrosion-resistant, which is more expensive and not conducive to industrial production. According to the embodiment, the total yield of the crude L-carnosine product is calculated to be 82.0%, and the yield is also relatively low. Summary of the invention

[0013] The invention aims at the deficiencies in the prior art and provides a method for preparing L-carnosine. The L-carnosine product prepared by the preparation method has high yield, high purity and low single impurity content and is more suitable for industrial production.

[0014] The technical solution of the present invention to solve the above-mentioned technical problems is as follows: a preparation method of L-carnosine, wherein the preparation method is:

[0015] S1, after compound I reacts with a base, it reacts with compound II to obtain compound III;

[0016] The compound I is , the compound II is , the compound III is ;

[0017] S2, compound III is dissolved in an organic solvent, a dehydrating agent is added, and dehydration is performed to prepare compound IV;

[0018] The compound IV is ;

[0019] S3, compound IV is dissolved in an organic solvent, trimethyl sulfoxide iodide and hydrochloric acid are added to react to obtain compound V, and then formamidine acetate is added to the system to react to obtain compound VI;

[0020] The compound V is , X is I or Cl; the compound VI is ;

[0021] S4. Compound VI is dissolved in water, a reduction catalyst is added, and the target product L-carnosine is obtained by hydrogenation reduction.

[0022] Furthermore, the specific operation of step S1 is: dissolving compound I and a base in an organic solvent, stirring evenly, to obtain a reaction solution A; dissolving compound II in an organic solvent, to obtain a reaction solution B; and slowly dropping the reaction solution B into the reaction solution A to obtain compound III.

[0023] Further, in step S1, the organic solvent is any one of acetonitrile and tetrahydrofuran;

[0024] In step S1, the base is any one of triethylamine and N,N-diisopropylethylamine.

[0025] Furthermore, in step S1, the reaction temperature of adding the reaction solution B to the reaction solution A for reaction is 10-20°C.

[0026] Further, in step S2, the organic solvent is dichloromethane or tetrahydrofuran;

[0027] In step S2, the dehydration reagent is at least one of oxalyl chloride and phosphorus trichloride.

[0028] Furthermore, the specific operation of step S2 is: controlling the temperature to 0-10° C., adding a dehydrating agent dropwise to the organic solvent of compound III, and after the addition is complete, heating the mixture to 20-30° C. for reaction.

[0029] In step S3, trimethylsulfoxide iodide and potassium carbonate are dissolved in an organic solvent to obtain a reaction solution C; compound IV is dissolved in an organic solvent to obtain a reaction solution D. The reaction solution C is added dropwise to the reaction solution D, hydrochloric acid is added dropwise to obtain compound V, and formamidine acetate is added to react to obtain compound VI.

[0030] Further, in step S3, the organic solvent is N,N-dimethylformamide;

[0031] In step S3, the reaction temperature is controlled at 0-10°C when preparing compound V; and the reaction temperature is controlled at 70-80°C when adding formamidine acetate to prepare compound VI.

[0032] Furthermore, in step S3, the molar ratio of trimethyl sulfoxide iodide to compound IV is (1.0-1.2):1; the molar ratio of formamidine acetate to compound IV is (1.0-1.3):1.

[0033] Furthermore, in step S3, the reduction catalyst is a mixture of palladium carbon and boron trifluoride tetrahydrofuran, wherein the molar ratio of compound VI to the palladium element and the boron element in the reduction catalyst is 1: (0.0005-0.0020): (0.0001-0.0003), and the molar ratio of the palladium element and the boron element in the reduction catalyst is (5-8): 1.

[0034] In step S4, the reaction temperature of the hydrogenation reduction is 60-70° C. and the pressure is 0.5-2.0 MPa.

[0035] The beneficial effects of the present invention are:

[0036] 1) Through chemical synthesis of L-carnosine, impurities in each step can be effectively controlled. The maximum single impurity content in the obtained L-carnosine is ≤0.03%, and the total impurity content is ≤0.2%, which meets the high purity requirements for raw materials in the field of pharmaceutical technology.

[0037] 2) This synthesis method has a high yield, with an overall yield of >93%, and is suitable for industrial production.

[0038] 3) In the preparation method of the present invention, since no protecting group is used, the corresponding impurities generated by the introduction and removal of the protecting group are avoided: phthalimide derivative impurities, hydrazine, tert-butyloxycarbonyl derivative impurities, benzyl derivative impurities, fluorenylmethyloxycarbonyl derivative impurities, trimethylsilane impurities, etc.

[0039] 4) The materials used in the preparation method of the present invention are cheap and easy to obtain: the starting materials, reagents, and solvents are all commercial bulk products with a wide supply and sufficient supply. No controlled reagents (such as explosive and highly toxic reagents) are used, which has little pollution and is environmentally friendly. ① L-aspartic acid is used as the raw material, and the price of commercial bulk goods is much lower than L-histidine; ② No amino protecting groups (phthalic anhydride, Boc anhydride, benzyl chloroformate, benzyl halide, fluorenylmethoxycarbonyl chloride, etc.) are used, saving the corresponding auxiliary materials and labor costs generated by the introduction and removal of protecting groups; ③ The reaction conditions are mild, the time is short, and there are no special requirements for production equipment, which is suitable for industrial production.

[0040] 5) The elemental impurities and solvent residues of the product obtained by the preparation method of the present invention meet the ICH guidelines. The stability test results show that the product has good stability. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 The HPLC detection spectrum of L-carnosine in Example 1;

[0042] Figure 2 The MS detection spectrum of L-carnosine in Example 1;

[0043] Figure 3 The H-NMR detection spectrum of L-carnosine in Example 1;

[0044] Figure 4 The HPLC detection spectrum of L-carnosine in Example 2;

[0045] Figure 5 The HPLC detection spectrum of L-carnosine in Example 3;

[0046] Figure 6 The HPLC detection spectrum of L-carnosine in Example 4. DETAILED DESCRIPTION

[0047] The specific implementation of the present invention is described in detail below. The present invention can be implemented in many other ways than those described herein, and those skilled in the art can make similar improvements without violating the connotation of the present invention, so the present invention is not limited by the specific embodiments disclosed.

[0048] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art to which the present invention belongs. The terms used are only for describing specific embodiments and are not intended to limit the present invention.

[0049] A preparation method of L-carnosine, the preparation method comprising:

[0050] S1, compound I (L-aspartic acid) reacts with a base and then reacts with compound II (2-cyanoacetyl chloride) to obtain compound III ((S)-2-(2-cyanoacetylamino)succinic acid); the specific reaction principle is:

[0051] ;

[0052] S2. Compound III is dissolved in an organic solvent, and a dehydrating agent is added to dehydrate to prepare compound IV ((S)-2-(2-cyanoacetylamino)succinic anhydride); the specific reaction principle is:

[0053] ;

[0054] S3, compound IV is dissolved in an organic solvent, trimethyl sulfoxide iodide and hydrochloric acid are added to react to obtain compound V ((S)-5-halogeno-2-(2-cyanoacetamido)-4-oxopentanoic acid), and then formamidine acetate is added to the system to react to obtain compound VI (cyanoacetyl-L-histidine); the specific reaction principle is:

[0055] Wherein X is I or Cl;

[0056] S4, compound VI is dissolved in water, a reduction catalyst is added, and the target product L-carnosine is obtained by hydrogenation reduction. The specific reaction principle is:

[0057] .

[0058] Specifically, the specific operation of step S1 is: dissolving compound I and a base in an organic solvent, stirring evenly, to obtain a reaction solution A; dissolving compound II in an organic solvent, to obtain a reaction solution B; and slowly dropping the reaction solution B into the reaction solution A, to obtain compound III.

[0059] Specifically, in step S1, the organic solvent is any one of acetonitrile and tetrahydrofuran;

[0060] In step S1, the base is any one of triethylamine and N,N-diisopropylethylamine.

[0061] Specifically, the reaction temperature of step S1 is 10-20° C. The temperature is controlled at 10-20° C., and the reaction solution B is added dropwise to the reaction solution A, and the temperature is stirred at 20-25° C. after the addition is completed.

[0062] Preferably, in reaction solution A, the mass ratio of the organic solvent to compound I is preferably 3:1, and the molar ratio of the base to compound I is preferably 2:1; in reaction solution B, the mass ratio of the organic solvent to compound II is preferably 2:1.

[0063] Specifically, in step S2, the organic solvent is dichloromethane or tetrahydrofuran;

[0064] In step S2, the dehydration reagent is at least one of oxalyl chloride and phosphorus trichloride.

[0065] Preferably, in step S2, when the dehydrating agent is oxalyl chloride, the molar ratio of oxalyl chloride to compound III is 1.3:1; when the dehydrating agent is phosphorus trichloride, the molar ratio of phosphorus trichloride to compound III is preferably 1.1:1.

[0066] Specifically, the specific operation of step S2 is: controlling the temperature to 0-10°C, adding a dehydrating agent dropwise to the organic solvent of compound III, reacting at 0-10°C after the addition is complete, and then heating to 20-30°C for reaction.

[0067] Specifically, the specific operation of step S3 is: dissolving trimethylsulfoxide iodide and potassium carbonate in an organic solvent to obtain a reaction solution C; dissolving compound IV in an organic solvent to obtain a reaction solution D. Adding the reaction solution C dropwise to the reaction solution D, adding hydrochloric acid dropwise to obtain compound V, adding formamidine acetate, and reacting to obtain compound VI.

[0068] Specifically, in step S3, the organic solvent is N,N-dimethylformamide;

[0069] In step S3, the reaction temperature is controlled at 0-10°C when preparing compound V; and the reaction temperature is controlled at 70-80°C when preparing compound VI.

[0070] Specifically, in step S3, the molar ratio of trimethyl sulfoxide iodide to compound IV is (1.0-1.2):1; the molar ratio of hydrochloric acid to compound IV is (1.0-1.5):1; and the molar ratio of formamidine acetate to compound IV is (1.0-1.5):1.

[0071] Preferably, in step S3, the molar ratio of trimethyl sulfoxide iodide to compound IV is 1.1:1; the molar ratio of the halide salt to compound IV is 1.1:1; and the molar ratio of formamidine acetate to compound IV is 1.1:1.

[0072] Specifically, in step S4, the reduction catalyst is a mixture of palladium carbon and boron trifluoride tetrahydrofuran, wherein the molar ratio of compound VI to the palladium element and the boron element in the reduction catalyst is 1: (0.0005-0.0020): (0.0001-0.0003), and the molar ratio of the palladium element and the boron element in the reduction catalyst is (5-8): 1.

[0073] Specifically, in step S4, the reaction temperature of the hydrogenation reduction is 60-70° C. and the pressure is 0.5-2.0 MPa.

[0074] The raw material compound II (2-cyanoacetyl chloride) involved in the embodiments of the present invention can be obtained by the following conventional method: using 2-cyanoacetic acid as a raw material, chlorinating the carboxyl group with a chlorination reagent such as thionyl chloride, oxalyl chloride, phosphorus trichloride, etc. to obtain 2-cyanoacetyl chloride.

[0075] Using one of the chlorination reagents, the specific preparation process of thionyl chloride is as follows:

[0076] Synthesis example 1

[0077] ;

[0078] Add 50.0kg 2-cyanoacetic acid to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, and introduce nitrogen protection. Add 50.0kg dichloromethane and 84.0kg thionyl chloride to the kettle, stir at 30-35℃ for 1h, add 0.5kg N,N-dimethylformamide dropwise, continue to stir at this temperature for 3h, remove the solvent under reduced pressure, and obtain 60.7kg 2-cyanoacetyl chloride, which is kept in the kettle for later use.

[0079] Example 1

[0080] S1. Preparation of Compound III

[0081] 70.9 kg of L-aspartic acid was added to a glass-lined kettle, nitrogen was replaced until the oxygen content was ≤6%, nitrogen protection was introduced, and then 212.8 kg of acetonitrile and 107.9 kg of triethylamine were added, and the mixture was stirred until all the solids were dissolved to obtain a reaction solution A. 141.8 kg of acetonitrile was added to the glass-lined kettle containing 2-cyanoacetyl chloride to be used in Synthesis Example 1, and the mixture was stirred evenly to obtain a reaction solution B. The temperature of the reaction solution was controlled to be 10-15°C, and the reaction solution B was added dropwise to the reaction solution A. After the addition was completed, the mixture was stirred at 20-25°C for 1 hour, cooled to 15-20°C, stirred for 1 hour, centrifuged, and continued to cool to 0-5°C, stirred for 1 hour, centrifuged, and dried to obtain 105.9 kg of compound III with a yield of 99.3% and a purity of 99.5%.

[0082] S2. Preparation of Compound IV

[0083] 105.0 kg of compound III was added to a glass-lined kettle, and nitrogen was replaced until the oxygen content was ≤6%. Nitrogen protection was introduced, and then 525.0 kg of dichloromethane was added. The temperature of the reaction liquid was controlled at 0-5°C, and 86.6 kg of oxalyl chloride was added dropwise to the kettle. After the addition was completed, it was stirred at 0-5°C for 1 h, heated to 20-25°C, stirred for 2 h, centrifuged, rinsed with 0-5°C dichloromethane, and dried to obtain 95.1 kg of compound IV with a yield of 99.5%.

[0084] S3. Preparation of Compound VI

[0085] Add 113.6 kg of trimethyl sulfoxide iodide and 35.7 kg of potassium carbonate to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 235.0 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 0-5°C to obtain reaction solution C. Add 94.0 kg of compound IV to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 235.0 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 0-5°C to obtain reaction solution D. Control the temperature of the reaction solution to 0-5°C, add reaction solution C dropwise to reaction solution D, and stir at 0-5°C for 2 hours after the addition. Control the temperature of the reaction solution to 0-5°C, and add 62.8 kg of hydrochloric acid dropwise to the kettle. Stir for 2 hours after the addition is completed. Then add 59.1 kg of formamidine acetate to the kettle, heat the reaction solution to 70-75°C, and stir for 3 hours. 500 kg of water and 500 kg of ethyl acetate were added to the kettle. After stirring and standing at room temperature, the aqueous phase was separated and concentrated under reduced pressure to 200 kg. The temperature of the concentrated liquid was controlled at 5-10 ° C. 350 kg of methanol was added dropwise, filtered, rinsed with methanol, and dried to obtain 110.1 kg of compound VI with a yield of 96.0% and a purity of 99.7%.

[0086] S4. Preparation of L-carnosine

[0087] Add 109.0kg of compound VI to a stainless steel kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, add 445.0kg of water to the kettle, stir until completely dissolved, transfer to a stainless steel hydrogenation kettle, and draw a mixed solution of 4.197kg 5% palladium carbon, 0.007kg 46% boron trifluoride tetrahydrofuran and 100.0kg water under negative pressure, replace the air in the kettle 3 times with nitrogen, and replace the nitrogen in the kettle 3 times with hydrogen. Raise the kettle temperature to 60-65℃, pass hydrogen, stir at 0.5-1.5Mpa for 6h, lower the kettle temperature to room temperature, replace with nitrogen to normal pressure, filter through a titanium rod and filter element filter, and remove the catalyst. The filtrate is transferred to a stainless steel kettle and concentrated under reduced pressure to 200kg. Control the temperature at 50-55℃, add 100kg of methanol to the kettle, add 0.555kg of L-carnosine seeds, grow crystals for 30 minutes, and add 200kg of methanol. The temperature was lowered to 10-15°C, stirred for 6 hours, centrifuged and filtered, washed with methanol, and dried to obtain 109.8 kg of L-carnosine. The yield was 98.5%, the purity was 99.87%, and the total yield was 93.4%.

[0088] The HPLC detection spectrum of L-carnosine is as follows Figure 1 As shown, the HPLC detection spectrum data is shown in Table 1.

[0089] Table 1 HPLC detection spectrum data of L-carnosine in Example 1

[0090]

[0091] The MS detection spectrum of L-carnosine is as follows Figure 2 As shown, the H-NMR detection spectrum of L-carnosine is as follows Figure 3 As shown. The test conditions for the H-NMR detection spectrum are:

[0092] Pulse sequence: zg30, 30 degree excitation;

[0093] Number of sampling points: 65536;

[0094] Solvent: deuterated heavy water;

[0095] Repeat scanning times: 8 times;

[0096] Spectral width: 10330.578 Hz;

[0097] Spectral resolution: 0.157632 Hz;

[0098] Spectrometer frequency: 500.1300 MHz.

[0099] The elemental impurities in L-carnosine prepared in this example were determined, wherein the limits were based on ICH Q3D (R1): Elemental Impurities Guidelines, and the elemental impurities are as shown in Table 2:

[0100] Table 2 Elemental impurities in L-carnosine obtained in Example 1

[0101]

[0102] The residual solvent of L-carnosine prepared in this example was determined according to the ICH Q3C (R8): Impurities: Residual Solvents Guidelines. The specific results are shown in Table 3 below.

[0103] Table 3 Solvent residues in L-carnosine obtained in Example 1

[0104]

[0105] The L-carnosine prepared in this example was subjected to a stability test. The experimental conditions were formulated with reference to the ICH tripartite coordination guidelines (International Council for Registration of Pharmaceuticals for Human Use) Stability Tests for New APIs and Preparations Q1A (R2) 2.1.2 Strong Test. The specific results are shown in Table 4 below.

[0106] Table 4 Stability test of L-carnosine obtained in Example 1

[0107]

[0108] The experimental results show that the L-carnosine prepared by the method described in this article still has good stability even under relatively harsh conditions.

[0109] Example 2

[0110] S1. Preparation of Compound III

[0111] 70.9 kg of L-aspartic acid was added to a glass-lined kettle, nitrogen was replaced until the oxygen content was ≤6%, nitrogen protection was introduced, and then 212.8 kg of acetonitrile and 107.8 kg of triethylamine were added, and the solid was stirred until all the solids were dissolved to obtain reaction solution A. 141.8 kg of acetonitrile was added to the glass-lined kettle containing 2-cyanoacetyl chloride to be used as described in Synthesis Example 1, and the reaction solution B was obtained after stirring evenly. The temperature of the reaction solution was controlled to be 15-20°C, and the reaction solution B was added dropwise to the reaction solution A. After the addition was completed, the reaction solution was stirred at 20-25°C for 1 hour, cooled to 15-20°C, stirred for 1 hour, centrifuged, and continued to cool to 0-5°C, stirred for 1 hour, centrifuged, and dried to obtain 105.8 kg of compound III with a yield of 99.2% and a purity of 99.5%.

[0112] S2. Preparation of Compound IV

[0113] 105.0 kg of compound III was added to a glass-lined kettle, and nitrogen was replaced until the oxygen content was ≤6%. Nitrogen protection was introduced, and then 525.0 kg of dichloromethane was added. The temperature of the reaction liquid was controlled at 5-10 ° C. 86.6 kg of oxalyl chloride was added dropwise to the kettle. After the addition was completed, it was stirred at 5-10 ° C for 1 h, heated to 25-30 ° C, stirred for 2 h, centrifuged, rinsed with 0-5 ° C dichloromethane, and dried to obtain 95.2 kg of compound IV with a yield of 99.6%.

[0114] S3. Preparation of Compound VI

[0115] Add 136.4 kg of trimethyl sulfoxide iodide and 42.8 kg of potassium carbonate to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 235.0 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 5-10 ° C to obtain reaction solution C. Add 94.0 kg of compound IV to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 235.0 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 5-10 ° C to obtain reaction solution D. Control the temperature of the reaction solution at 5-10 ° C, add reaction solution C dropwise to reaction solution D, and stir at 5-10 ° C for 2 hours after the dropwise addition. Control the temperature of the reaction solution at 5-10 ° C, and add 62.8 kg of hydrochloric acid dropwise to the kettle. Stir for 2 hours after the dropwise addition. Then add 59.1 kg of formamidine acetate to the kettle, raise the temperature of the reaction solution to 75-80 ° C, and stir for 3 hours. 500 kg of water and 500 kg of ethyl acetate were added to the kettle. After stirring and standing at room temperature, the aqueous phase was separated and concentrated under reduced pressure to 200 kg. The temperature of the concentrated liquid was controlled at 5-10 ° C. 350 kg of methanol was added dropwise, filtered, rinsed with methanol, and dried to obtain 110.0 kg of compound VI with a yield of 95.9% and a purity of 99.7%.

[0116] S4. Preparation of L-carnosine

[0117] Add 109.0kg of compound VI to a stainless steel kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, add 445.0kg of water to the kettle, stir until completely dissolved, transfer to a stainless steel hydrogenation kettle, and draw a mixed solution of 3.119kg 5% palladium carbon, 0.022kg 46% boron trifluoride tetrahydrofuran and 100.0kg water under negative pressure, replace the air in the kettle 3 times with nitrogen, and replace the nitrogen in the kettle 3 times with hydrogen. Raise the kettle temperature to 65-70℃, pass hydrogen, stir at 1.0-2.0Mpa for 6h, lower the kettle temperature to room temperature, replace with nitrogen to normal pressure, filter through a titanium rod and filter element filter to remove the catalyst. The filtrate is transferred to a stainless steel kettle and concentrated under reduced pressure to 200kg. Control the temperature at 50-55℃, add 100kg of methanol to the kettle, add 0.545kg of L-carnosine seeds, grow crystals for 30 minutes, and add 200kg of methanol. Cool down to 10-15℃, stir for 6h, centrifuge, rinse with methanol, dry to obtain 110.0kg L-carnosine. Yield 98.7%, purity 99.92%. Total yield 93.5%. The HPLC detection spectrum of L-carnosine is as follows Figure 4 The specific data are shown in Table 5 below.

[0118] Table 5 HPLC detection spectrum data of L-carnosine of Example 2

[0119]

[0120] The elemental impurities in L-carnosine prepared in this example were determined, wherein the limits were based on ICH Q3D (R1): Elemental Impurities Guidelines, and the elemental impurities are as shown in Table 6:

[0121] Table 6 Elemental impurities in L-carnosine obtained in Example 2

[0122]

[0123] The residual solvent of L-carnosine prepared in this example was determined according to the ICH Q3C (R8): Impurities: Residual Solvents Guidelines. The specific results are shown in Table 7 below.

[0124] Table 7 Solvent residues in L-carnosine obtained in Example 2

[0125]

[0126] The L-carnosine prepared in this example was subjected to a stability test. The experimental conditions were formulated with reference to the ICH tripartite coordination guidelines (International Council for Registration of Pharmaceuticals for Human Use) Stability Tests for New APIs and Preparations Q1A (R2) 2.1.2 Strong Test. The specific results are shown in Table 8 below.

[0127] Table 8 Stability test of L-carnosine obtained in Example 2

[0128]

[0129] The experimental results show that the L-carnosine prepared by the method described in this article still has good stability even under relatively harsh conditions.

[0130] Example 3

[0131] S1. Preparation of Compound III

[0132] 70.9 kg of L-aspartic acid was added to a glass-lined kettle, nitrogen was replaced until the oxygen content was ≤6%, nitrogen protection was introduced, and then 248.2 kg of tetrahydrofuran and 137.8 kg of N, N-diisopropylethylamine were added, and the solid was stirred until all the solids were dissolved to obtain reaction solution A. 141.8 kg of tetrahydrofuran was added to the glass-lined kettle containing 2-cyanoacetyl chloride to be used as described in Synthesis Example 1, and the reaction solution B was obtained after stirring. The temperature of the reaction solution was controlled to 10-15 ° C, and the reaction solution B was added dropwise to the reaction solution A. After the addition was completed, the reaction solution was stirred at 20-25 ° C for 1 hour. After stirring, the reaction solution was concentrated under reduced pressure to 200 kg, 300 kg of water was added, and the reaction solution was stirred at 0-5 ° C for 1 hour, centrifuged and dried to obtain 104.8 kg of compound III, with a yield of 98.3% and a purity of 99.6%. The solvent tetrahydrofuran was collected and recycled for reuse.

[0133] S2. Preparation of Compound IV

[0134] 104.0 kg of compound III was added to a glass-lined kettle, and nitrogen was replaced until the oxygen content was ≤6%. Nitrogen protection was introduced, and then 520.0 kg of tetrahydrofuran was added. The temperature of the reaction liquid was controlled at 0-5°C, and 78.5 kg of phosphorus trichloride was added dropwise to the kettle. After the addition was completed, it was stirred at 0-5°C for 1 hour, heated to 20-25°C, stirred for 3 hours, centrifuged, rinsed with 0-5°C tetrahydrofuran, and dried to obtain 94.4 kg of compound IV with a yield of 99.7%.

[0135] S3. Preparation of Compound VI

[0136] Add 123.7 kg of trimethyl sulfoxide iodide and 38.8 kg of potassium carbonate to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 232.5 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 0-5°C to obtain reaction solution C. Add 93.0 kg of compound IV to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 232.5 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 0-5°C to obtain reaction solution D. Control the temperature of the reaction solution to 0-5°C, add reaction solution C dropwise to reaction solution D, and stir at 0-5°C for 2 hours after the dropwise addition. Control the temperature of the reaction solution to 0-5°C, and add 62.1 kg of hydrochloric acid dropwise to the kettle. Stir for 2 hours after the dropwise addition. Then add 63.8 kg of formamidine acetate to the kettle, raise the temperature of the reaction solution to 70-75°C, and stir for 3 hours. 500 kg of water and 500 kg of ethyl acetate were added to the kettle. After stirring and standing at room temperature, the aqueous phase was separated and concentrated under reduced pressure to 200 kg. The temperature of the concentrated liquid was controlled at 5-10 ° C. 350 kg of methanol was added dropwise, filtered, rinsed with methanol, and dried to obtain 108.7 kg of compound VI with a yield of 96.2% and a purity of 99.4%.

[0137] S4. Preparation of L-carnosine

[0138] Add 108.0kg of compound VI to a stainless steel kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 440.0kg of water, stir until completely dissolved, transfer to a stainless steel hydrogenation kettle, and draw a mixed solution of 1.679kg 5% palladium carbon, 0.007kg 46% boron trifluoride tetrahydrofuran and 100kg water under negative pressure, replace the air in the kettle with nitrogen 3 times, and replace the nitrogen in the kettle with hydrogen 3 times. Raise the kettle temperature to 60-65℃, pass hydrogen, stir at 0.5-1.5Mpa for 6h, lower the kettle temperature to room temperature, replace with nitrogen to normal pressure, filter through a titanium rod and filter element filter to remove the catalyst. The filtrate is transferred to a stainless steel kettle and concentrated under reduced pressure to 200kg. Control the temperature at 50-55℃, add 100kg of methanol to the kettle, add 0.540kg of L-carnosine seeds, grow crystals for 30 minutes, and add 200kg of methanol. Cool down to 10-15℃, stir for 6h, centrifuge, rinse with methanol, dry to obtain 109.0kg L-carnosine. Yield 98.7%, purity 99.89%. Total yield 93.1%. The HPLC detection spectrum of L-carnosine is as follows Figure 5 The specific data are shown in Table 9 below.

[0139] Table 9 HPLC detection spectrum data of L-carnosine of Example 3

[0140]

[0141] The elemental impurities in L-carnosine prepared in this example were determined, wherein the limits were based on ICH Q3D (R1): Elemental Impurities Guidelines, and the elemental impurities are as shown in Table 10:

[0142] Table 10 Elemental impurities in L-carnosine obtained in Example 3

[0143]

[0144] The residual solvent of L-carnosine prepared in this example was determined according to the ICH Q3C (R8): Impurities: Residual Solvents Guidelines. The specific results are shown in Table 11 below.

[0145] Table 11 Solvent residues in L-carnosine obtained in Example 3

[0146]

[0147] The L-carnosine prepared in this example was subjected to a stability test. The experimental conditions were formulated with reference to the ICH tripartite coordination guidelines (International Council for Registration of Pharmaceuticals for Human Use) Stability Testing for New APIs and Preparations Q1A (R2) 2.1.2 Strong Test. The specific results are shown in Table 12 below.

[0148] Table 12 Stability test of L-carnosine obtained in Example 3

[0149]

[0150] The experimental results show that the L-carnosine prepared by the method described in this article still has good stability even under relatively harsh conditions.

[0151] Example 4

[0152] S1. Preparation of Compound III

[0153] 70.9 kg of L-aspartic acid was added to a glass-lined kettle, nitrogen was replaced until the oxygen content was ≤6%, nitrogen protection was introduced, and then 248.2 kg of tetrahydrofuran and 137.8 kg of N, N-diisopropylethylamine were added, and the solid was stirred until all the solids were dissolved to obtain reaction solution A. 141.8 kg of tetrahydrofuran was added to the glass-lined kettle containing 2-cyanoacetyl chloride to be used as described in Synthesis Example 1, and the reaction solution B was obtained after stirring. The temperature of the reaction solution was controlled to 15-20°C, and the reaction solution B was added dropwise to the reaction solution A. After the addition was completed, the reaction solution was stirred at 20-25°C for 1 hour. After stirring, the reaction solution was concentrated under reduced pressure to 200 kg, 300 kg of water was added, and the reaction solution was stirred at 0-5°C for 1 hour, centrifuged and dried to obtain 105.0 kg of compound III with a yield of 98.5% and a purity of 99.6%. The solvent tetrahydrofuran was collected and recycled for reuse.

[0154] S2. Preparation of Compound IV

[0155] 104.0 kg of compound III was added to a glass-lined kettle, and nitrogen was replaced until the oxygen content was ≤6%. Nitrogen protection was introduced, and then 520.0 kg of tetrahydrofuran was added. The temperature of the reaction liquid was controlled at 0-5 ° C. 78.5 kg of phosphorus trichloride was added dropwise to the kettle. After the addition was completed, it was stirred at 0-5 ° C for 1 h, heated to 20-25 ° C, stirred for 3 h, centrifuged, rinsed with 0-5 ° C tetrahydrofuran, and dried to obtain 94.1 kg of compound IV with a yield of 99.4%.

[0156] S3. Preparation of Compound VI

[0157] Add 123.7 kg of trimethyl sulfoxide iodide and 38.8 kg of potassium carbonate to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 232.5 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 5-10 ° C to obtain reaction solution C. Add 93.0 kg of compound IV to a glass-lined kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 232.5 kg of N, N-dimethylformamide, stir for 1 hour, and lower the kettle temperature to 5-10 ° C to obtain reaction solution D. Control the temperature of the reaction solution to 0-5 ° C, add reaction solution C dropwise to reaction solution D, and stir at 0-5 ° C for 2 hours after the dropwise addition. Control the temperature of the reaction solution to 5-10 ° C, and add 62.1 kg of hydrochloric acid dropwise to the kettle. Stir for 2 hours after the dropwise addition. Then add 69.1 kg of formamidine acetate to the kettle, raise the temperature of the reaction solution to 75-80 ° C, and stir for 3 hours. 500 kg of water and 500 kg of ethyl acetate were added to the kettle. After stirring and standing at room temperature, the liquid was separated and the aqueous phase was retained. The concentrated liquid was concentrated to 200 kg under reduced pressure. The temperature of the concentrated liquid was controlled at 5-10 ° C. 350 kg of methanol was added dropwise, filtered, rinsed with methanol, and dried to obtain 108.8 kg of compound VI with a yield of 96.3% and a purity of 99.4%.

[0158] S4. Preparation of L-carnosine

[0159] Add 108.0kg of compound VI to a stainless steel kettle, replace with nitrogen until the oxygen content is ≤6%, pass nitrogen protection, then add 440.0kg of water, stir until completely dissolved, transfer to a stainless steel hydrogenation kettle, and draw a mixed solution of 4.197kg 5% palladium carbon, 0.018kg 46% boron trifluoride tetrahydrofuran and 100kg water under negative pressure, replace the air in the kettle with nitrogen 3 times, and replace the nitrogen in the kettle with hydrogen 3 times. Raise the kettle temperature to 65-70℃, pass hydrogen, stir at 1.0-2.0Mpa for 6h, lower the kettle temperature to room temperature, replace with nitrogen to normal pressure, filter through a titanium rod and filter element filter to remove the catalyst. The filtrate is transferred to a stainless steel kettle and concentrated under reduced pressure to 200kg. Control the temperature at 50-55℃, add 100kg of methanol to the kettle, add 0.540kg of L-carnosine seeds, grow the crystal for 30 minutes, and add 200kg of methanol. Cool down to 10-15℃, stir for 6h, centrifuge, rinse with methanol, dry to obtain 109.2kg L-carnosine. Yield 98.9%, purity 99.91%. Total yield 93.2%. The HPLC detection spectrum of L-carnosine is as follows Figure 6 The specific data are shown in Table 13 below.

[0160] Table 13 HPLC detection spectrum data of L-carnosine of Example 4

[0161]

[0162] The elemental impurities in L-carnosine prepared in this example were determined, wherein the limits were based on ICH Q3D (R1): Elemental Impurities Guidelines, and the elemental impurities are as shown in Table 14:

[0163] Table 14 Elemental impurities in L-carnosine obtained in Example 4

[0164]

[0165] The residual solvent of L-carnosine prepared in this example was determined according to the ICH Q3C (R8): Impurities: Residual Solvents Guidelines. The specific results are shown in Table 15 below.

[0166] Table 15 Solvent residues in L-carnosine obtained in Example 4

[0167]

[0168] The L-carnosine prepared in this example was subjected to a stability test. The experimental conditions were formulated with reference to the ICH tripartite coordination guidelines (International Council for Registration of Pharmaceuticals for Human Use) Stability Tests for New APIs and Preparations Q1A (R2) 2.1.2 Strong Test. The specific results are shown in Table 16 below.

[0169] Table 16 Stability test of L-carnosine obtained in Example 4

[0170]

[0171] The experimental results show that the L-carnosine prepared by the method described in this article still has good stability even under relatively harsh conditions.

[0172] Comparative Example 1

[0173] Compound III was prepared by the same method as in Example 1, except that, in step S1, reaction solution A was added dropwise to reaction solution B (different from the present invention in which reaction solution B was added dropwise to reaction solution A).

[0174] The purity of compound IV prepared by this comparative example is 83.5%. Changing the feeding method to make the local acyl chloride relatively excessive, dehydrating with carboxylic acid to form anhydride, and then reacting with the primary amine of L-aspartic acid that does not participate in the condensation reaction to form the corresponding dipeptide or polypeptide derivative, leads to a decrease in the purity of compound IV.

[0175] Comparative Example 2

[0176] Compound III was prepared by the same method as in Example 1, except that in step S1, when reaction solution B was added dropwise to reaction solution A, the temperature of the reaction solution was controlled to be -10 to 0°C (lower than the temperature specified in the present invention).

[0177] The yield of compound IV prepared by this comparative example is 99.2% and the purity is 99.5%. Compared with Example 1, there is no significant change in yield and purity. However, lowering the reaction temperature requires a lower condensing medium and a longer cooling time, resulting in a waste of energy and time.

[0178] Comparative Example 3

[0179] Compound III was prepared by the same method as in Example 1, except that in step S1, reaction solution B was added dropwise to reaction solution A, and the temperature of the reaction solution was controlled to be 30 to 40° C. (higher than the temperature specified in the present invention).

[0180] The purity of compound IV prepared by this comparative example is 87.3%. During the dropwise addition process, when the reaction temperature is raised to 30-40°C, part of the acyl chloride and carboxylic acid undergo dehydration to form anhydride, which continues to react with the primary amine of L-aspartic acid that does not participate in the condensation reaction to form the corresponding dipeptide or polypeptide derivative. This results in a decrease in the purity of compound IV.

[0181] Among them, the relevant conditions and product data of Comparative Examples 1-3 and Example 1 are shown in Table 17 below.

[0182] Table 17 Investigation of factors affecting the yield and purity of compound III in step S1

[0183]

[0184] Comparative Example 4

[0185] Compound IV was prepared by the same method as in Example 1, except that in step S2, the temperature of the reaction solution was controlled at -20°C to -10°C, and a dehydrating agent (lower than the temperature specified in the present invention) was added dropwise to compound III.

[0186] The yield of compound IV prepared by this comparative example is 99.6%. There is no significant change in yield relative to Example 1. However, lowering the reaction temperature requires a lower condensing medium and a longer cooling time, resulting in a waste of energy and time.

[0187] Comparative Example 5

[0188] Compound IV was prepared by the same method as in Example 1, except that in step S2, the temperature of the reaction solution was controlled at 30-40° C., and a dehydrating agent (higher than the temperature specified in the present invention) was added dropwise to compound III.

[0189] The yield of compound IV prepared by this comparative example is 87.6%. The yield is lower than that of Example 1. At a higher temperature, the cyano group may react more easily with the chlorination reagent to produce new impurities.

[0190] Among them, the relevant conditions and product data of Comparative Examples 4-5 and Example 1 are shown in Table 18 below.

[0191] Table 18 Investigation of factors affecting the yield of compound IV in step S2

[0192]

[0193] Comparative Example 6

[0194] Compound VI was prepared by the same method as in Example 1, except that in step S3, compound IV, trimethylsulfoxide iodide, potassium carbonate and an organic solvent were mixed, and then hydrochloric acid was added.

[0195] Compound VI prepared by this comparative example cannot be obtained by the same method as in Example 1. Changing the feeding method will cause the nucleophilic reagent trimethylsulfoxide iodide to attack multiple reaction sites of compound IV, produce a large amount of impurities, and wrap the product. During the post-treatment process, it is impossible to separate and purify the target intermediate VI.

[0196] Comparative Example 7

[0197] Compound VI was prepared by the same method as in Example 1, except that in step S3, reaction solution C was added dropwise to reaction solution D, and the temperature of the reaction solution was controlled to be -20°C to -10°C (lower than the temperature specified in the present invention).

[0198] The yield of compound VI prepared by this comparative example was 96.0% and the purity was 99.7%. There was no significant change in yield and purity compared with Example 1. However, lowering the reaction temperature requires a lower condensing medium and a longer cooling time, resulting in a waste of energy and time.

[0199] Comparative Example 8

[0200] Compound VI was prepared by the same method as in Example 1, except that in step S3, reaction solution C was added dropwise to reaction solution D, and the temperature of the reaction solution was controlled to be 30 to 40° C. (higher than the temperature specified in the present invention).

[0201] The yield of compound VI prepared by this comparative example is 65.2% and the purity is 47.6%. Compared with Example 1, the yield and purity are significantly reduced. Increasing the dropping temperature will also cause the nucleophilic reagent trimethyl sulfoxide iodide to attack multiple reaction sites of compound IV, produce impurities, and affect the high purity and high chiral compound VI.

[0202] Among them, the relevant conditions and product data of Comparative Examples 6-8 and Example 1 are shown in Table 19 below.

[0203] Table 19 Investigation of factors affecting the yield and purity of compound VI in step S3

[0204]

[0205] Comparative Example 9

[0206] L-carnosine was prepared by the same method as in Example 1, except that in step S4, the reaction temperature of the hydrogenation reduction was 40-50° C. (lower than the temperature specified in the present invention).

[0207] After 6 hours of reaction, the substrate was not completely reacted, and 8.84% of compound VI remained. If the reaction is to be complete, the reaction time needs to be extended, which increases the time consumption; or the catalyst dosage needs to be increased, which increases the cost of auxiliary materials.

[0208] Comparative Example 10

[0209] L-carnosine was prepared by the same method as in Example 1, except that in step S4, the reaction temperature of the hydrogenation reduction was 80-90° C. (higher than the temperature specified in the present invention).

[0210] The purity of L-carnosine prepared by this comparative example is 95.4%. Increasing the reaction temperature will enhance the hydrogenation reduction ability, so that the unsaturated bonds in the substrate and product structure participate in the reduction, generate corresponding reduction byproducts, and reduce the purity of the product L-carnosine.

[0211] Among them, the relevant conditions and product data of Comparative Examples 9-10 and Example 1 are shown in Table 20 below.

[0212] Table 20 Effect of step S4 reaction temperature on L-carnosine yield and purity

[0213]

[0214] By comparing the experimental results of Example 1 and Comparative Examples 9 and 10, it can be seen that if the temperature of the hydrogenation reduction reaction is lowered, the substrate reaction is incomplete, resulting in a decrease in the product yield; if the temperature of the hydrogenation reduction reaction is increased, the impurity level in the product will increase, resulting in a decrease in the product purity. Therefore, the reaction temperature defined in the present invention is more conducive to obtaining a high-purity, high-yield product.

[0215] Comparative Example 11

[0216] L-carnosine was prepared by the same method as in Example 1, except that in step S4, no boron element (boron trifluoride tetrahydrofuran) was added.

[0217] After 6 hours of reaction, the substrate was not completely reacted, and the remaining amount of compound VI was 7.01%. If the reaction is to be completed, the reaction time needs to be extended, which increases the time consumption; or the catalyst dosage needs to be increased, which increases the cost of auxiliary materials.

[0218] Comparative Example 12

[0219] L-carnosine was prepared by the same method as in Example 1, except that in step S4, the amount of palladium carbon and boron trifluoride tetrahydrofuran added was adjusted so that the molar ratio of compound VI to the palladium element and the boron element in the reduction catalyst was 1:0.0020:0.0001, and the molar ratio of the palladium element and the boron element in the reduction catalyst was 20:1.

[0220] After 6 hours of reaction, the substrate was not completely reacted, and 5.03% of compound VI remained. If the reaction is to be completed, the reaction time needs to be extended, which increases the time consumption; or the catalyst dosage needs to be increased, which increases the cost of auxiliary materials.

[0221] Comparative Example 13

[0222] L-carnosine was prepared by the same method as in Example 1, except that in step S4, the amounts of palladium carbon and boron trifluoride tetrahydrofuran added were adjusted so that the molar ratio of compound VI to the palladium element and the boron element in the reduction catalyst was 1:0.0020:0.0001, and the molar ratio of the palladium element and the boron element in the reduction catalyst was 2:1.

[0223] The purity of L-carnosine prepared by this comparative example is 98.4%. Increasing the reaction temperature will enhance the hydrogenation reduction ability, so that the unsaturated bonds in the substrate and product structures participate in the reduction, generate corresponding reduction byproducts, and reduce the purity of the product L-carnosine.

[0224] Among them, the relevant conditions and product data of Comparative Examples 11-13 and Examples 1-4 are shown in Table 21 below.

[0225] Table 21 Effect of step S4 catalyst composition on L-carnosine yield and purity

[0226]

[0227] By comparing the experimental results of Examples 1-4 and Comparative Examples 11-13, it can be seen that not adding boron or reducing the proportion of boron in the catalyst will result in incomplete substrate reaction due to decreased catalytic capacity; increasing the proportion of boron in the catalyst beyond the scope described in this patent application will result in excessive catalytic capacity, which will produce over-reduction impurities that are difficult to remove, indirectly leading to decreased product purity and yield. In view of the above research, it is shown that in the process of hydrogenation reduction reaction, in addition to the reaction temperature, the ratio of palladium and boron in the catalyst is also crucial, affecting the product quality of L-carnosine.

[0228] It can be seen from the above examples and comparative examples that examples 1-4 use the preparation method of the present invention to prepare L-carnosine: L-aspartic acid is used as the starting material, reacted with cyanoacetyl chloride to obtain (S)-2-(2-cyanoacetylamino)succinic acid, and under the action of a dehydrating agent, the corresponding acid anhydride is obtained, and then reacted with trimethylsulfoxide iodide, hydrochloric acid, acetic acid formamidine cyclization, and finally hydrogenation reduction to obtain the target product L-carnosine. The L-carnosine prepared by the above method has the characteristics of high purity, high yield, low raw material cost, and low energy consumption, and the prepared product has high stability and low single impurity content, and is more suitable for industrial production.

[0229] The technical features of the above-described embodiments may be arbitrarily combined. To make the description concise, not all possible combinations of the technical features in the above-described embodiments are exhaustively listed. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0230] For those skilled in the art, several modifications and improvements may be made without departing from the concept of the present invention, all of which belong to the protection scope of the present invention. The protection scope of the present invention shall be based on the attached claims.

Claims

1. A method for preparing L-carnosine, It is characterized in that The preparation method is: S1, after compound I reacts with a base, it reacts with compound II to obtain compound III; The compound I is , the compound II is , the compound III is ; S2, compound III is dissolved in an organic solvent, a dehydrating agent is added, and dehydration is performed to prepare compound IV; The compound IV is ; S3, compound IV is dissolved in an organic solvent, trimethyl sulfoxide iodide and hydrochloric acid are added to react to obtain compound V, and then formamidine acetate is added to the system to react to obtain compound VI; The compound V is , X is I or Cl; the compound VI is ; S4. Compound VI is dissolved in water, a reduction catalyst is added, and the target product L-carnosine is obtained by hydrogenation reduction; the reduction catalyst is a mixture of palladium carbon and boron trifluoride tetrahydrofuran.

2. A method for preparing L-carnosine according to claim 1, It is characterized in that The specific operation of step S1 is: dissolving compound I and a base in an organic solvent, stirring evenly to obtain a reaction solution A; dissolving compound II in an organic solvent to obtain a reaction solution B; and slowly dropping the reaction solution B into the reaction solution A to obtain compound III.

3. A method for preparing L-carnosine according to claim 2, It is characterized in that In step S1, the organic solvent is any one of acetonitrile and tetrahydrofuran; The base is any one of triethylamine and N,N-diisopropylethylamine.

4. A method for preparing L-carnosine according to claim 2, It is characterized in that In step S1, the reaction solution B is added to the reaction solution A for reaction at a temperature of 10-20°C.

5. A method for preparing L-carnosine according to claim 1, It is characterized in that In step S2, the organic solvent is dichloromethane or tetrahydrofuran; The dehydration agent is at least one of oxalyl chloride and phosphorus trichloride.

6. A method for preparing L-carnosine according to claim 1, It is characterized in that The specific operation of step S2 is: controlling the temperature to 0-10° C., adding a dehydrating agent dropwise to the organic solvent of compound III, and after the addition is complete, heating the mixture to 20-30° C. to carry out the reaction.

7. A method for preparing L-carnosine according to claim 1, It is characterized in that The specific operation of step S3 is: dissolving trimethyl sulfoxide iodide and potassium carbonate in an organic solvent to obtain a reaction solution C; dissolving compound IV in an organic solvent to obtain a reaction solution D; adding the reaction solution C dropwise to the reaction solution D, adding hydrochloric acid dropwise to obtain compound V, adding formamidine acetate, and reacting to obtain compound VI.

8. A method for preparing L-carnosine according to claim 7, It is characterized in that In step S3, the organic solvent is N,N-dimethylformamide; When preparing compound V, the reaction temperature is controlled at 0-10°C; when adding formamidine acetate to prepare compound VI, the reaction temperature is controlled at 70-80°C; In step S3, the molar ratio of trimethyl sulfoxide iodide to compound IV is (1.0-1.2):1; the molar ratio of formamidine acetate to compound IV is (1.0-1.3):

1.

9. A method for preparing L-carnosine according to claim 1, It is characterized in that In step S4, the molar ratio of compound VI to palladium and boron in the reduction catalyst is 1:(0.0005-0.0020):(0.0001-0.0003), and the molar ratio of palladium and boron in the reduction catalyst is (5-8):

1.

10. A method for preparing L-carnosine according to claim 1, It is characterized in that In step S4, the reaction temperature of the hydrogenation reduction is 60-70° C. and the pressure is 0.5-2.0 MPa.

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