A method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid

Abstract

The present invention relates to a method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid. The method mainly solves the technical problem that there is currently a lack of a method suitable for amplifying the synthesis of (2S, 4R)-2-amino-4-methylnonanoic acid. The synthesis of the present invention is carried out through eight steps, namely, (1) coupling of heptanoic acid with a chiral auxiliary group (R)-4-benzyl-2-oxazolidinone; (2) asymmetric methylation under the control of a chiral auxiliary group; (3) reduction to obtain the corresponding alcohol; (4) reaction of the alcohol with methanesulfonyl chloride to obtain the corresponding mesylate; (5) conversion of the mesylate to the corresponding iodide; (6) condensation of the iodide with diethyl acetamidomalonate; (7) hydrolysis and decarboxylation of the substituted diethyl acetamidomalonate; and (8) enzymatic separation by L-AcylaRe to obtain the final product. The target product with an optical purity of more than 99% is obtained by eight-step reaction. The present invention has the advantages of high yield, low cost, simple operation and purification, and is suitable for amplified synthesis.

Description

Technical Field

[0001] The present invention relates to a method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid. (2S, 4R)-2-amino-4-methylnonanoic acid is a non-natural chiral amino acid, an intermediate for synthesizing polypeptide drugs, and belongs to the technical field of drug synthesis chemical industry. Background Art

[0002] Peptide drugs are a new type of drug molecule between small molecule drugs and protein drugs, and they have some characteristics of both. The quality control level is close to that of small molecule drugs, and the activity is close to that of protein drugs. It combines the advantages of both and demonstrates its superiority in clinical application and production preparation. Therefore, peptide drug molecules have broad application prospects. However, peptide drugs also encounter problems such as poor stability and poor blood-brain barrier permeability during clinical application. In response to these problems, researchers have designed and screened peptide drugs with clinical application value by introducing non-natural chiral amino acids into peptide drugs for structural modification. With the development of new peptide drugs, the use of non-natural chiral amino acids in the medical field is increasing. Therefore, it is of great significance to study the synthesis of non-natural chiral amino acids.

[0003] The American Journal of Chemistry (J. Am. Chem. Soc. 1999, 121, 6197-6205, Dale L. Boger) first reported the total synthesis of a cyclic heptapeptide HUN-7293, an effective inhibitor of cell adhesion molecule expression with anti-inflammatory properties. It contains the unnatural amino acid (2S, 4R)-2-amino-4-methyloctanoic acid; the structural formula involved is as follows:

[0004] .

[0005] The Journal of Natural Products (J. Nat. Prod. 2006, 69, 736–741) disclosed that four peptaibolsculicinins with strong anticancer activity were isolated from the insect pathogenic fungus Culicinomyces clavisporus strain LL-12I252 (Figure 2). These compounds contain a new amino acid (2S)-2-amino-4-methyldecanoic acid, which is the first amino acid found in peptaibols. The Journal of Peptide Science (Journal of Peptide Science, 2011, 17(8), 576-580, Zhang, Wei) disclosed the chiral standard samples synthesized by Tong, and carefully examined and compared them with the nuclear magnetic resonance data of natural products, confirming that the stereochemistry of the amino acid at C-4 is R; the involved structural formula is as follows:

[0006] .

[0007] (2S, 4R)-2-Amino-4-methyl non-natural amino acids are a very important class of compounds in the synthesis of peptide drugs. Summary of the invention

[0008] The purpose of the present invention is to disclose a method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid which is simple, efficient, mild and suitable for scaled-up production. The main purpose is to solve the technical problem that there is currently a lack of a method suitable for scaled-up synthesis of (2S, 4R)-2-amino-4-methylnonanoic acid.

[0009] The technical scheme of the present invention is: a method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid, characterized in that it comprises the following steps:

[0010] Step 1: Add (R)-4-benzyl-2-oxazolidinone to heptanoic acid in the presence of pivaloyl chloride, triethylamine and lithium chloride to obtain (R)-4-benzyl-3-heptanoyl-2-oxazolidinone;

[0011] Step 2: Methylate the (R)-4-benzyl-3-heptanoyl-2-oxazolidinone obtained in step 1 with iodomethane in a 2-methyltetrahydrofuran solution in the presence of lithium diisopropylamide to obtain (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone;

[0012] Step 3: reducing the (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone obtained in step 2 in the presence of lithium borohydride to obtain (R)-2-methyl-1-heptanol;

[0013] Step 4: Add methanesulfonyl chloride to the (R)-2-methyl-1-heptanol obtained in step 3 in the presence of triethylamine to obtain (R)-methanesulfonic acid-2-methyl-1-heptyl ester;

[0014] Step 5: Add sodium iodide to the (R)-methanesulfonic acid-2-methyl-1-heptyl ester obtained in step 4 in an acetone solution to obtain (R)-1-iodo-2-methylheptane through reaction;

[0015] Step 6: Add diethyl acetamidomalonate to the (R)-1-iodo-2-methylheptane obtained in step 5 in the presence of a base to obtain diethyl (R)-2-acetamido-2-(2-methylheptyl)malonate;

[0016] Step 7: hydrolyze and decarboxylate the (R)-2-acetylamino-2-(2-methylheptyl)malonic acid diethyl ester obtained in step 6 in the presence of sodium hydroxide to obtain (4R)-2-acetylamino-4-methylnonanoic acid;

[0017] Step 8: The (4R)-2-acetylamino-4-methylnonanoic acid obtained in step 7 is hydrolyzed by L-Acylase to obtain (2S, 4R)-2-amino-4-methylnonanoic acid;

[0018] The reaction formula is as follows:

[0019] .

[0020] In the above reaction, the base in the sixth step is one of potassium tert-butoxide or sodium ethoxide, and preferably the base is potassium tert-butoxide.

[0021] The beneficial effects of the present invention are as follows: the present invention uses heptanoic acid as a raw material, obtains the chirality of the methyl group through asymmetric alkylation controlled by a chiral auxiliary group, then undergoes condensation with diethyl acetamidomalonate, hydrolysis, decarboxylation, and finally undergoes enzymatic hydrolysis and separation by L-Acylase to obtain the final product. The raw materials of this method are cheap and easy to obtain, the reaction conditions are mild, the method is simple and efficient, and the quality and yield are high, and it is economical and effective. DETAILED DESCRIPTION

[0022] Embodiment 1:

[0023] Step 1: Coupling of heptanoic acid with (R)-4-benzyl-2-oxazolidinone

[0024] Add 2-methyltetrahydrofuran (12 L), heptanoic acid (954 g, 7.33 mol) and triethylamine (1480 g, 14.66 mol) to a 20 L three-necked flask. Replace nitrogen, cool to -10 degrees Celsius, and then drop pivaloyl chloride (952g, 7.90 mol). After addition, stir at the same temperature for 2 hours. Add anhydrous lithium chloride (286 g, 6.75 mol) and (R)-4-benzyl-2-oxazolidinone (1.0 kg, 5.64 mol). After addition, slowly warm the reaction mixture to room temperature and stir overnight. Quench the reaction with 1 NHCl (6 L) and separate the layers. Wash the organic phase with 5% sodium hydroxide aqueous solution (6 L), wash with saturated brine (6 L), and dry with anhydrous sodium sulfate. The product was filtered and the filtrate was directly spin-dried to obtain (R)-4-benzyl-3-heptanoyl-2-oxazolidinone (1485 g, yield 91%).

[0025] Step 2: Methylation of (R)-4-benzyl-3-heptanoyl-2-oxazolidinone

[0026] Add 2-methyltetrahydrofuran (12 L) and (R)-4-benzyl-3-heptanoyl-2-oxazolidinone (1447 g, 5.0 mol) to a 20 L three-necked flask. Replace nitrogen, cool to -78 degrees Celsius, and then drop 2.0 M lithium diisopropylamide (LDA) tetrahydrofuran / n-heptane / ethylbenzene solution (3 L, 6.0 mol). After addition, stir at the same temperature for 0.5 hour. Add iodomethane (852 g, 6.0 mol) dropwise. After addition, slowly warm the reaction mixture to room temperature and stir overnight. Quench the reaction with 0.5 N HCl (6 L) and separate the layers. Wash the organic phase with saturated brine (6 L) and dry over anhydrous sodium sulfate. The mixture was filtered and the filtrate was directly dried by rotary evaporation to obtain a crude product, which was then purified by column chromatography to obtain (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone (1001 g, yield 66%).

[0027] Step 3: Reduction of (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone

[0028] Anhydrous tetrahydrofuran (10 L), methanol (211 g, 6.6 mol), (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone (1001 g, 3.3 mol) were added to a 20 L three-necked flask. The nitrogen was replaced, the mixture was cooled to 0 degrees Celsius, and then 2.0 M lithium borohydride tetrahydrofuran (3 L, 6.0 mol) was added dropwise. After the addition, the mixture was stirred at the same temperature for 0.5 hours. The reaction mixture was then slowly warmed to room temperature and stirred for 3 hours. The reaction was quenched with aqueous ammonium chloride solution (10 L) and extracted with ethyl acetate (10 L). The organic phase was washed with saturated brine (10 L) and dried over anhydrous sodium sulfate. The filtrate was filtered and directly dried to obtain the crude product, which was then purified by column chromatography to obtain (R)-2-methyl-1-heptanol (352 g, yield 82%), [a] D = +12.6(C= 1.0, CHCl3).

[0029] 1 H NMR (400 MHz, CDCl3): δ3.56-3.38 (m, 2 H), 1.67-1.56 (m, 1 H), 1.45-1.03 (m, 8 H), 0.96-0.87 (m, 6 H).

[0030] Step 4: Synthesis of (R)-2-methyl-1-heptyl methanesulfonate

[0031] Add methyl tert-butyl ether (MTBE) (3500 mL) and (R)-2-methyl-1-heptanol (352 g, 2.7 mol) to a 10 L three-necked flask, replace nitrogen, and cool to -10 degrees Celsius. Add methanesulfonyl chloride (332 g, 2.9 mol), and then add triethylamine (327 g, 3.24 mol) dropwise. After the addition, stir at the same temperature for 0.5 hours. Then the reaction mixture is slowly warmed to room temperature and stirred for 1 hour. Quench the reaction with 1 N HCl (1000 mL) and separate the liquids. Wash the organic phase with saturated brine (1000 mL) and dry over anhydrous sodium sulfate. Filter and spin-dry the filtrate to obtain (R)-methanesulfonic acid-2-methyl-1-heptyl ester (499 g, yield 89%).

[0032] Step 5: Synthesis of (R)-1-iodo-2-methylheptane

[0033] Add acetone (5000 mL), (R)-methanesulfonic acid-2-methyl-1-heptyl ester (499 g, 2.4 mol) and sodium iodide (540 g, 3.6 mol) to a 10 L three-necked flask. Heat and reflux overnight. Cool to room temperature. Filter and spin-dry the filtrate. Add MTBE (2000 mL), stir for 0.5 hours, filter, and spin-dry the filtrate to obtain (R)-1-iodo-2-methylheptane (535 g, yield 93%).

[0034] 1 H NMR (400 MHz, CDCl3): δ3.25 (dd, J 1 = 4.8 Hz, J 2 = 9.6 Hz, 1H),3.16 (dd, J 1 = 6.0 Hz, J 2= ​​9.6 Hz, 1H), 1.45-1.20 (m, 8H), 0.97 (d, J = 6.4Hz, 3H), 0.91 (t, J = 7.2 Hz, 3H).

[0035] Step 6: Synthesis of diethyl (R)-2-acetylamino-2-(2-methylheptyl)malonate

[0036] Add N,N-dimethylformamide (DMF) (4000 mL) and diethyl acetamidomalonate (521 g, 2.4 mol) to a 10 L three-necked flask. Replace nitrogen, cool to 0 degrees Celsius, and then add potassium tert-butoxide (296 g, 2.64 mol) in batches. After addition, stir at the same temperature for 0.5 hours. Then add (R)-1-iodo-2-methylheptane (530 g, 2.2 mol). After addition, the reaction mixture is heated to 80 degrees Celsius and stirred for 16 hours. Cool to room temperature. Dilute with water and extract with ethyl acetate. The organic phase is washed with saturated brine (1000 mL) and dried over anhydrous sodium sulfate. Filter, and the filtrate is directly spin-dried to obtain (R)-2-acetamido-2-(2-methylheptyl) diethyl malonate (692 g, yield 95%).

[0037] Step 7: Hydrolysis of diethyl (R)-2-acetylamino-2-(2-methylheptyl)malonate

[0038] Add ethanol (2000 mL), water (2000 mL) and (R)-2-acetylamino-2-(2-methylheptyl)malonate (691 g, 2.1 mol) to a 10 L three-necked flask. Cool to 0 degrees Celsius, then add sodium hydroxide (128 g, 3.2 mol) in batches. After addition, heat to reflux overnight. Cool to room temperature. Dilute with water and extract with ethyl acetate. Wash the organic phase with saturated brine (1000 mL) and dry over anhydrous sodium sulfate. Filter and spin dry the filtrate to obtain (4R)-2-acetylamino-4-methylnonanoic acid (229 g, yield 71%).

[0039] 1 H NMR (400 MHz, DMSO- d 6): δ12.45 (brs, 1H), 8.13-8.08 (m, 1H), 4.22-4.02 (m, 1H), 1.83-1.82 (m, 3H), 1.61-1.56 (m, 1H), 1.46-1.40 (m, 2H), 1.39-1.06 (m, 8H), 0.88-0.80 (m, 6H).

[0040] Step 8: Enzymatic separation of (4R)-2-acetylamino-4-methylnonanoic acid

[0041] Add pure water (5000 mL) and (4R)-2-acetylamino-4-methylnonanoic acid (229 g, 1.5 mol) to a 10 L three-necked flask. Adjust pH = 8~9 with 1 M sodium hydroxide aqueous solution. Heat to 37 degrees Celsius and add L-Acylase (Amano enzyme, 10 g). Maintain the temperature at 37 degrees Celsius and pH at 8~9 and stir for 24 hours. Adjust pH = 3 with hydrochloric acid and extract with ethyl acetate. Adjust the aqueous phase to pH = 6.5 and concentrate to a small volume. Collect the precipitated solid by filtration, wash with cold water, and dry to obtain (2S, 4R)-2-amino-4-methylnonanoic acid (112 g, yield 79%).

[0042] 1 H NMR (400 MHz, D2O): δ3.14 (dd, J 1 = 5.2 Hz, J 2= ​​8.8 Hz, 1H), 1.43-1.27 (m, 3H), 1.17-1.06 (m, 8H), 0.79-0.76 (m, 6H).

[0043] In Example 2, the base used in the sixth step is sodium ethoxide, and the rest is the same as in Example 1.

Claims

1. A method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid, characterized in that: The following steps are involved: Step 1: Add (R)-4-benzyl-2-oxazolidinone to heptanoic acid in the presence of pivaloyl chloride, triethylamine and lithium chloride to obtain (R)-4-benzyl-3-heptanoyl-2-oxazolidinone; Step 2: Methylate the (R)-4-benzyl-3-heptanoyl-2-oxazolidinone obtained in step 1 with iodomethane in a 2-methyltetrahydrofuran solution in the presence of lithium diisopropylamide to obtain (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone; Step 3: reducing the (R)-4-benzyl-3-((R)-2-methylheptanoyl)-2-oxazolidinone obtained in step 2 in the presence of lithium borohydride to obtain (R)-2-methyl-1-heptanol; Step 4: Add methanesulfonyl chloride to the (R)-2-methyl-1-heptanol obtained in step 3 in the presence of triethylamine to obtain (R)-methanesulfonic acid-2-methyl-1-heptyl ester; Step 5: Add sodium iodide to the (R)-methanesulfonic acid-2-methyl-1-heptyl ester obtained in step 4 in an acetone solution to obtain (R)-1-iodo-2-methylheptane through reaction; Step 6: Add diethyl acetamidomalonate to the (R)-1-iodo-2-methylheptane obtained in step 5 in the presence of a base to obtain diethyl (R)-2-acetamido-2-(2-methylheptyl)malonate; Step 7: hydrolyze and decarboxylate the (R)-2-acetylamino-2-(2-methylheptyl)malonic acid diethyl ester obtained in step 6 in the presence of sodium hydroxide to obtain (4R)-2-acetylamino-4-methylnonanoic acid; Step 8: The (4R)-2-acetylamino-4-methylnonanoic acid obtained in step 7 is hydrolyzed by L-Acylase to obtain (2S, 4R)-2-amino-4-methylnonanoic acid; the reaction formula is as follows: 。 2. A method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid according to claim 1, characterized in that: The base in the sixth step is potassium tert-butoxide or sodium ethoxide.

3. A method for synthesizing (2S, 4R)-2-amino-4-methylnonanoic acid according to claim 2, characterized in that: The base is potassium tert-butoxide.

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