Synthetic method of deoxycholic acid

Abstract

The invention relates to the technical field of deoxycholic acid synthesis, in particular to a synthetic method of deoxycholic acid. The synthetic method of deoxycholic acid has the advantages of short synthetic route, high yield, short production period and simplicity in operation.

Description

Technical Field

[0001] The invention relates to the technical field of deoxycholic acid synthesis, and in particular to a method for synthesizing deoxycholic acid. Background Art

[0002] The study of bile acid and bile acid chemistry has been going on for more than half a century. Bile acid accounts for the largest proportion of animal bile, accounting for more than 50% of the total amount of animal bile. Bile acid is a complex organic compound with cyclopentane and polyhydrophenanthrene as the basic parent nucleus. Its structural characteristics are that it consists of two parts: a rigid steroid nucleus and an aliphatic side chain. The 10th and 13th positions of the fused ring are linked to an aliphatic angular methyl structure, of which 3, 6, 7, and 12 can all be replaced by active oxygen-containing functional groups. Due to the differences in the number and position of the functional group substitutions, different levels of bile acid are formed. Normal human bile acid is divided into free bile acid and conjugated bile acid according to its structure. Among them, cholic acid, chenodeoxycholic acid, and deoxycholic acid are called free bile acids. The products of free bile acid combined with glycine or taurine, such as glycocholic acid, taurochenodeoxycholic acid, etc., are called conjugated bile acids. Moreover, free and conjugated bile acids mainly exist in the form of stable salts, so they can also be called bile salts. Conjugated bile acids are easily soluble in water, and the pH value in the aqueous solution is low, which makes bile salts exist in a more stable form. It is not easy to dissociate and precipitate solids in an acidic environment or in the presence of metal ions. From the source, it is divided into primary bile acid, secondary bile acid, and tertiary bile acid. Primary bile acid is produced from cholesterol in the liver, represented by cholic acid and chenodeoxycholic acid. After primary bile acid enters the intestine, it undergoes a deoxygenation reaction under the action of intestinal flora to generate secondary bile acid, mainly deoxycholic acid and ketolithocholic acid. After the secondary bile acid participates in the body's circulation and is reused in the body, the final product of metabolism in the intestine is called tertiary bile acid, which includes ursodeoxycholic acid and sulfolithocholic acid.

[0003] Deoxycholic acid, also known as deoxycholate and cholanic acid, is an injectable cell-dissolving drug with a chemical name of 3α, 12α-dihydroxy-5β-cholanic acid. It was first developed by Kaisera Biopharmaceuticals and approved by the U.S. FDA on April 29, 2015 under the trade name Belkyra. Deoxycholic acid injection can destroy cell membranes and cause cell lysis when injected into body tissues. It is used to improve moderate to severe contour protrusion or excessive facial fullness of submandibular fat in adults. It is not recommended for use in any other parts except submandibular fat cells.

[0004] There are currently two main strategies for the synthesis of deoxycholic acid:

[0005] 1. A semi-synthetic strategy using cholic acid as the starting material. Cholic acid undergoes sarret oxidation to selectively oxidize the 7-hydroxyl group to a carbonyl group, and then the 17-substituent is removed by Huang Minglong reduction to obtain the target product, deoxycholic acid. This synthesis method has few reaction steps and a simple process route. However, there are obvious problems: first, the currently commercialized cholic acid is extracted from animal liver, which contains animal-derived viruses and may cause allergies and other high safety hazards; second, the selectivity of the oxidation reaction during the synthesis process is poor, and the intermediate needs to be purified by column chromatography. In addition, the Huang Minglong reduction method has a high reaction temperature and rapidly releases gas, which is not conducive to industrial production safety.

[0006] 2. Using products obtained by fermentation and other operations of plant-extracted sterols, such as 9-hydroxyandrostenedione, cortisone and hydrocortisone, as starting materials, deoxycholic acid is prepared through a semi-synthetic strategy. This method fundamentally solves the problem of biosafety of synthetic raw materials and deoxycholic acid APIs. Using hydrocortisone as the starting material requires 17 steps of reaction, which is longer than 9-hydroxyandrostenedione, and undergoes multiple steps of hydrogenation, oxidation and reduction reactions. One of the reduction steps requires the control of low temperature of -78°C, which is difficult to operate in industrial production, and the cost of synthesis is high. 9-Hydroxyandrostenedione is a chemical raw material that has been commercialized on a large scale. It has a wide range of uses in many fields and has a stable supply from manufacturers. Therefore, most deoxycholic acid syntheses use 9-hydroxyandrostenedione as the raw material.

[0007] In WO 2008157635, Kaisela Biopharmaceuticals reported a method for synthesizing deoxycholic acid using 9-hydroxyandrostenedione as a raw material; Nanjing Norit Pharmaceuticals Co., Ltd. reported a new method for synthesizing deoxycholic acid in Chinese patent application CN106146593A. The method made the following improvements based on the original research: (1) the carbonyl at position 17 was protected by ketalization, and when the chiral hydroxyl at position 3 was constructed, the competitive reaction of the carbonyl at position 17 being reduced to hydroxyl was avoided, and the low-temperature reaction was also avoided; (2) the asymmetric hydrogenation at position 5 was adjusted to be carried out after the carbonyl at position 17 was protected by ketalization, avoiding the generation of byproducts of the hydrogenation of the carbonyl at position 17. However, both of the above methods inevitably have the disadvantages of long routes, low yields, long production cycles, and cumbersome operations. In particular, when the conjugated double bond of the carbonyl at position 12 is reduced, it is inevitable to use dry palladium carbon, which is very easy to ignite, and the reaction proceeds slowly. There are also a large number of byproducts of excessive reduction of the carbonyl, which need to be oxidized. In the experiment, the hydrogenation and oxidation reactions need to be repeated 5-7 times, and the purity of the product is not high. In production, this step of reaction alone consumes a large amount of dry palladium-carbon catalyst, the operation is cumbersome, the risk factor is high, and the production cycle is several times that of the same type of reaction, which greatly increases the production cost.

[0008] Therefore, developing a method for synthesizing deoxycholic acid with simple operation and low production cost has great application prospects. Summary of the invention

[0009] The present invention provides a method for synthesizing deoxycholic acid, comprising the following steps:

[0010] S1. Compound A11 reacts with methyl acrylate in the presence of a Lewis acid to produce compound A12, namely 3α-benzoyloxy-5β-chole-9(11),16-diene-24-oic acid methyl ester;

[0011] The A11 compound is (Z)-3α-benzoyloxy-5β-pregna-9(11),17(20)-diene.

[0012]

[0013] S2. In the presence of a first catalyst, compound A12 reacts with hydrogen to produce A13, ie, 3α-benzoyloxy-5β-chole-9(11)-ene-24-oic acid methyl ester;

[0014]

[0015] S3. Compound A13 reacts with an oxidant to generate A14, i.e., 3α-benzoyloxy-5β-chole-9(11)-ene-12-one-24-oic acid methyl ester;

[0016]

[0017] S4. Compound A14 is hydrolyzed with a base to generate A15, i.e., 3α-hydroxy-5β-chole-9(11)-ene-12-one-24-oic acid;

[0018]

[0019] S5. In the presence of a second catalyst, compound A15 is hydrogenated and reduced to produce A16, i.e., 3α-hydroxy-5β-cholane-12-one-24-acid;

[0020]

[0021] S6. In the presence of a third catalyst, compound A16 undergoes an esterification reaction with methanol to produce A17, i.e., 3α-hydroxy-5β-cholane-12-one-24-acid methyl ester;

[0022]

[0023] S7. In the presence of a chiral reducing agent, compound A17 is chirally reduced to generate A18, i.e., methyl deoxycholate;

[0024]

[0025] S8. Compound A18 is hydrolyzed with alkali to generate DCA, i.e., deoxycholic acid;

[0026]

[0027] The synthesis method of the deoxycholic acid is as follows:

[0028]

[0029] As an embodiment, the Lewis acid is selected from at least one of AlCl3, EtAlCl2, and TiCl4.

[0030] As an embodiment, the first catalyst in S2 is selected from at least one of Pd / C, Pd(OH)2 / C, and PtO2 / C.

[0031] As an embodiment, the solvent in S2 is selected from at least one of tetrahydrofuran, methanol, ethanol, ethyl acetate, and N-methylpyrrolidone.

[0032] As an embodiment, the oxidant in S3 is selected from at least one of tert-butyl peroxide, hydrogen peroxide, peracetic acid, chromium trioxide, pyridinium chlorochromate, sodium hypochlorite, 2-iodoacetylbenzoic acid and Dess-Martin periodinane.

[0033] As an embodiment, the base in S4 is selected from at least one of sodium hydroxide, potassium hydroxide and lithium hydroxide.

[0034] As an embodiment, the solvent in S4 is selected from at least one of tetrahydrofuran, methanol, ethanol and acetone.

[0035] As an embodiment, the second catalyst in S5 is selected from at least one of Pd / C, dry palladium carbon, Pd(OH)2 / C, PtO2 / C and Raney nickel.

[0036] As an embodiment, the pressure of hydrogenation in S5 is 0.3MPa-3.0MPa, and the reaction temperature of hydrogenation is 20°C-80°C.

[0037] As an embodiment, the solvent in S5 is selected from at least one of tetrahydrofuran, methanol, ethanol, ethyl acetate, and N-methylpyrrolidone.

[0038] As an embodiment, the third catalyst in S6 is selected from at least one of concentrated sulfuric acid, hydrochloric acid, thionyl chloride, and phosphorus trichloride.

[0039] As an embodiment, the chiral reducing agent in S7 is selected from at least one of LiAl(Ot-Bu)3H, sodium triacetoxyborohydride, and diisopinocampheyl chloroborane.

[0040] As an embodiment, the base in S8 is selected from at least one of sodium hydroxide, potassium hydroxide, and lithium hydroxide.

[0041] As an embodiment, the solvent in S8 is selected from at least one of tetrahydrofuran, methanol, ethanol and acetone.

[0042] Compared with the prior art, the present invention has the following beneficial effects:

[0043] 1. Compared with the reported methods, the present invention focuses on optimizing the key step of hydrogenating the carbonyl conjugated double bond at position 12, effectively avoiding the need to use dry palladium carbon and the generation of carbonyl reduction by-products.

[0044] 2. Compared with the previously reported methods, the present invention avoids the oxidation process of carbonyl reduction byproducts, significantly shortens the production cycle, and effectively reduces production costs.

[0045] 3. The total yield of deoxycholic acid prepared by the present invention reaches 44.14%, which is greatly improved in actual production compared with the reported methods.

[0046] 4. The deoxycholic acid synthesis method of the present invention obtains deoxycholic acid with a purity of more than 99.8%, and the product quality is significantly better than that of the reported method.

[0047] 5. The synthesis method of deoxycholic acid described in the present invention has the advantages of short synthesis route, high yield, short production cycle and simple operation. The production cost is greatly reduced compared with the reported methods and is suitable for large-scale industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 is the HNMR spectrum of compound A11.

[0049] Figure 2 This is the HPLC spectrum of compound A11.

[0050] Figure 3 This is the HNMR spectrum of deoxycholic acid prepared in Example 1.

[0051] Figure 4 This is the HPLC detection spectrum of deoxycholic acid prepared in Example 1.

[0052] Figure 5 This is the HPLC detection spectrum of deoxycholic acid prepared in Example 2.

[0053] Figure 6This is the HPLC detection spectrum of deoxycholic acid prepared in Example 3. DETAILED DESCRIPTION

[0054] The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0055] The material information used in the present invention is as follows:

[0056] Ethyl aluminum dichloride: CAS number: 563-43-9;

[0057] Pd / C: CAS No.: 7440-05-3;

[0058] PCC: CAS No.: 26299-14-9;

[0059] PtO2 / C: CAS No.: 1314-15-4;

[0060] Lithium tri-tert-butoxyaluminum hydride: CAS number: 17476-04-9.

[0061] The preparation method of the A11 compound (Z)-3α-benzoyloxy-5β-pregnane-9(11),17(20)-diene in the present invention is as follows:

[0062]

[0063] (1) Preparation of intermediate I-2

[0064] Add 8 L of dichloromethane to a 20 L three-necked flask, add 1.0 kg of SM1 raw material under stirring, stir at room temperature until it is completely dissolved, slowly add 487 g of concentrated sulfuric acid, after the concentrated sulfuric acid is added, heat to reflux, react for 5 hours, and TLC detection shows no SM1 raw material.

[0065] The reaction system was cooled to 0°C, 2N sodium hydroxide solution was added dropwise to a pH of 7-8, the liquids were separated, the aqueous phase was back-extracted twice with DCM, the combined organic phases were washed once with purified water and saturated sodium chloride, the organic phase was distilled under reduced pressure to obtain a light yellow solid, methyl tert-butyl ether was added and slurried for 3 h, the filter was filtered, and the filter cake was washed with methyl tert-butyl ether to obtain an off-white solid, which was dried at 45°C to obtain 870 g of an off-white solid, with a yield of 90% and a purity of 99.55%.

[0066] (2) Preparation of intermediate I-3

[0067] Add 6960 ml of toluene to a 20L three-necked flask, add 870 g of I-2 material and 29.1 g of p-toluenesulfonic acid monohydrate under stirring, heat to reflux, open the water separator to receive the azeotrope of toluene and water, maintain reflux, react until TLC detects no I-2 raw material, and stop the reaction.

[0068] The reaction mixture was cooled to room temperature, 1740 ml of ethyl acetate was added, washed with 2 L of purified water and saturated sodium chloride, the organic phase was concentrated to dryness, and then 1740 ml of n-hexane was added. The temperature was lowered to 10°C, stirred, and filtered. The filter cake was washed with 500 ml of n-hexane mixed solution to obtain an off-white solid. The solid was dried at 45°C for 10 h to obtain 930 g of an off-white solid with a yield of 91.7% and a purity of 98.59%.

[0069] (3) Preparation of intermediate I-4

[0070] Add 2920 ml of N-methylpyrrolidone (NMP) to a 5 L hydrogenation reactor, add 730 g of I-3 material under stirring, replace hydrogen, maintain the pressure at 0.5 MPa, react at 20 ° C for 16 hours, and TLC detection shows no raw material.

[0071] The reaction solution was filtered, and the solid was rinsed with a small amount of NMP to obtain a light yellow filtrate. 5840 ml of purified water was added dropwise under stirring and filtered to obtain a white solid. The filter cake was rinsed with 2190 ml of purified water to obtain a white solid. The filter cake was dried at 45°C for 16 h to obtain 662 g of a white solid with a yield of 90% and a purity of 94.79%.

[0072] (4) Preparation of intermediate I-5

[0073] Add 3010 ml of tetrahydrofuran to a 10L three-necked flask, add 602 g of I-4 material under stirring, replace nitrogen, cool to -10°C under nitrogen protection, slowly add 2000 ml of lithium tri-tert-butoxyaluminum hydride tetrahydrofuran solution (1 mol / L), after the addition is complete, keep at -10°C for 2 hours, and TLC detection shows that there is no I-4 raw material.

[0074] 3010 ml of hydrochloric acid (6 mol / L) was slowly added dropwise under ice bath. After the addition was completed, the temperature was raised to 20°C and reacted for 2 h. TLC detected that there was no starting material.

[0075] 1800 ml of purified water and 1200 ml of ethyl acetate were added to the reaction solution, stirred for 30 minutes, and allowed to stand for stratification. The aqueous phase was extracted twice with 1800 ml of ethyl acetate, the organic phases were combined, the organic phase was washed with 2400 ml of purified water and once with 2400 ml of saturated sodium chloride, dried over anhydrous sodium sulfate, filtered, and the filtrate was distilled under reduced pressure to obtain a white solid, which was recrystallized using 1200 ml of a mixed solution of acetone: n-hexane = 1:5, filtered, and the filter cake was washed with 300 ml of a mixed solution of acetone: n-hexane = 1:8 to obtain a white solid, and dried at 45°C for 6 h to obtain 463 g of a white solid with a yield of 88.3% and a purity of 98.52%.

[0076] (5) Preparation of (Z)-3a-benzoyloxy-5β-pregna-9(11),17(20)-diene

[0077] Add 4716 ml of tetrahydrofuran to a 10L three-necked flask, add 1113 g of triphenylethylphosphonium bromide under stirring, replace nitrogen, cool to -10°C under nitrogen protection, add 336 g of potassium tert-butoxide, keep at -10°C for 1 h after the addition is completed, add 393 g of I-5, and react for 8 h after the addition is completed. TLC detection shows that there is no raw material.

[0078] 17.1 g of 4-dimethylaminopyridine was added to the reaction solution, and 620 g of triethylamine was slowly added dropwise. After the addition of triethylamine was completed, 767 g of benzoyl chloride was slowly added dropwise. After the addition was completed, the temperature was raised to 20-30°C and the reaction was carried out for 16 hours. TLC detection showed that there was no starting material.

[0079] 7800 ml of purified water was slowly added dropwise to the reaction solution, and then 1560 ml of ether was added for extraction. The aqueous phase was extracted twice with 800 ml of ether respectively, and the organic phases were combined and washed once with 400 ml of saturated sodium chloride. The organic phase was distilled under reduced pressure until no liquid flowed out to obtain a brown solid-liquid mixture.

[0080] The crude product was recrystallized with 2000 ml of isopropanol and filtered. The filter cake was rinsed with 800 ml of 0°C isopropanol to obtain a white solid. The white solid was dried at 45°C for 16 h to obtain 475 g of white solid with a yield of 86.8% and a purity of 98.94%.

[0081] 1H NMR(400MHz,Chloroform-d)δ8.14–7.94(m,2H),7.58–7.48(m,1H),7.42(dd,J=8.4, 7.0Hz,2H),5.51–5.34(m,1H),5.20(tdd,J=7.1,5.0,2.1Hz,1H),5.00(ddd,J=10.9,5 .2,2.8Hz,1H),2.48–2.33(m,3H),2.25(dq,J=8.3,3.8,3.0Hz,1H),2.18–1.96(m,3H ),1.92–1.82(m,1H),1.82–1.61(m,9H),1.50–1.15(m,5H),1.11(s,3H),0.82(s,3H).

[0082] LCMS: 405.2789 [M+H] +

[0083] MP=109.2℃-111.7℃

[0084] HPLC=98.94%

[0085] HNMR spectrum see Figure 1 , HPLC spectrum see Figure 2 .

[0086] Example 1

[0087] This embodiment provides a method for synthesizing deoxycholic acid, comprising the following steps:

[0088] S1. Preparation of intermediate A12 (3α-benzoyloxy-5β-chole-9(11),16-diene-24-oic acid methyl ester)

[0089] Add 5000mL of dichloromethane to a 10L three-necked flask, add 500g of A11 material under stirring, stir at room temperature to dissolve the solid, evacuate, replace nitrogen, cool to -10°C under nitrogen protection, add 233.82g of methyl acrylate dropwise, after the addition is completed, keep -10°C to react for 1h, add 1569.49g of dichloroethylaluminum n-hexane solution (25%wt), after the addition is completed, heat to 25°C and react for 24h.

[0090] The A11 compound is (Z)-3α-benzoyloxy-5β-pregna-9(11),17(20)-diene;

[0091] The above reaction solution was slowly added dropwise to 10 L of ice water. After the addition was completed, the mixture was allowed to stand and separate into layers. The aqueous phase was extracted twice with dichloromethane. The organic phases were combined, washed with water and saturated sodium chloride. The organic phase was distilled under reduced pressure until no liquid flowed out. The solid was recrystallized from isopropanol / water = 5 / 1 to obtain a white solid. The solid was dried at 45°C with air for 16 h to obtain 540 g of a white to off-white solid with a yield of 89.1%.

[0092] S2. Preparation of intermediate A13 (3α-benzoyloxy-5β-chole-9(11)-ene-24-oic acid methyl ester)

[0093] Add 1940 mL of tetrahydrofuran to a 5 L hydrogenation reactor, add 485 g of A12 material under stirring, the solid is dissolved, add 33.95 g of 10% Pd / C, replace the hydrogen, maintain the pressure at 0.2 MPa, and react for 16 h.

[0094] The reaction solution was filtered and rinsed with 500 mL of tetrahydrofuran. The filtrate was distilled under reduced pressure until no liquid flowed out to obtain a colorless oil. The oil was heated with 2 L of acetonitrile until it was completely dissolved. The temperature was lowered to 0°C, and a large amount of solid precipitated. The temperature was maintained to continue crystallization for 4 h. The filter cake was filtered with suction, and 485 mL of acetonitrile (0°C) was used to rinse the filter cake. The wet product was placed at 45°C and dried with air for 16 h to obtain 447 g of a white solid product with a yield of 91.8%.

[0095] S3. Preparation of intermediate A14 (3α-benzoyloxy-5β-chole-9(11)-ene-12-one-24-oic acid methyl ester)

[0096] Add 3200 mL of acetonitrile to a 10 L three-necked flask, add 320 g of A13 material under stirring, raise the temperature to 45°C, add 86.59 g of cuprous iodide, slowly dropwise add 146.50 g of tert-butyl peroxide, and maintain the reaction at 45°C for 16 h.

[0097] The temperature was lowered to 30°C, 160.00 g of diatomaceous earth was added to the reaction solution, filtered, rinsed with ethyl acetate, the collected filtrate was washed once with 1280 mL of 10% sodium sulfite aqueous solution, and the combined organic phases were washed once with 640 mL of saturated saline solution.

[0098] The organic phase was transferred to a 10 L three-necked flask, and 140.00 g of PCC was added under stirring. The mixture was reacted at 30° C. for 16 h. TLC detection (PE:EA=8:1, 254 nm) showed that no raw material remained.

[0099] The reaction solution was filtered, rinsed with 1600 mL of ethyl acetate, and the collected filtrate was washed once with 1280 mL of 10% sodium sulfite aqueous solution, and then washed once with 640 mL of water and saturated saline solution. The organic phase was distilled under reduced pressure to remove dichloromethane, and the solid was recrystallized from n-heptane to obtain a white solid, which was dried at 45°C with air blast to obtain 272 g of a white solid with a yield of 82.5%.

[0100] S4. Preparation of intermediate A15 (3α-hydroxy-5β-chole-9(11)-ene-12-one-24-oic acid)

[0101] Take a 5L three-necked flask, add 450mL of tetrahydrofuran and methanol respectively, add 180g of A14 raw material under stirring, dissolve under stirring, add 710mL of 2N lithium hydroxide aqueous solution, heat to 50℃ and react for 8h. TLC detection shows that no raw material remains.

[0102] The reaction solution was cooled to 20°C, and methanol and tetrahydrofuran were distilled off under reduced pressure to obtain a colorless liquid. 2N dilute hydrochloric acid was added dropwise to adjust the pH to 2-3. A large amount of white solid precipitated. The mixture was stirred for crystallization for 2 h, filtered, and washed with 900 mL of purified water to obtain a white solid. The mixture was dried with air at 45°C for 16 h to obtain 101.5 g of a white solid with a yield of 93.8%.

[0103] S5. Preparation of Intermediate A16 (3α-Hydroxy-5β-cholane-12-one-24-acid) Solution

[0104] Add 700 mL of methanol to the hydrogenation kettle, add 70 g of A15 raw material under stirring, completely dissolve, add 30.9 g of palladium carbon, replace with nitrogen and then replace with hydrogen three times, pressurize the hydrogen to 0.4 MPa, heat to 65 ° C and react for 16 hours. Cool to 20 ° C, filter, rinse, and the solution is directly used for the next reaction.

[0105] S6. Preparation of intermediate A17 (3α-hydroxy-5β-cholan-12-one-24-oic acid methyl ester)

[0106] Add 3.5 g of concentrated sulfuric acid to the A16 solution, heat to 60°C, react for 4 h, and stop the reaction after TLC detection. Add 700 mL of purified water, a large amount of solid precipitates, cool by 20°C, continue stirring and crystallizing for 5 h, filter to obtain a white solid, rinse with 700 mL of purified water, dry the solid at 45°C, collect the material to obtain a white solid, dry at 45°C for 16 h, and obtain 66 g of a white solid with a yield of 91.0%.

[0107] S7. Preparation of intermediate A18 (methyl deoxycholate)

[0108] Take a 2L three-necked flask, add 400mL of tetrahydrofuran, add 40.0g of A17 solid under stirring, cool to -10℃, slowly add 200mL of lithium tri-tert-butoxyaluminum hydride tetrahydrofuran solution dropwise, release heat in small amount, control the internal temperature not higher than 10℃, after the addition is complete, keep the reaction warm for 16h.

[0109] Add 1000 mL of 2N / L dilute hydrochloric acid to the reaction solution at low temperature, control the internal temperature not higher than 20°C, add 1000 mL of ethyl acetate, stir, separate the layers, extract the aqueous phase once with 500 mL of ethyl acetate, combine the organic phases, wash once with 1000 mL of purified water and 500 mL of saturated brine respectively, distill the organic phase under reduced pressure to obtain a colorless oil, and use the crude product directly for the next step reaction without purification.

[0110] S8. Preparation of target DCA

[0111] Transfer the crude oil A18 from the previous step into a 1L flask, add 100mL of methanol and 100mL of tetrahydrofuran, and dissolve under stirring. Slowly add 100mL of 2N / L NaOH solution, raise the temperature to 40°C and react for 2h. Detect by TLC. The reaction is complete when no A18 is found.

[0112] To the reaction solution, 400 mL of purified water was added, methanol and tetrahydrofuran were removed by distillation under reduced pressure, and 2N / L hydrochloric acid was added dropwise with stirring to adjust the pH to 1-2 (internal temperature was controlled at 15-25°C). A large amount of solid precipitated, and the mixture was stirred at 20°C for 4 h. The mixture was filtered and washed with 800 mL of purified water until the filtrate was neutral to obtain a white solid. The solid was dried at 45°C for 24 h to obtain 40.0 g of crude DCA (yield 99.5%).

[0113] Take a 500mL three-necked flask, add 60mL of methanol, add 30.0g of crude DCA (deoxycholic acid) under stirring, the solid is completely dissolved, slowly add 450mL of dichloromethane, the reaction solution is clarified, heated to 40°C, distilled, the solid slowly precipitates, take the supernatant of the reaction solution, GC detection, control the methanol content in the reaction solution at 3-4%, stop adding dichloromethane and distillation, keep the internal temperature at 20°C, stir and crystallize for 4h, filter, rinse with 150mL of dichloromethane to obtain a white solid, dry at 45°C for 8h, and obtain 25.6g of the product as a white solid with a yield of 85.3%.

[0114] Take a 1000mL three-necked flask, add 138mL of purified water, add 2.76g of sodium hydroxide under stirring, dissolve under stirring (control the internal temperature not higher than 20°C), add 23g of DCA solid in batches, and all the solids are dissolved (control the internal temperature not higher than 20°C), add 0.69g of activated carbon, heat to 80°C and stir for 2h, filter, cool to 10°C, add about 35mL of 2N / L dilute hydrochloric acid dropwise, adjust the pH to 1-2 (control the internal temperature not higher than 20°C), stir at 20°C for 2h, filter, rinse with 460mL of purified water until the filtrate is neutral, and obtain a white solid, dry at 45°C with air blast for 16-24h to obtain 20.76g of white solid, with a yield of 90.3% and a purity (CAD) of 99.8% ( Figure 4 ).

[0115] Example 2

[0116] This embodiment provides a method for synthesizing deoxycholic acid, comprising the following steps:

[0117] S1. Preparation of intermediate A12 (3α-benzoyloxy-5β-chole-9(11),16-diene-24-oic acid methyl ester): same as Example 1.

[0118] S2. Preparation of intermediate A13 (3α-benzoyloxy-5β-chole-9(11)-ene-24-oic acid methyl ester)

[0119] Add 500 mL of methanol to a 1 L hydrogenation reactor, add 100 g of A12 material under stirring, add 7.0 g of 10% Pd / C, replace the hydrogen, maintain the pressure at 0.2 MPa, and react for 24 hours.

[0120] The reaction solution was filtered and washed with methanol. The filtrate was concentrated to obtain a colorless oil. The oil was heated with acetonitrile until it was completely dissolved. The temperature was lowered to 0°C, and a large amount of solid precipitated. The temperature was maintained to continue crystallization for 4 hours. The product was filtered with suction, and the filter cake was washed with acetonitrile (0°C). The wet product was placed at 45°C and dried with air for 16 hours to obtain 84.7 g of a white solid product with a yield of 84.3%.

[0121] S3. Preparation of intermediate A14 (3α-benzoyloxy-5β-chole-9(11)-ene-12-one-24-oic acid methyl ester)

[0122] Add 1 L of glacial acetic acid to a 21 L three-necked flask, add 100 g of A13 material under stirring, then add 100 g of chromium trioxide, raise the temperature to 65 °C, and react for 24 h.

[0123] After the reaction was completed, most of the glacial acetic acid was removed by vacuum distillation, and then 2L of purified water was slowly added dropwise, a large amount of solid precipitated, filtered, and the solid was washed with water until the filtrate was colorless, and dried at 45°C to obtain a crude product. The obtained crude product was recrystallized from n-heptane to obtain a white solid, and dried at 45°C to obtain 68.5g of a white solid, with a yield of 66.5%.

[0124] S4. Preparation of intermediate A15 (3α-hydroxy-5β-chole-9(11)-ene-12-one-24-oic acid)

[0125] Take a 1L three-necked flask, add 450mL of tetrahydrofuran and methanol respectively, add 180g of A14 raw material under stirring, dissolve under stirring, add 710mL of 2N lithium hydroxide aqueous solution, react at 25°C for 24h, and TLC detection shows no raw material remaining.

[0126] The reaction solution was cooled to 20°C, and methanol and tetrahydrofuran were removed by distillation under reduced pressure to obtain a colorless liquid. 2N dilute hydrochloric acid was added dropwise to adjust the pH to 2-3, and a large amount of white solid precipitated. The solution was heated to 75°C, stirred and hot-slurried for 2-3 hours, filtered, filtered, and the solid was slurried once with 75°C hot water, filtered, and washed with 900 mL of purified water to obtain a white solid. The solution was dried at 45°C for 16 hours to obtain 99.8 g of a white solid with a yield of 92.2%.

[0127] S5. Preparation of Intermediate A16 (3α-Hydroxy-5β-cholane-12-one-24-acid) Solution

[0128] Add 700 mL of methanol to the hydrogenation kettle, add 70 g of A15 raw material under stirring, completely dissolve, add 18.5 g of platinum carbon, replace with nitrogen and then replace with hydrogen three times, pressurize the hydrogen to 1.5 MPa, react at room temperature for 32 hours, filter, rinse, and directly proceed to the next step of reaction.

[0129] S6. Preparation of intermediate A17 (3α-hydroxy-5β-cholan-12-one-24-oic acid methyl ester): same as Example 1.

[0130] S7. Preparation of intermediate A18 (methyl deoxycholate)

[0131] Take a 2L three-necked flask, add 400mL of tetrahydrofuran, add 40.0g of A17 solid under stirring, cool to -10℃, slowly add 185mL of diisopinocampheyl chloroborane tetrahydrofuran solution, and after the addition is complete, keep the reaction warm for 48h.

[0132] Water was added to the reaction solution at low temperature to quench the reaction, the internal temperature was controlled not higher than 20°C, 1000 mL of ethyl acetate was added, stirred, and layers were separated. The aqueous phase was extracted once with 500 mL of ethyl acetate, the organic phases were combined, and washed once with 1000 mL of purified water and 500 mL of saturated brine respectively. The organic phase was distilled under reduced pressure to obtain a colorless oil, and the crude product was directly used for the next step reaction without purification.

[0133] S8. Preparation of target DCA

[0134] The crude oil A18 from the previous step was transferred to a 1L flask, and 100mL of methanol and 100mL of tetrahydrofuran were added to dissolve under stirring. 100mL of 2N / L LiOH solution was slowly added, and the reaction was allowed to react at room temperature for 12h. The reaction was terminated when no A18 was found by TLC.

[0135] Add 400 mL of purified water to the reaction solution, remove methanol and tetrahydrofuran by distillation under reduced pressure, add 2N / L hydrochloric acid dropwise with stirring to adjust the pH to 1-2 (internal temperature controlled at 15-25°C), a large amount of solid precipitates, heat to 80°C and stir for 4 h, filter, wash with 800 mL of hot water until the filtrate is neutral, and obtain a white solid, which is dried at 45°C with air for 24 h to obtain 39.8 g of crude DCA (yield 98.5%).

[0136] Heat 30 g of crude DCA to 35-45°C with 400 ml of 4% methanol in dichloromethane solution, keep reflux and slurry for 8 h, cool to 20-30°C and stir for 1-2 h, filter, wash with 150 mL of dichloromethane to obtain a white solid, dry with air at 45°C for 8 h to obtain 25.8 g of the product as a white solid, with a yield of 86.0%.

[0137] Take a 1000mL three-necked flask, add 138mL of purified water, add 2.76g of sodium hydroxide while stirring, dissolve while stirring (control the internal temperature not higher than 20℃), add 23g of DCA solid in batches, and dissolve all the solid (control the internal temperature not higher than 20℃), add 0.69g of activated carbon, heat to 80℃ and stir for 2h, filter, cool to 10℃, add about 35mL of 2N / L dilute hydrochloric acid, adjust the pH to 1-2 (control the internal temperature not higher than 20℃), stir at 80℃ for 2h, filter, rinse with hot water until the filtrate is neutral, and obtain a white solid. Dry at 45℃ with air blast for 16-24h to obtain 21.5g of white solid, with a yield of 93.5% and a purity (CAD) of 99.85%. ( Figure 5 )

[0138] Example 3

[0139] This embodiment provides a method for synthesizing deoxycholic acid, comprising the following steps:

[0140] S1. Preparation of intermediate A12 (3α-benzoyloxy-5β-chole-9(11),16-diene-24-oic acid methyl ester): same as Example 1.

[0141] S2. Preparation of intermediate A13 (3α-benzoyloxy-5β-chole-9(11)-ene-24-oic acid methyl ester): same as Example 1.

[0142] S3. Preparation of intermediate A14 (3α-benzoyloxy-5β-chole-9(11)-ene-12-one-24-oic acid methyl ester): same as Example 1.

[0143] S4. Preparation of intermediate A15 (3α-hydroxy-5β-chole-9(11)-ene-12-one-24-oic acid)

[0144] Take a 1L three-necked flask, add 500mL of acetone, add 100g of A14 raw material under stirring, dissolve under stirring, add 500mL of 2N sodium hydroxide aqueous solution, react at 45°C for 6h, and TLC detection shows that no raw material remains.

[0145] The reaction solution was cooled to 20°C, acetone was removed by distillation under reduced pressure, 2N dilute hydrochloric acid was added dropwise to adjust the pH to 2-3, a large amount of white solid precipitated, the temperature was raised to 75°C, stirred and hot slurried for 2-3h, filtered, filtered, the solid was slurried once with 75°C hot water, filtered, and washed with purified water to obtain a white solid, which was dried at 45°C for 16h to obtain 54.4g of white solid with a yield of 90.5%.

[0146] S5. Preparation of Intermediate A16 (3α-Hydroxy-5β-cholane-12-one-24-acid) Solution

[0147] Add 700 mL of ethyl acetate to the hydrogenation kettle, add 70 g of A15 raw material under stirring, add 35 g of Pd(OH)2 / C, replace with nitrogen and then replace with hydrogen three times, pressurize the hydrogen to 2.0 MPa, and react at 45°C for 24 hours. After cooling to room temperature, filter, rinse, and concentrate the solution to obtain a solid, then add 700 ml of methanol to dissolve it and directly proceed to the next step of reaction.

[0148] S6. Preparation of intermediate A17 (3α-hydroxy-5β-cholan-12-one-24-oic acid methyl ester): same as Example 1.

[0149] S7. Preparation of intermediate A18 (methyl deoxycholate): Same as Example 1

[0150] S8. Preparation of target DCA

[0151] The crude oil A18 from the previous step was transferred to a 1 L flask, 200 mL of acetone was added, 100 mL of 2N / L LiOH solution was slowly added, and the mixture was reacted at 45°C for 6 h. The reaction was terminated when no A18 was found by TLC.

[0152] Add 400 mL of purified water to the reaction solution, remove acetone by distillation under reduced pressure, and add 2N / L hydrochloric acid dropwise while stirring to adjust the pH to 1-2 (internal temperature controlled at 15-25°C). A large amount of solid precipitates. Keep stirring at 20°C for 4 h, filter, and wash with 800 mL of purified water until the filtrate is neutral to obtain a white solid. Dry with air at 45°C for 24 h to obtain 36.9 g of crude DCA (yield 91.3%).

[0153] Heat 30 g of crude DCA to 35-45°C with 400 ml of 4% methanol in dichloromethane solution, keep reflux and beat for 8 h, cool to 20-30°C and stir for 1-2 h, filter, wash with 150 mL of dichloromethane to obtain a white solid, dry with air at 45°C for 8 h to obtain 27.1 g of the product as a white solid with a yield of 90.3%.

[0154] Take a 1000mL three-necked flask, add 138mL of purified water, add 2.5g of sodium hydroxide while stirring, dissolve while stirring (control the internal temperature not higher than 20℃), add 20g of DCA solid in batches, and completely dissolve the solid (control the internal temperature not higher than 20℃), add 0.65g of activated carbon, heat to 40℃ and stir for 2h, filter, cool to 10℃, add about 35mL of 2N / L dilute hydrochloric acid dropwise, adjust the pH to 1-2 (control the internal temperature not higher than 20℃), stir at 20℃ for 2h, filter, rinse with 460mL of purified water until the filtrate is neutral, and obtain a white solid. Dry with air at 45℃ for 24h to obtain 20.9g of white solid, with a yield of 90.9% and a purity (CAD) of 99.6%. ( Figure 6 )

[0155] 1 H NMR (400MHz, DMSO-d6) δ11.92 (s, 1H), 4.45 (d, J = 4.2Hz, 1H), 4.20 (d, J = 4.0Hz, 1H),3.79(q,J=3.2Hz,1H),3.32(s,1H),2.22(ddd,J=15.1,9.6,5.2Hz,1H),2.0 9(ddd,J=15.6,9.1,6.8Hz,1H),1.85–1.43(m,10H),1.40–1.13(m,11H),1.02( ddd,J=25.3,12.9,4.5Hz,2H),0.91(d,J=6.4Hz,3H),0.84(s,4H),0.59(s,3H).

[0156] LCMS: 415.3 [M+Na] +

[0157] mp=172.1℃-173.6℃

[0158] The HNMR spectrum of deoxycholic acid is shown in Figure 3 As can be seen from the figure, the deoxycholic acid synthesized by the present invention is consistent with the data reported in the literature, and the purity detected by HPLC is as high as 99.6% (CAD detector) or more, and the quality is qualified.

[0159] The above is a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A method for synthesizing deoxycholic acid, characterized in that: The steps include: S1. Compound A11 reacts with methyl acrylate in the presence of a Lewis acid to produce compound A12; S2. In the presence of a first catalyst, compound A12 reacts with hydrogen to produce A13; S3. Compound A13 reacts with an oxidant to generate A14; S4. Compound A14 is hydrolyzed with a base to generate A15; S5. In the presence of a second catalyst, compound A15 is hydrogenated and reduced to generate A16; S6. In the presence of a third catalyst, compound A16 undergoes an esterification reaction with methanol to produce A17; S7. In the presence of a chiral reducing agent, compound A17 is chirally reduced to generate A18; S8. Compound A18 is hydrolyzed with alkali to generate DCA, i.e., deoxycholic acid; The A11 compound is (Z)-3α-benzoyloxy-5β-pregna-9(11),17(20)-diene; The synthesis method of the deoxycholic acid is as follows:

2. The synthesis method according to claim 1, characterized in that The Lewis acid is selected from at least one of AlCl3, EtAlCl2, and TiCl4.

3. The synthesis method according to claim 1, characterized in that The first catalyst in S2 is selected from at least one of Pd / C, Pd(OH)2 / C, and PtO2 / C.

4. The synthesis method according to claim 1, characterized in that The oxidant in S3 is selected from at least one of tert-butyl peroxide, hydrogen peroxide, peracetic acid, pyridinium chlorochromate, sodium hypochlorite, 2-iodoacetylbenzoic acid and Dess-Martin periodinant.

5. The synthesis method according to claim 1, characterized in that The alkali in S4 is selected from at least one of sodium hydroxide, potassium hydroxide and lithium hydroxide.

6. The synthesis method according to claim 1, characterized in that The second catalyst in S5 is selected from at least one of Pd / C, dry palladium carbon, Pd(OH)2 / C, PtO2 / C and Raney nickel.

7. The synthesis method according to claim 1, characterized in that The pressure of hydrogenation in S5 is 1.5MPa-3.0MPa, and the reaction temperature of hydrogenation is 20°C-80°C.

8. The synthesis method according to claim 1, characterized in that The third catalyst in S6 is selected from at least one of concentrated sulfuric acid, hydrochloric acid, thionyl chloride, and phosphorus trichloride.

9. The synthesis method according to claim 1, characterized in that The chiral reducing agent in S7 is selected from at least one of LiAl(Ot-Bu)3H, sodium triacetoxyborohydride, and diisopinoceneyl chloroborane.

10. The synthesis method according to claim 1, characterized in that The alkali in S8 is selected from at least one of sodium hydroxide, potassium hydroxide and lithium hydroxide.

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