Flavonoid eutectic compound as well as preparation method and application thereof

Abstract

The invention discloses a flavonoid eutectic compound as well as a preparation method and application thereof, and relates to the technical field of medicinal chemistry, and the flavonoid eutectic compound is a baicalin-amine eutectic. The preparation method of the flavonoid eutectic compound comprises the following steps: carrying out coordination reaction on baicalin and amine in a solvent, concentrating a reaction solution after the reaction is finished, cooling, crystallizing, filtering and drying to obtain a baicalin-amine eutectic substance. According to the present invention, the baicalin and the amine are adopted as the starting raw materials, the baicalin-amine eutectic is obtained through the coordination reaction of the baicalin and the amine, the solubility of the baicalin-amine eutectic in water is far higher than the solubility of the baicalin, and the stability of the product under conditions of high temperature, high humidity and illumination can be significantly improved.

Description

Technical Field

[0001] The present invention relates to the technical field of medicinal chemistry, and in particular to a flavonoid eutectic compound and a preparation method and application thereof. Background Art

[0002] Scutellaria baicalensis, the dried root of Scutellaria baicalensis Georgi, belongs to the dicotyledonous Lamiaceae family. It is a traditional Chinese medicine first documented in Shennong's Herbal Classic. It is commonly used to clear heat and dampness, purge fire and detoxify, and stop bleeding and stabilize pregnancy. Baicalin (BA), one of the main active ingredients in Scutellaria baicalensis, belongs to the flavonoid class and is chemically named 5,6-dihydroxy-4-oxo-2-phenyl-4H-1-benzopyran-7-yl-β-D-pyranoglucuronic acid. Its structural formula is as follows:

[0003]

[0004] Baicalin has multiple effects, including antioxidant, anti-tumor, anti-inflammatory, antibacterial, and antiviral, and is widely used clinically. Studies have found that when the pH value of baicalin is 7.4-8.0, the baicalin aglycone nucleus rapidly opens in alkaline solution to form chalcone. The mechanism is as follows:

[0005]

[0006] Therefore, baicalin is extremely unstable in alkaline solutions and its stability decreases at pH values ​​above 3, complicating pharmaceutical production and storage. Baicalin exhibits poor stability in biological samples, particularly in liver homogenates, likely due to the presence of numerous and diverse hepatic drug-producing enzymes in liver tissue. Therefore, the impact of enzymes should be considered during in vivo analysis. Furthermore, baicalin's poor water solubility results in low oral bioavailability, which in turn affects its efficacy and limits its application in pharmaceutical formulations. Therefore, the development of new baicalin APIs with improved solubility, stability, and bioavailability is of great significance. Summary of the Invention

[0007] The technical problem to be solved by the present invention is to provide a method for preparing a flavonoid eutectic compound, in which baicalin and amine are subjected to a coordination reaction to obtain a baicalin-amine eutectic, thereby improving the stability of baicalin while simplifying the synthesis process and improving the environmental friendliness of the process.

[0008] The technical problem to be solved by the present invention is achieved by adopting the following technical solutions:

[0009] One of the purposes of the present invention is to provide a flavonoid cocrystal compound, which is a baicalin-amine cocrystal.

[0010] Furthermore, the amine is at least one of tromethamine, choline, and an amino acid, preferably tromethamine. Tromethamine molecules contain amino and hydroxyl groups, which can form a multiple hydrogen bond network with the phenolic hydroxyl and carboxyl groups in the baicalin molecule, stabilizing the eutectic structure, thereby increasing the solubility and dissolution rate of baicalin, enhancing bioavailability, improving drug stability, and maintaining or enhancing pharmacological activity.

[0011] The second object of the present invention is to provide a method for preparing the flavonoid cocrystal compound, comprising the steps of: coordinating baicalin with an amine in a solvent; concentrating the reaction solution after the reaction is completed, cooling the solution for crystallization, filtering the solution, and drying the solution to obtain a baicalin-amine cocrystal.

[0012] Furthermore, the molar ratio of baicalin to amine is 1:(1-1.5), preferably 1:(1-1.2).

[0013] Furthermore, the solvent is at least one of water and ethanol, preferably water.

[0014] Furthermore, the reaction temperature of the coordination reaction is 20-80°C, preferably 40-60°C.

[0015] The third object of the present invention is to provide the use of the flavonoid cocrystal compound in the preparation of antioxidants, anti-tumor drugs, anti-inflammatory drugs, antibacterial drugs, and antiviral drugs.

[0016] A fourth object of the present invention is to provide a pharmaceutical composition comprising a pharmaceutically effective amount of the flavonoid co-crystal compound.

[0017] The beneficial effects of the present invention are as follows: baicalin and amine are used as starting raw materials, and baicalin-amine eutectic is obtained through the coordination reaction of baicalin and amine; the synthetic method has readily available raw materials, is simple to operate, and is green and environmentally friendly; the solubility of the prepared baicalin-amine eutectic in water is much higher than that of baicalin, and the stability of the product under high temperature, high humidity and light conditions can be significantly improved. BRIEF DESCRIPTION OF THE DRAWINGS

[0018] Figure 1 This is the hydrogen spectrum of baicalin-tromethamine cocrystal;

[0019] Figure 2 This is the carbon spectrum of baicalin-tromethamine cocrystal;

[0020] Figure 3 is the hydrogen spectrum of baicalin;

[0021] Figure 4 is the SDT diagram of baicalin-tromethamine cocrystal;

[0022] Figure 5XRD patterns of baicalin, tromethamine, baicalin-tromethamine cocrystal, and physical mixture of baicalin and tromethamine;

[0023] Figure 6 This is the FT-IR spectrum of baicalin-tromethamine cocrystal;

[0024] Figure 7 is the FT-IR spectrum of baicalin;

[0025] Figure 8 is the FT-IR spectrum of tromethamine;

[0026] Figure 9 FT-IR spectrum of the physical mixture of baicalin and tromethamine;

[0027] Figure 10 This is the hydrogen spectrum of baicalin-arginine cocrystal. DETAILED DESCRIPTION

[0028] In order to make the technical means, creative features, objectives and effects achieved by the present invention easier to understand, the present invention is further described below with reference to specific embodiments and illustrations.

[0029] Determination of baicalin content in baicalin-amine eutectic:

[0030] Test solution: Accurately weigh an appropriate amount of baicalin-amine eutectic (equivalent to 10 mg of baicalin) and place it in a 20 mL volumetric flask, add 10 mL of methanol for ultrasonic dissolution, then add 5 mL of water to dissolve, and dilute with methanol to the scale to obtain the test sample mother solution; Accurately measure 2 mL of the test sample mother solution and place it in a 25 mL volumetric flask, dilute with methanol to the scale to obtain the test sample solution.

[0031] Reference substance solution: Accurately weigh 10 mg of baicalin reference substance and place it in a 20 mL volumetric flask, add 10 mL of methanol and ultrasonically dissolve it, then add 5 mL of water to dissolve it and dilute it to the scale with methanol to obtain the reference substance mother solution; accurately measure 2 mL of the reference substance mother solution and place it in a 25 mL volumetric flask, dilute it to the scale with methanol to obtain the reference substance solution.

[0032] Chromatographic conditions: octadecylsilane bonded silica gel as the filler (Amethyst C18-H, 4.6 mm × 250 mm, 5 μm); 0.1% phosphoric acid solution as mobile phase A, acetonitrile as mobile phase B, gradient elution according to Table 1; column temperature, 30°C; flow rate, 1 mL / min; detection wavelength, 278 nm; injection volume, 10 μL.

[0033] Table 1

[0034]

[0035] Example 1

[0036] Baicalin (13.38 g, 30 mmol), tromethamine (3.63 g, 30 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 8 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 88.81%, and the baicalin content was 78.1% (theoretical value 78.6%).

[0037] Baicalin-tromethamine cocrystal: 1 H NMR(400MHz,DMSO-d6)δ8.12-8.03(m,2H),7.65-7.55(m,3H),7.05(s,1H),7.00(s,1H),5.10(brs,1H),5 .04(d,J=7.2Hz,1H),3.68(d,J=9.7Hz,1H),3.45(s,6H),3.35(dt,J=18.3,8.9Hz,3H),3.31-3.23(m,2H). 13 C NMR (101 MHz, DMSO-d6) δ 183.04, 172.76, 163.93, 152.25, 147.08, 132.48, 131.29, 131.28, 129.65, 126.85, 105.16, 101.23, 94.79, 76.30, 74.68, 73.42, 72.50, 61.05, 60.18 *3. See for details Figure 1 and Figure 2 .

[0038] Baicalin: 1 H NMR (400 MHz, DMSO-d6) δ 13.86 (brs, 1H), 12.61 (s, 1H), 8.71 (s, 1H), 8.13-8.06 (m, 2H), 7.69-7.56 (m, 3H), 7.06 (s, 1H), 7.03 (s, 1H), 5.53 (d, J = 4.3 Hz, 1H), 5.47 (brs, 1H), 5.34 (brs, 1H), 5.26 (d, J = 7.4 Hz, 1H), 4.08 (d, J = 9.5 Hz, 1H), 3.55-3.37 (m, 5H). See for details. Figure 3 .

[0039] There are obvious active hydrogen signal peaks at 12.61 and 8.71 ppm in the hydrogen spectrum of baicalin. These active hydrogen signals basically disappear in the hydrogen spectrum of baicalin-tromethamine cocrystal. The six hydrogen atoms at 3.45 ppm come from tromethamine. It is calculated that the molar ratio of baicalin to tromethamine in the cocrystal molecule is 1:1.

[0040] From the SDT diagram ( Figure 4 ) It can be seen that the melting point of baicalin-tromethamine eutectic is 195-200°C, and it loses 5.442% of its weight within 124°C, indicating that the molecule contains water.

[0041] Depend on Figure 5 It can be seen that the XRD pattern of baicalin shows characteristic peaks at 2θ angles of 4.461°, 4.940°, 5.579°, 6.901°, 8.141°, 9.300°, 9.882°, 11.179°, 11.720°, 12.595°, 13.359°, 14.859°, 16.221°, 17.282°, 17.681°, 18.420°, 19.799°, 20.159°, 20.680°, 21.180°, 23.322°, 23.859°, 24.822°, 26.580°, and 28.719°. The XRD pattern of tromethamine showed characteristic peaks at 2θ angles of 10.840°, 14.299°, 15.379°, 18.279°, 20.262°, 21.681°, 22.640°, 25.342°, 25.980°, 30.600°, 32.141°, 34.120°, and 38.582°. However, the XRD pattern of baicalin-tromethamine cocrystal showed no characteristic peaks, which preliminarily indicated the formation of a new cocrystal.

[0042] FT-IR images ( Figures 6 to 9 ) analysis revealed that the infrared characteristic peaks of the baicalin-tromethamine cocrystal molecule had changed significantly compared with baicalin. Specifically, some characteristic peaks of baicalin showed shifts, weakened intensities, and morphological changes. As shown in Table 2, the characteristic peak of the -OH band in baicalin is at 3289 cm -1 (ν O-H ), while in the eutectic molecules these characteristic peaks move to the low frequency direction to 3372 cm -1 (ν O-H The reason for this change is that after the cocrystal is formed, the -NH in tromethamine forms a strong NH…O hydrogen bond with the -OH in baicalin, which leads to a decrease in the vibration frequency of -OH and -NH and a broadening of the spectral band, thereby causing the characteristic peak of baicalin to undergo varying degrees of red shift.

[0043] Table 2

[0044]

[0045] Example 2

[0046] Baicalin (13.38 g, 30 mmol), choline (3.63 g, 30 mmol), and 200 mL of water were heated to 60°C and stirred for 2 h. After the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, stirred for 3 h, and filtered. The filter cake was dried at 55°C to obtain a baicalin-choline cocrystal. The yield was 75.48%, and the baicalin content was 57.2% (theoretical value 78.6%). HPLC analysis showed that the baicalin content was far lower than the theoretical value, possibly due to the strong alkalinity of choline, which destroyed the baicalin.

[0047] Example 3

[0048] Baicalin (13.38 g, 30 mmol), γ-aminobutyric acid (3.09 g, 30 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to crystallize. The mixture was filtered, and the filter cake was dried at 55°C to obtain baicalin-γ-aminobutyric acid cocrystal. The yield was 75.73%, and the baicalin content was 90.5% (theoretical value 81.2%), indicating that the molar ratio of baicalin to γ-aminobutyric acid is not fixed.

[0049] Example 4

[0050] Baicalin (13.38 g, 30 mmol), arginine (5.23 g, 30 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After the reaction, the reaction mixture was cooled naturally, stirred for 3 hours, and then filtered. The filter cake was dried at 55°C to obtain a baicalin-arginine cocrystal. The yield was 87.10%, and the baicalin content was 70.5% (theoretical value 71.9%), indicating a 1:1 molar ratio of baicalin to arginine.

[0051] 1 H NMR (400 MHz, DMSO-d6) δ 8.93 (brs, 1H), 8.06 (dd, J = 7.9, 1.9 Hz, 2H), 7.83 (brs, 4H), 7.62-7.52 (m, 3H), 7.03 (s, 1H), 6.98 (s, 1H), 5.06 (d, J = 7.2 Hz, 1H), 3.71 (d, J = 9.5 Hz, 1H), 3.43-3.24 (m, 5H), 3.06 (h, J = 7.1 Hz, 2H), 1.87-1.46 (m, 4H) ppm. Figure 10 .

[0052] Example 5

[0053] Baicalin (13.38 g, 30 mmol), lysine (4.38 g, 30 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After the reaction, the reaction mixture was cooled naturally, stirred for 3 hours, and then filtered. The filter cake was dried at 55°C to obtain a baicalin-lysine cocrystal. The yield was 86.20%, and the baicalin content was 80.2% (theoretical value 75.34%), indicating that the molar ratio of baicalin to lysine is not fixed.

[0054] Example 6

[0055] Baicalin (13.38 g, 30 mmol), tromethamine (3.99 g, 33 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 91.92%, and the baicalin content was 78.0%.

[0056] Example 7

[0057] Baicalin (13.38 g, 30 mmol), tromethamine (4.36 g, 36 mmol), and 200 mL of water were heated to 60°C and stirred for 2 hours. After the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 92.04%, and the baicalin content was 77.4%.

[0058] Example 8

[0059] Baicalin (13.38 g, 30 mmol), tromethamine (3.63 g, 30 mmol), and 200 mL of water were heated to 20°C and stirred for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 88.22%, and the baicalin content was 78.2%.

[0060] Example 9

[0061] Baicalin (13.38 g, 30 mmol), tromethamine (3.63 g, 30 mmol), and 200 mL of water were heated to 80°C and stirred for 2 hours. After completion of the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 88.16%, and the baicalin content was 77.7%.

[0062] Example 10

[0063] Baicalin (13.38 g, 30 mmol), tromethamine (3.63 g, 30 mmol), and 200 mL of 20% v / v ethanol were heated to 60°C and stirred for 2 hours. After the reaction, the reaction solution was concentrated under reduced pressure to 40 mL, cooled naturally, and stirred for 3 hours to allow crystallization. The mixture was filtered and the filter cake was dried at 55°C to obtain a baicalin-tromethamine cocrystal. The yield was 88.75%, and the baicalin content was 77.8%.

[0064] Example 11

[0065] Solubility test:

[0066] Baicalin passed through a 100-mesh sieve, the baicalin-tromethamine cocrystal prepared in Example 1, and the baicalin-arginine cocrystal prepared in Example 4 were added to a certain volume of water, and sonicated at room temperature for 2-5 min. When no solute particles or droplets were visible, the solution was considered to be completely dissolved.

[0067] Test results showed that the solubility of baicalin-tromethamine cocrystals was greater than 800 mg / mL, while that of baicalin-arginine cocrystals was approximately 100 mg / mL. The solubility of baicalin was 0.160 mg / mL, making the solubility of baicalin-tromethamine cocrystals in water over 5,000 times that of baicalin. Baicalin is classified as Class IV by the BCS, indicating low solubility and low permeability. The solubility of baicalin-tromethamine cocrystals was significantly improved, facilitating absorption and thus enhancing bioavailability.

[0068] Example 12

[0069] Moisture absorption test:

[0070] 1. Take a dry stoppered glass weighing bottle (outer diameter 50mm, height 15mm) and place it in a constant temperature desiccator at 25±1℃ (place ammonium chloride or saturated ammonium sulfate solution at the bottom) one day before the test. Accurately weigh the total weight m1.

[0071] 2. Take 1g of the test sample and spread it evenly in the above weighing bottle, and accurately weigh the total weight m2.

[0072] (1) Experimental group: baicalin-tromethamine cocrystal prepared in Example 1 and baicalin-arginine cocrystal prepared in Example 4;

[0073] (2) Control group: baicalin.

[0074] 3. Open the weighing bottle and place it together with the bottle cap in a constant temperature dryer for 24 hours.

[0075] 4. Cover the weighing bottle and accurately weigh the total weight m3.

[0076] Weight gain percentage = (m3-m2) / (m2-m1)×100%

[0077] 5. The results of the moisture absorption test are shown in Table 3.

[0078] Table 3

[0079] Inspection conditions Baicalin Baicalin-tromethamine cocrystal Baicalin-arginine cocrystal 0h 1.00g 1.00g 1.00g 24h 1.01g 1.01g 1.06g

[0080] As shown in Table 4, the hygroscopicity of the baicalin-tromethamine cocrystal prepared according to the present invention remained unchanged under high humidity conditions, while that of the baicalin-arginine cocrystal significantly increased by 6%. Therefore, the low hygroscopicity of the baicalin-tromethamine cocrystal according to the present invention indicates its high stability.

[0081] Example 13

[0082] Stability test:

[0083] 1. Test sample

[0084] (1) Experimental group: baicalin-tromethamine cocrystal prepared in Example 1 and baicalin-arginine cocrystal prepared in Example 4;

[0085] (2) Control group: baicalin.

[0086] 2. Stability inspection conditions

[0087] (1) Thermal degradation: Take 1.0 g of the test sample and place it in a drying oven at 60°C;

[0088] (2) Photodegradation: Take 1.0 g of the test sample and place it in an environment with an illumination of 4500 ± 500 lx;

[0089] (3) High humidity degradation: Take 1.0 g of the test sample and place it in a desiccator containing saturated potassium nitrate solution.

[0090] 3. The stability test results are shown in Tables 4 and 5.

[0091] Table 4

[0092]

[0093] Table 5

[0094]

[0095] As shown in Tables 4 and 5, compared with baicalin, the baicalin-tromethamine cocrystal and baicalin-arginine cocrystal prepared by the present invention have less changes in the baicalin content and the content of related substances under high temperature, high humidity and light conditions, and their properties do not change significantly, indicating that their quality is stable and controllable.

[0096] The basic principles, main features, and advantages of the present invention are shown and described above. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention. Such changes and modifications are intended to fall within the scope of the present invention. The scope of protection claimed in the present invention is defined by the appended claims and their equivalents.

Claims

1. A flavonoid eutectic compound, characterized in that: The flavonoid cocrystal compound is baicalin-amine cocrystal.

2. The flavonoid eutectic compound according to claim 1, characterized in that: The amine is at least one of tromethamine, choline, and amino acid, preferably tromethamine.

3. The method for preparing the flavonoid eutectic compound according to claim 1 or 2, characterized in that: The baicalin and amine are coordinated in a solvent. After the reaction is completed, the reaction solution is concentrated, cooled for crystallization, filtered, and dried to obtain a baicalin-amine eutectic.

4. The preparation method according to claim 3, wherein: The molar ratio of baicalin to amine is 1:(1-1.5), preferably 1:(1-1.2).

5. The preparation method according to claim 3, wherein: The solvent is at least one of water and ethanol, preferably water.

6. The preparation method according to claim 3, wherein: The reaction temperature of the coordination reaction is 20-80°C, preferably 40-60°C.

7. Use of the flavonoid eutectic compound according to any one of claims 1 to 2 or the flavonoid eutectic compound obtained by the preparation method according to any one of claims 3 to 6 in the preparation of antioxidants, anti-tumor drugs, anti-inflammatory drugs, antibacterial drugs, and antiviral drugs.

8. A pharmaceutical composition comprising a pharmaceutically effective amount of the flavonoid co-crystal compound according to any one of claims 1 to 2 or the flavonoid co-crystal compound obtained by the preparation method according to any one of claims 3 to 6.

Images