Silybin compound and preparation method thereof

Abstract

The invention discloses a silibinin compound and a preparation method thereof. The method comprises the following steps: mixing pretreated quercetin and silibinin according to a formula ratio, uniformly stirring in a solution containing beta-cyclodextrin and vitamin C, slowly cooling under a control condition to promote the formation of eutecticum, and finally carrying out spray drying to obtain powder. According to the method, the solubility and the stability of the silibinin are remarkably improved, meanwhile, the biological activity of the silibinin is enhanced, and the method is suitable for clinical application and has wide application prospects. The control conditions comprise the temperature of 45-55 DEG C and the pH value of 6.5-7.5.

Description

Technical Field

[0001] The invention relates to the technical field of silibinin, and in particular to a silibinin complex and a preparation method thereof. Background Art

[0002] Silybin is a natural flavonoid compound derived from milk thistle (Silybum marianum), which is mainly used to treat health problems such as liver disease. With the deepening of modern pharmacological research, researchers have found that silybin has multiple biological activities such as antioxidant, anti-inflammatory, and anti-fibrosis, and its application prospects are increasingly valued in the medical field. However, the clinical application of silybin is limited by its poor water solubility and low bioavailability. In order to solve this problem, researchers have tried to improve its physicochemical properties and biological activity through various drug modification technologies.

[0003] In recent years, cocrystal technology has attracted extensive attention as an effective strategy to improve drug solubility and bioavailability. Cocrystals are solid materials formed by two or more molecules interacting with each other through non-covalent bonds in a certain ratio. Compared with traditional salt formation or complex compounds, cocrystals can significantly improve the solubility, stability and bioavailability of drugs without changing the molecular structure of drugs. Although cocrystal technology has shown great potential in improving drug performance, how to select suitable cocrystal formers and optimal cocrystal preparation conditions in the application of silybin remains a technical challenge.

[0004] Existing silibinin preparations mostly use simple solvent precipitation or physical mixing methods. Although these methods are simple to operate, they often cannot effectively solve the problems of poor solubility and low bioavailability. In addition, the existing technology also shows certain limitations in improving the stability of silibinin, such as being easily affected by light, heat and pH. In contrast, our invention uses cocrystal technology to screen suitable cocrystal formers, such as certain natural flavonoids or other bioactive molecules, and optimize the cocrystal preparation process, which can not only significantly improve the solubility and stability of silibinin, but also enhance its biological activity in the form of cocrystals, providing a new solution for clinical application. The development of this technology fills the gaps in the existing technology and opens up a new path for the development and application of silibinin and related drugs. Summary of the invention

[0005] Existing silibinin preparations mostly use simple solvent precipitation or physical mixing methods. Although these methods are simple to operate, they often cannot effectively solve the problems of poor solubility and low bioavailability. In addition, the existing technology also shows certain limitations in improving the stability of silibinin, such as being easily affected by light, heat and pH. In contrast, our invention uses cocrystal technology to screen suitable cocrystal formers, such as certain natural flavonoids or other bioactive molecules, and optimize the cocrystal preparation process, which can not only significantly improve the solubility and stability of silibinin, but also enhance its biological activity in the form of cocrystals, providing a new solution for clinical application. The development of this technology fills the gaps in the existing technology and opens up a new path for the development and application of silibinin and related drugs.

[0006] The present application provides a method for preparing a silybin complex, comprising the following steps:

[0007] S1. The pretreated quercetin and silybin were added to 90 vol% ethanol according to the formula ratio and mixed in a high-speed stirrer;

[0008] S2. Then add the mixed powder ethanol solution and vitamin C to the β-cyclodextrin aqueous solution and stir evenly to ensure complete dissolution, and slowly cool;

[0009] S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions;

[0010] S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 60-70°C.

[0011] By adopting the above technical scheme, first, under high-speed stirring, quercetin and silybin are fully mixed in ethanol to form a preliminary intermolecular interaction. Secondly, β-cyclodextrin includes silybin, vitamin C protects the active ingredient, and stirring ensures uniform dissolution. Then, a stable eutectic structure is formed by slow cooling, and hydrogen bonds and π-π interactions are used to enhance the stability of molecules. Next, ethanol is volatilized at low temperature by reduced pressure rotary evaporation to avoid high temperature damage and protect the structure and activity of quercetin and silybin. Finally, spray drying ensures that the solution is quickly converted into a uniform powder, thereby improving the solubility and bioavailability of the complex. The above technical scheme significantly improves the solubility and stability of the silybin complex through the formation of eutectic technology and inclusion complexes, enhances its biological activity, and is suitable for industrial production and clinical application.

[0012] As a preferred technical solution for the preparation of a silybin complex, the pre-treated quercetin is processed as follows:

[0013] Mix quercetin powder with an appropriate amount of ethanol, filter the solution using a microporous filter or fine-pore filter paper to remove undissolved solid impurities to obtain a clear quercetin solution, and slowly heat the solution in a constant temperature water bath at a temperature not exceeding 30°C to evaporate most of the solvent, transfer the almost dry quercetin to a drying oven, and continue to dry it at a temperature not exceeding 40°C until it is completely dry. After drying, the quercetin can be optionally subjected to ultraviolet irradiation for 5 to 15 minutes.

[0014] By adopting the above technical scheme, quercetin powder is mixed with an appropriate amount of ethanol, the purpose of which is to remove impurities and low-molecular-weight inactive components that may exist in the raw materials, which helps to improve the purity of quercetin and thus enhance its effect in the co-crystal. The above solution is filtered using a microporous filter or fine-pore filter paper to remove undissolved solid impurities to obtain a clear quercetin solution. Low-temperature evaporation helps to prevent structural damage or reduced activity of quercetin that may occur due to high temperature. The dried quercetin powder should have good fluidity and uniformity and be suitable for subsequent co-crystal preparation. The chemical activity of quercetin is enhanced by a slight photochemical reaction, thereby improving the strength and stability of its interaction when forming a co-crystal with silybin.

[0015] As a preferred technical solution for the preparation of a silybin complex, the controlled conditions are: temperature 45-55° C., pH 6.5-7.5.

[0016] By adopting the above technical scheme, the successful formation of the co-crystal is ensured by controlling the temperature within the range of 45-55°C and the pH within the range of 6.5-7.5.

[0017] As a preferred technical solution for the preparation of a silybin complex, the mass ratio of pretreated quercetin to silybin is (1 to 3):1.

[0018] By adopting the above technical scheme, the mass ratio between pretreated quercetin and silybin is optimized.

[0019] As a preferred technical solution for the preparation of a silybin complex, the mass ratio of β-cyclodextrin to silybin is (2 to 4):1.

[0020] By adopting the above technical solution, the mass ratio between β-cyclodextrin and silybin is optimized.

[0021] As a preferred technical solution for the preparation of a silybin complex, the mass ratio of vitamin C to silybin is (0.1 to 3):1.

[0022] By adopting the above technical solution, the mass ratio between vitamin C and silybin is optimized.

[0023] The silybin complex prepared by the preparation method of the present application forms a co-crystal structure with the quercetin, and the hydrophobic inner cavity of the beta-cyclodextrin encapsulates the silybin molecules.

[0024] The silybin complex and the preparation method thereof provided by the present invention have the following beneficial effects:

[0025] 1. Significantly improve solubility: Through co-crystal technology, silybin and quercetin form a co-crystal structure, and β-cyclodextrin includes silybin molecules, which significantly improves the solubility of silybin in water and helps to improve its bioavailability.

[0026] 2. Enhanced stability: Vitamin C in the complex acts as an antioxidant, effectively protecting silybin and quercetin from oxidative damage, maintaining their activity, and ensuring the stability of the product under different environmental conditions.

[0027] 3. Improve bioavailability: By optimizing the co-crystal preparation process and screening suitable co-crystal formers, the absorption rate of silybin in the body can be significantly improved, thereby enhancing its clinical efficacy.

[0028] 4. Easy to operate and suitable for industrial production: It adopts simple and easy process steps such as high-speed stirring and spray drying. It is easy to operate, suitable for large-scale industrial production, and has high economic value.

[0029] 5. Environmental protection and safety: The solvents used in the preparation process are ethanol and water, which are environmentally friendly and safe, avoiding the use of toxic and harmful solvents and complying with the principles of green chemistry. DETAILED DESCRIPTION

[0030] In order to make the above-mentioned objects, features and advantages of the present invention more obvious and easy to understand, the specific implementation methods of the present invention are described in detail below in conjunction with the embodiments of the specification.

[0031] In the following description, many specific details are set forth to facilitate a full understanding of the present invention, but the present invention may also be implemented in other ways different from those described herein, and those skilled in the art may make similar generalizations without violating the connotation of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0032] Secondly, the term "one embodiment" or "embodiment" as used herein refers to a specific feature, structure, or characteristic that may be included in at least one implementation of the present invention. The term "in one embodiment" that appears in different places in this specification does not necessarily refer to the same embodiment, nor does it refer to a separate or selective embodiment that is mutually exclusive with other embodiments.

[0033] Preparation Example 1

[0034] Mix quercetin powder with an appropriate amount of ethanol, filter the solution using a microporous filter or fine-pore filter paper to remove undissolved solid impurities to obtain a clear quercetin solution, and slowly heat the solution in a constant temperature water bath at a temperature not exceeding 30°C to evaporate most of the solvent, transfer the almost dry quercetin to a drying oven, and continue to dry it at a temperature not exceeding 40°C until it is completely dry. After drying, the quercetin can be subjected to ultraviolet irradiation for 10 minutes.

[0035] Example 1

[0036] S1. The pretreated quercetin and silybin were added to ethanol in a mass ratio of 1:1 and mixed in a high-speed stirrer;

[0037] S2. The mixed powder ethanol solution and vitamin C are added to the β-cyclodextrin aqueous solution and stirred to ensure complete dissolution; wherein the mass ratio between β-cyclodextrin and silybin is 2:1, and the mass ratio between vitamin C powder and silybin is 0.1:1;

[0038] S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions; wherein the controlled conditions are: temperature 45°C, pH 6.5;

[0039] S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 60°C.

[0040] Example 2

[0041] S1. The pretreated quercetin and silybin were added to ethanol in a mass ratio of 2:1 and mixed in a high-speed stirrer;

[0042] S2. The mixed powder ethanol solution and vitamin C are added to the β-cyclodextrin aqueous solution and stirred to ensure complete dissolution; wherein the mass ratio between β-cyclodextrin and silybin is 3:1, and the mass ratio between vitamin C powder and silybin is 0.5:1;

[0043] S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions; wherein the control conditions are: temperature 50°C, pH 7.0;

[0044] S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 65°C.

[0045] Example 3

[0046] S1. The pretreated quercetin and silybin were added to ethanol in a mass ratio of 3:1 and mixed in a high-speed stirrer;

[0047] S2. The mixed powder ethanol solution and vitamin C are added to the β-cyclodextrin aqueous solution and stirred to ensure complete dissolution; wherein the mass ratio between β-cyclodextrin and silybin is 4:1, and the mass ratio between vitamin C powder and silybin is 1.5:1;

[0048] S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions; wherein the controlled conditions are: temperature 55°C, pH 7.5;

[0049] S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 70°C.

[0050] Example 4

[0051] S1. The pretreated quercetin and silybin were added to ethanol in a mass ratio of 2:1 and mixed in a high-speed stirrer;

[0052] S2. The mixed powder ethanol solution and vitamin C are added to the β-cyclodextrin aqueous solution and stirred to ensure complete dissolution; wherein the mass ratio between β-cyclodextrin and silybin is 3:1, and the mass ratio between vitamin C powder and silybin is 2:1;

[0053] S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions; wherein the control conditions are: temperature 55°C, pH 7.0;

[0054] S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 70°C.

[0055] Comparative Example 1

[0056] The difference between this comparative example and Example 2 is that this comparative example uses an equal amount of untreated quercetin instead of pretreated quercetin.

[0057] Comparative Example 2

[0058] The difference between this comparative example and Example 2 is that an equal amount of vitamin C is used to replace β-cyclodextrin.

[0059] Comparative Example 3

[0060] The difference between this comparative example and Example 2 is that vitamin C is replaced by an equal amount of β-cyclodextrin.

[0061] Performance Testing

[0062] To verify the solubility, stability and bioavailability of the pre-treated quercetin and silybin complex, the following test methods can be used:

[0063] 1. Solubility test

[0064] method:

[0065] Sample preparation: Take a certain amount of silybin complex powder (about 50 mg) and add it to 10 ml of deionized water.

[0066] Dissolution process: Shake at 100 rpm in a constant temperature shaker (37°C) for 2 hours.

[0067] Determination method: The concentration of silybin in the solution was determined using high performance liquid chromatography (HPLC). Quantitative analysis can be performed using either the external standard method or the internal standard method.

[0068] Evaluation criteria:

[0069] The solubility of silybin (mg / ml) was calculated and compared with that of pure silybin.

[0070] 2. Stability test

[0071] method:

[0072] Thermal stability:

[0073] Sample preparation: A certain amount of silybin complex powder was placed in a drying oven at 60°C.

[0074] Time point sampling: samples were collected at 0 hours, 24 hours, 48 ​​hours and 72 hours.

[0075] Determination method: HPLC was used to determine the silybin content in the samples at each time point.

[0076] Evaluation criteria: Calculate the degradation rate of silybin and evaluate the thermal stability of the complex.

[0077] Light stability:

[0078] Sample preparation: A certain amount of silybin complex powder was exposed to ultraviolet light.

[0079] Time point sampling: samples were collected at 0 hours, 12 hours, 24 hours and 48 hours.

[0080] Determination method: HPLC was used to determine the silybin content in the samples at each time point.

[0081] Evaluation criteria: Calculate the degradation rate of silybin and evaluate the photostability of the complex.

[0082] 3. Bioavailability testing

[0083] Animal Experimentation:

[0084] Experimental design: Select appropriate animal models (such as rats), randomly divide them into 7 groups, and take the complexes of Examples 1 to 4 and Comparative Examples 1 to 3 respectively.

[0085] Administration: The dosage is calculated based on body weight and administered in equal amounts of silybin (mg / kg).

[0086] Blood sample collection: Blood samples were collected at different time points after administration (e.g., 0.5 hours, 1 hour, 2 hours, 4 hours, 6 hours, 8 hours).

[0087] Determination method:

[0088] Liquid chromatography-mass spectrometry (LC-MS / MS) was used to measure the concentration of silybin in plasma.

[0089] The blood drug concentration-time curve was plotted and pharmacokinetic parameters such as Cmax (maximum blood drug concentration), Tmax (time to reach maximum blood drug concentration) and AUC (area under the curve) were calculated.

[0090] Evaluation criteria:

[0091] The pharmacokinetic parameters were compared to evaluate the bioavailability-enhancing effect of the silybin complex.

[0092] Table 1

[0093]

[0094] It can be seen from Examples 1 to 4 and Table 1 that the silybin complex prepared by the scheme of the present application has the advantages of good solubility, high thermal stability, high photostability and good bioavailability.

[0095] Combining Example 2, Comparative Example 1 and Table 1, it can be seen that the solubility, thermal stability, photostability and bioavailability of Example 2 are better than those of Comparative Example 1, indicating that the pretreated quercetin significantly improves the solubility, thermal stability, photostability and bioavailability of the silybin complex.

[0096] Combining Example 2, Comparative Example 2, Comparative Example 3 and Table 1, it can be seen that the solubility, thermal stability, photostability and bioavailability of Example 2 are better than those of Comparative Example 2 and Comparative Example 3, indicating that vitamin C and β-cyclodextrin have synergistic effects in enhancing the solubility, thermal stability, photostability and bioavailability of the silybin complex.

[0097] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention is described in detail with reference to the aforementioned embodiments, those skilled in the art can still modify the technical solutions described in the aforementioned embodiments or replace some of the technical features therein by equivalents. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for preparing a silybin complex, It is characterized in that Includes the following S1. The pretreated quercetin and silybin were added to 90 vol% ethanol according to the formula ratio and mixed in a high-speed stirrer; S2. Then add the mixed powder ethanol solution and vitamin C to the β-cyclodextrin aqueous solution and stir evenly to ensure complete dissolution; S3. The solution in step S2 is evaporated by a rotary evaporator under reduced pressure to evaporate ethanol, and slowly cooled under controlled temperature conditions; S4. Dry the solution into powder by spray drying, and the spray drying temperature is controlled at 60-70°C.

2. The method for preparing the silybin complex according to claim 1, It is characterized in that The processing steps of pre-treated quercetin are: Mix quercetin powder with an appropriate amount of ethanol, filter the solution using a microporous filter or fine-pore filter paper to remove undissolved solid impurities to obtain a clear quercetin solution, and slowly heat the solution in a constant temperature water bath at a temperature not exceeding 30°C to evaporate most of the solvent, transfer the almost dry quercetin to a drying oven, and continue to dry it at a temperature not exceeding 40°C until it is completely dry. After drying, the quercetin can be optionally subjected to ultraviolet irradiation for 5 to 15 minutes.

3. The method for preparing the silybin complex according to claim 2, It is characterized in that The controlled conditions are: temperature 45-55°C, pH 6.5-7.

5.

4. The method for preparing the silybin complex according to claim 1, It is characterized in that The mass ratio between pretreated quercetin and silybin was (1 to 3):

1.

5. The method for preparing the silybin complex according to claim 1, It is characterized in that The mass ratio between β-cyclodextrin and silybin is (2 to 4):

1.

6. The method for preparing the silybin complex according to claim 1, It is characterized in that The mass ratio between vitamin C and silybin is (0.1 to 3):

1.

7. A silybin complex prepared by the preparation method according to any one of claims 1 to 6, It is characterized in that The silybin forms a co-crystal structure with the quercetin, and the hydrophobic inner cavity of the β-cyclodextrin encapsulates the silybin molecule.

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