Extraction method and preparation method of low-residue silymarin

Abstract

The invention discloses an extraction method and a preparation method of low-residue silymarin, and the extraction method of the low-residue silymarin comprises the following steps: elution of grease: crushing silymarin seeds, soaking the crushed silymarin seeds with n-hexane, and eluting the grease to obtain a soaking solution; ethanol replacement: adding ethanol into the soak solution, standing, filtering, concentrating the filtrate, adding water, standing, and taking the lower solution and precipitate; and crystallizing and drying: concentrating the lower-layer solution and the precipitate, recrystallizing, filtering, and drying and crystallizing to obtain the silymarin. The extraction method of the low-residue silymarin, provided by the invention, has the effect of reducing solvent residues in the silymarin.

Description

Technical Field

[0001] The present invention belongs to the technical field of plant extract extraction, and specifically relates to a method for extracting low-residue silymarin and its preparation method. Background Art

[0002] Silymarin is a mixture of flavonolignan compounds extracted from the dried seeds and fruits of the Compositae plant Silybum marianum, and its main components include silybin, isosilybin, silydianin, etc. Silymarin is a light yellow powder, odorless, slightly bitter in taste, and hygroscopic. It is insoluble in water and chloroform, soluble in alkaline aqueous solutions, soluble in methanol and ethanol, and soluble in organic solvents such as acetone and ethyl acetate. Silymarin is considered a safe natural compound and has the effects of protecting the liver, antioxidation, anti-inflammation, etc., and is mainly used for treating hepatobiliary diseases such as acute and chronic hepatitis, fatty liver, liver cirrhosis, alcoholic liver injury, metabolic toxic liver injury, and gallstones.

[0003] There are various methods for extracting silymarin, and common methods include organic solvent extraction method, supercritical fluid extraction method, enzymatic extraction, ultrasonic-assisted extraction, microwave-assisted extraction, etc.

[0004] The organic solvent extraction method is the most traditional extraction method, and usually uses organic solvents such as ethanol, acetone, n-hexane, and ethyl acetate. The specific steps are as follows: ① After crushing the silymarin seeds, mix them with the organic solvent.

[0005] ② Extract silymarin by heating under reflux or maceration at room temperature.

[0006] ③ The extract is concentrated, filtered and other steps to remove impurities to obtain a crude extract of silymarin.

[0007] ④ Finally, further purification is carried out by methods such as crystallization or column chromatography to improve the purity of silymarin.

[0008] The advantages of the organic solvent extraction method for silymarin are its high efficiency, simple operation, low cost, short extraction time and high extraction rate. These advantages give it significant economic advantages in industrial production, especially in scenarios with high requirements for extraction efficiency and cost control. However, in the process of extracting silymarin with organic solvents, it is difficult to completely remove the organic solvents, and solvent residue is an important quality control issue. Especially in the fields of pharmaceuticals, health products, and food additives, solvent residue may have a significant impact on the safety, effectiveness, and quality of products. Summary of the Invention

[0009] Aiming at the deficiencies of the prior art, the purpose of the present invention is to provide a method for extracting low-residue silymarin and its preparation method.

[0010] To achieve the foregoing invention object, the technical solution adopted by the present invention includes: An extraction method of silymarin with low residue, comprising the following steps: Eluting grease: After crushing the milk thistle seeds, soak them with n-hexane and elute the grease to obtain a soaking solution; Ethanol replacement: Add ethanol to the soaking solution, let it stand and then filter. Concentrate the filtrate, add water and let it stand, and take the lower layer solution and precipitate; Crystallizing and drying: Concentrate the lower layer solution and precipitate, then crystallize and filter, and then dry the crystals to obtain silymarin.

[0011] In addition to silymarin, the milk thistle seeds also contain a large amount of lipids, fatty acids and other fat-soluble components. In the present invention, n-hexane, as a non-polar solvent, can effectively dissolve the fat-soluble components in the milk thistle seeds, while silymarin itself is a polar substance and is insoluble in n-hexane.

[0012] In the present invention, ethanol can dissolve silymarin, and ethanol has good compatibility with n-hexane, and silymarin can be dissolved in the mixed solution of ethanol and n-hexane.

[0013] In the present invention, during the ethanol replacement process, after concentrating the filtrate and adding water and letting it stand, cyclohexane is immiscible with water and floats on the water surface. Ethanol and water can be mixed in any ratio, and a large amount of ethanol dissolves in water, reducing the ethanol content in cyclohexane and facilitating the separation of silymarin from the cyclohexane layer into the water-ethanol mixed solution. After part of the silymarin separates from cyclohexane, it cannot be completely dissolved in the water-ethanol mixed solution and crystallizes to form a precipitate.

[0014] In the present invention, in the crystallizing and drying step, the lower layer solution and precipitate cannot be evaporated and crystallized. During the evaporation and crystallization process, the impurities dissolved in water will be mixed into the obtained silymarin, affecting the purity of the obtained silymarin.

[0015] Preferably, in the step of eluting grease, after crushing the milk thistle seeds, soak them with n-hexane for 6 - 10 h, and the dosage of n-hexane is 4 - 6 times the mass of the milk thistle seeds; after the soaking is completed, remove the upper layer liquid and then replenish n-hexane with the same volume as the removed upper layer liquid, soak again for 2 - 3 h, repeat the operation until the upper layer liquid is colorless and transparent after the soaking is completed, and then remove the upper layer liquid to obtain the soaking solution.

[0016] In the present invention, the process of eluting grease can effectively remove the fat-soluble components in the milk thistle seeds and retain silymarin, which is beneficial to reducing the difficulty of subsequent extraction of silymarin.

[0017] Preferably, the mass of the soaking solution is 1.5 - 2 times the mass of the milk thistle seeds.

[0018] Preferably, in the ethanol replacement step, the amount of ethanol used is 0.5 to 1 times the mass of the soaking solution.

[0019] Preferably, in the ethanol replacement step, the volume of the filtrate after concentration is 0.1 to 0.25 times that before concentration.

[0020] The boiling point of ethanol is 78.4 °C, and the boiling point of n - hexane is 69 °C. In the present invention, during the concentration of the filtrate, since the boiling point of n - hexane is lower than that of ethanol, the evaporation rate of n - hexane is greater than that of ethanol, so that the mass ratio of ethanol to n - hexane increases after the filtrate is concentrated, thereby achieving a large amount of removal of n - hexane. This is conducive to reducing the volume of the n - hexane - ethanol phase in the subsequent process, and thus conducive to enabling silymarin to enter the water - ethanol phase as much as possible from the n - hexane - ethanol phase, and further conducive to reducing the difficulty of extracting silymarin.

[0021] Preferably, in the ethanol replacement step, the filtrate is evaporated and concentrated at 40 °C to 60 °C.

[0022] Preferably, in the ethanol replacement step, the mass of water is 8 to 12 times the mass of the filtrate after concentration.

[0023] In the present invention, when the amount of water used is increased, it is conducive to increasing the amount of ethanol dissolved in water. At the same time, the concentration of ethanol in the ethanol - water phase decreases, and the ability of the ethanol - water phase to dissolve silymarin decreases, which is conducive to reducing the difficulty of extracting silymarin.

[0024] Preferably, in the crystallization and drying step, the lower - layer solution and the precipitate are evaporated and concentrated at 80 °C to 85 °C.

[0025] In the present invention, when the lower - layer solution and the precipitate are evaporated and concentrated at 80 °C to 85 °C, the ethanol contained therein can be fully evaporated, which is thus conducive to reducing the solvent residue in the obtained silymarin.

[0026] Compared with the prior art, the advantages of the present invention include: (1) The extraction method of silymarin with low residue provided by the present invention is simple to operate, without requiring harsh operating conditions or the participation of strong acids or strong bases; (2) The extraction method of silymarin with low residue provided by the present invention, in the ethanol replacement step, silymarin is removed from n - hexane and transferred to the water phase, converting the extraction solvent of silymarin from n - hexane to ethanol and water, which is thus conducive to reducing the n - hexane residue in the obtained silymarin; (3) The extraction method of silymarin with low residue provided by the present invention, during the crystallization and drying process, ethanol is volatile, which is thus conducive to reducing the solvent residue in the obtained silymarin. Specific embodiments

[0027] To enable those skilled in the art to understand the features and effects of this application, the following provides a general description and definition of the terms and phrases mentioned in the specification and claims. Unless otherwise specified, all technical and scientific terms used herein shall have the ordinary meaning understood by those skilled in the art with respect to this application. In case of conflict, the definition in this specification shall prevail.

[0028] The theories or mechanisms described and disclosed herein, whether right or wrong, shall not in any way limit the scope of this application, that is, the content of this application can be implemented without being limited by any specific theory or mechanism.

[0029] In this document, "this application" is the same as "this invention" and "this disclosure".

[0030] In this document, the terms "a", "an", "one", or similar expressions are used to describe the components and technical features of this application. Such descriptions are merely for convenience of expression and give a general meaning to the scope of this application. Therefore, such descriptions should be understood to include one or at least one, and the singular also includes the plural, unless clearly referring to something else.

[0031] In this document, "or a combination thereof" means "or any combination thereof", and "any one", "any kind", "any one" means "any one", "any kind", "any one".

[0032] In this document, the terms "comprising", "including", "having", "containing", or any other similar terms are all open-ended transitional phrases, which are intended to cover non-exclusive inclusions. For example, a composition or its product containing multiple elements is not limited to only the elements listed herein, but may also include other elements that are usually inherent in the composition or its product but not explicitly listed. In addition, unless there is a clear contrary statement, the term "or" refers to the inclusive "or", rather than the exclusive "or". For example, any of the following situations satisfies the condition "A or B": A is true (or exists) and B is false (or does not exist), A is false (or does not exist) and B is true (or exists), A and B are both true (or exist). In addition, in this document, the interpretations of the terms "comprising", "including", "having", "containing" should be regarded as having specifically disclosed and simultaneously covering closed transitional words such as "consisting of", "composed of", "the balance being", etc., and connecting words such as "substantially consisting of", "mainly consisting of", "mainly composed of", "basically containing", "basically consisting of", "basically composed of", "essentially containing".

[0033] In this text, all features or conditions defined in the form of numerical ranges or percentage ranges, such as numerical values, quantities, contents, and concentrations, are only for the sake of brevity and convenience. Accordingly, the description of a numerical range or percentage range should be regarded as having covered and specifically disclosed all possible sub-ranges and individual numerical values within the range (including integers and fractions), especially integer values. For example, a range description such as "1.0 to 8.0" or "between 1.0 and 8.0" or "between 1.0 and 8.0" should be regarded as having specifically disclosed all sub-ranges such as 1.0 to 8.0, 1.0 to 7.0, 2.0 to 8.0, 2.0 to 6.0, 3.0 to 6.0, 4.0 to 8.0, 3.0 to 8.0, etc., and should be regarded as covering the endpoint values, especially the sub-ranges defined by integer values, and should be regarded as having specifically disclosed individual numerical values within the range such as 1.0, 2.0, 3.0, 4.0, 5.0, 6.0, 7.0, 8.0, etc. Unless otherwise specified, the foregoing interpretation method applies to all contents of this application throughout the text, regardless of the scope.

[0034] If a quantity, concentration, or other numerical value or parameter is expressed as a range, a preferred range (or a better range), or a series of upper and lower limits, it should be understood that all ranges formed by any pair of the upper limit or preferred value (or better value) and the lower limit or preferred value (or better value) of the range have been specifically disclosed herein, regardless of whether these ranges are separately disclosed. In addition, when a numerical range is mentioned in this text, unless otherwise stated, the range should include its endpoints and all integers and fractions within the range.

[0035] In this text, on the premise that the purpose of the invention can be achieved, a numerical value should be understood to have the precision of the significant digits of that numerical value. For example, the number 40.0 should be understood to cover the range from 39.50 to 40.49.

[0036] Unless otherwise specified, in this application, a compound refers to a chemical substance formed by the connection of two or more elements through chemical bonds, including small molecule compounds and high molecular compounds, and is not limited thereto. In this text, a compound is not only limited to a single chemical substance when being interpreted, but can also be interpreted as the same kind of chemical substances with the same composition or the same properties.

[0037] Unless otherwise specified, in this application, parts by weight represent the relative weight parts in a composition, which can be any weight unit, such as but not limited to weight units such as kilograms, kilograms, grams, pounds, etc. For example, 100 parts by weight of polyphenylene ether resin means that it can be 100 kilograms of polyphenylene ether resin or 100 pounds of polyphenylene ether resin.

[0038] It should be understood that the features disclosed in each embodiment of this text can be arbitrarily combined to form the technical solutions of this application, as long as there is no contradiction in the combination of these features.

[0039] The present application will be described below with specific embodiments and examples. It should be understood that these specific embodiments and examples are merely illustrative and are not intended to limit the scope and use of the present application.

[0040] Unless otherwise specified, the methods, reagents, and conditions used in the preparation examples, comparative examples, and examples below are conventional methods, reagents, and conditions in the art.

[0041] Preparation Example Preparation Example 1 The raw materials used in this preparation example include: 10 kg of milk thistle seeds.

[0042] The preparation method of this preparation example includes: Z1. Put the milk thistle seeds into 20 kg of water and ultrasonically clean them for 30 min, then take them out, repeat the operation 3 times, and then dry them; Z2. Put the dried milk thistle seeds into a grinder and grind them to a particle size of 400 mesh; Z3. Dry the ground milk thistle seeds at 80 °C for 5 h to obtain milk thistle powder.

[0043] Example Example 1 The preparation method of this example includes: S1.1. At 25 °C, soak 1 kg of the milk thistle powder obtained in Preparation Example 1 in 4 kg of n-hexane for 6 h to obtain a primary immersion liquid; S1.2. After pouring out the upper n-hexane solution of the primary immersion liquid, the amount poured out is half of the volume of the primary immersion liquid, and then replenish n-hexane with the same volume as the poured-out primary immersion liquid, soak for another 2 h, repeat the operation until the primary immersion liquid is colorless and transparent, and then remove the upper primary immersion liquid to obtain an immersion liquid, and the mass of the immersion liquid is 1.5 kg; S2. Add 0.75 kg of ethanol to the immersion liquid, let it stand for 2 h and then filter, concentrate the filtrate at 40 °C to 0.25 times the volume; then slowly add the concentrated filtrate to water, the mass of water is 8 times the mass of the concentrated filtrate, let it stand for 0.5 h, and take the lower-layer solution and precipitate; S3. Evaporate and concentrate the lower-layer solution and precipitate at 80 °C to 0.2 times the volume of the liquid, then cool to 2 °C for crystallization and filtration, and then dry at 80 °C for 4 h to obtain silymarin.

[0044] Example 2 The difference between this example and Example 1 is that: the mass of the immersion liquid obtained in step S1.2 is 1.8 kg, and the amount of ethanol used in step S2 is 0.9 kg.

[0045] Example 3 The difference between this example and Example 1 is that: the mass of the soaking solution obtained in step S1.2 is 2.0 kg, and the amount of ethanol used in step S2 is 1 kg.

[0046] Example 4 The preparation method of this example includes: S1.1. At 25 °C, soak 1 kg of the silymarin powder obtained in Preparation Example 1 with 5 kg of n-hexane for 8 h to obtain a primary soaking solution; S1.2. After pouring out the upper n-hexane solution of the primary soaking solution, the amount poured out is half of the volume of the primary soaking solution, and then add n-hexane with the same volume as the poured-out primary soaking solution, soak for 3 h again, repeat the operation until the primary soaking solution is colorless and transparent, and then remove the upper primary soaking solution to obtain a soaking solution, and the mass of the soaking solution is 1.5 kg; S2. Add 1.2 kg of ethanol to the soaking solution, let it stand for 2 h and then filter, concentrate the filtrate to 0.1 times the volume at 50 °C; then slowly add the concentrated filtrate to water, the mass of water is 12 times the mass of the concentrated filtrate, let it stand for 0.5 h and then take the lower layer solution and precipitate; S3. Evaporate and concentrate the lower layer solution and precipitate to 0.2 times the volume of the liquid at 80 °C, then cool to 2 °C for crystallization and filtration, and then dry at 85 °C for 4 h to obtain silymarin.

[0047] Example 5 The difference between this example and Example 4 is that: the amount of ethanol used in step S2 is 1.5 kg.

[0048] Example 6 The preparation method of this example includes: S1.1. At 25 °C, soak 1 kg of the silymarin powder obtained in Preparation Example 1 with 6 kg of n-hexane for 10 h to obtain a primary soaking solution; S1.2. After pouring out the upper n-hexane solution of the primary soaking solution, the amount poured out is half of the volume of the primary soaking solution, and then add n-hexane with the same volume as the poured-out primary soaking solution, soak for 2 h again, repeat the operation until the primary soaking solution is colorless and transparent, and then remove the upper primary soaking solution to obtain a soaking solution, and the mass of the soaking solution is 1.8 kg; S2. Add 0.9 kg of ethanol to the soaking solution, let it stand for 2 h and then filter, concentrate the filtrate to 0.1 times the volume at 60 °C; then slowly add the concentrated filtrate to water, the mass of water is 10 times the mass of the concentrated filtrate, let it stand for 0.5 h and then take the lower layer solution and precipitate; S3. Evaporate and concentrate the lower layer solution and precipitate to 0.2 times the volume of the liquid at 80 °C, then cool to 2 °C for crystallization and filtration, and then dry at 80 °C for 4 h to obtain silymarin.

[0049] Example 7 The difference between this example and Example 6 is that in step S2, the filtrate is concentrated to 0.25 times the volume.

[0050] Example 8 The preparation method of this example includes: S1.1. At 25°C, soak 1 kg of the silybum marianum powder obtained in Preparation Example 1 with 5 kg of n-hexane for 8 h to obtain a primary leaching solution; S1.2. After pouring out the upper n-hexane solution of the primary leaching solution, pour out half of the volume of the primary leaching solution, then supplement n-hexane with the same volume as the poured-out primary leaching solution, soak for another 2 h, repeat the operation until the primary leaching solution is colorless and transparent, and then remove the upper primary leaching solution to obtain a soaking solution, and the mass of the soaking solution is 1.5 kg; S2. Add 0.75 kg of ethanol to the soaking solution, let it stand for 2 h and then filter, concentrate the filtrate to 0.15 times the volume at 40°C; then slowly add the concentrated filtrate to water, and the mass of water is 8 times the mass of the concentrated filtrate, let it stand for 0.5 h and then take the lower-layer solution and precipitate; S3. Evaporate and concentrate the lower-layer solution and precipitate to 0.2 times the liquid volume at 80°C, then cool to 2°C for crystallization and filter, and then dry at 80°C for 4 h to obtain silymarin.

[0051] Example 9 The difference between this example and Example 8 is that in step S2, the mass of water is 12 times the mass of the concentrated filtrate.

[0052] Table 1 Dosages of some reagents and some process parameters in Examples 1 - 9 Test Example Example 1 Example 2 Example 3 Example 4 Example 5 Example 6 Example 7 Example 8 Example 9 Silybum marianum powder / kg 1 1 1 1 1 1 1 1 1 Duration of n - hexane soaking in Step S1.1 / h 6 6 6 8 8 10 10 8 8 Dosage of n - hexane in Step S1.1 / kg 4 4 4 5 5 6 6 5 5 Mass of soaking solution / kg 1.5 1.8 2 1.5 1.5 1.8 1.8 1.5 1.5 <![CDATA[m 乙醇 / m 浸泡液 > 0.5 0.5 0.5 0.8 1 0.5 0.5 0.5 0.5 Concentration temperature in Step S2 / °C 40 40 40 50 50 60 60 40 40 Concentration multiple of filtrate 0.25 0.25 0.25 0.1 0.1 0.1 0.25 0.15 0.15 <![CDATA[V 水 / V 浓缩后的滤液 > 8 8 8 12 12 10 10 8 12 Evaporation and concentration temperature in Step S3 / °C 80 80 80 85 85 80 80 80 80 Comparative example Comparative Example 1 The preparation method of this comparative example includes: D1.1. At 25°C, soak 1 kg of the silybum marianum powder obtained in Preparation Example 1 with 4 kg of n-hexane for 6 h to obtain a primary leaching solution; D1.2. After pouring out the upper n-hexane solution of the primary leaching solution, pour out half of the volume of the primary leaching solution, then supplement n-hexane with the same volume as the poured-out primary leaching solution and soak for 2 h, repeat the operation until the primary leaching solution is colorless and transparent, and then remove the upper primary leaching solution to obtain a soaking solution, and the mass of the soaking solution is 1.5 kg; D2. Dry the soaking solution at 50°C and then add 0.75 kg of ethanol, let it stand for 2 h and then filter, and take the upper clear liquid; D3. Evaporate and concentrate the upper clear liquid to 0.2 times the liquid volume at 80°C, then cool to 2°C for crystallization and filter, and then dry at 80°C for 4 h to obtain silymarin.

[0053] Experimental results The silymarin obtained in Examples 1 to 9 and Comparative Example 1 was subjected to content detection, ethanol residue detection, and n-hexane residue detection. The detection method was the United States Pharmacopoeia, and the detection procedure was CGJT / JFB21002.

[0054] Table 2 Parameters of the silymarin obtained in Examples 1 to 9 and Comparative Example 1 Test Example Residual amount of n - hexane / ppm Residual amount of ethanol / ppm Purity of silymarin % Example 1 33 78.65 46.47 Example 2 37 78.00 46.55 Example 3 39 78.65 46.66 Example 4 24 74.10 48.31 Example 5 16 75.40 48.34 Example 6 42 84.50 47.67 Example 7 34 79.95 47.61 Example 8 38 80.60 46.52 Example 9 36 78.65 47.93 Comparative Example 1 106 114.00 29.73 It should be understood that the above embodiments are only to illustrate the technical concept and characteristics of the present invention, and the purpose is to enable those who are familiar with this technology to understand the content of the present invention and implement it accordingly. It cannot be determined that the specific implementation of the present invention is only limited to these descriptions. For those of ordinary skill in the technical field to which the present invention belongs, without departing from the concept of the present invention, several simple deductions or substitutions can also be made. Any equivalent changes or modifications made according to the spirit and essence of the present invention should be covered within the protection scope of the present invention.

Claims

1. A method for extracting silymarin with low residue, characterized in that, It includes the following steps: Eluting grease: After crushing the silybum marianum seeds, soak them with n-hexane and elute the grease to obtain a soaking solution; Ethanol replacement: Add ethanol to the soaking solution, let it stand and then filter. Concentrate the filtrate, add water and let it stand, and take the lower-layer solution and the precipitate; Crystallizing and drying: Concentrate the lower-layer solution and the precipitate, crystallize again and then filter, and then dry the crystals to obtain silymarin.

2. The extraction method of silymarin with low residue according to claim 1, characterized in that: In the step of eluting grease, after crushing the silybum marianum seeds, soak them with n-hexane for 6 - 10 h, and the dosage of n-hexane is 4 - 6 times the mass of the silybum marianum seeds; after the soaking is completed, remove the upper-layer liquid and then replenish n-hexane with the same volume as the removed upper-layer liquid, soak again for 2 - 3 h, repeat the operation until the upper-layer liquid is colorless and transparent after the soaking is completed, and then remove the upper-layer liquid to obtain the soaking solution.

3. A method for extracting silymarin with low residue according to any one of claims 1 or 2, characterized in that: The mass of the soaking solution is 1.5 - 2 times the mass of the silybum marianum seeds.

4. The extraction method of silymarin with low residue according to claim 1, characterized in that: In the step of ethanol replacement, the dosage of ethanol is 0.5 - 1 times the mass of the soaking solution.

5. The extraction method of silymarin with low residue according to claim 1, characterized in that: In the step of ethanol replacement, the volume of the filtrate after concentration is 0.1 - 0.25 times that before concentration.

6. The extraction method of silymarin with low residue according to claim 1, characterized in that: In the step of ethanol replacement, the filtrate is evaporated and concentrated at 40°C - 60°C.

7. The extraction method of silymarin with low residue according to claim 1, wherein: In the step of ethanol replacement, the mass of water is 8 - 12 times the mass of the filtrate after concentration.

8. The extraction method of silymarin with low residue according to claim 1, characterized in that: In the step of crystallizing and drying, the lower-layer solution and the precipitate are evaporated and concentrated at 80°C - 85°C.