Method for purifying oleanolic acid from Tibetan capillary artemisia
High-purity oleanolic acid was extracted from Artemisia capillaris by combining alkaline dissolution with acid precipitation, activated carbon reflux decolorization, and methanol recrystallization. This method solved the problems of high difficulty and high cost in oleanolic acid purification and achieved efficient and low-cost oleanolic acid extraction.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-12-22
- Publication Date
- 2026-03-24
AI Technical Summary
In the existing technology, oleanolic acid is difficult to purify, resulting in its high price, and there are no relevant literature reports on the purification process of oleanolic acid from Artemisia capillaris.
A method combining alkali dissolution and acid precipitation with activated carbon reflux decolorization and recrystallization was adopted. This method involves purifying the crude extract of Artemisia capillaris by alkali dissolution and acid precipitation, followed by multiple alkali dissolution and acid precipitation and activated carbon reflux decolorization, and then recrystallization with methanol to obtain high-purity oleanolic acid.
This study achieved efficient extraction of high-purity oleanolic acid from Artemisia capillaris, with a yield and purity of 98.35%, solving the problem of difficult purification of oleanolic acid and reducing production costs.
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Abstract
Description
Technical Field
[0001] This invention relates to a method for purifying oleanolic acid from Artemisia capillaris. Background Technology "Tibetan Artemisia capillaris," also known as "Sichuan Swertia," is a plant belonging to the genus Swertia in the family Gentianaceae. It has a wide range of uses in Tibetan medicine, and the main detectable chemical substances include oleanolic acid, swertiamarin, mangiferin, and gentiopicrin. Oleanolic acid is a pentacyclic triterpenoid compound with lipid-lowering, anti-inflammatory, hepatoprotective, antioxidant, and anti-tumor effects. Currently, oleanolic acid is mainly isolated from natural plants such as oleanolic fruit, privet fruit, and achyranthes bidentata. Natural oleanolic acid has high safety and few toxic side effects, showing good prospects for clinical application. However, due to limited raw materials and difficulties in purifying oleanolic acid, the price of high-purity oleanolic acid remains high. Therefore, expanding the source of oleanolic acid raw materials and developing its purification process is a meaningful endeavor. Literature indicates that Tibetan Artemisia capillaris contains a relatively high amount of oleanolic acid, but no literature reports on its purification process have been found to date.
[0002] The content of the background section is merely the technology known to the inventor and does not necessarily represent the prior art in this field. Summary of the Invention
[0003] In view of the above problems, the present invention discloses a method for purifying oleanolic acid from Artemisia capillaris, comprising the following steps: (1) The crude extract of Artemisia capillaris was purified by alkali dissolution and acid precipitation to obtain crude oleanolic acid. The crude oleanolic acid was washed to obtain the sample after alkali dissolution and acid precipitation. (2) Dissolve the sample after alkali dissolution and acid precipitation to obtain a solution; (3) Reflux the dissolved solution with activated carbon to decolorize it, filter it, and wait for the filtrate to crystallize to obtain the oleanolic acid product.
[0004] Furthermore, in step (1), the crude extract of Artemisia capillaris is obtained by ultrasonic extraction of Artemisia capillaris.
[0005] Furthermore, in step (1), the alkaline dissolution and acid precipitation process is as follows: the crude extract of Artemisia capillaris is alkalized with concentrated alkaline solution, refluxed and filtered, adjusted to acidity, crystallized and filtered to obtain crude oleanolic acid.
[0006] Furthermore, in step (1), before the alkali dissolution and acid precipitation, the crude extract of Artemisia capillaris is decolorized by reflux with activated carbon; and / or The filtrate obtained after crystallization and filtration is concentrated, cooled, crystallized again, and filtered to obtain another portion of crude oleanolic acid; and / or The concentrated alkaline solution is alkalized to pH 11; and / or The acid used to adjust the pH to acidity is concentrated hydrochloric acid, adjusted to pH=1.
[0007] Furthermore, in step (1), the alkaline dissolution and acid precipitation are repeated 2-4 times. Before repeating the alkaline dissolution and acid precipitation, the sample after the previous alkaline dissolution and acid precipitation is dissolved.
[0008] Furthermore, in step (1), the sample dissolved after the previous alkali dissolution and acid precipitation is dissolved in anhydrous ethanol; and / or The cleaning solution for washing crude oleanolic acid is water; and / or After the previous alkaline dissolution and acid precipitation, the sample was dissolved and then decolorized by activated carbon reflux, followed by repeated alkaline dissolution and acid precipitation.
[0009] Furthermore, in step (2), the sample after alkali-soluble acid precipitation is dissolved in anhydrous methanol until the mass concentration is 0.1 g·L⁻¹. -1 ; and / or In step (3), the sample after alkali dissolution and acid precipitation is dissolved and heated to 50-70℃, followed by activated carbon reflux decolorization; and / or In step (3), the crystallization temperature of the filtrate is 10-30℃.
[0010] Furthermore, in step (3), the filtration is vacuum filtration; and / or In step (3), the sample after alkali dissolution and acid precipitation is dissolved and heated to 60°C, followed by activated carbon reflux decolorization; and / or In step (3), the crystallization temperature of the filtrate is 15°C.
[0011] Furthermore, the oleanolic acid product obtained in step (3) is repeated 2-4 times, including the dissolution in step (2) and the steps of refluxing the dissolved solution with activated carbon for decolorization, filtering, and crystallizing the filtrate.
[0012] Furthermore, in the activated carbon reflux decolorization process, the mass of activated carbon added is 2-3 times the total mass of the solid. Attached Figure Description
[0013] The accompanying drawings are provided to further illustrate the invention and form part of the specification. They are used in conjunction with embodiments of the invention to explain the invention and do not constitute a limitation thereof. In the drawings: Figure 1 HPLC chromatogram of crude extract of Artemisia capillaris (A); Figure 2 HPLC chromatogram of sample (B) after the first alkaline dissolution and acid precipitation; Figure 3 HPLC chromatogram of sample (C) after second alkaline dissolution and acid precipitation; Figure 4 HPLC chromatogram of sample (D) after the first recrystallization; Figure 5HPLC chromatogram of oleanolic acid product (E) after three recrystallizations; Figure 6 HPLC chromatogram of oleanolic acid standard (F); Figure 7 UV-Vis spectrum of oleanolic acid product; Figure 8 1H NMR spectrum of oleanolic acid products; Figure 9 Structural diagram of oleanolic acid; Figure 10 FT-IR image of oleanolic acid product. Detailed Implementation
[0014] In the following description, only certain exemplary embodiments are depicted simply. As those skilled in the art will recognize, the described embodiments can be modified in various ways without departing from the spirit or scope of the invention. Therefore, the drawings and description are considered to be exemplary in nature and not restrictive.
[0015] Unless otherwise defined herein, scientific and technical terms used in conjunction with this invention will have the meaning commonly understood by one of ordinary skill in the art. Furthermore, unless the context otherwise requires, singular terms shall include plural forms, and plural terms shall include singular forms. More specifically, as used in this specification and the appended claims, unless the context explicitly indicates otherwise, the singular forms “a,” “an,” and “the” include plural indicators. In this application, unless otherwise stated, the use of “or” means “and / or.” Furthermore, the use of the term “comprising” and other forms such as “including” and “containing” is not limiting. Moreover, the scope provided in the specification and the appended claims includes all values between endpoints. Preferred embodiments of the invention are described below; it should be understood that the preferred embodiments described herein are for illustration and explanation only and are not intended to limit the invention.
[0016] Example The invention generally described herein will be more readily understood by referring to the following examples, which are provided by way of illustration and are not intended to limit the invention. Furthermore, unless otherwise specified, the experimental methods in the following examples are conventional methods. Unless otherwise specified, the raw materials, reagents, and other materials used in the following examples are commercially available products.
[0017] 1. Instruments and reagents 1.1 Instruments Agilent 1260 High Performance Liquid Chromatograph (HPLC), Agilent Technologies Inc.; XH-2008DE Ultrasonic Synthesis Extraction System, Beijing Xianghu Technology Development Co., Ltd.; 765 UV-Vis Spectrophotometer, Shanghai Instrument & Electronics Analytical Instrument Co., Ltd.; DZF-6050 Vacuum Drying Oven, Shanghai Boxun Industrial Co., Ltd. Medical Equipment Factory; RE-5203 Rotary Evaporator, Shanghai Yarong Biochemical Instrument Factory; TD4 Benchtop Low-Speed Centrifuge, Hunan Instrument & Meter General Factory Centrifuge Factory; HTY-800g Traditional Chinese Medicine Pulverizer, Yongkang Hongtaiyang Electromechanical Co., Ltd.
[0018] 1.2 Test Drugs The Tibetan Artemisia capillaris, originating from Sichuan, was identified by Professor Zhang Jixiang of the School of Chemistry and Materials Science at Langfang Normal University as the dried whole herb of *Swertia spp.*, a plant belonging to the Gentianaceae family. Oleanolic acid standard (≥98%, Shanghai Aladdin Biochemical Technology Co., Ltd.); dimethyl sulfoxide-d6 (99.9% + 0.03% TMS, Shanghai Maclean Biochemical Technology Co., Ltd.); potassium bromide (spectrally pure, Shanghai Maclean Biochemical Technology Co., Ltd.); acetonitrile and phosphoric acid (chromatographically pure, Tianjin Chemical Reagent Supply and Marketing Co., Ltd.); all other reagents were of analytical grade.
[0019] 2. Methods and Results Example 1 2.1 Preparation of oleanolic acid products 2.1.1 Ultrasonic coarse extraction of oleanolic acid from Artemisia capillaris: A certain amount of Artemisia capillaris was crushed using a pulverizer and passed through an 80-mesh sieve for later use.
[0020] 2.1.2 Ultrasonic Extraction: Using 85% ethanol solution at a material-to-liquid ratio of 1:16, ultrasonic extraction was performed at approximately 60°C and 800W for 40 minutes. The mixture was then filtered under reduced pressure. The residue was extracted using the same method, and the two filtrates were combined. The filtrate was concentrated under reduced pressure to obtain an extract. The extract was dissolved in deionized water, boiled for 30 minutes, and then centrifuged (4000 r·min). -1 Centrifuge for 5 min, discard the supernatant, combine the precipitates and completely dissolve them with anhydrous ethanol to obtain the crude extract of Artemisia capillaris, for later use.
[0021] 2.2 Preliminary separation of oleanolic acid from Artemisia capillaris by alkaline dissolution and acid precipitation method Take the above-mentioned crude extract of Artemisia capillaris in a round-bottom flask, add activated carbon granules three times the total solid mass of the sample, reflux for decolorization, filter while hot, alkalize with concentrated alkali to pH=11, reflux for 30 min, filter under vacuum to obtain sodium oleanolic acid solution, adjust pH to=1 with concentrated hydrochloric acid, place in a refrigerator for 24 h to crystallize, filter to obtain a portion of crude oleanolic acid and mother liquor, concentrate the mother liquor, cool, allow to crystallize, filter to obtain another portion of crude oleanolic acid, combine the above crude products, dry the crude products, wash with an appropriate amount of deionized water to obtain the sample after the first alkali dissolution and acid precipitation. Weigh the sample, dissolve in a small amount of anhydrous ethanol, add activated carbon twice the total solid mass of the sample, reflux for decolorization, filter while hot, repeat the above alkali dissolution and acid precipitation steps to obtain the sample after the second alkali dissolution and acid precipitation.
[0022] 2.3 Recrystallization yields Artemisia capillaris and oleanolic acid products. After the second alkaline dissolution and acid precipitation, the oleanolic acid sample was weighed and dissolved in anhydrous methanol to prepare a solution with a mass concentration of 0.1 g·L⁻¹. -1 The solution was heated to 60℃, and activated carbon twice the total solid mass of the sample was added. The mixture was then refluxed for decolorization, vacuum filtered, and the filtrate was allowed to crystallize naturally at room temperature (15℃) to obtain the first recrystallized sample. The above steps were repeated three times, involving activated carbon decolorization and methanol recrystallization, to obtain a high-purity oleanolic acid product.
[0023] Example 2 High Performance Liquid Chromatography Analysis 2.4.1 Chromatographic conditions: The chromatographic column was SB-C18 (150 mm × 4.6 mm, 5 µm); the mobile phase was chromatographically pure acetonitrile-water solution (80:20, v / v), and the flow rate was 1.0 mL·min. -1 The column temperature was 40 ℃, the detection wavelength was 210 nm, and the injection volume was 10 µL.
[0024] 2.4.2 Standard Solution: Accurately weigh 7.5 mg of oleanolic acid standard and place it in a 25 mL volumetric flask. Dilute to 25 mL with anhydrous methanol (chromatographic grade) to obtain an oleanolic acid standard solution with a concentration of 0.3 mg / mL. -1 Filter using a 0.45µm syringe-type microporous membrane and store at around 4℃ until use.
[0025] 2.4.3 Accurately weigh 7.5 mg of the oleanolic acid product from Artemisia capillaris and oleanolic acid samples from other purification steps (samples after the first alkaline dissolution and acid precipitation, the second alkaline dissolution and acid precipitation, and the first recrystallization). Place each sample in a 25 mL volumetric flask and dilute to 25 mL with anhydrous methanol (chromatographic grade) to obtain product and other sample solutions with a concentration of 0.3 mg / mL. -1The oleanolic acid product and other oleanolic acid samples from purification steps (samples after the first alkali dissolution and acid precipitation, samples after the second alkali dissolution and acid precipitation, and samples after the first recrystallization) were filtered through a 0.45µm syringe-type microporous membrane and stored at around 4℃ for later use.
[0026] 2.4.4 Linearity Examination: Take 0.5 mL, 1.0 mL, 1.5 mL, 2.0 mL, 2.5 mL, and 3.0 mL of oleanolic acid standard solution with a concentration of 0.3 mg / mL as described in "2.4.2", and add them to different 10 mL volumetric flasks. Dilute to 10 mL with chromatographic grade methanol. After filtration through a 0.45 μm syringe filter, inject the solution using high-performance liquid chromatography (HPLC) under the conditions described in "2.4.1". Perform linear regression with the oleanolic acid standard concentration as the abscissa and the oleanolic acid peak area as the ordinate. The regression equation for oleanolic acid is: Y = 3750.25X + 2.19 r=0.9996 The results showed that the concentration of oleanolic acid was 0.015-0.09 mg·mL. -1 The mass concentration and peak area have a good linear relationship.
[0027] 2.4.5 Precision Test: The oleanolic acid standard solution under section “2.4.2” was diluted 5 times to obtain a concentration of 0.06 mg·mL. -1 Six injections were performed consecutively under the chromatographic conditions described in "2.4.1". The peak area was measured, and the RSD of the oleanolic acid peak area was calculated to be 1.0%, indicating that the instrument has good precision and is suitable for the measurement of oleanolic acid in Artemisia capillaris.
[0028] 2.4.6 Stability test: At 0, 6, 12, 24, 48 and 72 h after preparation, 10 µL of the same oleanolic acid product solution was accurately pipetted and injected according to the chromatographic conditions in section “2.4.1”. The RSD of the oleanolic acid peak area was calculated to be 1.5%, indicating that the oleanolic acid product solution was stable within 72 h.
[0029] 2.4.7 Liquid Chromatography Analysis of Sample Solutions The product and other sample solutions, as well as the crude extract of Artemisia capillaris, were continuously injected and analyzed under the chromatographic conditions described in "2.4.1".
[0030] HPLC detection showed that... Figure 6 It can be seen that the retention time of oleanolic acid is 13.6 min. At this retention time, [the following is a description of a specific product / process]... Figures 1-6 It can be seen that characteristic peaks (i.e., peak marked 1 in the figure) were observed in all samples, indicating the presence of oleanolic acid. Comparison revealed that the crude extract of Artemisia capillaris (…) Figure 1The peak intensity of the target component was significantly lower than that of other components, indicating that the content of oleanolic acid was low and further separation and purification were needed. Figure 2 and Figure 3 The image shows the sample after alkali dissolution and acid precipitation. As can be seen from the figure, the alkali dissolution and acid precipitation method removed a significant amount of substances with retention times of approximately 3 minutes and 5 minutes. Before alkali dissolution, activated carbon removed some pigments and other impurities. Oleanolic acid, being a weak acid, dissolves in the alkali solution as sodium oleanolicate, while some impurities are insoluble in the alkali solution and are removed by filtration. During acid precipitation, sodium oleanolicate combines with the acid to form oleanolic acid. In an acidic environment, oleanolic acid precipitates due to its reduced solubility. Some neutral impurities remain soluble in water. After filtration, a sample of oleanolic acid with a certain purity is obtained. Figure 3 It was found that after two alkaline dissolution and acid precipitation processes, some impurities with acidity close to oleanolic acid remained unremoved, and the sample color was relatively dark, indicating that some pigments were still present. Therefore, we further purified the oleanolic acid sample by activated carbon reflux decolorization followed by methanol recrystallization. Figure 4 It can be seen that activated carbon decolorization followed by methanol recrystallization can effectively purify oleanolic acid. Figure 5 It can be seen that after three rounds of activated carbon decolorization and methanol recrystallization, a high-purity oleanolic acid product was obtained.
[0031] Example 3: Ultraviolet-Visible Spectroscopic Analysis of Oleanolic Acid Products 2.5 Take a small amount of Artemisia capillaris oleanolic acid product, dissolve it in chromatographically pure methanol, and perform ultraviolet-visible spectroscopy analysis.
[0032] Depend on Figure 7 It can be seen that the oleanolic acid product of Artemisia capillaris has a maximum absorption peak at around 210 nm, which is consistent with the value found in the literature, further indicating that the product is oleanolic acid.
[0033] For details on the ultraviolet-visible spectroscopy analysis method and the literature values consulted, please refer to: Li Bibo, Zhao Ling, Zhang Xuemei, et al. Determination of oleanolic acid sustained-release pill content and in vitro release rate by ultraviolet spectroscopy [J]. Journal of Chongqing Medical University, 2009, 34(4):447; Di Jinming, Liu Binbin, Zhao Jing, et al. Comparative study of five methods for determining oleanolic acid content [J]. Northwest Pharmaceutical Journal, 2013, 28(5):483.
[0034] Example 4: 1H NMR Spectroscopy Analysis of Oleanolic Acid Products 2.6 Take a small amount of Artemisia capillaris oleanolic acid product and analyze it using dimethyl sulfoxide-d6 as solvent by 1H NMR spectrum.
[0035] Depend on Figure 8The chemical shifts and corresponding peak assignments in the 1H NMR spectrum are as follows: δ: 12.01 (1H, s, carboxyl hydrogen peak), 5.16 (1H, brs, H-12), 4.28 (1H, d, H-18), 3.32 (hydroxyl hydrogen peak), 1.24, 1.09, 0.89, 0.87, 0.85, 0.72, 0.68 (each 3H, s, 7×CH3), 2.5 is the solvent peak of DMSO-d6, and 0 is the peak of the standard substance TMS. These data are consistent with literature reports; therefore, this product is identified as oleanolic acid.
[0036] For details on the nuclear magnetic resonance hydrogen spectroscopy analysis method and the literature values consulted, please refer to: Tian Guanghui, Zhao Hua, Liu Cunfang, et al. Study on fat-soluble components in Evodia rutaecarpa fruit [J]. Chinese Medicinal Herbs, 2008, 31(3):382; Zhang Xiuyan, Wang Xiaoqin, Wen Aiping. Study on chemical components of Gentiana macrophylla flower [J]. Chinese Traditional and Herbal Drugs, 2017, 48(2):241.
[0037] Figure 9 This is a structural diagram of oleanolic acid.
[0038] Example 5 Infrared Spectroscopic Analysis of Oleanolic Acid Products 2.7 Take a small amount of Tibetan Artemisia capillaris oleanolic acid product, add an appropriate amount of spectrally pure potassium bromide, stir evenly, grind into powder using a mortar and pestle, compress into tablets, and perform infrared spectroscopy determination.
[0039] Figure 10 The infrared spectrum of the product is shown in Table 1, and the assignments of its main absorption peaks are listed. Infrared spectral analysis shows that the measurement results are consistent with... Figure 9 The structural formula of oleanolic acid is consistent with that of oleanolic acid, and the above data further confirms that the product is oleanolic acid.
[0040] For details on the infrared spectroscopy analysis method and the literature values consulted, please refer to: Wu Lianzhong, Ke Weizhong. Verification of the structural characteristics of effective components of traditional Chinese medicine by laser Raman spectroscopy [J]. Spectroscopy and Spectral Analysis, 1995, 15(6):51.
[0041] Table 1 Infrared spectral analysis of oleanolic acid products
[0042] Example 6: Calculation of Oleanolic Acid Product Yield 2.8 Accurately weigh 100.0000 g of dried Artemisia capillaris raw material. After purification using the above optimized process, place the final product in a vacuum drying oven (60℃) until constant weight. The mass of the purified product was determined to be 0.0770 g using an analytical balance. Substituting this mass into the following formula [Dai Fucai, Zhao Liyan, Cao Xiangming, et al. Purification and structural identification of chlorogenic acid in Chrysanthemum indicum [J]. Food Industry, 2020, 41(10):213], the yield was calculated to be 0.077%.
[0043] Y = (m / G) × 100% In the formula: Y is the yield of oleanolic acid extracted from Artemisia capillaris, %; m is the mass of oleanolic acid product, g; G is the mass of dried Artemisia capillaris powder used, g.
[0044] Example 7: Repeatability test of purification process and calculation of product purity 2.9 Accurately weigh 3 portions of Artemisia capillaris powder, purify the oleanolic acid product according to methods “2.1-2.3”, prepare the Artemisia capillaris oleanolic acid product solution according to method “2.4.3”, dilute the solution 5 times, and inject it according to the chromatographic conditions under “2.4.1”. The average purity of the oleanolic acid product was found to be 98.35%. The RSD was 0.33%, indicating that the purification method has good repeatability.
[0045] Table 2 Process Validation Results
[0046] During the alkali dissolution and acid precipitation of Artemisia capillaris samples containing oleanolic acid, white particulate matter precipitated during the first acid precipitation step. Liquid chromatography analysis revealed that these white particulate matter contained only a small amount of oleanolic acid, while the majority of the oleanolic acid was present in the brown precipitate sample containing more impurities. Analysis indicates that the oleanolic acid content in Artemisia capillaris is low. During the first alkali dissolution and acid precipitation, oleanolic acid precipitated along with pigments and other impurities, while the white particulate matter was sodium chloride generated during pH adjustment. Therefore, after the first alkali dissolution and acid precipitation, the sample can be filtered, dried, and washed with a small amount of deionized water. Since sodium chloride is soluble in water while oleanolic acid is insoluble, the sodium chloride is removed. Furthermore, since oleanolic acid is soluble in ethanol, drying the sample to remove the ethanol before washing with water can effectively reduce the loss of oleanolic acid.
[0047] The amount of activated carbon used can be adjusted according to the sample color. Activated carbon mainly removes impurities such as pigments. If the sample color is dark, the amount can be increased by 1-2 times.
[0048] During the first recrystallization of the oleanolic acid sample from Artemisia capillaris with methanol, liquid chromatography revealed a slight tailing of the oleanolic acid peak. This was likely due to the presence of a small amount of ursolic acid in the product. After three recrystallizations, the tailing disappeared, yielding a high-purity oleanolic acid product.
[0049] Finally, it should be noted that the above descriptions are merely preferred embodiments of the present invention and are not intended to limit the present invention. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art can still modify the technical solutions described in the foregoing embodiments or make equivalent substitutions for some of the technical features. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention. Furthermore, although the elements of the present invention can be described or claimed individually, it is also conceivable to have multiple elements, unless explicitly limited to a single element.
Claims
1. A method for purifying oleanolic acid from Artemisia capillaris, characterized in that, Includes the following steps: (1) The crude extract of Artemisia capillaris was purified by alkali dissolution and acid precipitation to obtain crude oleanolic acid. The crude oleanolic acid was washed to obtain the sample after alkali dissolution and acid precipitation. (2) Dissolve the sample after alkali dissolution and acid precipitation to obtain a solution; (3) Reflux the dissolved solution with activated carbon to decolorize it, filter it, and wait for the filtrate to crystallize to obtain the oleanolic acid product.
2. The method for purifying oleanolic acid from Artemisia capillaris according to claim 1, characterized in that, In step (1), the crude extract of Artemisia capillaris is obtained by ultrasonic extraction of Artemisia capillaris.
3. The method for purifying oleanolic acid from Artemisia capillaris according to claim 1, characterized in that, In step (1), the alkaline dissolution and acid precipitation process is as follows: the crude extract of Artemisia capillaris is alkalized with concentrated alkaline solution, refluxed and filtered, adjusted to acidity, crystallized and filtered to obtain crude oleanolic acid.
4. The method for purifying oleanolic acid from Artemisia capillaris according to claim 3, characterized in that, In step (1), before the alkali dissolution and acid precipitation, the crude extract of Artemisia capillaris is decolorized by reflux with activated carbon; and / or The filtrate obtained after crystallization and filtration is concentrated, cooled, crystallized again, and filtered to obtain another portion of crude oleanolic acid; and / or The concentrated alkaline solution is alkalized to pH 11; and / or The acid used to adjust the pH to acidity is concentrated hydrochloric acid, adjusted to pH=1.
5. The method for purifying oleanolic acid from Artemisia capillaris according to claim 3, characterized in that, In step (1), the alkaline dissolution and acid precipitation are repeated 2-4 times. Before repeating the alkaline dissolution and acid precipitation, the sample after the previous alkaline dissolution and acid precipitation is dissolved.
6. The method for purifying oleanolic acid from Artemisia capillaris according to claim 5, characterized in that, In step (1), anhydrous ethanol is used to dissolve the sample after the previous alkaline dissolution and acid precipitation; and / or The cleaning solution for washing crude oleanolic acid is water; and / or After the previous alkaline dissolution and acid precipitation, the sample was dissolved and then decolorized by activated carbon reflux, followed by repeated alkaline dissolution and acid precipitation.
7. The method for purifying oleanolic acid from Artemisia capillaris according to claim 1, characterized in that, In step (2), the sample after alkali-soluble acid precipitation is dissolved in anhydrous methanol until the mass concentration is 0.1 g·L⁻¹. -1 ; and / or In step (3), the sample after alkali dissolution and acid precipitation is dissolved and heated to 50-70℃, followed by activated carbon reflux decolorization; and / or In step (3), the crystallization temperature of the filtrate is 10-30℃.
8. The method for purifying oleanolic acid from Artemisia capillaris according to claim 7, characterized in that, In step (3), the filtration is vacuum filtration; and / or In step (3), the sample after alkali dissolution and acid precipitation is dissolved and heated to 60°C, followed by activated carbon reflux decolorization; and / or In step (3), the crystallization temperature of the filtrate is 15°C.
9. The method for purifying oleanolic acid from Artemisia capillaris according to claim 1, characterized in that, Repeat the steps of dissolving in step (2) and decolorizing by refluxing activated carbon and filtering the dissolved solution in step (3) and crystallizing the filtrate 2-4 times for the oleanolic acid product obtained in step (3).
10. The method for purifying oleanolic acid from Artemisia capillaris according to any of the preceding claims, characterized in that, In the activated carbon reflux decolorization process, the mass of activated carbon added is 2-3 times the total mass of the solid.