A deep eutectic solvent for extracting polysaccharides from evening primrose seeds and an ultrasonic method thereof
By combining eutectic solvents with ultrasound, the problems of long extraction time and high energy consumption of evening primrose seed polysaccharides have been solved, achieving efficient and low-cost polysaccharide extraction with a significantly improved extraction rate.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- SHENYANG AGRI UNIV
- Filing Date
- 2025-12-04
- Publication Date
- 2026-07-03
AI Technical Summary
Existing methods for extracting polysaccharides from evening primrose seeds are time-consuming, energy-intensive, prone to loss of active components, and have low yields, lacking efficient and rapid extraction methods.
A method combining eutectic solvent (DES) and ultrasound was employed. By selecting a specific molar ratio of hydrogen bond acceptor and hydrogen bond donor combination, the DES was prepared. Combined with ultrasound treatment of evening primrose seeds, the extraction process was optimized, including the preparation of the DES, sample processing, and ultrasonic extraction steps.
It significantly improves the extraction rate of evening primrose seed polysaccharides, with the extraction rate increasing by up to 800%. It is low-cost, environmentally friendly, and suitable for widespread application.
Smart Images
Figure CN121226580B_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of polysaccharide extraction technology, specifically relating to a eutectic solvent for extracting polysaccharides from evening primrose seeds and its ultrasonic method. Background Technology
[0002] Evening Primrose ( Oenothera biennis L. Evening primrose (Oenothera spp.) is a perennial herb belonging to the Onagraceae family and the Oenothera genus. Native to North America, it was initially introduced to Europe and subsequently spread rapidly to temperate and subtropical regions worldwide. It is cultivated in Northeast, North, East, and Southwest China and widely used in medicine, cosmetics, and chemical industries, making it one of the most promising species for development. Evening primrose seeds are rich in mucilage, primarily composed of polysaccharides. However, there are currently no reports on evening primrose seed polysaccharides. Polysaccharides are information molecules widely found in nature and organisms, possessing immunomodulatory, antioxidant, antiviral, and antitumor effects. They can also lower blood lipids, delay aging, and combat fatigue. Therefore, the extraction and property study of polysaccharides are crucial for the in-depth development and comprehensive utilization of evening primrose seed polysaccharides.
[0003] Traditional polysaccharide extraction methods, which use water as a medium, suffer from drawbacks such as time consumption, high energy consumption, easy loss of active components, and low yield. Therefore, it is necessary to establish an efficient and rapid method for extracting evening primrose seed polysaccharides, which has important theoretical and practical significance for the functional research and comprehensive development and utilization of evening primrose seed polysaccharides.
[0004] Deep eutectic solvents (DES) are a green and novel medium that can dissolve more active substances through hydrogen bonding. They have advantages such as non-toxicity, low volatility, high thermal stability, low energy consumption, and simple synthesis. Currently, there is no ultrasonic-assisted method using DES for the extraction of evening primrose seed polysaccharides. Summary of the Invention
[0005] To address the shortcomings of existing technologies, the present invention aims to provide a low-eutectic solvent and its ultrasonic method for extracting polysaccharides from evening primrose seeds. By comparing the polysaccharide yields from evening primrose seeds extracted with water and different combinations of low-eutectic solvents, a high-yield DES was screened, and the ultrasonic-assisted extraction technology was optimized. This extraction method is convenient, fast, and inexpensive, making it suitable for widespread application.
[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows:
[0007] A eutectic solvent for extracting polysaccharides from evening primrose seeds comprises a hydrogen bond acceptor and a hydrogen bond donor in a molar ratio of 1:(1~2); wherein the hydrogen bond acceptor is choline chloride, and the hydrogen bond donor is a single component or a two-component component; the single component comprises any one of glycerol, malonic acid, lactic acid, 1,3-butanediol, butylene glycol, D-sorbitol, citric acid, and ethanol; the two-component component is 1,3-butanediol and butylene glycol, wherein the molar ratio of 1,3-butanediol and butylene glycol is 2:(0.5~1);
[0008] The above-mentioned ultrasonic method for extracting eutectic solvents from evening primrose seeds includes the following steps:
[0009] (1) Preparation of eutectic solvent: The hydrogen bond acceptor and hydrogen bond donor are mixed in an Erlenmeyer flask and stirred under constant temperature water bath conditions until a clear and transparent viscous liquid is obtained. Water is added to mix and the eutectic solvent DES is obtained.
[0010] (2) Sample preparation: The dried evening primrose seeds were ground into powder and stored at -20℃ for later use;
[0011] (3) Ultrasonic-assisted extraction: Add the eutectic solvent DES to the evening primrose seed powder, mix well and then extract by ultrasonication;
[0012] (4) After ultrasonication, filter the supernatant and add anhydrous ethanol to the supernatant until the volume fraction of anhydrous ethanol is 80%. Let it stand at 4~7℃, then centrifuge, discard the supernatant to obtain the precipitate, and dry the precipitate to obtain evening primrose seed polysaccharide extract.
[0013] The constant temperature range mentioned in step (1) is 55~75℃;
[0014] The amount of water added in step (1) is 30-50% of the mass of the eutectic solvent DES;
[0015] The ratio of evening primrose seed powder to eutectic solvent DES in step (3) is 1:(20~80)g / ml;
[0016] The ultrasonic time in step (3) is 20~70 min, the ultrasonic power is 20~90 W, and the extraction temperature is 30~80℃;
[0017] The settling time mentioned in step (4) is 12~24 h;
[0018] The centrifugation temperature in step (4) is 10~20℃, and the centrifugation speed is 3600~5000 r·min. -1 The centrifugation time is 5-10 min.
[0019] Compared with the prior art, the beneficial effects of the present invention are as follows:
[0020] 1. This invention specifically explores the characteristics of evening primrose seeds and extracts them by combining eutectic solvent treatment and ultrasonic processes in a specific order. By setting different hydrogen bond donors in the eutectic solvent and matching them with specific hydrogen bond acceptors, the extraction rate of evening primrose seed polysaccharides is significantly improved, greatly enhancing the resource utilization rate of evening primrose.
[0021] 2. In this invention, by optimizing and improving the extraction process, ultrasonic treatment is used during the extraction process to prevent the structure of polysaccharide compounds in evening primrose from being destroyed due to excessively high extraction temperature. Therefore, the technical solution of this invention can improve the extraction rate of evening primrose seed polysaccharides.
[0022] 3. The method of this invention is convenient and quick, and can extract evening primrose seed polysaccharides in a short time without the need for expensive instruments and equipment; DES has the good properties of ionic liquids, excellent solubility and extractability, high extraction rate, up to 800% higher than conventional water extraction, low cost, non-toxic, environmentally friendly, and suitable for widespread application. Attached Figure Description
[0023] Figure 1 Surface scanning electron microscope image of evening primrose seed polysaccharide extracted by ultrasonication in eutectic solvent, scale bar 5 μm;
[0024] Figure 2 Surface scanning electron microscope image of evening primrose seed polysaccharide extracted by ultrasonication in eutectic solvent, scale bar 20 μm;
[0025] Figure 3 The graph shows the effect of different eutectic solvent systems on the polysaccharide content of evening primrose seeds. Detailed Implementation
[0026] Example 1
[0027] Preparation of Evening Primrose Seed Polysaccharide Extract
[0028] Choline chloride and glycerol were placed in an Erlenmeyer flask at a molar ratio of 1:2 and stirred in a constant temperature water bath at 65°C until a clear, transparent, viscous liquid was obtained. After cooling to room temperature, no crystals precipitated. Water was added to a moisture content of 30% to obtain DES. Dried evening primrose seeds were ground into powder. An appropriate amount of evening primrose seed powder was weighed and the prepared DES was added to it at a material-to-liquid ratio of 1:20. After mixing, the mixture was sonicated for 30 min at a power of 90 W at a temperature of 60°C. After sonication, the mixture was filtered, and the supernatant was retained. Anhydrous ethanol was added to the supernatant to a volume fraction of 80%. The mixture was allowed to stand at 4°C for 24 h and then subjected to further treatment at 20°C and 3600 r·min. -1 Centrifuge for 5 minutes, discard the supernatant to obtain the precipitate, remove ethanol and dry to obtain evening primrose seed polysaccharide extract.
[0029] In this embodiment, the yield of evening primrose seed polysaccharide was 18.35% ± 0.31%, which was 608.40% higher than that of conventional water extraction.
[0030] Example 2
[0031] Preparation of Evening Primrose Seed Polysaccharide Extract
[0032] Same as Example 1, except that glycerol is replaced with butanediol.
[0033] In this embodiment, the yield of evening primrose seed polysaccharide was 19.84% ± 0.12%, which was 665.92% higher than that obtained by conventional water extraction.
[0034] Example 3
[0035] Preparation of Evening Primrose Seed Polysaccharide Extract
[0036] Same as Example 1, except that glycerol is replaced with citric acid.
[0037] In this embodiment, the yield of evening primrose seed polysaccharide was 12.48% ± 0.15%, which was 381.85% higher than that of conventional water extraction.
[0038] Example 4
[0039] Preparation of Evening Primrose Seed Polysaccharide Extract
[0040] Same as Example 1, except that glycerol is replaced with D-sorbitol.
[0041] In this embodiment, the yield of evening primrose seed polysaccharide was 23.34% ± 0.03%, which was 801.16% higher than that of conventional water extraction. During the combined experiment, the D-sorbitol filtration process was relatively slow, and the separation efficiency between solvent and powder was low.
[0042] Example 5
[0043] Preparation of Evening Primrose Seed Polysaccharide Extract
[0044] Same as Example 1, except that glycerol is replaced with malonic acid.
[0045] In this embodiment, the yield of evening primrose seed polysaccharide was 16.22% ± 0.18%, which was 526.25% higher than that of conventional water extraction.
[0046] Example 6
[0047] Preparation of Evening Primrose Seed Polysaccharide Extract
[0048] Same as Example 1, except that 1-propanetriol is replaced with lactic acid.
[0049] In this embodiment, the yield of evening primrose seed polysaccharide was 17.05% ± 0.14%, which was 558.3% higher than that of conventional water extraction.
[0050] Example 7
[0051] Preparation of Evening Primrose Seed Polysaccharide Extract
[0052] Same as Example 1, except that glycerol is replaced with ethanol.
[0053] In this embodiment, the yield of evening primrose seed polysaccharide was 19.40% ± 0.18%, which was 649.03% higher than that of conventional water extraction.
[0054] Example 8
[0055] Preparation of Evening Primrose Seed Polysaccharide Extract
[0056] Same as Example 1, except that glycerol is replaced with 1,3-butanediol.
[0057] In this embodiment, the yield of evening primrose seed polysaccharide was 22.94% ± 0.06%, which was 781.16% higher than that obtained by conventional water extraction.
[0058] Example 9
[0059] Preparation of Evening Primrose Seed Polysaccharide Extract
[0060] Choline chloride, 1,3-butanediol, and butanediol were placed in an Erlenmeyer flask at a molar ratio of 1:2:0.5 and stirred in a constant temperature water bath at 65°C until a clear, transparent, viscous liquid was obtained. After cooling to room temperature, no crystals precipitated. Water was added to a moisture content of 30% to obtain DES. Dried evening primrose seeds were ground into powder. An appropriate amount of evening primrose seed powder was weighed and the prepared DES was added to it at a material-to-liquid ratio of 1:20. After mixing, the mixture was sonicated for 30 min at a power of 90 W at a temperature of 60°C. After sonication, the mixture was filtered, and the supernatant was retained. Anhydrous ethanol was added to the supernatant to a volume fraction of 80%. The mixture was allowed to stand at 4°C for 24 h and then extracted at 20°C and 3600 r·min. -1 Centrifuge for 5 minutes, discard the supernatant to obtain the precipitate, remove ethanol and dry to obtain evening primrose seed polysaccharide extract.
[0061] In this embodiment, the yield of evening primrose seed polysaccharide was 23.08% ± 0.13%, which was 791.25% higher than that of conventional water extraction.
[0062] Comparative Example 1
[0063] Grind dried evening primrose seeds into powder. Weigh an appropriate amount of the evening primrose seed powder and add ultrapure water at a material-to-liquid ratio of 1:30. Mix well and then sonicate for 30 min at a power of 90 W at a temperature of 60°C. After sonication, filter the mixture and retain the supernatant. Add anhydrous ethanol to the supernatant until the volume fraction reaches 80%. Let it stand at 4°C for 24 h, and then extract at 20°C and 3600 r·min.-1 Centrifuge for 5 min, discard the supernatant to obtain the precipitate, remove ethanol, and dry to obtain evening primrose seed polysaccharide extract. In this comparative example, the yield of evening primrose seed polysaccharide was only 2.59% ± 0.10%.
[0064] Method for determining polysaccharide content: Accurately weigh 10 mg of glucose, dissolve it in purified water at room temperature, transfer it to a 100 mL volumetric flask, and dilute to volume to prepare a 0.1 mg / mL solution. -1 Standard glucose solutions were prepared. 0, 0.2, 0.4, 0.6, 0.8, and 1.0 mL of glucose standard solution were placed in test tubes, respectively, and the volume was brought to 2.0 mL with distilled water. Then, 1 mL of 6% phenol solution and 5 mL of concentrated sulfuric acid solution were added. The mixtures were heated in a boiling water bath for 1 min, and the absorbance was measured. A standard curve was plotted with absorbance on the ordinate and concentration on the abscissa, and the polysaccharide content of the samples was calculated based on the standard curve.
[0065] Table 1. Comparison of the yield of evening primrose seed polysaccharides from different hydrogen bond donor extractants and water extraction.
[0066]
[0067] Example 10
[0068] Preparation of Evening Primrose Seed Polysaccharide Extract
[0069] Same as Example 9, except that the moisture content is changed to 10%.
[0070] In this embodiment, the yield of evening primrose seed polysaccharide was 15.47% ± 0.50%, which was 497.06% higher than that of conventional water extraction.
[0071] Example 11
[0072] Preparation of Evening Primrose Seed Polysaccharide Extract
[0073] Same as Example 9, except that the molar ratio of 1,3-butanediol and butanediol is changed to 2:1.
[0074] In this embodiment, the yield of evening primrose seed polysaccharide was 15.47% ± 0.50%, which was 497.06% higher than that of conventional water extraction.
[0075] Example 12
[0076] Preparation of Evening Primrose Seed Polysaccharide Extract
[0077] Same as Example 9, except that the molar ratio of 1,3-butanediol and butanediol is changed to 2:1, and the ultrasonic time is changed to 50 min.
[0078] In this embodiment, the yield of evening primrose seed polysaccharide was 21.22% ± 0.40%, which was 719.31% higher than that obtained by conventional water extraction.
[0079] Depend on Figure 3 As shown in Table 1, the polysaccharide yield is much lower when water is used as the extraction solvent than when DES is used. This is because the cell walls of evening primrose seeds are not fully opened, indicating that the water extraction method is insufficient for extracting polysaccharides from evening primrose seeds. Figure 1 and Figure 2 As shown, under the defined extraction conditions, the DES in this application completely opens the cell walls of evening primrose seeds. The DES can then interact with the cell walls of the evening primrose seeds, disrupting or dissolving them, allowing the extract to escape. Figure 1 and Figure 2 As can be seen, polysaccharides have larger pores and more typical vessel-like morphology, which promotes the dissolution of contents, thus indicating that the DES designed in this application has a higher and more thorough extraction degree.
[0080] The eutectic solvent DES of this application has hydrogen bond acceptors and hydrogen bond donors that are linked together by hydrogen bonds, exhibiting the excellent properties of ionic liquids. It is also characterized by non-toxicity, low volatility, and high thermal stability. Compared with water extraction, the high surface tension, high density, and high polarity of DES give it superior solubility and extractability for evening primrose seed polysaccharides. Different DES and their combinations significantly affect the extraction yield, purity, and activity of evening primrose seed polysaccharides. Compared with water extraction, it has a lower extraction temperature, shorter extraction time, and higher extraction rate.
Claims
1. A method for extracting evening primrose seed polysaccharides using an ultrasound-assisted eutectic solvent, characterized in that, Includes the following steps: (1) Preparation of eutectic solvent: The hydrogen bond acceptor and hydrogen bond donor are mixed in an Erlenmeyer flask and stirred under constant temperature water bath conditions until a clear and transparent viscous liquid is obtained. Water is added to mix and the eutectic solvent DES is obtained. (2) Sample preparation: The dried evening primrose seeds were ground into powder and stored at -20℃ for later use; (3) Ultrasonic-assisted extraction: Add the eutectic solvent DES to the evening primrose seed powder, mix well and then extract by ultrasonication; (4) After ultrasonication, filter the supernatant and add anhydrous ethanol to the supernatant until the volume fraction of anhydrous ethanol is 80%. Let it stand at 4~7℃, then centrifuge, discard the supernatant to obtain the precipitate, and dry the precipitate to obtain evening primrose seed polysaccharide extract. The eutectic solvent is a hydrogen bond acceptor and a hydrogen bond donor with a molar ratio of 1:(1~2); The hydrogen bond acceptor is choline chloride; The hydrogen bond donor is a 1,3-hydroxyl group with a molar ratio of 2:(0.5~1). Butylene glycol and butylene glycol; In step (1), the amount of water added is 30-50% of the mass of the eutectic solvent DES; in step (3), the ultrasonic time is 20-70 min and the ultrasonic power is 20-90 W.
2. The method according to claim 1, characterized in that, The constant temperature range in step (1) is 55~75℃.
3. The method according to claim 1, characterized in that, The ratio of evening primrose seed powder to eutectic solvent DES in step (3) is 1:(20~80)g / ml.
4. The method according to claim 1, characterized in that, The extraction temperature in step (3) is 30~80℃.
5. The method according to claim 1, characterized in that, The settling time mentioned in step (4) is 12~24 h.
6. The method according to claim 1, characterized in that, The centrifugation temperature in step (4) is 10~20℃, and the centrifugation speed is 3600~5000 r·min. -1 The centrifugation time is 5-10 min.