White paeony root polysaccharide with function of reducing blood fat as well as preparation method and application of white paeony root polysaccharide
By extracting and preparing white peony polysaccharide from white peony, the problem of large side effects of existing lipid-lowering drugs has been solved, and effective lipid-lowering effects have been achieved in cell and animal models, providing a safe and effective natural lipid-lowering drug solution.
Patent Information
- Application Number
- CN202511519562.1
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-23
- Publication Date
- 2026-02-27
AI Technical Summary
Existing lipid-lowering drugs have significant side effects and poor adherence, and long-term use may cause damage to the liver and kidneys.
Peony polysaccharides with lipid-lowering function were extracted and prepared from white peony. Through multiple heating extractions with ethanol and water, centrifugation, and purification with ion exchange resin, low-molecular-weight and high-molecular-weight white peony polysaccharides were obtained for the preparation of lipid-lowering drugs.
Paeonia lactiflora polysaccharide has shown significant lipid and cholesterol-lowering effects in cell and animal models, with high safety, making it suitable for the preparation of natural lipid-lowering drugs.
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Figure CN121574271A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of natural products, and in particular to a white peony polysaccharide with lipid-lowering function, its preparation method and application. Background Technology
[0002] White peony root has a long history of use in Traditional Chinese Medicine (TCM) for treating various diseases. For example, it is frequently used to treat gynecological conditions such as menstrual disorders and various gynecological inflammatory diseases. Furthermore, white peony root is also used to treat liver diseases; studies have shown that its total glycosides have a protective effect against liver damage in diabetic rats, possibly related to its inhibition of endoplasmic reticulum stress. In cardiovascular diseases, white peony root is also used; paeoniflorin is considered to have the potential to treat atherosclerosis, and its mechanism may be related to regulating inflammation and immune dysfunction. Besides these diseases, white peony root can also be used to treat abdominal pain and diarrhea.
[0003] The main active components of white peony root include monoterpenes and their glycosides, such as paeoniflorin and paeoniflorin lactone. Paeoniflorin is the main bioactive component of white peony root, possessing analgesic, anti-inflammatory, and sedative effects. In addition, white peony root also contains various phenolic compounds. The processing method of white peony root also significantly affects its efficacy and quality; different processing methods produce different effects.
[0004] The existing first-line lipid-lowering drugs are mainly statins (such as atorvastatin), fibrates (such as fenofibrate), and cholesterol absorption inhibitors (such as ezetimibe). Although they can quickly lower blood lipid levels, long-term use can easily cause serious side effects. Statins may cause elevated liver enzymes, muscle pain, or even rhabdomyolysis. Fibrates have the potential to damage kidney function, and some patients discontinue treatment due to intolerance to side effects, resulting in poor adherence. Summary of the Invention
[0005] The purpose of this invention is to overcome the shortcomings and deficiencies of the prior art and to provide a white peony polysaccharide with lipid-lowering function, its preparation method and application.
[0006] The objective of this invention is achieved through the following technical solution: A method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function includes the following steps: (1) White peony root was extracted with ethanol by heating, the ethanol extract was separated, and the white peony root was dried; (2) Add water to the dried white peony root, heat and extract, repeat twice, combine the water extracts and concentrate; (3) Add ethanol to the concentrated aqueous extract, stir, let stand, centrifuge, and take the supernatant; (4) adding ethanol into the supernatant, stirring, standing, centrifuging, taking the precipitate, drying, dissolving in water, adding ion exchange resin, collecting the leakage, pure water washing liquid and 0.1% sodium chloride washing liquid, combining, concentrating, freeze-drying to obtain the white peony root polysaccharide with the function of lowering blood lipid.
[0007] The adding amount of the ethanol in step (1) is 3-5 times of the mass of the white peony root.
[0008] The heating extraction condition in step (1) is 50-70℃ extraction for 5-20 min.
[0009] The adding amount of the water in step (2) is 3-5 times of the mass of the white peony root.
[0010] The heating extraction condition in step (2) is 50-70℃ extraction for 5-20 min.
[0011] The concentration in step (2) is concentrated to 1 / 10-1 / 20 of the original volume.
[0012] The adding amount of the ethanol in step (3) is 1-1.5 times of the water extract after concentration.
[0013] The centrifuging condition in step (3) is 4000-6000r / min, 5-15 min.
[0014] The adding amount of the ethanol in step (4) is 2-4 times of the supernatant.
[0015] The centrifuging condition in step (4) is 4000-6000r / min, 5-15 min.
[0016] The ion exchange resin in step (4) is DEAE weak anion exchange resin.
[0017] The white peony root polysaccharide with the function of lowering blood lipid is prepared by the above preparation method.
[0018] The application of the above white peony root polysaccharide with the function of lowering blood lipid in the preparation of blood lipid lowering drugs.
[0019] The application of the above white peony root polysaccharide with the function of lowering blood lipid in the preparation of high blood lipid treatment drugs.
[0020] The present application has the following advantages and effects relative to the prior art: This invention is the first to extract a peony polysaccharide with lipid-lowering function from peony root. Cell experiments have verified that the polysaccharide can reduce lipid production in a hyperlipidemic cell model. Further animal model experiments have shown that the polysaccharide can also achieve lipid-lowering effects in hyperlipidemic animal models, proving that the polysaccharide can be used to prepare lipid-lowering drugs. Moreover, the polysaccharide is a natural component with high safety and broad application prospects. Attached Figure Description
[0021] Figure 1 This is an image showing the Oil Red O staining results of the hyperlipidemia cell model experiment in Example 2.
[0022] Figure 2 This is a graph showing the total triglyceride (TG) detection results in the hyperlipidemia cell model experiment in Example 2.
[0023] Figure 3 This is a graph showing the total cholesterol (TC) detection results from the hyperlipidemia cell model experiment in Example 2.
[0024] Figure 4 This is the chromatogram of the standard used in the polysaccharide characterization experiment in Example 3.
[0025] Figure 5 This is a chromatogram of the sample from the polysaccharide characterization experiment in Example 3.
[0026] Figure 6 This is a graph showing the changes in blood lipid-related indicators in Example 4.
[0027] Figure 7 This is a photograph of a section of liver tissue stained with Red O in Example 4. Detailed Implementation
[0028] The present invention will be further described in detail below with reference to the embodiments and accompanying drawings, but the embodiments of the present invention are not limited thereto.
[0029] Unless otherwise specified in the following implementation plan, the test conditions are generally as per standard test conditions or the test conditions recommended by the reagent company. Unless otherwise specified, all materials and reagents used are commercially available.
[0030] Example 1 1.1 Preparation of low molecular weight polysaccharides from Paeonia lactiflora Take 100 grams of white peony root, add 3-5 times the amount of 95% ethanol, heat to 60°C and extract for 10 minutes. The alcohol-containing liquid is used to extract paeoniflorin and other components. Dry the white peony root, add 5 times the amount of pure water, heat to 60°C and extract for 10 minutes, repeat 2 times. Concentrate the water extract to 50 ml, add 1.5 times the amount of 95% ethanol, stir, let stand and centrifuge for 10 minutes at 5000 r / min, discard the precipitate and take the supernatant. Add 3 times the amount of 95% ethanol to the supernatant, stir, let stand and centrifuge for 10 minutes at 5000 r / min, take the precipitate (containing medium and small molecular weight polysaccharides). Dry the precipitate at 40°C, dissolve in 10 ml of pure water, load onto a glass column (1.5 cm in diameter and 30 cm in height) containing 50 ml of DEAE-52 weak anion exchange resin, take the leakage liquid and 50 ml of pure water rinse liquid and 50 ml of 0.1% sodium chloride rinse liquid, combine, concentrate and freeze-dry to obtain white peony root low molecular polysaccharides.
[0031] 1.2 Preparation of white peony root high molecular polysaccharides Take 100 grams of white peony root, add 3-5 times the amount of 95% ethanol, heat to 60°C and extract for 10 minutes. The alcohol-containing liquid is used to extract paeoniflorin and other components. Dry the white peony root, add 5 times the amount of pure water, heat to 60°C and extract for 10 minutes, repeat 2 times. Concentrate the water extract to 50 ml, add 1 times the amount of 95% ethanol, stir, let stand and centrifuge for 10 minutes at 5000 r / min, take the precipitate (containing high molecular weight polysaccharides), dry the precipitate at 40°C, dissolve in 10 ml of pure water, load onto a glass column (1.5 cm in diameter and 30 cm in height) containing 50 ml of DEAE-52 weak anion exchange resin, take the leakage liquid and 50 ml of pure water rinse liquid and 50 ml of 0.1% sodium chloride rinse liquid, combine, concentrate and freeze-dry to obtain white peony root high molecular polysaccharides.
[0032] Example 2 High-fat cell model experiment 2.1 Model establishment Take HepG2 cells in the logarithmic growth phase to simulate liver cells, inoculate 5x10 5 cells per well in a six-well plate. After the cells are completely adherent, add oleic acid at a concentration of 100 umol / L (0.1 mmol / L) and palmitic acid at a concentration of 50 umol / L (0.05 mmol / L) to the model group for lipid induction for 24 hours. When the cells become round and there are bright lipid droplets in the middle, it indicates that the model has been successfully established.
[0033] 2.2 Experimental setup After the model was established, six groups of normal, model, high concentration of high molecular weight polysaccharide from Radix Paeoniae Alba (300 ug / ml), low concentration of high molecular weight polysaccharide from Radix Paeoniae Alba (100 ug / ml), high concentration of low molecular weight polysaccharide from Radix Paeoniae Alba (300 ug / ml), low concentration of low molecular weight polysaccharide from Radix Paeoniae Alba (100 ug / ml) were set. After 24 hours of action, the supernatant and cell layer were collected, and oil red O staining was used to show the intracellular lipid storage. Then the cells were collected, ultrasonically broken in ice bath, centrifuged at 6000 r / min for 10 min, and the supernatant was taken to detect total triglyceride (TG) and total cholesterol (TC) using the kit from Nanjing Jiancheng.
[0034] 2.3 Experimental results The experimental results are shown in Figures 1-3 It can be seen that the low molecular component extracted from the polysaccharide of Radix Paeoniae Alba has better effect than the high molecular component, and has more significant effect on reducing TG and TC in the cell model. Therefore, the low molecular component is further characterized and researched.
[0035] Example 3 Characterization of polysaccharide composition (1) Determination of monosaccharide composition The chromatography system used is Thermo ICS 5000+ ion chromatography system (ICS 5000+, Thermo Fisher Scientific, USA), which uses an electrochemical detector to analyze and detect monosaccharide components.
[0036] Dionex™ CarboPac™ PA20 (150*3.0mm, 10μm) liquid chromatography column was used; the injection volume was 5μl. The mobile phase A (H2O), the mobile phase B (0.1M NaOH), the mobile phase C (0.1M NaOH, 0.2M NaAc), the flow rate was 0.5ml / min; the column temperature was 30℃; the elution gradient was: 0min A phase / B phase / C phase (95:5:0, V / V), 26min A phase / B phase / C phase (85:5:10, V / V), 42min A phase / B phase / C phase (85:5:10, V / V), 42.1min A phase / B phase / C phase (60:0:40, V / V), 52min A phase / B phase / C phase (60:40:0, V / V), 52.1min A phase / B phase / C phase (95:5:0, V / V), 60min A phase / B phase / C phase (95:5:0, V / V).
[0037] (2) Determination of polysaccharide molecular weight The chromatography system adopts a gel chromatography-differential-multiple angle laser light scattering system, the liquid phase system is U3000 (Thermo, USA), the differential detector is Optilab T-rEX (Wyatt technology, CA, USA), and the laser light scattering detector is DAWN HELEOS II (Wyatt technology, CA, USA). The specific chromatographic column and elution conditions are as follows: A gel exclusion chromatography column Ohpak SB-805 HQ (300x8mm) and Ohpak SB-803 HQ (300x8mm) are used in series. The column temperature is 45℃, the sample injection amount is 100 μL, the mobile phase A (0.02% NaN3, 0.1M NaNO3) has a flow rate of 0.6mL / min, and the elution gradient is isocratic for 75min. The gel chromatography can elute the molecules in the solvent according to the weight or size, and the larger molecules can only enter the larger gel pores, while the smaller molecules can enter more gel particles. Thus, the larger molecules move a shorter distance in the gel bed, and the smaller molecules move a longer distance. The larger molecules are eluted first, and the smaller molecules are eluted later, thereby achieving the separation effect and the purpose of separating according to the molecular size. Then, the differential detector detects the concentration information of the sample according to the refractive intensity, the multi-angle laser light scattering instrument detects the light scattering information of the macromolecules, and the molecular weight corresponding to each component is calculated according to the Mark-Houwink Equation.
[0038] (3) Test results Through detection, the average molecular weight MP of the low molecular weight polysaccharide in white peony root is 3000, and combined with the fact that the monosaccharide in the previous sample is mainly glucose, it can be known that the white peony root polysaccharide obtained by extraction and purification this time is a glucan.
[0039] Table 1 Content detection results of different monosaccharides in white peony root polysaccharide Sample name Fuc (ug / mg) Ara (ug / mg) Rha (ug / mg) Gal (ug / mg) Glc (ug / mg) Xyl (ug / mg) Man (ug / mg) Baishao polysaccharide 0 9.7249 2.1706 8.9892 387.2421 0 2.4569 Sample name Fru (ug / mg) Rib (ug / mg) Gal-UA (ug / mg) Gul-UA (ug / mg) Glc-UA (ug / mg) Man-UA (ug / mg) Sum of each component content Baishao polysaccharide 0 0 0 0 0 0 410.5838 Example 4 Verification of the effect of white peony root polysaccharide on reducing blood lipids 4.1 Animal grouping 48 C57BL / 6 mice aged 6-8 weeks and weighing 16-18 g were selected, and after adaptive feeding for 1 week, they were randomly divided into normal group, model group, and white peony root polysaccharide group (white peony root low molecular weight polysaccharide aqueous solution 100mg / kg) according to the body weight, 6 in each group.
[0040] 4.2 Modeling and drug administration Each group was fed regular feed, distilled water, and peony polysaccharide daily. Administration was administered via gavage for 5 days. On the third day of administration, except for the normal control group, the model group and the administered group underwent intramuscular injection of Triton WR-1339 (10 mL / kg) to establish the model. Mice were fasted for 12 hours prior to sample collection. Two hours after the last administration, blood samples were collected from the mice's eyeballs, centrifuged at 3000 rpm for 15 minutes, and then used for serum index testing. The mice were dissected, and their livers were collected, washed with physiological saline, and weighed. A portion of the liver was placed in 4% histocellular fixative for HE and Oil Red O staining; another portion was placed in 1.5 mL EP tubes for relevant index detection. Daily mouse weight was recorded during model establishment and administration.
[0041] 2.3 Experimental Results Experimental results are as follows Figures 6-7 As shown, where Figure 6 The results of the study on changes in blood lipid-related indicators showed that peony polysaccharide could significantly reduce total cholesterol, triglycerides, and low-density lipoprotein, while also reducing liver AST and ALT, suggesting a protective effect on liver cells.
[0042] Figure 7 Hematoxylin and eosin (HE) stained sections of liver tissue showed that, due to the dissolution of lipid droplets by organic solvents such as xylene during paraffin sectioning, round or oval transparent vacuoles appeared in the cytoplasm of hepatocytes. The proportion of hepatocytes containing vacuoles was significantly higher in the model group than in the normal group, and the arrangement of liver lobules was disordered. Treatment with Paeonia lactiflora polysaccharide significantly reduced these symptoms. Oil Red O, as a lipid-soluble dye, can directly bind to neutral fats (triglycerides), making the lipid droplets in hepatocytes appear red. The red staining area was significantly higher in the model group than in the normal group, but this was significantly reduced after treatment with Paeonia lactiflora polysaccharide.
[0043] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.
Claims
1. A method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function, characterized in that... Includes the following steps: (1) White peony root was extracted with ethanol by heating, the ethanol extract was separated, and the white peony root was dried; (2) Add water to the dried white peony root, heat and extract, repeat twice, combine the water extracts and concentrate; (3) Add ethanol to the concentrated aqueous extract, stir, let stand, centrifuge, and take the supernatant; (4) Add ethanol to the supernatant, stir, let stand, centrifuge, take the precipitate, dry, add water to dissolve, add ion exchange resin, collect the leakage liquid, pure water rinsing liquid and 0.1% sodium chloride rinsing liquid, combine, concentrate, freeze dry, and obtain white peony polysaccharide with lipid-lowering function.
2. The method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function according to claim 1, characterized in that: The amount of ethanol added in step (1) is 3 to 5 times the mass of white peony root; The heating extraction conditions described in step (1) are 50-70℃ for 5-20 minutes.
3. The method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function according to claim 1, characterized in that: The amount of water added in step (2) is 3 to 5 times the mass of the white peony root; The heating extraction conditions described in step (2) are 50-70℃ for 5-20 minutes; The concentration mentioned in step (2) is to concentrate to 1 / 10 to 1 / 20 of the original volume.
4. The method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function according to claim 1, characterized in that: The amount of ethanol added in step (3) is 1 to 1.5 times that of the concentrated water extract; The centrifugation conditions described in step (3) are 4000-6000 r / min for 5-15 min.
5. The method for preparing Paeonia lactiflora polysaccharide with lipid-lowering function according to claim 1, characterized in that: The amount of ethanol added in step (4) is 2 to 4 times that of the supernatant; The centrifugation conditions described in step (4) are 4000–6000 r / min for 5–15 min; The ion exchange resin mentioned in step (4) is a DEAE weak anion exchange resin.
6. A white peony polysaccharide with lipid-lowering function, characterized in that: It is prepared by any of the preparation methods described in claims 1 to 5.
7. The use of the white peony polysaccharide with lipid-lowering function as described in claim 6 in the preparation of lipid-lowering drugs.
8. The use of the white peony polysaccharide with lipid-lowering function as described in claim 6 in the preparation of drugs for treating hyperlipidemia.