Lithocarpus litseifolius modified acidic polysaccharide as well as preparation method and application thereof

Abstract

The invention provides modified acidic polysaccharide of lithocarpus litseifolius. The modified acidic polysaccharide is prepared from natural acidic polysaccharide of lithocarpus litseifolius through a chemical method or an enzyme method. Every 100 mg of the modified acidic polysaccharide of lithocarpus litseifolius contains 89.75 mg + / -1.04 mg to 90.67 mg + / -1.46 mg of total polysaccharide. The total uronic acid content is 23.70 mg + / -1.58 mg to 71.74 mg + / -2.07 mg; total bound phenol: 2.06 mg of GAE + / -0.06 mg of GAE-4.56 mg of GAE + / -0.04 mg of GAE (equivalent weight of gallic acid); the esterification degree is 3.72% + / -0.90% to 34.98% + / -0.62%; the molecular weight is (0.562 + / -0.012) * 10 < 4 > Da-(4.268 + / -0.055) * 10 < 4 > Da; the proportion of the galacturonic acid glycan (HG) is 15.28% to 65.02%; the ratio of the type I rhamnogalacturonan (RG-I) is 28.47% to 72.37%; and the length of a side chain is 0.44 to 10.31. The invention further provides a preparation method and application of the lithocarpus litseifolius modified acidic polysaccharide. The lithocarpus litseifolius modified acidic polysaccharide prepared by the invention has remarkable immunoregulatory activity, and especially has better immunocompetence when being used for removing arabinose side chains and reducing galactose side chains.

Description

Technical Field

[0001] The invention relates to a modified acidic polysaccharide of Artemisia selengensis leaves and belongs to the field of health-care foods or medicines. Background Art

[0002] Immunity is a physiological process that helps the body eliminate pathogens (bacteria, viruses, etc.) and tumor cells and maintain the body's health. It is of indispensable importance to the physiological health of the human body. Polysaccharides are considered to be natural immunomodulators or immunopotentiators because of their significant immunostimulatory activity. Polysaccharides can play an immunomodulatory role by activating immune cell surface receptors, promoting the release of cytokines, enhancing the proliferation and differentiation of immune cells, strengthening the functions of phagocytes, natural killer cells, and lymphocytes, and promoting the growth of immune organs and the secretion of immunoglobulins, thereby improving the body's immunity.

[0003] Ligusticum chuanxiong is an evergreen tree belonging to the genus Ligusticum, Fagaceae, Fagaceae, also known as sweet tea, and is widely distributed in southern China. Ligusticum chuanxiong has three functions: tea, sugar, and medicine. It has long been used to prevent and treat diabetes. It will be listed as a new food material in 2017, opening up its application in health foods and ordinary foods. Ligusticum chuanxiong contains a variety of bioactive compounds, such as polysaccharides, phenolic acids, and flavonoids, which have potential pharmacological effects. Ligusticum chuanxiong polysaccharide is one of the most important bioactive ingredients of Ligusticum chuanxiong, with significant antioxidant, anti-inflammatory, anti-diabetic and immune-regulating effects, and has broad development prospects in the field of functional health foods.

[0004] Fu Mengqian, Extraction optimization, structural characterization and in vitro bioactivity evaluation of polysaccharides from Ligusticum chuanxiong, Sichuan Agricultural University, June 2023. This article reports the conditions of pressurized hot water-assisted extraction technology and microwave-assisted low eutectic solvent extraction technology of polysaccharides from Ligusticum chuanxiong, and analyzes the structural characteristics of the purified Ligusticum chuanxiong polysaccharides. The extracted Ligusticum chuanxiong polysaccharides showed good in vitro antioxidant activity (ABTS free radical scavenging activity, DPPH free radical scavenging activity and NO free radical scavenging activity, as well as total reducing power), anti-glycosylation activity, hypoglycemic activity (α-glucosidase inhibitory activity), immunomodulatory activity and in vitro microbial beneficial activity. Patent application number 202110615589.6, invention name: Sweet tea polysaccharide extract with high antioxidant activity and extraction method thereof; The present invention discloses a sweet tea polysaccharide extract with high antioxidant activity and an extraction method thereof. The present invention adopts a low eutectic solvent-microwave assisted extraction method to extract polysaccharides from sweet tea, effectively improving the solvent's solubility for polysaccharides, achieving efficient extraction of polysaccharides and the prepared sweet tea polysaccharides having high antioxidant activity. It has a series of advantages such as high extraction efficiency, low extraction cost, simple extraction process, stable and reliable, and the extracted sweet tea polysaccharides having high antioxidant activity. Summary of the invention

[0005] The present invention, on the basis of extracting natural Zinnia zingiberensis acidic polysaccharides with a low eutectic solvent, modifies the natural Zinnia zingiberensis acidic polysaccharides (chemical and enzymatic modification), and compares the in vitro immunomodulatory activity of Zinnia zingiberensis acidic polysaccharides before and after modification, which can clearly affect the key effector structure and potential mechanism of the in vitro immunomodulatory effect of Zinnia zingiberensis acidic polysaccharides, and is conducive to more fully utilizing Zinnia zingiberensis resources, and providing a reference for the exploration of the immunomodulatory effect and mechanism of Zinnia zingiberensis polysaccharides, and providing theoretical and technical support for the development of Zinnia zingiberensis polysaccharide-related functional foods. Compared with natural Zinnia zingiberensis acidic polysaccharides, the modified Zinnia zingiberensis acidic polysaccharides have better in vitro immune activity.

[0006] The invention provides a modified acidic polysaccharide from the leaves of Alpinia zingiberensis, and also provides a preparation method and application of the modified acidic polysaccharide from the leaves of Alpinia zingiberensis.

[0007] The present invention provides a modified acidic polysaccharide from the leaves of the elm, which is a modified acidic polysaccharide prepared from natural acidic polysaccharide from the leaves of the elm by a chemical method or an enzymatic method;

[0008] The total sugar content of the modified acidic polysaccharide of Artemisia selengensis leaf per 100 mg is: 89.75 mg ± 1.04 mg–90.67 mg ± 1.46 mg; total uronic acid: 23.70 mg ± 1.58 mg–71.74 mg ± 2.07 mg; total protein: 1.53 mg ± 0.02 mg–3.16 mg ± 0.07 mg; total bound phenol: 2.06 mg GAE ± 0.06 mg GAE–4.56 mg GAE ± 0.04 mg GAE (gallic acid equivalent); degree of esterification: 3.72% ± 0.90%–34.98% ± 0.62%;

[0009] Molecular weight w :(0.562±0.012)×10 4 Da–(4.268±0.055)×10 4 Da; dispersion coefficient M w / M n : 1.163–1.729;

[0010] The sugar components of the modified acidic polysaccharide from the leaves of Artemisia selengensis mainly include the following monosaccharides: galacturonic acid, galactose, arabinose, rhamnose, xylose, glucose, glucuronic acid, and mannose. The molar percentages of each monosaccharide are: (21.16%–70.56%): (17.26%–33.14%): (0.14%–27.48%): (5.53%–7.29%): ( 1.36%–6.85%):(1.71%–2.27%):(1.79%–2.89%):(0.83%–1.21%); among them, galacturonan (HG) accounted for 21.94%–65.02%, type I rhamnogalacturonan (RG-Ⅰ) accounted for 28.47%–72.37%, and the side chain length was 0.44–10.31.

[0011] More preferably,

[0012] The total polysaccharide content of modified acidic polysaccharide from Artemisia selengensis leaf is 89.75mg±1.04mg–90.09mg±1.07mg per 100mg; total uronic acid: 49.57mg±0.94mg–71.74mg±2.07mg; total protein: 3.03mg±0.07mg–3.16mg±0.07mg; total bound phenol: 2.06mg GAE±0.06mg GAE–2.69mg GAE±0.10mg GAE (gallic acid equivalent); degree of esterification: 7.74%±0.40%–14.54%±0.84%;

[0013] Molecular weight w :(0.562±0.012)×10 4 Da–(1.317±0.018)×10 4 Da; dispersion coefficient M w / M n : 1.163–1.183;

[0014] The sugar components of the modified acidic polysaccharide from the leaves of Artemisia selengensis mainly include the following monosaccharides: galacturonic acid, galactose, arabinose, rhamnose, xylose, glucose, glucuronic acid, and mannose. The molar percentages of each monosaccharide are: (55.89%–70.56%): (17.26%–28.99%): (0.14%–0.47%): (5.53%–6.46%): ( 1.36%–2.40%):(1.71%–1.78%):(2.22%–2.89%):(1.12%–1.21%); among them, galacturonan (HG) accounted for 49.44%–65.02%, type I rhamnogalacturonan (RG-Ⅰ) accounted for 28.47%–42.37%, and the side chain length was 0.44–0.86.

[0015] Wherein, the natural Zinnia zingiberensis acidic polysaccharide is prepared by a low eutectic solvent-assisted heating extraction method;

[0016] The chemically modified acidic polysaccharide is an acidic polysaccharide prepared by modification with sodium hydroxide, hydrochloric acid and trifluoroacetic acid; the enzymatically modified acidic polysaccharide is an acidic polysaccharide prepared by modification with pectinase derived from Aspergillus aculeatus.

[0017] The present invention also provides a method for preparing the modified acidic polysaccharide from Ligusticum chuanxiong, which comprises the following steps:

[0018] a. Using deep eutectic solvent to assist heating to extract natural acidic polysaccharides from Zingiber officinale;

[0019] b. The natural acidic polysaccharide of Zingiber officinale prepared in step a is modified by chemical or enzymatic methods.

[0020] Wherein, the method for modifying polysaccharides in step b is:

[0021] 0.5% (w / v) of natural Ziziphus truncatula acidic polysaccharide was added to a sodium hydroxide solution with a pH value of 12.0, and stirred at 4°C for 30 min. Subsequently, hydrochloric acid (1 M) was added until the pH value was 7.0, and the solution was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0 kDa and freeze-dried to obtain a Ziziphus truncatula modified acidic polysaccharide (STP-AD) with esterification removed.

[0022] 0.5% (w / v) of natural Ziziphus truncatula acidic polysaccharide was added to 0.5M trifluoroacetic acid solution to make the final concentration of trifluoroacetic acid in the system 0.25M, and the mixture was reacted at 95°C for 1.5h and 4h, respectively. Subsequently, sodium hydroxide solution (1M) was added until the pH value was 7.0, and the mixture was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0kDa and freeze-dried to obtain Ziziphus truncatula modified acidic polysaccharide (STP-AH1) with arabinose side chains removed and Ziziphus truncatula modified acidic polysaccharide (STP-AH2) with arabinose side chains removed and galactose side chains reduced, respectively; wherein, under the condition of reaction for 1.5h, the Ziziphus truncatula modified acidic polysaccharide (STP-AH1) with arabinose side chains removed was prepared; under the condition of reaction for 4h, the Ziziphus truncatula modified acidic polysaccharide (STP-AH2) with arabinose side chains removed and galactose side chains reduced was prepared;

[0023] 0.5% (w / v) of natural Artemisia selengensis acidic polysaccharide was added to 0.3U / mL pectinase solution, reacted at 40°C for 6h, inactivated, centrifuged and the supernatant was taken; the supernatant was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0kDa and freeze-dried to obtain Artemisia selengensis modified acidic polysaccharide (STP-E) with reduced galacturonic acid polysaccharide ratio.

[0024] The present invention also provides the use of the modified acidic polysaccharide of Artemisia selengensis in preparing health food that helps to enhance immunity.

[0025] The present invention also provides the use of the modified acidic polysaccharide of Ligusticum chuanxiong in preparing a medicine with immunomodulatory effect.

[0026] Polysaccharides are a class of biologically active substances with complex and diverse structures. Polysaccharides with different structures exhibit different biological activities. Plant acidic polysaccharides are mainly pectin polysaccharides, and the main structural domains of pectin polysaccharides are galacturonan (HG) and type I rhamnogalacturonan (RG-I). A large number of studies have shown that the HG / RG I ratio, molecular weight, branch length, degree of esterification, glycosidic bonds and constituent monosaccharides of pectin polysaccharides are the key chemical structures of polysaccharides with health-care effects, and these structures are closely related to their biological activities. Different modification methods will produce different degrees of substitution, and the effects on activity are also different, which can make the modified polysaccharides exhibit different physicochemical properties and biological activities. The present invention can clarify the key effect structure and potential mechanism of the in vitro immunomodulatory effect of the acidic polysaccharide of the leaf branch of the ginger. The natural ginger leaf branch acidic polysaccharide of the present invention has an immune-enhancing effect, and the natural ginger leaf branch acidic polysaccharide is modified and modified, and the structural characteristics of the modified acidic polysaccharide of the leaf branch of the ginger and its immunostimulatory effect and potential mechanism on RAW264.7 cells are analyzed. The modified acidic polysaccharide of Zingiber officinale prepared by the invention has significant immunomodulatory activity, especially the modified acidic polysaccharide of Zingiber officinale with arabinose side chains removed and galactose side chains reduced has better immunomodulatory activity. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 High performance gel exclusion chromatogram of modified acidic polysaccharide from Ligusticum chuanxiong (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural acidic polysaccharide from Ligusticum chuanxiong, modified acidic polysaccharide from Ligusticum chuanxiong with esterification removed, modified acidic polysaccharide from Ligusticum chuanxiong with arabinose side chain removed, modified acidic polysaccharide from Ligusticum chuanxiong with arabinose side chain removed and galactose side chain reduced, and modified acidic polysaccharide from Ligusticum chuanxiong with reduced galacturonic acid polysaccharide ratio, respectively);

[0028] Figure 2 Monosaccharide composition chromatogram of modified acidic polysaccharide from Ligusticum chuanxiong (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural acidic polysaccharide from Ligusticum chuanxiong, modified acidic polysaccharide from Ligusticum chuanxiong with esterification removed, modified acidic polysaccharide from Ligusticum chuanxiong with arabinose side chain removed, modified acidic polysaccharide from Ligusticum chuanxiong with arabinose side chain removed and galactose side chain reduced, and modified acidic polysaccharide from Ligusticum chuanxiong with reduced galacturonic acid polysaccharide ratio, respectively);

[0029] Figure 3Fourier transform infrared spectra of modified acidic polysaccharides from Ligusticum chuanxiong (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural acidic polysaccharides from Ligusticum chuanxiong, modified acidic polysaccharides from Ligusticum chuanxiong with esterification removed, modified acidic polysaccharides from Ligusticum chuanxiong with arabinose side chains removed, modified acidic polysaccharides from Ligusticum chuanxiong with arabinose side chains removed and galactose side chains reduced, and modified acidic polysaccharides from Ligusticum chuanxiong with reduced galacturonic acid polysaccharides, respectively);

[0030] Figure 4 Modified acidic polysaccharide from Pinellia ternata 1 H NMR spectra (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural Z. zingiberensis acidic polysaccharide, Z. zingiberensis modified acidic polysaccharide with esterification removed, Z. zingiberensis modified acidic polysaccharide with arabinose side chain removed, Z. zingiberensis modified acidic polysaccharide with arabinose side chain removed and galactose side chain reduced, and Z. zingiberensis modified acidic polysaccharide with reduced galacturonic acid polysaccharide ratio, respectively);

[0031] Figure 5 Modified acidic polysaccharide from Pinellia ternata 13 C NMR spectra (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural Z. zingiberensis acidic polysaccharide, Z. zingiberensis modified acidic polysaccharide with esterification removed, Z. zingiberensis modified acidic polysaccharide with arabinose side chain removed, Z. zingiberensis modified acidic polysaccharide with arabinose side chain removed and galactose side chain reduced, and Z. zingiberensis modified acidic polysaccharide with reduced galacturonic acid polysaccharide ratio, respectively);

[0032] Figure 6 Effects of modified acidic polysaccharides from Ligusticum wallichii on cytotoxicity (A), production of nitric oxide (NO) (B), production of tumor necrosis factor-α (TNF-α) (C), and production of interleukin-6 (IL-6) (D) in RAW264.7 macrophages (Note: STP, STP-AD, STP-AH1, STP-AH2, and STP-E represent natural acidic polysaccharides from Ligusticum wallichii, modified acidic polysaccharides from Ligusticum wallichii with esterification removed, modified acidic polysaccharides from Ligusticum wallichii with arabinose side chains removed, modified acidic polysaccharides from Ligusticum wallichii with arabinose side chains removed and galactose side chains reduced, and modified acidic polysaccharides from Ligusticum wallichii with reduced galacturonic acid polysaccharides; error bars represent standard deviations; significant (p<0.05) differences between different samples are indicated by data with different letters; significant differences between the tested samples and the blank control are indicated by *p<0.05 and **p<0.01);

[0033] Figure 7Effect of modified acidic polysaccharide from Ligusticum chuanxiong on RAW The effects of 264.7 on the production of nitric oxide (NO) (A), tumor necrosis factor-α (TNF-α) (B) and interleukin-6 (IL-6) (C) secretion in macrophages, as well as the effects of modified acidic polysaccharides from Ligusticum chuanxiong on protein expression (D) (Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural acidic polysaccharides from Ligusticum chuanxiong, modified acidic polysaccharides from Ligusticum chuanxiong with esterification removed, modified acidic polysaccharides from Ligusticum chuanxiong with arabinose side chains removed, modified acidic polysaccharides from Ligusticum chuanxiong with arabinose side chains removed and galactose side chains reduced, and modified acidic polysaccharides from Ligusticum chuanxiong with reduced galacturonic acid polysaccharides, respectively; error bars represent standard deviations; significant (p<0.05) differences between different samples are represented by data with different letters; significant differences between the tested samples and the blank control are represented by *p<0.05 and **p<0.01). DETAILED DESCRIPTION

[0034] Example 1 Preparation of modified acidic polysaccharide from Ligusticum chuanxiong

[0035] 1. Preparation of natural acidic polysaccharide from Zingiber officinale:

[0036] a. Freeze-dry the harvested Litsea cubeba leaves, grind into powder and sieve:

[0037] b. Degreasing: Degrease the powder of Ligusticum chuanxiong leaves with 80% ethanol, centrifuge and discard the supernatant to obtain the precipitate;

[0038] c. Low eutectic solvent-assisted extraction: Use a low eutectic solvent with a water content of 30% to assist in heating extraction, centrifuge, and reserve the supernatant;

[0039] d. Alcohol precipitation: Use 75% (v / v) ethanol for fractionation precipitation, then centrifuge and collect the precipitate;

[0040] e. Redissolve: Redissolve the precipitate in step d with ultrapure water;

[0041] f. Starch removal: Use α-amylase and saccharifying enzyme to remove starch in the complex solution, centrifuge, and keep the supernatant for later use;

[0042] g. Ultrafiltration: ultrafiltration the supernatant, collect the fractions with molecular weight between 3 kDa and 100 kDa with the ultrafiltration membrane, and freeze-dry them to obtain natural S. zingiberensis acidic polysaccharide (STP).

[0043] 2. Preparation of deesterified modified acidic polysaccharide from Ligusticum chuanxiong using low temperature alkaline deesterification technology:

[0044] a. Take 0.5% (w / v) of the natural Zingiber officinale acidic polysaccharide described in 1 and add it to a sodium hydroxide solution with a pH value of 12.0, and stir at 4°C for 30 minutes;

[0045] b. Add hydrochloric acid (1 M) to the solution in step a until the pH value is 7.0, and ultrafilter through an ultrafiltration membrane with a molecular weight limit of 3.0 kDa and freeze-dry to obtain a modified acidic polysaccharide of S. truncatula (STP-AD) with esterification removed.

[0046] 3. Preparation of modified acidic polysaccharides of Ligusticum wallichii with arabinose side chains removed and galactose side chains reduced by controlled different acid hydrolysis technologies:

[0047] a. Take 0.5% (w / v) of the natural Zingiber officinale acidic polysaccharide described in 1 and add 0.5M trifluoroacetic acid solution to make the final concentration of trifluoroacetic acid in the system 0.25M, and react at 95°C for 1.5h and 4h respectively;

[0048] b. Add sodium hydroxide solution (1M) to the solution of step a until the pH value is 7.0, and ultrafiltration and freeze drying are performed through an ultrafiltration membrane with a molecular weight limit of 3.0 kDa to obtain a modified acidic polysaccharide (STP-AH1) of the leaves of the ginger with arabinose side chains removed and a modified acidic polysaccharide (STP-AH2) of the leaves of the ginger with arabinose side chains removed and galactose side chains reduced. Wherein, under the condition of 1.5h of reaction, the modified acidic polysaccharide (STP-AH1) of the leaves of the ginger with arabinose side chains removed is prepared; under the condition of 4h of reaction, the modified acidic polysaccharide (STP-AH2) of the leaves of the ginger with arabinose side chains removed and galactose side chains reduced is prepared.

[0049] 4. Preparation of modified acidic polysaccharide of Ligustrum lucidum with reduced galacturonic acid polysaccharide ratio by directional enzyme cleavage technology a. Take 0.5% (w / v) of natural Ligustrum lucidum acidic polysaccharide described in 1 and add it to 0.3U / mL pectinase solution, react at 40°C for 6h, inactivate, centrifuge and take the supernatant;

[0050] b. The supernatant was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0 kDa and freeze-dried to obtain a galacturonic acid polysaccharide modified from S. zingiberensis leaves (STP-E).

[0051] Example 2 Characterization of the physicochemical properties of the modified acidic polysaccharide of Ligusticum chuanxiong

[0052] The total polysaccharide, total uronic acid, total bound phenol and total protein contents of modified acidic polysaccharide of Ligusticum chuanxiong were determined by colorimetry. The molecular weight and dispersity of modified acidic polysaccharide of Ligusticum chuanxiong were determined by SEC-MALLS-RID (Wyatt Technology Co., Santa Barbara, CA, USA). The monosaccharide composition of modified acidic polysaccharide of Ligusticum chuanxiong was determined by high performance liquid chromatography (L-20A, Shimadzu, Japan) combined with 1-phenyl-3-methyl-5-pyrazolone (PMP) pre-column derivatization. The functional groups and degree of esterification of modified acidic polysaccharide of Ligusticum chuanxiong were analyzed by Fourier transform infrared spectroscopy (PerkinElmer, Waltham, MA, USA). The glycosidic bonds of modified acidic polysaccharide of Ligusticum chuanxiong were analyzed by nuclear magnetic resonance spectroscopy (Bruker, Rheinstetten, Germany). Specific method reference: Author: Ding-Tao Wu, Meng-Xi Fu, Huan Guo, Yi-Chen Hu, Xiao-Qin Zheng, Ren-You Gan and Liang Zou; Title: Microwave-Assisted Deep Eutectic Solvent Extraction, Structural Characteristics, and Biological Functions of Polysaccharides from Sweet Tea (Lithocarpus litseifolius) Leaves; Journal Name: ANTIOXIDANTS, Volume 11, Issue 8; DOI: 10.3390 / antiox11081578.

[0053] 1Analysis of chemical components of modified acidic polysaccharides from the leaves of Alpinia zeylanicum

[0054] Table 1 summarizes the chemical components of natural Z. zingiberensis acidic polysaccharide (STP) and modified Z. zingiberensis acidic polysaccharides (STP-AD, STP-AH1, STP-AH2 and STP-E). The results showed that there was no significant difference in the total polysaccharide content between natural Z. zingiberensis acidic polysaccharide and modified Z. zingiberensis acidic polysaccharide. The content of uronic acid can affect the biological activity of polysaccharides to a certain extent. Generally, the higher the content of uronic acid, the better the biological activity of polysaccharides. The total uronic acid content of STP was 29.49 mg / 100 mg, the total uronic acid content of STP-AD decreased slightly to 26.71 mg / 100 mg, the total uronic acid content of STP-E decreased significantly to 23.70 mg / 100 mg, and the total uronic acid content of STP-AH1 and STP-AH2 were significantly increased to 49.57 mg / 100 mg and 71.74 mg / 100 mg, respectively. And the total protein content of STP-E, STP-AH1 and STP-AH2 increased significantly to 1.85mg / 100mg-3.16mg / 100mg. There was no significant difference between the total bound phenol content of STP-E and the total bound phenol content of STP. The total bound phenol of STP-AD, STP-AH1 and STP-AH2 decreased to 3.12mg GAE / 100mg, 2.69mg GAE / 100mg and 2.06mg GAE / 100mg (gallic acid equivalent), respectively. The degree of esterification will also affect the activity of polysaccharides. Generally, the lower the degree of esterification, the better the biological activity of the polysaccharide. As can be seen from Table 1, after mild alkaline deesterification, the esterification degree of STP (38.51%) decreased to 3.72% (STP-AD), and after low-concentration acid hydrolysis for different times, the esterification degree decreased to 14.54% (STP-AH1) and 7.74% (STP-AH2). After pectinase degradation, the esterification degree decreased to 34.98% (STP-E).

[0055] Table 1 Chemical components of modified acidic polysaccharides from Zingiber officinale

[0056]

[0057] Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E respectively represent natural acidic polysaccharide of Z. zingiberensis, modified acidic polysaccharide of Z. zingiberensis with esterification removed, modified acidic polysaccharide of Z. zingiberensis with arabinose side chain removed, modified acidic polysaccharide of Z. zingiberensis with arabinose side chain removed and galactose side chain reduced, and modified acidic polysaccharide of Z. zingiberensis with reduced galacturonic acid polysaccharide ratio. The results in the table are expressed as (mean ± standard deviation); superscript ae indicates significant difference (p<0.05); statistical significance was tested by ANOVA.

[0058] 2 Structural characteristics of modified acidic polysaccharides from Alpinia officinalis

[0059] (1) Molecular weight distribution

[0060] Molecular weight is often related to the activity of polysaccharides. Generally speaking, polysaccharides with lower molecular weight have better activity. Figure 1 AF. The results showed that STP, STP-AD, STP-AH1 and STP-AH2 all presented symmetrical elution curves. The molecular weights of STP and STP-AD were 4.472×10 4 Da and 4.268×10 4 Da, and the elution curves of STP and STP-AD were highly overlapped, and the retention times were almost the same. The retention times of STP-AH1, STP-AH2, and STP-E shifted later, and the molecular weights were significantly reduced to 1.317×10 4 Da, 0.562×10 4 Da and 1.458×10 4 In addition, the polydispersity (M w / M n ) is between 1.163 and 1.749, which is consistent with the trend of molecular weight change.

[0061] Table 2 Molecular weight (M) of modified acidic polysaccharides from Ligusticum chuanxiong w ), dispersion coefficient (M w / M n ), monosaccharide composition molar ratio

[0062]

[0063] Note: STP, STP-AD, STP-AH1, STP-AH2 and STP-E represent natural acidic polysaccharide from Pinellia ternata, modified acidic polysaccharide from Pinellia ternata with esterification removed, modified acidic polysaccharide from Pinellia ternata with arabinose side chains removed, modified acidic polysaccharide from Pinellia ternata with arabinose side chains removed and galactose side chains reduced, and modified acidic polysaccharide from Pinellia ternata with reduced galacturonic acid polysaccharide ratio, respectively; HG(%)=GalA(%)-Rha(%); RG-I(%)=2Rha(%)+Gal(%)+Ara(%); Side chain length=(Ara(%)+Gal(%)) / Rha(%); Superscript ae indicates significant difference (p<0.05); Statistical significance was tested by ANOVA.

[0064] (2) Monosaccharide composition analysis

[0065] In order to reveal the effects of different modification methods on the chemical structure of natural acidic polysaccharides from the leaves of Pinellia ternata, the monosaccharide composition, chemical groups and glycosidic bonds of modified acidic polysaccharides from the leaves of Pinellia ternata were systematically studied. Figure 2 As shown in AF, the monosaccharide compositions of STP, STP-AD, STP-AH1, STP-AH2 and STP-E are the same, including rhamnose, mannose, glucuronic acid, galacturonic acid, glucose, galactose, xylose and arabinose, indicating that the monosaccharide composition of the modified acidic polysaccharide of Pinellia ternata leaf is generally stable. As shown in Table 2, after alkaline deesterification, the molar percentages of the constituent monosaccharides in STP-AD and STP are very similar. Compared with STP, the galacturonic acid content of STP-E by enzymatic hydrolysis was significantly reduced to 21.26%, the arabinose content of STP-AH1 by acid hydrolysis was significantly reduced to 0.47%, and the galacturonic acid content was significantly increased to 55.89%. With the extension of acid hydrolysis time, the arabinose content and galactose content of STP-AH2 were further reduced to 0.14% and 17.26%, and the galacturonic acid content was further increased to 70.56%. The HG ratio, RG-I ratio and RG-I side chain length can be estimated by the content of galacturonic acid, rhamnose, arabinose and galactose. Generally speaking, the HG ratio, RG-I ratio and RG-I side chain length will affect the activity of polysaccharides. Generally, the higher the proportion of RG-I domain, the stronger the activity, and the lower the side chain length, the enhanced the activity. The HG ratio, RG-I ratio and RG-I side chain length of the modified acidic polysaccharide of Ligusticum chuanxiong are shown in Table 2. The HG ratio, RG-I ratio and RG-I side chain length of STP are 25.23%, 64.86% and 8.06, respectively. The HG ratio, RG-I ratio and RG-I side chain length of STP-AD are 21.94%, 69.22% and 7.49, respectively. The results show that mild alkali treatment has little effect on the HG ratio, RG-I ratio and RG-I side chain length of the acidic polysaccharide of Ligusticum chuanxiong, and is generally stable. After enzymatic hydrolysis, the HG content of STP-E was significantly reduced to 15.28%, and the RG-I side chain length increased to 10.31. Pectinase significantly destroyed the HG domain of natural Z. zingiberensis acidic polysaccharide. This shows that pectinase treatment successfully prepared Z. zingiberensis modified acidic polysaccharide with low HG content. After low-concentration acid hydrolysis, the HG content of Z. zingiberensis modified acidic polysaccharide significantly increased to 49.44% (STP-AH1) and 65.02% (STP-AH2), and the RG-I side chain length was significantly reduced to 0.86 (STP-AH1) and 0.44 (STP-AH2), which significantly destroyed the side chain of Z. zingiberensis acidic polysaccharide, indicating that acid hydrolysis successfully prepared Z. zingiberensis modified acidic polysaccharide with reduced side chain length.

[0066] (3) Infrared spectral characteristics

[0067] like Figure 3 As shown in the figure, compared with STP, the number of main characteristic absorption bands of modified acidic polysaccharides from Pinus armeniaca (STP-AD, STP-AH1, STP-AH2 and STP-E) did not change significantly, including 3397.9 cm-1 (OH vibration), 2923.8cm -1 (CH vibration), 1738.8cm -1 (COO-CH3 vibration), 1626.7cm -1 (COO-H vibration), 1437.7cm -1 (C-OH vibration), 1239.1cm -1 (COC vibration), 1073.9cm -1 (CO vibration) and 1023.9cm -1 (CC vibration). Compared with STP, STP-AD, STP-AH1, STP-AH2 and STP-E have a -1 The peak intensity around 1626.7 cm -1 The peak intensity around 1738.8cm -1 The peak area of ​​1738.8cm -1 and 1626.7cm -1 The average ratio of the sum of the peak areas was used to estimate the esterification degree of the acidic polysaccharides from the leaves of Pinus armeniaca. Figure 3 Medium STP-AD at 1738.8cm -1 The peak almost disappeared, and the esterification degree was reduced to 3.72%, indicating that the modified acidic polysaccharide of Pinus armeniaca with low esterification degree was successfully prepared by low-concentration alkaline deesterification. The detailed results are shown in Table 1.

[0068] (4) Nuclear magnetic resonance spectroscopy

[0069] The effects of different modification methods on the chemical groups and glycosidic bonds of natural Zingiber officinale polysaccharides were evaluated by one-dimensional nuclear magnetic resonance analysis. Figure 4 , 5 As shown in Figure 2, the 1D NMR spectra of STP and modified acidic polysaccharides from the leaves of Z. zingiberensis (STP-AD, STP-AH1, STP-AH2, and STP-E) are similar. Typical signals of HG and RG-I domains were found in STP ( Figure 4 , 5). For example, the signals corresponding to 1,4-α-D-GalAMep, 1,4-α-D-GalAp, 1,5-α-L-Araf, T-α-L-Araf, 1,3-α-L-Araf, 1,2,4-α-L-Rhap, and 1,2-α-L-Rhap residues were determined in the range of 4.96 to 5.30 ppm. The signals corresponding to 1,3,6-β-D-Galp and 1,3-β-D-Galp residues were detected in the range of 4.46 and 4.54 ppm. In addition, the C1 signals corresponding to 1,4-α-D-GalAp, 1,4-α-D-GalAMep, 1,3,6-β-D-Galp / 1,3-β-D-Galp, 1,5-α-L-Araf, and T-α-L-Araf residues in the range of 99.50 to 109.15 ppm were also determined. In addition, at 3.81ppm( 1 H) and 52.81ppm( 13 The typical signal at C) indicates the presence of methyl esterification on the carboxyl group of GalAp residue (GalA-OCH 3 ), at 2.07 to 2.18 ppm ( 1 H) range indicates the presence of acetylation of GalAp residues. The typical signal at 170.67ppm indicates the presence of α-1,4-D-GalAMep residues. However, compared with STP, the signal intensity of STP-AD at about 3.81ppm, 20.02ppm, 23.03ppm, 52.81ppm and 170.67ppm weakened or even disappeared, indicating that this study can successfully reduce or remove the methylation and acetylation degree of GalAp residues in natural Zingiber officinale acidic polysaccharides through mild alkali treatment. The signal intensity of STP-AH1 and STP-AH2 weakened or even disappeared at about 2.07ppm, 2.18ppm, 3.81ppm, 5.10ppm, 5.16ppm, 5.18ppm, 52.81ppm, 103.09ppm, 107.28ppm and 109.15ppm, and can be reduced or removed at 5.08ppm ( 1 H) was observed for 1,4-α-D-GalAMep residues of STP-AH1. This indicates that this study can successfully reduce or remove Araf and Galp residues of natural Zingiber officinale acidic polysaccharides through low-concentration acid hydrolysis, and can partially reduce the methylation and acetylation degrees of GalAp residues. The signal intensity of STP-E weakened or even disappeared at 5.04ppm and 52.81ppm, indicating that this study can reduce the GalAp residues of natural Zingiber officinale acidic polysaccharides through low-concentration pectin enzymatic hydrolysis.

[0070] The beneficial effects of the present invention are demonstrated by the following efficacy tests.

[0071] Experimental Example 1: Immunological structure-activity relationship and potential mechanism of action of modified acidic polysaccharide from Pinellia ternata

[0072] The present invention adopts an in vitro model system to evaluate the influence of natural Zingiber officinale acidic polysaccharides and modified Zingiber officinale acidic polysaccharides on immune enhancement and their potential mechanisms of action, and clarifies the key effector structures and potential mechanisms that affect the in vitro immunomodulatory effects of Zingiber officinale acidic polysaccharides. The results of the invention are helpful to promote its application in the food / functional food industry.

[0073] 1 Experimental Materials and Reagents

[0074] Table 3 Materials and reagents

[0075]

[0076] 2 Experimental methods

[0077] 2.1 Modification of acidic polysaccharides from Zingiber officinale

[0078] Extraction and modification of natural Zingiber officinale acidic polysaccharide prepared according to the method of Example 1

[0079] 2.2 In vitro immunological activity and structure-activity relationship analysis of modified acidic polysaccharides from the leaves of Pinellia ternata

[0080] 2.2.1 Cytotoxicity assay

[0081] 5×10 4 100 μL of RAW 264.7 macrophages with a cell concentration of 100 μL / well were placed in a 96-well microplate and cultured overnight in a 37°C incubator. The supernatant was aspirated, and 100 μL of modified acidic polysaccharides (STP, STP-AD, STP-AH1, STP-AH2 and STP-E) of different concentrations (50, 100, 200 μg / mL) of Artemisia selengensis leaves were added to the wells. The blank culture medium was used as the blank control, and LPS (1 μg / mL) was used as the positive control. The culture was cultured for 24 hours. The supernatant was aspirated, and 100 μL of MTT solution (1 mg / mL) was added to the wells and cultured for 4 hours. The supernatant was aspirated, and 100 μL of DMSO was added to the wells, and the OD was measured at 570 nm. The formula for calculating the cell proliferation rate is as follows:

[0082] Cell proliferation rate (%) = 1-(A1-A2) / A1×100% (2.1)

[0083] A1 is the absorbance value of the blank group, and A2 is the absorbance value of the sample.

[0084] 2.2.2 Determination of NO and cytokine (TNF-α, IL-6) content

[0085] 1×10 51 mL of RAW 264.7 macrophages with a cell concentration of 100 μg / well was placed in a 24-well microplate and cultured overnight in a 37°C incubator. The supernatant was aspirated, and 1 mL of STP, STP-AD, STP-AH1, STP-AH2 and STP-E at different concentrations (50, 100, 200 μg / mL) were added to the wells. The blank medium was used as the blank control, and LPS (1 μg / mL) was used as the positive control. The cells were cultured for 48 hours. The supernatant was aspirated, and the contents of NO and cytokines (TNF-α, IL-6) were determined according to the instructions of the ELISA kit.

[0086] 2.3 Potential cellular immune mechanism of modified acidic polysaccharides from Pinus armeniaca leaves

[0087] 1×10 5 1 mL of RAW 264.7 macrophages with a cell concentration of 1 μg / well was placed in a 24-well microplate and cultured overnight in a 37°C incubator. The supernatant was aspirated, and 1 mL of culture medium with or without TAK-242 (1 μM) and with or without C29 (30 μM) was added to the wells for 4 h. The supernatant was aspirated, and 1 mL of STP, STP-AD, STP-AH1, STP-AH2 and STP-E of different concentrations (50, 100, 200 μg / mL) were added to the wells, with the culture medium blank as the blank control and LPS (1 μg / mL) as the positive control, and cultured for 48 h. The supernatant was aspirated, and the contents of NO and cytokines (TNF-α, IL-6) were determined according to the instructions of the ELISA kit. In addition, the acidic polysaccharide (STP-AH1) modified by removing the arabinose side chain and reducing the galactose side chain of the leaf of Ziziphus chinensis was selected. 200 μg / mL of STP-AH1 was incubated with RAW 264.7 cells in a six-well plate for 48 hours, and blank medium and LPS (1 μg / mL) were used as blank control and positive control, respectively. The supernatant was removed and the well plate was washed twice with PBS. 1 mL of PBS was added to each well, and the cells were gently blown and transferred to a 1.5 mL centrifuge tube. The cells were collected after centrifugation. RIPA cell lysis buffer (containing 0.1% PMSF and protein phosphatase inhibitors) was added and lysed on ice for 30 minutes. Capillary protein immunoblot analysis was performed on the Protein Simple Wes system using a 12–230 kDa separation module (SM-W004) and an anti-rabbit detection module (DM-001) according to the manufacturer's instructions. The data were analyzed using Compass software, and the appropriate exposure time was set to ensure maximum signal.

[0088] 3 Experimental results

[0089] 3.1 In vitro immunological activity and structure-activity relationship of modified acidic polysaccharides from the leaves of Pinellia ternata

[0090] Figure 6A shows the toxic effects of STP, STP-AD, STP-AH1, STP-AH2 and STP-E on RAW264.7 cells. The results show that the acidic polysaccharide modified from the leaves of Pinus armeniaca has no obvious toxic effect on cell viability at the experimental concentration. Figure 6 As shown in BD, different concentrations (50-200 μg / mL) of STP, STP-AD, STP-AH1, STP-AH2 and STP-E can stimulate the release of NO, TNF-α and IL-6 by RAW264.7 macrophages, and the increase is concentration-dependent, indicating that the modified acidic polysaccharides from the leaves of Pinus armeniaca have significant immunostimulatory effects in vitro. Figure 6 BD showed that the immunostimulatory effect of STP was the weakest, and the immunostimulatory effects of different modified acidic polysaccharides from the leaves of the tree of the elm were all enhanced, indicating that modification can improve the in vitro immunostimulatory effect of natural acidic polysaccharides from the leaves of the elm ... Figure 6 BD showed that the immunostimulatory effect of STP-AH1 and STP-AH2 was significantly enhanced, significantly higher than other groups, and the immunostimulatory effect of STP-AH2 was higher than that of STP-AH1. Among them, STP-AH1 removed the arabinose side chain, and STP-AH2 further reduced the length of the galactose side chain on the basis of removing the arabinose side chain, which showed that acid hydrolysis could reduce the length of the side chain of RG-Ⅰ acidic polysaccharide of Ligusticum chuanxiong, so that more main chains were exposed, and the immunostimulatory effect was enhanced. The molecular weight and esterification degree of STP-AH1 and STP-AH2 were significantly reduced, and the uronic acid content was significantly increased, which may also lead to the further enhancement of the immunostimulatory effect of STP-AH1 and STP-AH2. The results showed that the esterification degree, HG and RG-Ⅰ ratio, RG-Ⅰ side chain length, uronic acid and molecular weight of the acid polysaccharide from the leaves of the ginger all affected the in vitro immune effect of the acid polysaccharide from the leaves of the ginger. Among them, the esterification degree, RG-Ⅰ side chain length and HG ratio of the acid polysaccharide from the leaves of the ginger were negatively correlated with its immunostimulatory effect, while the exposure degree of the main chain, uronic acid content and RG-Ⅰ ratio were positively correlated with its immunostimulatory effect. The side chain length of RG-Ⅰ, i.e. the exposure degree of the main chain and uronic acid content, had a more significant effect on the in vitro immune activity of the acid polysaccharide from the leaves of the ginger.

[0091] 3.2 Potential cellular immune mechanism of modified acidic polysaccharides from Pinellia ternata leaves

[0092] To further reveal the potential mechanism of natural S. zingiberensis acidic polysaccharide (STP) and modified S. zingiberensis acidic polysaccharide (STP-AD, STP-AH1, STP-AH2, and STP-E) mediated macrophage activation, we investigated the effects of C29 (a selective intracellular TLR2 signaling inhibitor) and TAK-242 (a selective intracellular TLR4 signaling inhibitor) on NO, TNF-α, and IL-6 production in RAW 264.7 cells. Figure 7 As shown in AC, STP, STP-AD, STP-AH1, STP-AH2, and STP-E without C29 or TAK-242 treatment could significantly promote the production of NO, TNF-α, and IL-6 in RAW 264.7 cells. However, the addition of C29 (30 μM) or TAK-242 (1 μM) to STP, STP-AD, STP-AH1, STP-AH2, and STP-E could significantly inhibit the secretion of NO, TNF-α, and IL-6 by RAW 264.7 cells. And the inhibition rate of TAK-242 addition was significantly higher than that of C29. These results confirmed that STP, STP-AD, STP-AH1, STP-AH2, and STP-E could activate RAW 264.7 cells to exert immunostimulatory effects by interacting with TLR2 or TLR4 receptors on the surface of macrophages, and that TLR4 receptors had a greater effect on the immune effects of modified acidic polysaccharides from Ligusticum chuanxiong. In addition, the present invention also determined that the acidic polysaccharide of Zingiber officinale activated RAW264.7 cells to increase the expression levels of NF-κBp65 and NF-κBp-p65. STP-AH1 was selected for determination. Figure 7 D shows that STP-AH1 can significantly upregulate the protein expression level of NF-κB p-p65, indicating that STP-AH1 can activate the TLR4 / NF-κB signaling pathway and thus exert an immunostimulatory effect.

Claims

1. A modified acidic polysaccharide from Ligusticum chuanxiong, characterized in that: It is a modified acidic polysaccharide prepared by chemical or enzymatic methods from natural acidic polysaccharide of Zingiber officinale. The total polysaccharide content of modified acidic polysaccharide from Artemisia selengensis leaf is 89.75mg±1.04mg–90.67mg±1.46mg per 100mg; total uronic acid: 23.70mg±1.58mg–71.74mg±2.07mg; total protein: 1.53mg±0.02mg–3.16mg±0.07mg; total bound phenol: 2.06mg GAE±0.06mg GAE–4.56mg GAE±0.04mg GAE (gallic acid equivalent); degree of esterification: 3.72%±0.90%–34.98%±0.62%; Molecular weight w :(0.562±0.012)×10 4 Da–(4.268±0.055)×10 4 Da; dispersion coefficient M w / M n : 1.163–1.729; The sugar components of the modified acidic polysaccharide from the leaves of Artemisia selengensis mainly include the following monosaccharides: galacturonic acid, galactose, arabinose, rhamnose, xylose, glucose, glucuronic acid, and mannose. The molar percentages of each monosaccharide are: (21.16%–70.56%): (17.26%–33.14%): (0.14%–27.48%): (5.53%–7.29%): ( 1.36%–6.85%):(1.71%–2.27%):(1.79%–2.89%):(0.83%–1.21%); among them, galacturonan (HG) accounted for 21.94%–65.02%, type I rhamnogalacturonan (RG-Ⅰ) accounted for 28.47%–72.37%, and the side chain length was 0.44–10.

31.

2. The modified acidic polysaccharide of Artemisia selengensis according to claim 1, characterized in that: The total polysaccharide content of modified acidic polysaccharide from Artemisia selengensis leaf is 89.75mg±1.04mg–90.09mg±1.07mg per 100mg; total uronic acid: 49.57mg±0.94mg–71.74mg±2.07mg; total protein: 3.03mg±0.07mg–3.16mg±0.07mg; total bound phenol: 2.06mg GAE±0.06mg GAE–2.69mg GAE±0.10mg GAE (gallic acid equivalent); degree of esterification: 7.74%±0.40%–14.54%±0.84%; Molecular weight w :(0.562±0.012)×10 4 Da–(1.317±0.018)×10 4 Da; dispersion coefficient M w / M n : 1.163–1.183; The sugar components of the modified acidic polysaccharide from the leaves of Artemisia selengensis mainly include the following monosaccharides: galacturonic acid, galactose, arabinose, rhamnose, xylose, glucose, glucuronic acid, and mannose. The molar percentages of each monosaccharide are: (55.89%–70.56%): (17.26%–28.99%): (0.14%–0.47%): (5.53%–6.46%): ( 1.36%–2.40%):(1.71%–1.78%):(2.22%–2.89%):(1.12%–1.21%); among them, galacturonan (HG) accounted for 49.44%–65.02%, type I rhamnogalacturonan (RG-Ⅰ) accounted for 28.47%–42.37%, and the side chain length was 0.44–0.

86.

3. The modified acidic polysaccharide of Artemisia selengensis according to claim 1 or 2, characterized in that: The natural Zingiber officinale acidic polysaccharide is prepared by a low eutectic solvent-assisted heating extraction method; The chemically modified acidic polysaccharide is an acidic polysaccharide prepared by modification with sodium hydroxide, hydrochloric acid and trifluoroacetic acid; the enzymatically modified acidic polysaccharide is an acidic polysaccharide prepared by modification with pectinase derived from Aspergillus aculeatus.

4. A method for preparing the modified acidic polysaccharide of Ligusticum chuanxiong according to any one of claims 1 to 3, characterized in that: It includes the following steps: a. Using a low eutectic solvent to assist in the extraction of natural acidic polysaccharides from the leaves of Zingiber officinale; b. The natural acidic polysaccharide of Zingiber officinale prepared in step a is modified by chemical or enzymatic methods.

5. The method for preparing modified acidic polysaccharide of Ligusticum chuanxiong according to claim 4, characterized in that: The method for modifying the acidic polysaccharide in step b is: 0.5% (w / v) of natural Ziziphus truncatula acidic polysaccharide was added to a sodium hydroxide solution with a pH value of 12.0, and stirred at 4°C for 30 min. Subsequently, hydrochloric acid (1 M) was added until the pH value was 7.0, and the solution was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0 kDa and freeze-dried to obtain a modified acidic polysaccharide of Ziziphus truncatula (STP-AD) with esterification removed. 0.5% (w / v) of natural Ziziphus truncatula acidic polysaccharide was added to 0.5M trifluoroacetic acid solution to make the final concentration of trifluoroacetic acid in the system 0.25M, and the mixture was reacted at 95°C for 1.5h and 4h, respectively. Then, sodium hydroxide (1M) was added until the pH value was 7.0, and the mixture was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0kDa and freeze-dried to obtain Ziziphus truncatula modified acidic polysaccharide (STP-AH1) with arabinose side chains removed and Ziziphus truncatula modified acidic polysaccharide (STP-AH2) with arabinose side chains removed and galactose side chains reduced, respectively; wherein, under the condition of reaction for 1.5h, the Ziziphus truncatula modified acidic polysaccharide STP-AH1 with arabinose side chains removed was prepared; under the condition of reaction for 4h, the Ziziphus truncatula modified acidic polysaccharide STP-AH2 with arabinose side chains removed and galactose side chains reduced was prepared; 0.5% (w / v) of natural Artemisia selengensis acidic polysaccharide was added to 0.3U / mL pectinase solution, reacted at 40°C for 6h, inactivated, centrifuged and the supernatant was taken; the supernatant was ultrafiltered through an ultrafiltration membrane with a molecular weight limit of 3.0kDa and freeze-dried to obtain Artemisia selengensis modified acidic polysaccharide (STP-E) with reduced galacturonic acid polysaccharide ratio.

6. Use of the modified acidic polysaccharide from Ligusticum chuanxiong according to any one of claims 1 to 3 in the preparation of health food that helps to enhance immunity.

7. Use of the modified acidic polysaccharide of Ligusticum chuanxiong according to any one of claims 1 to 3 in the preparation of a drug having an immunomodulatory effect.

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