A method for extracting soluble dietary fiber from polygonatum residue

Abstract

This invention discloses a method for extracting soluble dietary fiber from Polygonatum sibiricum residue, belonging to the field of traditional Chinese medicine extraction technology. This invention provides a method for extracting soluble dietary fiber from Polygonatum sibiricum residue with high yield, and discovers that this soluble dietary fiber can effectively resist oxidation, significantly improve the richness and uniformity of gut microbiota, improve gut microbiota composition imbalance, promote the proliferation of beneficial bacteria, inhibit the excessive growth of harmful bacteria, and play a protective role against intestinal damage by regulating the structure of intestinal microbiota and maintaining microecological stability, showing broad application prospects.

Description

Technical Field

[0001] This invention discloses a method for extracting soluble dietary fiber from Polygonatum residue, belonging to the field of traditional Chinese medicine extraction technology. Background Technology

[0002] Polygonatum is a herbaceous plant belonging to the genus Polygonatum in the family Liliaceae. Polygonatum sibiricum It possesses the functions of replenishing qi and blood, regulating the spleen and stomach, and moisturizing the lungs. It can not only effectively slow down aging, improve immunity, and lower blood sugar levels, but also fight against various diseases such as cancer. Therefore, it is widely used in the pharmaceutical and food fields. In recent years, Polygonatum polysaccharides have been widely used in the prevention and treatment of cardiovascular diseases, alleviating signs of aging, and reducing the probability of developing tumors. Polygonatum saponins are more abundant in Polygonatum and Polygonatum yunnanensis. Domestically and internationally, approximately 80 steroidal saponin compounds have been extracted and isolated from more than 10 medicinal Polygonatum species for clinical use. The pharmacological effects of Polygonatum saponins are mainly reflected in antibacterial, hypoglycemic, hypolipidemic, immunomodulatory, memory improvement, and kidney protection. [4] Currently, Polygonatum is mainly processed through steaming and boiling, and consumed directly as candied fruit, tea, etc., or extracted to obtain Polygonatum extract, which is used in the production of traditional Chinese medicine or as a medicinal decoction piece in clinical practice.

[0003] In the deep processing of Polygonatum sibiricum, methods such as water extraction and alcohol extraction are often used to extract active ingredients such as polysaccharides and saponins, generating a large amount of processing residue. This residue is still rich in dietary fiber, hemicellulose, lignin, and some underutilized structural polysaccharides, especially soluble dietary fiber. Direct disposal of this residue not only wastes resources but may also impose an environmental burden. Therefore, the high-value utilization of Polygonatum sibiricum residue has significant economic and ecological value. Soluble dietary fiber is a key component of plant cell walls, mainly including cellulose, hemicellulose, and lignin, and has physiological functions such as promoting intestinal peristalsis, increasing fecal volume, regulating intestinal microecological balance, and preventing constipation. In recent years, with the increasing demand for functional foods, the development and utilization of dietary fiber has gradually become a research hotspot. Compared with dietary fiber from traditional sources, dietary fiber derived from by-products of medicinal and edible plants has both functional and natural advantages, and has good market development potential. However, the soluble dietary fiber in Polygonatum sibiricum residue is often tightly bound to proteins, polysaccharides, and other cell wall components, resulting in a dense structure and low direct utilization rate.

[0004] Therefore, it is of great significance to develop a method for extracting soluble dietary fiber from Polygonatum residue in high yield. Summary of the Invention

[0005] This invention provides a method for extracting soluble dietary fiber from Polygonatum sibiricum residue, the method comprising the following steps: (1) Take the powder of Polygonatum sibiricum residue, add water and sonicate; (2) Add α-amylase complex enzyme for incubation, wherein the α-amylase complex enzyme is composed of α-amylase and saccharifying enzyme; (3) Add papain and incubate. After incubation, inactivate the papain. (4) Add cellulase complex enzyme for incubation, and inactivate cellulase complex enzyme after incubation; the cellulase complex enzyme is composed of cellulase and xylanase; (5) Collect the supernatant, concentrate it, precipitate it with alcohol, filter it, and collect the precipitate.

[0006] Furthermore, the Polygonatum sibiricum residue powder is prepared by the following method: take dry Polygonatum sibiricum powder, extract with water, filter, collect the solid, dry, and sieve to obtain Polygonatum sibiricum residue powder.

[0007] Furthermore, in the preparation process of the Polygonatum sibiricum residue powder, the material-to-liquid ratio for water extraction is 1:8-12, preferably 1:10, and the water extraction conditions are boiling water extraction for 2-6 hours, preferably 4 hours.

[0008] Further, in step (1), the mass ratio of the Polygonatum sibiricum residue powder to water is 1:(20-40); the ultrasonic treatment power is 180-220W, the temperature is 50-70℃, and the time is 10-20 min; the ultrasonic treatment also includes adjusting the pH of the system to 5.5-6.5; In step (2), the mass ratio of α-amylase to saccharifying enzyme in the α-amylase complex enzyme is (3-5):1; the mass concentration of the α-amylase complex enzyme is 1.0%-2.0%; the incubation time is 1-3 h; the temperature is 50-70℃; and the method is water bath incubation. After the incubation is completed, the reaction solution is cooled to room temperature and the pH is adjusted to 6.5-7.5. In step (3), the mass concentration of papain is 0.15%-0.25%, the incubation time is 1-3 hours, the temperature is 45-65℃, and the method is water bath incubation; In step (4), the mass ratio of cellulase to xylanase in the cellulase complex is 1:(0.5-2), and the mass concentration of the cellulase complex is 0.20%-0.25%; the incubation time is 1-3 h, the temperature is 50-70℃, and the method is ultrasonic incubation; In step (5), the reagent for alcohol precipitation is 90% aqueous ethanol solution-100% ethanol, the temperature is 2-6℃, and the time is 24-72 h.

[0009] Further, in step (1), the mass ratio of the Polygonatum sibiricum residue powder to water is 1:30, the ultrasonic treatment power is 200 W, the temperature is 60 ℃, and the time is 15 min; the ultrasonic treatment also includes adjusting the pH of the system to 6.0; In step (2), the mass ratio of α-amylase to saccharifying enzyme in the α-amylase complex enzyme is 4:1; the mass concentration of the α-amylase complex enzyme is 1.5%; the incubation time is 2 h; the temperature is 60℃; after the incubation, the reaction solution is cooled to room temperature and the pH is adjusted to 7. In step (3), the mass concentration of papain is 0.15%, the incubation time is 2 h, and the temperature is 55℃; the temperature for inactivating papain is 100℃. In step (4), the mass ratio of cellulase to xylanase in the cellulase complex is 1:1, the mass concentration of the cellulase complex is 0.20%, the incubation time is 2 h, and the temperature is 60℃; the inactivation temperature of the cellulase complex is 100℃, and the time is 10 min. In step (5), the reagent for alcohol precipitation is a 95% aqueous ethanol solution, the temperature is 4°C, and the time is 48 h.

[0010] Furthermore, the method further includes the following steps: (6) Dissolve the precipitate collected in step (5) in water, dialyze it in water, and freeze dry it after dialysis. Furthermore, the method further includes the following steps: (7) The product after freeze-drying in step (6) was purified sequentially by DEAE Sepharose Fast Flow column and Sephadex G-100 column to obtain purified soluble dietary fiber.

[0011] Furthermore, during DEAE Sepharose Fast Flow column purification, the packing material used was DEAE Sepharose Fast Flow packing material that had been soaked and rinsed with 0.5 mol / mL hydrochloric acid. Gradient elution was performed sequentially using three column volumes of water, 0.2 M NaCl aqueous solution, 0.5 M NaCl aqueous solution, and 1.0 M NaCl aqueous solution as eluents. The collected component was the effluent when water was used as the eluent. When purifying with a Sephadex G-100 column, a 0.1 M NaCl aqueous solution was used as the eluent.

[0012] The present invention also provides soluble dietary fiber prepared according to the above method.

[0013] The present invention also provides the use of the soluble dietary fiber prepared according to the above method in the preparation of antioxidants.

[0014] The present invention also provides the use of the soluble dietary fiber prepared according to the above method in the preparation of a medicament for improving intestinal damage.

[0015] Furthermore, the drug is a drug that enhances the richness and uniformity of intestinal flora, improves the imbalance of flora composition, promotes the proliferation of beneficial bacteria, and inhibits the excessive growth of harmful bacteria; the drug is a drug that regulates the structure of intestinal flora and maintains the stability of the microecology; the drug is a drug that maintains intestinal structure and absorption function.

[0016] This invention provides a method for high-yield extraction of soluble dietary fiber from Polygonatum residue, and finds that the soluble dietary fiber can effectively resist oxidation, significantly improve the richness and uniformity of the gut microbiota, improve the imbalance of gut microbiota composition, promote the proliferation of beneficial bacteria, inhibit the excessive growth of harmful bacteria, and play a protective role against intestinal damage by regulating the structure of the gut microbiota and maintaining the stability of the microecology. It has broad application prospects.

[0017] The present invention will be described in detail below through specific embodiments. It should be understood that these embodiments are only for explaining the present invention and are not intended to limit the scope of the present invention. As will be known to those skilled in the art, modifications can be made to the present invention without departing from the spirit of the invention, and such modifications also fall within the scope of the present invention. Attached Figure Description

[0018] Figure 1. Effect of feed-to-liquid ratio on SDF yield.

[0019] Figure 2 Effect of α-amylase concentration on SDF yield.

[0020] Figure 3 Effect of papain concentration on SDF yield.

[0021] Figure 4 Effect of cellulose complex enzyme concentration on SDF yield.

[0022] Figure 5 Response surface plot showing the interaction between various factors and SDF yield.

[0023] Figure 6 Contour plot of the interaction of various factors on SDF yield Figure 7 Isolation and purification of soluble dietary fiber SDF from Polygonatum sibiricum residue. Eluent chromatogram of SDF on DEAE Sepharose Fast Flow ion exchange column (left); purification of SDF-1 on Sephadex G-100 column (right).

[0024] Figure 8 High-performance molecular inhibition chromatogram.

[0025] Figure 9 Effects of SDF-1 on the morphology of jejunum and ileum (100×).

[0026] Figure 10 Alpha diversity of gut microbiota in mice with intestinal injury.

[0027] Figure 11 β-diversity of gut microbiota in mice with intestinal injury.

[0028] Figure 12 Microbial composition at the phylum level.

[0029] Figure 13 Differences in microorganisms at the phylum level.

[0030] Figure 14 Microbial composition at the genus level.

[0031] Figure 15 Differences among microorganisms at the genus level. Detailed Implementation

[0032] The raw materials and equipment used in the specific embodiments of this invention are all known products, obtained by purchasing commercially available products. Specifically, α-amylase, saccharifying enzyme, papain, cellulase, and xylanase were all purchased from Shanghai Maclean Biochemical Technology Co., Ltd.

[0033] The Polygonatum residue used in this specific embodiment of the invention is the residue after Polygonatum has undergone the following processing: Polygonatum macrocephala is dried in an oven. 100g of dried Polygonatum powder is weighed and placed in a round-bottom flask. Water is used as the extraction solvent (solid-to-liquid ratio 1:10 (m / v, g / mL)). The mixture is boiled in a heating mantle for 4 hours (timing starts after the water boils). Extraction is performed twice, and the mixture is filtered after the reaction is complete. The Polygonatum residue from which the Polygonatum polysaccharides have been extracted is dried for 24 hours, pulverized, sieved, and stored in a desiccator.

[0034] In the specific embodiments of the present invention, the room temperature refers to 25±5℃.

[0035] The enzyme concentration described in the specific embodiments of the present invention is a mass concentration.

[0036] Example 1. Preparation of soluble dietary fiber (SDF) from Polygonatum sibiricum residue Preparation method: Place the Polygonatum sibiricum residue in a 60℃ oven until completely dry. Remove the dried residue, pulverize it, and pass it through an 80-mesh sieve. Weigh 10 g of the Polygonatum sibiricum residue powder into a 1000 mL glass beaker, add an appropriate amount of deionized water according to the set material-to-liquid ratio, and stir thoroughly to ensure uniform mixing. Then, sonicate the mixture at 200 W power and 60℃ for 15 min, adjusting the pH to 6. Add the set amount of α-amylase complex enzyme at a mass ratio of 4:1 (α-amylase to saccharifying enzyme), and incubate in a 60℃ water bath for 2 h to fully hydrolyze the starch. Next, cool the reaction solution to room temperature, adjust the pH to 7, add the set amount of papain, and incubate in a 55℃ water bath for 2 h to hydrolyze the protein. Inactivate the enzyme at 100℃, cool, and add the set amount of cellulase complex enzyme at a mass ratio of 1:1 (cellulase to xylanase), adjust the temperature to 60℃, and sonicate at 200 W power for 2 h. The enzyme was then inactivated by heating at 100℃ for 10 min, and the supernatant was collected by vacuum filtration. The supernatant was concentrated, and four times its volume of 95% ethanol was added. The mixture was allowed to stand at 4℃ for 48 h to precipitate the ethanol. The precipitate was then collected by filtration, centrifugation, and drying to obtain the product. The product was dissolved in hot water, cooled, and placed in a 3400 kDa dialysis bag. Dialysis with deionized water was performed for 48 h. After dialysis, the product was freeze-dried at -80℃ and weighed to obtain the soluble dietary fiber of Polygonatum sibiricum.

[0037] SDF yield calculation formula: (mass of soluble dietary fiber from Polygonatum sibiricum / mass of Polygonatum sibiricum residue powder) × 100% 1. Single-factor experiment 1.1 Effect of material-to-liquid ratio on the extraction rate of soluble dietary fiber from Polygonatum sibiricum residue Following the steps of the "Preparation Method" described above, the concentrations of α-amylase complex enzyme (1.5%), papain (0.15%), and cellulase complex enzyme (0.15%) were set. The effect of different solid-liquid ratios (m / v, g / mL) of 1:10, 1:20, 1:30, 1:40, and 1:50 on the SDF yield was investigated. For details on the effect of solid-liquid ratio on SDF yield, please refer to [link to relevant documentation]. Figure 1 .

[0038] Depend on Figure 1 It can be seen that as the material-to-liquid ratio increases, the SDF yield first increases and then decreases, and the SDF yield reaches its maximum when the material-to-liquid ratio is 1:30.

[0039] 1.2 Effect of α-amylase complex enzyme on the yield of soluble dietary fiber in Polygonatum sibiricum residue Following the steps of the "Preparation Method" described above, the material-to-liquid ratio (m / v) was set to 1:30, the papain concentration to be 0.15%, and the cellulase complex concentration to be 0.15%. The effect of α-amylase complex concentrations at 0.5%, 1.0%, 1.5%, 2.0%, and 2.5% on SDF yield was investigated. For details on the effect of α-amylase complex concentration on SDF yield, please refer to [link to relevant documentation]. Figure 2 .

[0040] Depend on Figure 2 It can be seen that as the concentration of α-amylase complex enzyme increases, the SDF yield shows a trend of first increasing and then decreasing, and the SDF yield reaches its maximum when the concentration of α-amylase complex enzyme is 1.5%.

[0041] 1.3 Effect of papain concentration on the yield of soluble dietary fiber in Polygonatum sibiricum residue Following the steps of the "Preparation Method" described above, the material-to-liquid ratio (m / v) was set to 1:30, the concentration of α-amylase complex enzyme was 1.5%, and the concentration of cellulase complex enzyme was 0.15%. The effect of papain concentrations of 0.05%, 0.10%, 0.15%, 0.20%, and 0.25% on SDF yield was investigated. For details on the effect of papain concentration on SDF yield, please refer to [link to relevant documentation]. Figure 3 .

[0042] Depend on Figure 3 It can be seen that with the increase of papain concentration, the SDF yield first increases and then stabilizes. When the concentration is 0.15%, the SDF yield reaches 18.4%. When the papain concentration exceeds 0.15%, the SDF yield does not change much.

[0043] 1.4 Effect of cellulase complex enzyme concentration on the yield of soluble dietary fiber in Polygonatum sibiricum residue Following the steps of the "Preparation Method" described above, the material-to-liquid ratio was set to m / v = 1:30, the concentration of α-amylase complex enzyme was 1.5%, and the concentration of papain was 0.15%. The effect of cellulase complex enzyme concentration on SDF yield was investigated at 0.05%, 0.10%, 0.15%, 0.20%, and 0.25%. For details on the effect of cellulase complex enzyme concentration on SDF yield, please refer to [link to relevant documentation]. Figure 4 .

[0044] As the concentration of cellulase complex increases, the SDF yield shows a trend of first increasing and then stabilizing. When the concentration of cellulase complex is 0.20%, the SDF yield reaches its maximum value of 18.73%.

[0045] 2. Optimization of soluble dietary fiber (SDF) extraction from Polygonatum sibiricum residue using response surface methodology. Based on the single-factor experiments, the material-to-liquid ratio (A), the dosage of α-amylase complex enzyme (B), the dosage of papain (C), and the dosage of cellulase complex enzyme (D) were selected as independent variables, with the SDF extraction rate (X) as the response value. Based on the Box-Behnken design principle, a four-factor, three-level response surface methodology was used to further optimize the process. The factor levels are shown in Table 1.

[0046] Table 1. Factor Levels and Coding in Box-Behnken Central Composite Experimental Design Table 2 Results of Response Surface Analysis Table 3 Response Surface Experimental Variance Analysis Note: Represents highly significant ( P <0.01); Indicates significance ( P <0.05).

[0047] As shown in Tables 2 and 3, the model is highly significant overall (F=336.27). P <0.0001), indicating that the established quadratic regression model can well reflect the relationship between each factor and the SDF yield. (Misfit term) P =0.1456, not significant, indicating that the model fits well, the experimental error is small, the regression equation is reliable, and it can be used for process optimization prediction. From the results of the single-factor significance analysis: A (material-to-liquid ratio) is extremely significant (P < 0.01); D (cellulase dosage) is extremely significant (P < 0.01). P <0.01); C (papain dosage) significantly ( P =0.0009); B (α-amylase dosage) was not significant ( P =0.4497). This indicates that the order of influence on SDF yield is: A≈D>C>B. The quadratic terms A², B², C², and D² are all highly significant. P <0.0001), indicating that there is a clear quadratic surface relationship between each factor and the response value, and the response surface shows a clear parabolic trend.

[0048] Analysis of variance showed that the interaction between BD and CD was highly significant (P < 0.01); the interaction between AC was significant (P < 0.01). P<0.05); AB was nearly significant (P=0.055); AD and BC were not significant. This indicates that there is a strong synergistic effect between cellulase and papain (CD); there is also a significant interaction effect between cellulase and α-amylase (BD); there is a certain synergistic effect between the material-liquid ratio and papain (AC); the other interactions are relatively weak.

[0049] Depend on Figure 5 and Figure 6 It can be seen that all response surfaces exhibit a clear parabolic structure with a "high in the middle and low on both sides," indicating the existence of an optimal process range. The contour plots are mostly elliptical, indicating interactions between factors. The contour ellipses of the CD and BD combination are relatively narrow, indicating a more significant interaction effect. When the feed-to-liquid ratio increases from 1:20 to 1:30, the SDF yield increases significantly, but decreases slightly when further increased to 1:40, indicating that a suitable feed-to-liquid ratio is beneficial for enzymatic reactions and mass transfer. The SDF yield reaches a high level when papain and cellulase are added at moderate amounts; excessively high or low amounts will lead to a decrease in yield.

[0050] The repeated experiments at the center point show that under the conditions of a material-to-liquid ratio of 1:30, an α-amylase complex enzyme concentration of 1.5%, a papain concentration of 0.15%, and a cellulase complex enzyme concentration of 0.20%, the SDF yield reached the highest value of 19.20%, and the repeatability was good (18.99%–19.20%), indicating that this condition is the optimal range.

[0051] The results indicate that the material-to-liquid ratio and the amount of cellulase complex enzyme have the most significant impact on SDF yield, followed by the amount of papain, while the amount of α-amylase complex enzyme has a relatively small impact. The interaction between CD and BD is extremely significant, indicating that multi-enzyme synergy has a significant enhancing effect, and a reasonable ratio of enzymes helps improve extraction efficiency. All quadratic terms are extremely significant, indicating that the influence of each factor on SDF yield is not a simple linear relationship, but rather that there exists an optimal range within a certain degree. Response surface methodology and contour line analysis further validate the reliability of the model and the synergistic effect among the factors, demonstrating that the established regression model can well reflect the actual extraction process.

[0052] Based on single-factor experiments, this invention optimized the SDF extraction process from Polygonatum sibiricum residue using the Box-Behnken response surface methodology. The results showed that the model was highly significant, with no significant lack-of-fit terms, indicating a good model fit and reliable prediction. The order of influence on SDF yield was: material-to-liquid ratio ≈ cellulase complex enzyme dosage > papain dosage > α-amylase complex enzyme dosage. Significant secondary effects were observed in all factors, and significant interactions existed between some factors, indicating that multi-enzyme synergy plays a crucial role in improving SDF yield. The optimized process conditions were: following the steps of the above-described "Preparation Method," controlling the material-to-liquid ratio at 1:30, the α-amylase complex enzyme concentration at 1.5%, the papain concentration at 0.15%, and the cellulase complex enzyme concentration at 0.20%. Under these conditions, the SDF yield reached 19.20%, with good repeatability. These results provide a theoretical basis and technical reference for the high-value utilization of Polygonatum sibiricum processing by-products.

[0053] Example 2: Purification and structural characterization of soluble dietary fiber from Polygonatum sibiricum residue The soluble dietary fiber SDF used in this embodiment is the Polygonatum sibiricum soluble dietary fiber prepared according to the optimal process conditions in Example 1.

[0054] 1. Purification of SDF from Polygonatum odoratum residue First, the DEAE Sepharose Fast Flow ion exchange column was soaked in 0.5 mol / mL hydrochloric acid for 1 h, and repeatedly rinsed to remove impurities. Then, the column was eluted with 4-5 times its volume of distilled water until neutral. The peristaltic pump flow rate was adjusted to 5 mL / min, and the column was equilibrated with distilled water for 2 h. 1 g of SDF was accurately weighed, dissolved in water, centrifuged, and the supernatant was added to the DEAE Sepharose Fast Flow ion exchange column (700 mm × 16 mm). The peristaltic pump flow rate was then adjusted to 15 mL / min, and the sample was eluted with three column volumes of distilled water, 0.2 M NaCl, 0.5 M NaCl, and 1.0 M NaCl solutions. The sample was analyzed using the phenol-sulfuric acid method at 490 nm, and each component was collected. The effluents eluted by water, 0.2 M NaCl, and 0.5 M NaCl were collected, concentrated, and dialyzed (MWCO 3500 Da), and then freeze-dried, and named A, B, and C respectively.

[0055] Fraction A, with higher yield and total sugar content, was further purified by gel column chromatography. The specific steps were as follows: 100 mg of fraction A was weighed, dissolved in 3 mL of distilled water, centrifuged, and the supernatant was loaded onto the sample. The conditions were as follows: the separation column was a Sepharose G-100 gel chromatography column (1.5 cm x 80 cm); 0.1 M NaCl solution was used as the mobile phase, and the flow rate was set to 2 mL / min; the sample was passed through a 0.22 pm membrane, and the loading volume was 3 mL; the collected sample was dialyzed, concentrated, and lyophilized to obtain soluble dietary fiber SDF-1 from Polygonatum sibiricum residue purified by Sephadex G-100 column separation.

[0056] The results of separating crude SDF using a DEAE Sepharose Fast Flow ion exchange column are as follows: Figure 7 As shown, the peaks from left to right correspond to the collections: A, B, and C. The yields of each component were 18.42%, 7.63%, and 5.79%, respectively. Component A was purified using a Sephadex G-100 column to obtain purified component SDF-1, with a yield of 15.60%. SDF-1 was collected for lyophilization and subsequent experiments.

[0057] The total sugar content of SDF-1 (2 mg / mL) was determined to be 90.23% ± 2.4% using the phenol-sulfuric acid method. No detectable protein or uronic acid was found. These results indicate that SDF-1 is mainly composed of carbohydrates.

[0058] 2. Monosaccharide composition analysis in SDF-1 Fucose and fructose in SDF-1 sample were analyzed and determined using high-performance liquid chromatography coupled with evaporative light scattering detector (HPLC-ELSD). Specific chromatographic conditions were as follows: an Agilent ZORBAX Original 70Å Carbohydrate Analysis column (4.6 mm × 250 mm, 5 μm); column temperature set at 30 ℃; mobile phase acetonitrile-water (85:15, v / v); flow rate 1.0 mL·min⁻¹. Detector parameters were: ELSD drift tube temperature 75 ℃, gas flow rate 2.5 L·min⁻¹, and gain 1. Precisely pipette 2 μL (or 5 μL) and 10 μL of reference solution were administered, and the sample volume was 10–20 μL. Quantification was performed using the external standard two-point method and calculated using a logarithmic equation. The average value was taken from three tests.

[0059] Table 4. Monosaccharide composition of SDF-1 in Polygonatum sibiricum residue (n=3) As shown in Table 4, the monosaccharide composition of SDF-1 in Polygonatum sibiricum residue includes rhamnose, fucose, arabinose, fructose, mannose, glucose and galactose.

[0060] 3. Molecular weight determination of SDF-1 The molecular weight distribution of SDF-1 was analyzed by high-performance gel permeation chromatography (HPLC). The chromatographic column was a TSKG3000PWXL gel column (300 mm × 7.8 mm, 10 μm); the detector was a Shodex RI-101 differential refractive index detector; the detection wavelength was 210 nm; the mobile phase was pure water; the flow rate was 0.5 mL / min; the column temperature was 0℃; and the injection volume was 20 μL. Accurately weigh 25 mg of each SDF-1 sample and place it in a 5 mL volumetric flask. Dissolve the sample in NaCl by heating, cool, and then dilute to the mark. After mixing, the sample is loaded onto the flask.

[0061] The molecular weight distribution of SDF-1 in Polygonatum sibiricum residue is as follows: Figure 8 As shown. A standard curve was plotted based on the molecular weight and relative retention time of the standard. The equation for the lgMw-RT calibration curve is: y = -0.3016x + 15.135 (R0). 2 =0.9978). Based on the retention time, the relative molecular mass of SDF-1 in Polygonatum sibiricum residue was calculated to be 41.53 kDa.

[0062] Example 3: Antioxidant Capacity Determination DPPH free radical scavenging rate determination: Prepare a 10 mg / mL SDF-1 sample solution, centrifuge, and take 1 mL of the supernatant. Add 3 mL of DPPH solution (0.04 mg / mL) prepared with anhydrous ethanol to each solution. React at room temperature in the dark for 30 min. Use anhydrous ethanol as a blank. Measure the absorbance A1 at 517 nm. Measure the absorbance A0 using distilled water instead of the sample solution. Measure the absorbance A2 by reacting 1 mL of sample solution with 3 mL of anhydrous ethanol solution. The scavenging rate is calculated according to the following formula. Ascorbic acid solution (10 mg / mL) is used as a positive control.

[0063] DPPH clearance rate = (A0 - A1 + A2) / A0 × 100% ABTS +Free radical scavenging rate determination: 2.5 mL of ABTS solution (7 mM) and 2.5 mL of potassium persulfate solution (7.35 mM) were precisely mixed, thoroughly shaken, and reacted at room temperature in the dark for 12 h. After the reaction, the resulting solution was diluted 50 times with anhydrous ethanol. The absorbance at 734 nm reached 0.70 ± 0.02, yielding the ABTS⁺ working solution. 0.2 mL of SDF-1 sample solutions of different concentrations were placed in dry colorimetric tubes, and 1 mL of ABTS⁺ working solution was added. The mixture was mixed and allowed to stand in the dark for 6 min. Anhydrous ethanol was used as a control, and the absorbance of the sample was measured as A. When anhydrous ethanol was used instead of the sample, the absorbance was measured as A0. + Self-radical scavenging rate is calculated using the following formula: ABTS+ clearance rate = (A0 - A) / A0 × 100% Table 5 DPPH free radical scavenging rate Table 5 shows that SDF-1 exhibited a certain scavenging ability against DPPH free radicals at different concentrations, with a significant increasing trend with increasing concentration. When the concentration increased from 0.6% to 4%, the scavenging rate of SDF-1 increased from 60.31% to 91.34%, indicating that its antioxidant activity has a significant dose-dependent effect. Compared with the positive control VC, the scavenging rate of VC was significantly higher than that of SDF-1 at 0.6% and 1% concentrations. P <0.05 indicates that SDF-1 has relatively weak antioxidant capacity under low concentration conditions. However, when the concentration increases to 4%, the scavenging rate of SDF-1 reaches 91.34%, which is not significantly different from that of VC. P =0.89), indicating that at higher concentrations, SDF-1 has shown antioxidant capacity close to that of vitamin C.

[0064] Table 6 ABTS+ free radical scavenging rate Table 6 shows that SDF-1 also exhibits strong scavenging ability against ABTS⁺ free radicals, with a significant increasing trend with increasing concentration. At concentrations of 0.6%, 1%, and 4%, the scavenging rates of SDF-1 were 63.15%, 78.665%, and 93.12%, respectively. This indicates that SDF-1 possesses good in vitro antioxidant activity. At concentrations of 0.6% and 1%, the scavenging ability of SDF-1 was significantly lower than that of VC ( ). P <0.05); when the concentration increased to 4%, the SDF-1 clearance rate was 93.12%, which was not significantly different from that of VC (<0.05); P =0.098), indicating that its antioxidant capacity is close to that of vitamin C at higher concentrations.

[0065] Example 4: Pharmacodynamic study of intestinal injury in mice 1. Experimental Methods Sixty mice were randomly divided into six groups of ten each after 7 days of acclimatization: normal control group (C group), model control group (CTX group), PC group, low-dose SDF-1 group (LP group), medium-dose SDF-1 group (MP group), and high-dose SDF-1 group (HP group). After the acclimatization period, from day 1 to day 21, the three dosage groups were administered SDF-1 at doses of 125, 250, and 500 mg / kg bw via gavage daily in the morning, respectively. The control and model groups were administered distilled water. From day 19 onwards, except for the control group, mice in the model and SDF-1 intervention groups were intraperitoneally injected with 80 mg / kg bw of CTX, while mice in the normal control group were injected with an equal volume of physiological saline. These injections were administered once daily for three days. Mice had free access to food and water during this period. The experimental design is shown in Table 7.

[0066] Table 7 Experimental Design Twenty-four hours after the last administration, all animals were fasted for 12 hours. Blood was collected from the eyeballs of mice, and the mice were then euthanized by cervical dislocation. Fresh blood samples were collected in additive-free blood collection tubes, allowed to stand at room temperature (26°C) for 30 min, centrifuged at 3000 g for 10 min at 4°C, and the serum was separated and stored at -80°C. The thymus and spleen of the mice were harvested and weighed; 3-5 cm sections of jejunum and ileum tissue were each cut and fixed in a general-purpose tissue fixative. The contents of the cecum were collected in a laminar flow hood. The remaining small intestine tissue and cecum contents were placed in sterile cryogenic tubes, flash-frozen in liquid nitrogen, and then stored at -80°C.

[0067] 2. Experimental Results 2.1 Effects of SDF-1 on body weight Mice were weighed using an electronic scale before gavage each day for 21 days. On day 22, mice were fasted for 12 hours but allowed free access to water before being weighed.

[0068] Table 8 Effects of SDF-1 on body weight As shown in Table 8, there was no significant difference in the initial body weight among the groups of mice. P =0.89). At the end of the trial, there was a significant difference in weight change ( P=0.04). The mice in group C had the highest final weight, while the mice in the cyclophosphamide model group (CTX group) had a significant decrease in weight, indicating that CTX treatment significantly inhibited the weight gain of mice. The final weight of mice in group PC and different doses of SDF-1 treatment groups (low dose L group, medium dose M group, and high dose H group) was higher than that of group C, but lower than that of group C, indicating that SDF-1 alleviated the inhibitory effect of CTX on weight gain to some extent.

[0069] 2.2 Effects of SDF-1 on the morphology of jejunum and ileum tissues After dissection, jejunal and ileal tissue samples were collected, fixed in 4% paraformaldehyde solution for 48 h, dehydrated with a gradient of alcohols, embedded in paraffin, sectioned, dewaxed, mounted, and stained with hematoxylin-eosin (HE). The sections were stained in hematoxylin solution for 4 min, then separated by acid and ammonia solution for 30 s, rinsed with running water for 60 min, dehydrated in 80% and 95% alcohol for 5 min, and stained with eosin solution for 4 min. The stained sections were then dehydrated three times with anhydrous alcohol for 5 min each time, and then immersed in xylene three times for 5 min each time. After the sections became clear, they were sealed with neutral resin.

[0070] Depend on Figure 9 It was found that the jejunal and ileal villi of the control group mice were neat, long, and orderly arranged with small gaps, which was a normal state. In the CTX group mice, the jejunal and ileal villi were loosely arranged, and many villi showed breakage, swelling, and shortening, indicating successful model establishment. Compared with the CTX group, the villi morphology of the medium- and high-dose SDF-1 groups was significantly improved, with longer and more tightly arranged villi, reduced villi breakage and shedding, and smaller villi gaps. This indicates that SDF-1 can improve the damage to the small intestinal villi structure caused by CTX and protect the intestinal barrier.

[0071] 2.3 Effects of SDF-1 on serum biochemical parameters Mouse whole blood was centrifuged (3000 g, 15 min), and the supernatant was collected to obtain serum samples. Jejunal tissue was thawed in an ice bath, ground into powder with liquid nitrogen, and collected in EP tubes. A certain mass of jejunal tissue powder was weighed, and 15 volumes of pre-chilled RIPA (strong) lysis buffer were added. The mixture was then lysed on ice for 30 min. The homogenate was obtained by centrifugation (4℃, 14000 r / min, 15 min), and the supernatant was collected. The protein concentration of the intestinal tissue homogenate was measured using a BCA kit and the mixture was leveled. Serum DAO, D-LA, LPS levels and mouse intestinal tissue homogenate SigA, β-DF, and LZM levels were measured using an ELISA kit. The kits were operated strictly according to the manufacturer's instructions.

[0072] Table 9 Effects of SDF-1 on serum biochemical parameters As shown in Table 9, there were significant differences in serum indicators among the groups. P <0.04). The levels of LPS, D-LA, and DAO in the CTX group were significantly higher than those in the C group, indicating that CTX treatment damaged the intestinal barrier function. The levels of LPS, D-LA, and DAO in the PC group and each dose of SDF-1 treated groups were significantly lower than those in the CTX group and close to those in the C group, indicating that SDF-1 can effectively alleviate CTX-induced intestinal barrier damage at low (L group), medium (M group), and high (H group) doses. Among them, the medium-dose group (M group) had the lowest LPS and D-LA levels, while DAO levels were also significantly improved, indicating that SDF-1 has a certain dose-dependent protective effect.

[0073] 2.4 Effects of SDF-1 on villus length and crypt depth in the jejunum and ileum After dissection, jejunal and ileal tissues were fixed for 24 h, dehydrated, and routinely processed, then embedded in paraffin and stained with hematoxylin and eosin (HE). Under an optical microscope, the five longest and most intact villi in the field of view were selected, and the villi length (from the villi tip to the villi base) and crypt depth (from the villi base to the base of the intestinal gland) were measured.

[0074] Table 10 Effects of SDF-1 on jejunal villus length and crypt depth Table 10 shows that different treatments had significant effects on the length of villi, crypt depth, and villi-to-crypt ratio in the mouse jejunum. P <0.01). In the CTX group, villus length was significantly shortened, crypt depth increased, and the villus-crypt ratio decreased to 2.69, indicating that CTX treatment damaged intestinal morphology. Compared with the CTX group, villus length was significantly increased and crypt depth decreased in the PC group and all doses of SDF-1 treated groups, with the villus-crypt ratio returning to near the control group level. The medium-dose SDF-1 group (M group) had the highest villus length and the lowest crypt depth, indicating that SDF-1 has a protective effect on intestinal structure. Low-dose (L group) and high-dose (H group) treatments also improved intestinal morphology, but the differences were not significant, indicating that SDF-1 has a stable protective effect on intestinal morphology within a certain range.

[0075] Table 11 Effects of SDF-1 on ileal villus length and crypt depth As shown in Figure 11, different treatments have a significant impact on ileal villus length and villus-to-hymen ratio. P= 0.02), while having no significant effect on crypt depth. CTX group mice showed significantly shortened villus length and a villus-crypt ratio decreased to 1.53, indicating that CTX treatment severely damaged the morphological structure of the ileum. Compared with the CTX group, mice in the PC group and each dose of SDF-1 treatment group showed significantly increased villus length and significantly improved villus-crypt ratio, indicating that the intestinal structure was restored. Specifically, the villus length in the medium-dose (M group) and high-dose (H group) SDF-1 groups were 360.34 µm and 370.35 µm, respectively, and the villus-crypt ratios were 2.10 and 2.13, respectively, both close to the control group levels, indicating that SDF-1 has a significant protective effect on the ileal villus structure.

[0076] 2.5 Protective effect of SDF-1 on the intestinal immune barrier 2.5.1 SDF-1 on immune organ indices The spleen index and thymus index are calculated using the following formulas: Spleen index (mg / g) = Spleen mass (mg) / Mouse body weight (g) Thymus index (mg / g) = thymus mass (mg) / mouse body weight (g) Table 12 Effects of SDF-1 on immune organ indices As shown in Table 12, there were significant differences in the thymus index and spleen index among the groups of mice. P＜ 0.05). The thymus index and spleen index of mice in the CTX group were significantly lower than those in the C group, indicating that CTX treatment led to atrophy of immune organs. The thymus index and spleen index of mice in the PC group and each dose of SDF-1 treatment group were higher than those in the CTX group, indicating that the damage to immune organs was alleviated to some extent. Among them, the thymus index of the medium-dose group (M group) and the high-dose group (H group) of SDF-1 were 1.26 mg / g and 1.35 mg / g, respectively, and the spleen index were 2.65 mg / g and 2.78 mg / g, respectively, which were close to the control group level. This indicates that SDF-1 can effectively improve CTX-induced atrophy of immune organs in mice and has a certain dose-dependent protective effect. 2.5.2 Effects of SDF-1 on the number of goblet cells and intraepithelial lymphocytes in the small intestine After dissection, jejunal and ileal tissues were fixed for 24 h, dehydrated, and routinely processed, then embedded in paraffin and stained with hematoxylin and eosin (HE). Under an optical microscope, the five longest and most intact villi in the field of view were selected, and the number of goblet cells and intraepithelial lymphocytes between columnar cells of each villi were counted.

[0077] Table 13 Effects of SDF-1 on the number of goblet cells and intraepithelial lymphocytes in jejunal epithelium As shown in Table 13, there were significant differences in the number of goblet cells and intraepithelial lymphocytes in the jejunal epithelium among the groups. P＜ (0.05). The number of goblet cells and intraepithelial lymphocytes in the CTX group were significantly lower than those in the C group, indicating that CTX treatment significantly impaired intestinal immune defense function. The number of goblet cells and lymphocytes in the PC group and each dose of SDF-1 treatment group was higher than that in the CTX group, indicating that intestinal immune cells were somewhat restored. Among them, the number of goblet cells and intraepithelial lymphocytes in the medium dose group (M group) and high dose group (H group) of SDF-1 were close to the control group, indicating that SDF-1 has a significant protective effect in maintaining the number of intestinal immune cells.

[0078] Table 14 Effects of SDF-1 on the number of goblet cells and intraepithelial lymphocytes in the ileum. As shown in Table 14, different treatments had a significant effect on the number of goblet cells and intraepithelial lymphocytes in the ileum. P <0.05). The number of goblet cells and intraepithelial lymphocytes in the ileum of mice in the CTX group were significantly lower than those in the C group, indicating that CTX treatment disrupted the ileal immune barrier function. Compared with the CTX group, the number of goblet cells and lymphocytes in mice in the PC group and each dose of SDF-1 treatment group was significantly increased. The number of goblet cells and intraepithelial lymphocytes in the medium-dose group (M group) and high-dose group (H group) of SDF-1 were close to the control group level, indicating that SDF-1 can effectively improve the reduction of ileal immune cells induced by CTX.

[0079] 2.5.3 Effects of SDF-1 on intestinal sIgA, β-DF and LZM Thaw jejunal tissue in an ice bath to prepare a 10% jejunal homogenate. Centrifuge at 3000 g for 5 min and collect the supernatant for analysis. Use an ELISA kit to measure the levels of sIgA, β-DF, and LZM in the intestinal tissue homogenate. Strictly follow the manufacturer's instructions for use.

[0080] Table 15 Effects of SDF-1 on gut sIgA, β-DF, and LZM Table 15 shows that there were significant differences in the levels of sIgA, β-DF, and LZM in the intestines of mice in each group. P<0.05). The levels of sIgA, β-DF, and LZM in the CTX group were significantly lower than those in the C group, indicating that CTX treatment significantly weakened the intestinal mucosal immune function in mice. The levels of intestinal immune factors in the PC group and each dose of SDF-1 treatment group were higher than those in the CTX group. In the high-dose SDF-1 group (H group), the levels of sIgA, β-DF, and LZM were close to those in the control group, indicating that SDF-1 can effectively restore the intestinal mucosal immune function damaged by CTX. Low-dose (L group) and medium-dose (M group) SDF-1 also showed significant improvement, but slightly lower than the high-dose group, indicating that SDF-1 has a dose-dependent protective effect on intestinal immune factors within a certain range.

[0081] 2.6 Effects of SDF-1 on the composition of volatile fatty acids in the intestine The content of SCFAs in cecal contents was determined by ether extraction. First, a standard curve for short-chain fatty acids was plotted. Six short-chain fatty acid standards, including acetic acid, were prepared into mixed standard sample solutions with concentrations of 6.25, 12.5, 25, 50, and 100 μg / mL using analytical grade ether. Ethylbutyric acid (1 mg / mL) internal standard solution was added, and the samples were analyzed using gas chromatography (Agilent 7890A). Accurately weighed mouse cecal contents samples were added to ultrapure water (0.1 g: 1 mL) and homogenized. Short-chain fatty acids and their salts were extracted into the water. The mixture was acidified with 1 / 10 volume of 1 mol / L hydrochloric acid, and then vortexed with an equal volume of ether to extract the short-chain fatty acids into the ether layer. The mixture was centrifuged at 4 ℃, 13000 g, and 15 min. The supernatant was collected, and ethylbutyric acid (1 mg / mL) internal standard was added before analysis. GC detection conditions: A Thermo Fisher Scientific TP-FFAP capillary column (30 m × 0.25 mm × 0.25 μm) was used, with high-purity nitrogen as the carrier gas and a flame ionization detector (FID). The injection volume was 1.0 μL, the initial column temperature was set to 80℃, and the temperature was increased to 165℃ at a rate of 20℃ / min and held for 3 minutes. The column flow rate was kept constant at 1 mL / min, the injection port and FID detector temperatures were both 220℃, and the split ratio was set to 20:1.

[0082] Table 16 Effects of SDF-1 on the composition of volatile fatty acids in the intestine (µg / mL) Table 16 shows that there were significant differences in the levels of acetic acid, propionic acid, butyric acid, and total VFA in the intestines of mice in each group. P <0.01), while the differences among isobutyric acid, isovaleric acid, and valeric acid were not significant ( P>0.05). In the CTX group, the levels of acetic acid, propionic acid, butyric acid (83.65 µg / mL), and total VFA were significantly lower than in the C group, indicating that CTX treatment significantly inhibited the production of short-chain fatty acids in the intestine. Compared with the CTX group, the VFA levels in the PC group and each dose of SDF-1 treatment group were increased. In the high-dose SDF-1 group (H group), the levels of acetic acid, propionic acid, butyric acid, and total VFA reached 170.54 µg / mL, 45.57 µg / mL, 178.65 µg / mL, and 403.2 µg / mL, respectively, close to the control group levels, indicating that SDF-1 can partially restore the intestinal VFA production inhibited by CTX. The low-dose (L group) and medium-dose (M group) SDF-1 groups also showed significant improvement, with a certain dose-dependent trend, but the degree of difference among the VFA components varied slightly.

[0083] 2.7 Effects of SDF-1 on gut microbiota Cecal contents samples were randomly selected from five mice each from groups C, CTX, and H for sequencing. First, total genomic DNA was extracted from the samples using the CTAB / SDS method. DNA purity was assessed by agarose gel electrophoresis and quantified using a UV spectrophotometer. Then, using the extracted genomic DNA as a template, the V3-V4 hypervariable region of bacterial 16S rDNA was amplified using universal primers 338F (5'-ACTCCTACGGGAGGCAGCA-3') and 806R (5'-GGACTACHVGGGTWTCTAAT-3'). The PCR amplification products from each sample were mixed, separated by 2% agarose gel electrophoresis, and purified using the AxyPrep DNA gel extraction kit. Finally, gene sequencing was performed on the Illumina platform (Illumina, San Diego, CA, USA) using the Illumina PE library construction method.

[0084] The results showed that group C had 486 unique OTUs, group H had 418, and group CTX had 92. This indicates that high-dose SDF-1 intervention can maintain the stability of the gut microbiota in mice and increase the diversity of gut microbiota.

[0085] Depend on Figure 10 It can be seen that there were no significant changes in alpha diversity among different groups of mice.

[0086] Depend on Figure 11 It can be seen that there are significant differences in the microbial composition among the three groups, indicating that SDF-1 can counteract the effects of CTX on the gut microbiota structure to a certain extent and restore the normal composition and structure of the gut microbiota.

[0087] Figure 12The community composition at the phylum level is shown for groups C, CTX, and H. Firmicutes was the dominant phylum in all groups, followed by Desulfobacterota, both constituting the main components of the community. In addition to these two phyla, Verrucomicrobiota, Actinobacteria, Bacteroidota, Patescibacteria, and Proteobacteria were also detected, but their relative abundance was low, representing a small proportion.

[0088] Depend on Figure 13 It can be seen that CTX treatment has a limited impact on the composition of gut microbiota at the phylum level, while SDF-1 intervention significantly increased the relative abundance of Firmicutes, Desulfobacterota and Actinobacteria, indicating that SDF-1 intervention can effectively reshape the gut microbiota structure after CTX perturbation.

[0089] Depend on Figure 14 It can be seen that the three groups of samples have basically the same compositional categories at the genus level. Among all samples, Desulfovibrio , Akkermansia , Lactobacillus , HT002 , Enterorhabdus as well as Lachnospiraceae_NK4A136_group This genus constitutes a relatively high proportion in different individuals and is the main component of fungi. In addition, it was also detected... Muribaculaceae_unclassified, Clostridium, Ruminococcus, Helicobacter, Anaerotignum, Eubacterium_siraeum_group, Lachnospiraceae_UCG-006, Bacteroides It belongs to the same genus, but its relative abundance varies among different samples.

[0090] Depend on Figure 15 It can be seen that CTX treatment significantly increased the abundance of g_Desulfovibrio, while significantly decreasing the abundance of g_Lachnospiraceae_NK4A136_group and g_Lactobacillus; and the microbial composition of group H was closer to that of group C, indicating that SDF-1 intervention alleviated the disturbance of gut microbiota by CTX to some extent.

[0091] The above results indicate that SDF-1 has a significant protective effect against CTX-induced intestinal damage in mice. CTX treatment significantly inhibited mouse weight gain, shortened jejunal and ileal villus length, increased crypt depth, and led to a decrease in the villus-crypt ratio, indicating impaired intestinal absorption. Simultaneously, the spleen and thymus indices of CTX-treated mice decreased, the number of goblet cells and intraepithelial lymphocytes in the jejunum and ileum decreased, and the levels of intestinal sIgA, β-defensin, and lysozyme were reduced, indicating disruption of the intestinal immune barrier. Furthermore, the levels of short-chain fatty acids in the cecum were significantly decreased, including acetic acid, propionic acid, butyric acid, and total VFA, indicating impaired intestinal microbial metabolic activity. These results collectively demonstrate that CTX treatment can lead to a decline in intestinal health in mice by damaging intestinal structure, weakening immune function, and inhibiting microbial metabolism. In addition, 16S rRNA results revealed that intestinal damage caused significant disruption of the intestinal community structure in mice, with reduced abundance of beneficial bacteria and relative enrichment of potentially harmful bacteria. SDF-1 intervention can effectively reverse the above abnormalities, significantly improve the richness and evenness of the gut microbiota, improve the imbalance of microbiota composition, promote the proliferation of beneficial bacteria, and inhibit the overgrowth of harmful bacteria. Furthermore, it plays a protective role against intestinal damage by regulating the structure of the gut microbiota and maintaining microecological stability.

[0092] SDF-1 intervention significantly improved various types of damage induced by CTX. Weight loss was alleviated, intestinal villus length increased, and the villous-to-crypto ratio was restored, indicating that SDF-1 can maintain intestinal structure and absorptive function. Regarding immune indicators, SDF-1 increased spleen and thymus indices, restored the number of immune cells in the jejunum and ileum, and significantly increased sIgA, β-defensin, and lysozyme levels, enhancing the intestinal mucosal immune barrier. Intestinal VFA levels were also significantly increased; in the high-dose SDF-1 group, acetic acid, propionic acid, butyrate, and total VFA levels were close to normal, suggesting that it can promote short-chain fatty acid production by providing dietary fiber substrates or regulating the gut microbiota. Therefore, SDF-1 has a dose-dependent protective effect against intestinal damage, possessing multiple effects including improving intestinal morphology, enhancing immune function, and regulating microbial metabolism.

Claims

1. A method for extracting soluble dietary fiber from Polygonatum sibiricum residue, characterized in that, The method includes the following steps: (1) Take the powder of Polygonatum sibiricum residue, add water and sonicate; (2) Add α-amylase complex enzyme for incubation, wherein the α-amylase complex enzyme is composed of α-amylase and saccharifying enzyme; (3) Add papain and incubate. After incubation, inactivate the papain. (4) Add cellulase complex enzyme for incubation, and inactivate cellulase complex enzyme after incubation; the cellulase complex enzyme is composed of cellulase and xylanase; (5) Collect the supernatant, concentrate it, precipitate it with alcohol, filter it, and collect the precipitate.

2. The method according to claim 1, characterized in that, In step (1), the mass ratio of the Polygonatum sibiricum residue powder to water is 1:(20-40); the ultrasonic treatment power is 180-220W, the temperature is 50-70℃, and the time is 10-20 min; the ultrasonic treatment also includes adjusting the pH of the system to 5.5-6.5; In step (2), the mass ratio of α-amylase to saccharifying enzyme in the α-amylase complex enzyme is (3-5):1; the mass concentration of the α-amylase complex enzyme is 1.0%-2.0%; the incubation time is 1-3 h; the temperature is 50-70℃; and the method is water bath incubation. After the incubation is completed, the reaction solution is cooled to room temperature and the pH is adjusted to 6.5-7.

5. In step (3), the mass concentration of papain is 0.15%-0.25%, the incubation time is 1-3 h, the temperature is 45-65℃, and the method is water bath incubation; In step (4), the mass ratio of cellulase to xylanase in the cellulase complex is 1:(0.5-2), and the mass concentration of the cellulase complex is 0.20%-0.25%; the incubation time is 1-3 h, the temperature is 50-70℃, and the method is ultrasonic incubation; In step (5), the reagent for alcohol precipitation is 90% aqueous ethanol solution-100% ethanol, the temperature is 2-6℃, and the time is 24-72h.

3. The method according to claim 2, characterized in that, In step (1), the mass ratio of the Polygonatum sibiricum residue powder to water is 1:30, the ultrasonic treatment power is 200 W, the temperature is 60 ℃, and the time is 15 min; the ultrasonic treatment also includes adjusting the pH of the system to 6.0; In step (2), the mass ratio of α-amylase to saccharifying enzyme in the α-amylase complex enzyme is 4:1; the mass concentration of the α-amylase complex enzyme is 1.5%; the incubation time is 2 h; the temperature is 60℃; after the incubation, the reaction solution is cooled to room temperature and the pH is adjusted to 7. In step (3), the mass concentration of papain is 0.15%, the incubation time is 2 h, and the temperature is 55℃; In step (4), the mass ratio of cellulase to xylanase in the cellulase complex is 1:1, the mass concentration of the cellulase complex is 0.20%, the incubation time is 2 h, and the temperature is 60℃. In step (5), the reagent for alcohol precipitation is a 95% aqueous ethanol solution, the temperature is 4°C, and the time is 48 h.

4. The method according to any one of claims 1-3, characterized in that, The method further includes the following steps: (6) Dissolve the precipitate collected in step (5) in water, dialyze it in water, and freeze dry it after the dialyzing is completed.

5. The method according to claim 4, characterized in that, The method further includes the following steps: (7) The product after freeze-drying in step (6) was purified sequentially by DEAE Sepharose Fast Flow column and Sephadex G-100 column to obtain purified soluble dietary fiber.

6. The method according to claim 5, characterized in that, For DEAE Sepharose Fast Flow column purification, the packing material used was DEAE Sepharose Fast Flow packing material that had been soaked and rinsed with 0.5 mol / mL hydrochloric acid. Gradient elution was performed sequentially using three column volumes of water, 0.2 M NaCl aqueous solution, 0.5 M NaCl aqueous solution and 1.0 M NaCl aqueous solution as eluents. The collected component was the effluent when water was used as the eluent. When purifying with a Sephadex G-100 column, a 0.1 M NaCl aqueous solution was used as the eluent.

7. Soluble dietary fiber prepared according to any one of claims 1-6.

8. Use of the soluble dietary fiber prepared according to any one of claims 1-6 in the preparation of antioxidants.

9. Use of the soluble dietary fiber prepared according to any one of claims 1-6 in the preparation of a medicament for improving intestinal damage.

10. The use according to claim 9, characterized in that, The drug is used to enhance the richness and uniformity of intestinal flora, improve the imbalance of flora composition, promote the proliferation of beneficial bacteria, and inhibit the excessive growth of harmful bacteria; the drug is used to regulate the structure of intestinal flora and maintain the stability of the microecology; the drug is used to maintain intestinal structure and absorption function.

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